1
|
Dall’Olio E, De Rensis F, Martignani E, Miretti S, Ala U, Cavalli V, Cipolat-Gotet C, Andrani M, Baratta M, Saleri R. Differential Expression of miR-223-3p and miR-26-5p According to Different Stages of Mastitis in Dairy Cows. Biomolecules 2025; 15:235. [PMID: 40001538 PMCID: PMC11853211 DOI: 10.3390/biom15020235] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/11/2025] [Revised: 02/03/2025] [Accepted: 02/04/2025] [Indexed: 02/27/2025] Open
Abstract
Mastitis is the leading cause of economic losses in dairy farming, significantly impairing animal welfare and the quality and quantity of milk production. MicroRNAs are increasingly gaining attention, in both human and veterinary medicine, as biomarkers for various diseases. This study evaluated the diagnostic potential of four circulating microRNAs (miR-26-5p, miR-142-5p, miR-146a, and miR-223-3p) by examining changes in their expression in milk samples from dairy cows at different immune-cell subpopulations correlated to different stage of mastitis with a validated method. Additionally, this study has analyzed the possible source of these circulating microRNAs by the measurement of their secretion from activated immune cells (lymphocytes, monocytes, and neutrophils). miR-223-3p has been significantly expressed in an acute stage of mastitis (p < 0.01) but not in the chronic or susceptible stages. Conversely, mir-26-5p has been significantly reduced in acute, chronic, and susceptible groups of animals. In immune-cell cultures, miR-26 has been shown to be down-regulated in lipopolysaccharide (LPS)-stimulated neutrophils, while miR-223 has been shown to be up-regulated in phytohemagglutinin (PHA)-stimulated lymphocytes. The differential expression of miR-223-3p and miR-26-5p, combined with differential and total somatic cell count, could serve as a useful tool for identifying the evolutionary stage of mastitis-related inflammatory pathology.
Collapse
Affiliation(s)
- Eleonora Dall’Olio
- Department of Veterinary Science, University of Parma, Strada del Taglio 10, 43126 Parma, Italy; (E.D.); (F.D.R.); (V.C.); (C.C.-G.); (M.A.); (R.S.)
| | - Fabio De Rensis
- Department of Veterinary Science, University of Parma, Strada del Taglio 10, 43126 Parma, Italy; (E.D.); (F.D.R.); (V.C.); (C.C.-G.); (M.A.); (R.S.)
| | - Eugenio Martignani
- Department of Veterinary Sciences, University of Torino, Largo Paolo Braccini 2, 10095 Grugliasco, Italy; (E.M.); (S.M.); (U.A.)
| | - Silvia Miretti
- Department of Veterinary Sciences, University of Torino, Largo Paolo Braccini 2, 10095 Grugliasco, Italy; (E.M.); (S.M.); (U.A.)
| | - Ugo Ala
- Department of Veterinary Sciences, University of Torino, Largo Paolo Braccini 2, 10095 Grugliasco, Italy; (E.M.); (S.M.); (U.A.)
| | - Valeria Cavalli
- Department of Veterinary Science, University of Parma, Strada del Taglio 10, 43126 Parma, Italy; (E.D.); (F.D.R.); (V.C.); (C.C.-G.); (M.A.); (R.S.)
| | - Claudio Cipolat-Gotet
- Department of Veterinary Science, University of Parma, Strada del Taglio 10, 43126 Parma, Italy; (E.D.); (F.D.R.); (V.C.); (C.C.-G.); (M.A.); (R.S.)
| | - Melania Andrani
- Department of Veterinary Science, University of Parma, Strada del Taglio 10, 43126 Parma, Italy; (E.D.); (F.D.R.); (V.C.); (C.C.-G.); (M.A.); (R.S.)
| | - Mario Baratta
- Department of Chemistry, Life Sciences and Environmental Sustainability, University of Parma, Parco Area delle Scienze 11a, 43124 Parma, Italy
| | - Roberta Saleri
- Department of Veterinary Science, University of Parma, Strada del Taglio 10, 43126 Parma, Italy; (E.D.); (F.D.R.); (V.C.); (C.C.-G.); (M.A.); (R.S.)
| |
Collapse
|
2
|
Puente-Rivera J, De la Rosa Pérez DA, Olvera SIN, Figueroa-Angulo EE, Saucedo JGC, Hernández-León O, Alvarez-Sánchez ME. The Circulating miR-107 as a Potential Biomarker Up-Regulated in Castration-Resistant Prostate Cancer. Noncoding RNA 2024; 10:47. [PMID: 39311384 PMCID: PMC11417898 DOI: 10.3390/ncrna10050047] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/11/2024] [Revised: 08/19/2024] [Accepted: 08/22/2024] [Indexed: 09/26/2024] Open
Abstract
Prostate cancer (PCa) is a prevalent malignancy in men globally. Current diagnostic methods like PSA testing have limitations, leading to overdiagnosis and unnecessary treatment. Castration-resistant prostate cancer (CRPC) emerges in some patients receiving androgen deprivation therapy (ADT). This study explores the potential of circulating microRNA-107 (miR-107) in liquid biopsies as a prognosis tool to differentiate CRPC from non-castration-resistant PCa (NCRPC). We designed a case-control study to evaluate circulating miR-107 in serum as a potential prognosis biomarker. We analyzed miR-107 expression in liquid biopsies and found significantly higher levels (p < 0.005) in CRPC patients, compared to NCRPC. Notably, miR-107 expression was statistically higher in the advanced stage (clinical stage IV), compared to stages I-III. Furthermore, CRPC patients exhibited significantly higher miR-107 levels (p < 0.05), compared to NCRPC. These findings suggest that miR-107 holds promise as a non-invasive diagnostic biomarker for identifying potential CRPC patients.
Collapse
Affiliation(s)
- Jonathan Puente-Rivera
- Posgrado en Ciencias Genómicas, Universidad Autónoma de la Ciudad de México (UACM), San Lorenzo 290, Col. Del Valle, México City 03100, Mexico; (J.P.-R.); (D.A.D.l.R.P.); (E.E.F.-A.)
- División de Investigación, Hospital Juárez de México, Instituto Politécnico Nacional 5160, Col. Magdalena de las Salinas, México City 07360, Mexico
| | - David Alejandro De la Rosa Pérez
- Posgrado en Ciencias Genómicas, Universidad Autónoma de la Ciudad de México (UACM), San Lorenzo 290, Col. Del Valle, México City 03100, Mexico; (J.P.-R.); (D.A.D.l.R.P.); (E.E.F.-A.)
| | - Stephanie I. Nuñez Olvera
- Departamento de Biología Celular y Fisiología, Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, México City 04510, Mexico;
| | - Elisa Elvira Figueroa-Angulo
- Posgrado en Ciencias Genómicas, Universidad Autónoma de la Ciudad de México (UACM), San Lorenzo 290, Col. Del Valle, México City 03100, Mexico; (J.P.-R.); (D.A.D.l.R.P.); (E.E.F.-A.)
| | - José Gadú Campos Saucedo
- Hospital Central Militar, Dirección General de Sanidad SEDENA, Blvd. Manuel Ávila Camacho S/N, Lomas de Sotelo, México City 11200, Mexico;
| | - Omar Hernández-León
- Servicio de Urología, Hospital Juárez de México, Instituto Politécnico Nacional 5160, Col. Magdalena de las Salinas, México City 07360, Mexico;
| | - María Elizbeth Alvarez-Sánchez
- Posgrado en Ciencias Genómicas, Universidad Autónoma de la Ciudad de México (UACM), San Lorenzo 290, Col. Del Valle, México City 03100, Mexico; (J.P.-R.); (D.A.D.l.R.P.); (E.E.F.-A.)
| |
Collapse
|
3
|
Pedersen OB, Hvas AM, Pasalic L, Kristensen SD, Grove EL, Nissen PH. Platelet Function and Maturity and Related microRNA Expression in Whole Blood in Patients with ST-Segment Elevation Myocardial Infarction. Thromb Haemost 2024; 124:192-202. [PMID: 37846463 DOI: 10.1055/s-0043-1776305] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/18/2023]
Abstract
BACKGROUND Reduced effect of antiplatelet therapy has been reported in patients with ST-segment elevation myocardial infarction (STEMI). MicroRNAs (miRs) may influence platelet function and maturity, and subsequently the effect of antiplatelet therapy. OBJECTIVES We aimed to explore the association between miR expression and platelet function and maturity in patients with acute STEMI and healthy individuals. METHODS We performed an observational study of STEMI patients admitted directly to primary percutaneous coronary intervention. Patients were treated with antiplatelet therapy according to guidelines. Within 24 hours after admission, blood samples were obtained to measure: the expression of 10 candidate miRs, platelet function markers using advanced flow cytometry, platelet aggregation, serum thromboxane B2, and platelet maturity markers. Furthermore, blood samples from healthy individuals were obtained to determine the normal variation. RESULTS In total, 61 STEMI patients and 50 healthy individuals were included. STEMI patients had higher expression of miR-21-5p, miR-26b-5p, and miR-223-3p and lower expression of miR-150-5p, miR423-5p, and miR-1180-3p than healthy individuals. In STEMI patients, the expression of miR-26b-5p showed the most consistent association with platelet function (all p-values <0.05, Spearman's rho ranging from 0.27 to 0.41), while the expression of miR-150-5p and miR-223-3p showed negative associations with platelet function. No association between miR expression and platelet maturity markers was observed. CONCLUSION In patients with STEMI, the expression of six miRs was significantly different from healthy individuals. The expression of miR-26b-5p may affect platelet function in acute STEMI patients and potentially influence the effect of antiplatelet therapy.
Collapse
Affiliation(s)
- Oliver Buchhave Pedersen
- Thrombosis and Haemostasis Research Unit, Department of Clinical Biochemistry, Aarhus University Hospital, Aarhus, Denmark
- Department of Cardiology, Aarhus University Hospital, Aarhus, Denmark
- Department of Clinical Medicine, Faculty of Health, Aarhus University, Aarhus, Denmark
| | | | - Leonardo Pasalic
- Institute of Clinical Pathology and Medical Research, Westmead Hospital, NSW Health Pathology, Sydney, Australia
- Westmead Clinical School, Faculty of Medicine and Health, University of Sydney, Sydney, Australia
| | - Steen Dalby Kristensen
- Department of Cardiology, Aarhus University Hospital, Aarhus, Denmark
- Department of Clinical Medicine, Faculty of Health, Aarhus University, Aarhus, Denmark
| | - Erik Lerkevang Grove
- Department of Cardiology, Aarhus University Hospital, Aarhus, Denmark
- Department of Clinical Medicine, Faculty of Health, Aarhus University, Aarhus, Denmark
| | - Peter H Nissen
- Thrombosis and Haemostasis Research Unit, Department of Clinical Biochemistry, Aarhus University Hospital, Aarhus, Denmark
- Department of Clinical Medicine, Faculty of Health, Aarhus University, Aarhus, Denmark
| |
Collapse
|
4
|
Khabou B, Fakhfakh R, Tahri S, Bahloul E, Hadj Kacem H, Belmabrouk S, Hachicha H, Sellami K, Turki H, Masmoudi H, Abida O. miRNA implication in the pathogenesis and the outcome of Tunisian endemic pemphigus foliaceous. Exp Dermatol 2023. [PMID: 37114366 DOI: 10.1111/exd.14821] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/30/2022] [Revised: 04/07/2023] [Accepted: 04/13/2023] [Indexed: 04/29/2023]
Abstract
Pemphigus foliaceous (PF) is a bullous autoimmune skin disease diagnosed through sera and skin analyses. PF severity is associated with maintained anti-Dsg1 sera levels and its prognosis is unpredictable. MicroRNA (miRNA), dynamic regulators of immune function, have been identified as potential biomarkers for some autoimmune diseases. This study aimed to assess the miRNA expression of miR-17-5p, miR-21-5p, miR-146a-5p, miR-155-5p and miR-338-3p using quantitative real-time PCR in peripheral blood mononuclear cells (PBMC) and lesional skin samples from untreated and treated PF patients (both remittent and chronic) over 3 months. Overall, miRNA expression was significantly higher in PBMC than in biopsy samples. Blood miR-21 expression was increased in untreated patients compared to controls and had a diagnostic value with an AUC of 0.78. After 6 weeks, it decreased significantly, similar to anti-Dsg1 antibodies and the PDAI score. In addition, a positive correlation was observed between cutaneous miR-21 expression and the disease activity score. Conversely, cutaneous expressions of miR-17, miR-146a and miR-155 were significantly higher in treated chronic patients compared to remittent ones. The cutaneous level of miR-155 positively correlated with pemphigus activity, making it a potential predictive marker for patients' clinical stratification with an AUC of 0.86.These findings suggest that blood miR-21 and cutaneous miR-155 can be used as supplemental markers for PF diagnosis and activity, respectively in addition to classical parameters.
Collapse
Affiliation(s)
- Boudour Khabou
- Autoimmunity, Cancer and Immunogenetics Research Laboratory (LR18SP12), Immunology Department, Habib Bourguiba Hospital, University of Sfax, Sfax, Tunisia
| | - Raouia Fakhfakh
- Autoimmunity, Cancer and Immunogenetics Research Laboratory (LR18SP12), Immunology Department, Habib Bourguiba Hospital, University of Sfax, Sfax, Tunisia
| | - Safa Tahri
- Autoimmunity, Cancer and Immunogenetics Research Laboratory (LR18SP12), Immunology Department, Habib Bourguiba Hospital, University of Sfax, Sfax, Tunisia
| | - Emna Bahloul
- Department of Dermatology, Hedi Chaker Hospital, University of Sfax, Sfax, Tunisia
| | - Hassen Hadj Kacem
- Department of Applied Biology College of Sciences, University of Sharjah, Sharjah, United Arab Emirates
- Research Institute of Science and Engineering, University of Sharjah, Sharjah, United Arab Emirates
| | - Sabrine Belmabrouk
- Research Institute of Science and Engineering, University of Sharjah, Sharjah, United Arab Emirates
| | - Hend Hachicha
- Autoimmunity, Cancer and Immunogenetics Research Laboratory (LR18SP12), Immunology Department, Habib Bourguiba Hospital, University of Sfax, Sfax, Tunisia
| | - Khadija Sellami
- Department of Dermatology, Hedi Chaker Hospital, University of Sfax, Sfax, Tunisia
| | - Hamida Turki
- Department of Dermatology, Hedi Chaker Hospital, University of Sfax, Sfax, Tunisia
| | - Hatem Masmoudi
- Autoimmunity, Cancer and Immunogenetics Research Laboratory (LR18SP12), Immunology Department, Habib Bourguiba Hospital, University of Sfax, Sfax, Tunisia
| | - Olfa Abida
- Autoimmunity, Cancer and Immunogenetics Research Laboratory (LR18SP12), Immunology Department, Habib Bourguiba Hospital, University of Sfax, Sfax, Tunisia
| |
Collapse
|
5
|
Speranza G, Mele GR, Favia P, Pederzolli C, Potrich C. Tuning Surface Properties via Plasma Treatments for the Improved Capture of MicroRNA Biomarkers. MATERIALS 2022; 15:ma15072641. [PMID: 35407971 PMCID: PMC9000635 DOI: 10.3390/ma15072641] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 02/26/2022] [Revised: 03/27/2022] [Accepted: 03/30/2022] [Indexed: 01/27/2023]
Abstract
Advanced materials could bring about fundamental improvements in the evolution of innovative analytical devices, i.e., biosensors or lab-on-a-chip devices, in particular in the context of liquid biopsies. Here, plasma deposition processes were tested for the introduction of primary amines on silicon surfaces by tuning the amounts and availability of amino-charged residues. Different binary (CH4/NH3) and ternary (CH4/NH3/H2 and CH4/NH3/N2) mixtures of gases were used as feeds for the plasma treatments. The obtained surfaces were fully characterized for their chemical and physical properties before their use as capture materials in a functional test. Synthetic and fluorescently conjugated microRNA-21 (miR-21) was selected as the target molecule. The capture of miR-21 increased linearly with the increase in amino nitrogen measured on surfaces. The surface showing the most promising performance was further analyzed in different conditions, i.e., varying pH and time of incubation, incubation with different microRNAs, and possible elution of captured microRNAs. The apparent pH range of primary amines present on the surfaces was around 3.5–4. Positively charged surfaces prepared via PE-CVD were, therefore, demonstrated as being suitable materials for the capture of microRNA biomarkers, paving the way for their inclusion in biomedical devices for the purification and analysis of circulating biomarkers.
Collapse
Affiliation(s)
- Giorgio Speranza
- Center for Sensors and Devices, Fondazione Bruno Kessler, Via Sommarive 18, 38123 Trento, Italy; (G.S.); (G.R.M.); (C.P.)
- Department of Industrial Engineering, University of Trento, v. Sommarive 9, 38123 Trento, Italy
- CNR-Istituto di Fotonica e Nanotecnologie, Via alla Cascata 56/C, 38123 Trento, Italy
| | - Gaetano Roberto Mele
- Center for Sensors and Devices, Fondazione Bruno Kessler, Via Sommarive 18, 38123 Trento, Italy; (G.S.); (G.R.M.); (C.P.)
- Department of Chemistry, CNR Inst. NANOTEC, University of Bari Aldo Moro, 70124 Bari, Italy;
| | - Pietro Favia
- Department of Chemistry, CNR Inst. NANOTEC, University of Bari Aldo Moro, 70124 Bari, Italy;
| | - Cecilia Pederzolli
- Center for Sensors and Devices, Fondazione Bruno Kessler, Via Sommarive 18, 38123 Trento, Italy; (G.S.); (G.R.M.); (C.P.)
| | - Cristina Potrich
- Center for Sensors and Devices, Fondazione Bruno Kessler, Via Sommarive 18, 38123 Trento, Italy; (G.S.); (G.R.M.); (C.P.)
- CNR-Istituto di Biofisica, Via alla Cascata 56/C, 38123 Trento, Italy
- Correspondence:
| |
Collapse
|
6
|
Pedersen OB, Hvas AM, Grove EL, Larsen SB, Pasalic L, Kristensen SD, Nissen PH. Association of whole blood microRNA expression with platelet function and turnover in patients with coronary artery disease. Thromb Res 2022; 211:98-105. [DOI: 10.1016/j.thromres.2022.01.026] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/08/2021] [Revised: 01/24/2022] [Accepted: 01/26/2022] [Indexed: 11/16/2022]
|
7
|
MiR-145-5p Inhibits the Invasion of Prostate Cancer and Induces Apoptosis by Inhibiting WIP1. JOURNAL OF ONCOLOGY 2021; 2021:4412705. [PMID: 34899906 PMCID: PMC8660234 DOI: 10.1155/2021/4412705] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 09/21/2021] [Revised: 10/21/2021] [Accepted: 10/25/2021] [Indexed: 11/26/2022]
Abstract
Prostate cancer (PCa) is a common malignant tumor of the male genitourinary system that seriously affects the quality of life of patients. Studying the pathogenesis and therapeutic targets of PCa is important. In this study, we investigated the role of miR-145-5p in PCa and its potential molecular mechanisms. The expression levels of miR-145-5p in PCa tissues and adjacent control tissues were detected by real-time quantitative polymerase chain reaction. The effects of miR-145-5p overexpression on PCa were studied using cell proliferation, migration, and invasion experiments. Furthermore, WIP1 was the target gene of miR-145-5p through the bioinformatics website and dual-luciferase reporter gene experiment. Further studies found that WIP1 downregulation could inhibit the proliferation, invasion, and cloning of PCa cells. Overexpression of WIP1 reversed the anticancer effects of miR-145. The anticancer effect of miR-145 was achieved by inhibiting the PI3K/AKT signaling pathway and upregulating ChK2 and p-p38MAPK. Taken together, these results confirmed that miR-145-5p inhibited the growth and metastasis of PCa cells by inhibiting the expression of proto-oncogene WIP1, thereby playing a role in tumor suppression in PCa and may become a potential therapeutic target for the treatment of PCa.
Collapse
|
8
|
Maryam Khorasani, Shahbazi S, Abolhasani M, Shahrokh H, Mahdian R. Expression Profile of MiR-200 Family Members and Their Targets in Prostate Cancer. CYTOL GENET+ 2021. [DOI: 10.3103/s009545272104006x] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
|
9
|
Du W, Li D, Xie J, Tang P. miR‑367‑3p downregulates Rab23 expression and inhibits Hedgehog signaling resulting in the inhibition of the proliferation, migration, and invasion of prostate cancer cells. Oncol Rep 2021; 46:192. [PMID: 34278506 PMCID: PMC8299014 DOI: 10.3892/or.2021.8143] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/16/2020] [Accepted: 04/02/2021] [Indexed: 12/13/2022] Open
Abstract
MicroRNAs play an important role in tumor cell proliferation, invasion, and Rab23 is a member of the Ras-related small GTPase family and plays a critical role in the progression of may types of tumors. The present study was designed to investigate the inhibitory effect of microRNA (miR)-367-3p on the proliferation, invasion, and metastasis of prostate cancer cells. qRT-PCR was used to detect the expression of miR-367-3p in prostate cancer and adjacent tissues. Cell proliferation, scratch, and Transwell assays were performed to verify the inhibitory effect of miR-367-3p overexpression or Ras-related protein Rab 23 (Rab23) knockdown on prostate cancer. Double luciferase reporter assay was utilized to verify whether miR-367-3p could target the Rab23 3′-untranslated region (UTR). The expression levels of Rab23, Gli1, and Gli2 in prostate cancer cells transfected with the miR-367-3p mimic were detected via qRT-PCR analysis. miR-367-3p expression in the prostate cancer tissues was downregulated compared with that in the para-cancer control tissues. miR-367-3p expression in DU145 and PC3 cells was also downregulated compared with that in the human prostate epithelial cell line RWPE-1. The overexpression of miR-367-3p or the knockdown of Rab23 inhibited the proliferation, invasion, and metastasis of prostate cancer cells. The results of the luciferase reporter assay confirmed that Rab23 was a target gene that was regulated by miR-367-3p. miR-367-3p specifically bound to the 3′-UTR of Rab23 mRNA. The overexpression of miR-367-3p inhibited Rab23 expression and the Hedgehog pathway. Cell function experiments confirmed that the overexpression of Rab23 reversed the anticancer effect of miR-367-3p. miR-367-3p was able to inhibit the Hedgehog pathway by targeting the expression of the Rab23 gene, thus inhibiting the proliferation, invasion, and metastasis of prostate cancer cells.
Collapse
Affiliation(s)
- Wei Du
- Department of Urology, The First Affiliated Hospital of Jinan University, Guangzhou, Guangdong 510630, P.R. China
| | - Dong Li
- Department of Urology, Nanhai Hospital of Guangdong Provincial People's Hospital, Foshan, Guangdong 528251, P.R. China
| | - Jianhao Xie
- Department of Clinical Laboratory, Nanhai Hospital of Guangdong Provincial People's Hospital, Foshan, Guangdong 528251, P.R. China
| | - Ping Tang
- Department of Urology, The First Affiliated Hospital of Jinan University, Guangzhou, Guangdong 510630, P.R. China
| |
Collapse
|
10
|
Bordanaba-Florit G, Royo F, Kruglik SG, Falcón-Pérez JM. Using single-vesicle technologies to unravel the heterogeneity of extracellular vesicles. Nat Protoc 2021; 16:3163-3185. [PMID: 34135505 DOI: 10.1038/s41596-021-00551-z] [Citation(s) in RCA: 167] [Impact Index Per Article: 41.8] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/27/2020] [Accepted: 03/31/2021] [Indexed: 12/12/2022]
Abstract
Extracellular vesicles (EVs) are heterogeneous lipid containers with a complex molecular cargo comprising several populations with unique roles in biological processes. These vesicles are closely associated with specific physiological features, which makes them invaluable in the detection and monitoring of various diseases. EVs play a key role in pathophysiological processes by actively triggering genetic or metabolic responses. However, the heterogeneity of their structure and composition hinders their application in medical diagnosis and therapies. This diversity makes it difficult to establish their exact physiological roles, and the functions and composition of different EV (sub)populations. Ensemble averaging approaches currently employed for EV characterization, such as western blotting or 'omics' technologies, tend to obscure rather than reveal these heterogeneities. Recent developments in single-vesicle analysis have made it possible to overcome these limitations and have facilitated the development of practical clinical applications. In this review, we discuss the benefits and challenges inherent to the current methods for the analysis of single vesicles and review the contribution of these approaches to the understanding of EV biology. We describe the contributions of these recent technological advances to the characterization and phenotyping of EVs, examination of the role of EVs in cell-to-cell communication pathways and the identification and validation of EVs as disease biomarkers. Finally, we discuss the potential of innovative single-vesicle imaging and analysis methodologies using microfluidic devices, which promise to deliver rapid and effective basic and practical applications for minimally invasive prognosis systems.
Collapse
Affiliation(s)
- Guillermo Bordanaba-Florit
- Exosomes Laboratory, Center for Cooperative Research in Biosciences (CIC bioGUNE), Basque Research and Technology Alliance (BRTA), Derio, Spain.
| | - Félix Royo
- Exosomes Laboratory, Center for Cooperative Research in Biosciences (CIC bioGUNE), Basque Research and Technology Alliance (BRTA), Derio, Spain.,Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (Ciberehd), Madrid, Spain
| | - Sergei G Kruglik
- Sorbonne Université, CNRS, Institut de Biologie Paris-Seine, Laboratoire Jean Perrin, Paris, France
| | - Juan M Falcón-Pérez
- Exosomes Laboratory, Center for Cooperative Research in Biosciences (CIC bioGUNE), Basque Research and Technology Alliance (BRTA), Derio, Spain. .,Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (Ciberehd), Madrid, Spain. .,Ikerbasque, Basque Foundation for Science, Bilbao, Spain.
| |
Collapse
|
11
|
Ghanbari S, Salimi A, Rahmani S, Nafissi N, Sharifi-Zarchi A, Mowla SJ. miR-361-5p as a promising qRT-PCR internal control for tumor and normal breast tissues. PLoS One 2021; 16:e0253009. [PMID: 34101749 PMCID: PMC8186776 DOI: 10.1371/journal.pone.0253009] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/14/2021] [Accepted: 05/27/2021] [Indexed: 11/19/2022] Open
Abstract
BACKGROUND One of the most widely used evaluation methods in miRNA experiments is qRT-PCR. However, selecting suitable internal controls (IC) is crucial for qRT-PCR experiments. Currently, there is no consensus on the ICs for miRNA qRT-PCR experiments in breast cancer. To this end, we tried to identify the most stable (the least expression alteration) and promising miRNAs in normal and tumor breast tissues by employing TCGA miRNA-Seq data and then experimentally validated them on fresh clinical samples. METHODS A multi-component scoring system was used which takes into account multiple expression stability criteria as well as correlation with clinical characteristics. Furthermore, we extended the scoring system for more than two biological sub-groups. TCGA BRCA samples were analyzed based on two grouping criteria: Tumor & Normal samples and Tumor subtypes. The top 10 most stable miRNAs were further investigated by differential expression and survival analysis. Then, we examined the expression level of the top scored miRNA (hsa-miR-361-5p) along with two commonly used ICs hsa-miR-16-5p and U48 on 34 pairs of Primary breast tumor and their adjacent normal tissues using qRT-PCR. RESULTS According to our multi-component scoring system, hsa-miR-361-5p had the highest stability score in both grouping criteria and hsa-miR-16-5p showed significantly lower scores. Based on our qRT-PCR assay, while U48 was the most abundant IC, hsa-miR-361-5p had lower standard deviation and also was the only IC capable of detecting a significant up-regulation of hsa-miR-21-5p in breast tumor tissue. CONCLUSIONS miRNA-Seq data is a great source to discover stable ICs. Our results demonstrated that hsa-miR-361-5p is a highly stable miRNA in tumor and non-tumor breast tissue and we recommend it as a suitable reference gene for miRNA expression studies in breast cancer. Additionally, although hsa-miR-16-5p is a commonly used IC, it's not a suitable one for breast cancer studies.
Collapse
Affiliation(s)
- Sogol Ghanbari
- Molecular Genetics Department, Biological Sciences Faculty, Tarbiat Modares University, Tehran, Iran
| | - Adel Salimi
- Computer Engineering Department, Sharif University of Technology, Tehran, Iran
| | - Saeid Rahmani
- Computer Engineering Department, Sharif University of Technology, Tehran, Iran
| | - Nahid Nafissi
- Surgical Department, School of Medicine, Iran University of Medical Sciences, Tehran, Iran
| | - Ali Sharifi-Zarchi
- Computer Engineering Department, Sharif University of Technology, Tehran, Iran
| | - Seyed Javad Mowla
- Molecular Genetics Department, Biological Sciences Faculty, Tarbiat Modares University, Tehran, Iran
- * E-mail:
| |
Collapse
|
12
|
MicroRNA and vascular endothelial growth factor (VEGF) as new useful markers in the diagnosis of benign prostatic hyperplasia in dogs. Theriogenology 2021; 171:113-118. [PMID: 34058504 DOI: 10.1016/j.theriogenology.2021.05.017] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/22/2021] [Revised: 04/13/2021] [Accepted: 05/15/2021] [Indexed: 11/22/2022]
Abstract
Numerous specific biomarkers with a prognostic and diagnostic value comparable to histopathological findings are now used for non-invasive diagnosis of prostate diseases in humans. Meanwhile, as far as dogs are concerned, the diagnosis of prostate disorders is based solely on clinical examination and ultrasound (USG). Therefore, the aim of the study was to assess the usefulness of two biomarkers, i.e. miRNA-129 and VEGF for the diagnosis of BPH in dogs. The study involved 40 dogs divided into three groups. Group I (n = 9) comprised healthy dogs up to the age of 5 years, Group II (n = 17) comprised dogs between the ages of 5-10 suffering from BPH as confirmed by the examination and Group III (n = 14) comprising dogs over 10 years of age, which also had BPH confirmed. The results demonstrated that dogs in group II and III exhibited a significant decrease in miRNA expression (P < 0.0001) and a significant increase in serum VEGF levels (P = 0.025) when compared to the dogs in group I. There was also a positive correlation between the prostate size and VEGF level. The findings led to the conclusion that the determination of miRNA-129 and VEGF can significantly contribute to the diagnosis of prostate disorders in dogs.
Collapse
|
13
|
Zheng H, Shi L, Tong C, Liu Y, Hou M. circSnx12 Is Involved in Ferroptosis During Heart Failure by Targeting miR-224-5p. Front Cardiovasc Med 2021; 8:656093. [PMID: 33969020 PMCID: PMC8097164 DOI: 10.3389/fcvm.2021.656093] [Citation(s) in RCA: 63] [Impact Index Per Article: 15.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/21/2021] [Accepted: 03/25/2021] [Indexed: 01/10/2023] Open
Abstract
Circular RNA (circRNA) is a subclass of non-coding RNAs that enables the circular transcripts resistant to the exonuclease digestion. Iron homeostasis is essential for the body to maintain normal physiological functions. At present, the relationship among circRNA, iron metabolism and heart failure remains largely unknown. This study aimed to explore the regulatory mechanism of circRNA and iron metabolism in heart failure. We obtained circRNA, miRNA and mRNA data from public databases and built a ceRNA network. The prediction results were verified in the myocardial tissues of pressure overload-induced heart failure mice through the use of histopathological staining methods, iron and malondialdehyde (MDA) measurement tests, quantitative real-time PCR (qRT-PCR), Western blot analysis and luciferase reporter assay. A total of 4 genes related to iron metabolism and oxidative stress were identified, and a ceRNA network involving 7 circRNAs, 7 miRNAs, and 4 mRNAs was constructed using bioinformatics tools. The results of qRT-PCR and Western blot analyses indicated that the expression level of FTH1 was similar with that predicted by bioinformatics analysis. Echocardiographic measurement showed that heart failure mice have lower fractional shortening and ejection fraction. Moreover, the myocardium of heart failure mice displayed obvious fibrosis as well as increased levels of iron and MDA compared to control mice. Besides, circSnx12 could act as an endogenous sponge to bind with miR-224-5p, and the 3'UTR region of FTH1 also had miRNA binding sites. A circRNA-miRNA-mRNA regulatory network was successfully constructed by identifying differentially expressed genes related to iron metabolism. This new approach reveals potential circRNA targets for the treatment of heart failure.
Collapse
Affiliation(s)
- Haoyuan Zheng
- Laboratory of Rescue Center of Severe Wound and Trauma Chinese People's Liberation Army, Department of Cardiovascular Surgery, General Hospital of Northern Theater Command of China Medical University, Shenyang, China
| | - Lin Shi
- Laboratory of Rescue Center of Severe Wound and Trauma Chinese People's Liberation Army, Emergency Medicine Department of General Hospital of Northern Theater Command, Shenyang, China
| | - Changci Tong
- Laboratory of Rescue Center of Severe Wound and Trauma Chinese People's Liberation Army, Emergency Medicine Department of General Hospital of Northern Theater Command, Shenyang, China
| | - Yunen Liu
- The Second Affiliated Hospital of Shenyang Medical College, The Veterans General Hospital of Liaoning Province, Shenyang, China.,Shenyang Medical College, Shenyang, China
| | - Mingxiao Hou
- Laboratory of Rescue Center of Severe Wound and Trauma Chinese People's Liberation Army, Department of Cardiovascular Surgery, General Hospital of Northern Theater Command of China Medical University, Shenyang, China.,The Second Affiliated Hospital of Shenyang Medical College, The Veterans General Hospital of Liaoning Province, Shenyang, China.,Shenyang Medical College, Shenyang, China
| |
Collapse
|
14
|
Diallo I, Ho J, Laffont B, Laugier J, Benmoussa A, Lambert M, Husseini Z, Soule G, Kozak R, Kobinger GP, Provost P. Altered microRNA Transcriptome in Cultured Human Liver Cells upon Infection with Ebola Virus. Int J Mol Sci 2021; 22:ijms22073792. [PMID: 33917562 PMCID: PMC8038836 DOI: 10.3390/ijms22073792] [Citation(s) in RCA: 13] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/01/2021] [Revised: 03/27/2021] [Accepted: 03/30/2021] [Indexed: 02/07/2023] Open
Abstract
Ebola virus (EBOV) is a virulent pathogen, notorious for inducing life-threatening hemorrhagic fever, that has been responsible for several outbreaks in Africa and remains a public health threat. Yet, its pathogenesis is still not completely understood. Although there have been numerous studies on host transcriptional response to EBOV, with an emphasis on the clinical features, the impact of EBOV infection on post-transcriptional regulatory elements, such as microRNAs (miRNAs), remains largely unexplored. MiRNAs are involved in inflammation and immunity and are believed to be important modulators of the host response to viral infection. Here, we have used small RNA sequencing (sRNA-Seq), qPCR and functional analyses to obtain the first comparative miRNA transcriptome (miRNome) of a human liver cell line (Huh7) infected with one of the following three EBOV strains: Mayinga (responsible for the first Zaire outbreak in 1976), Makona (responsible for the West Africa outbreak in 2013–2016) and the epizootic Reston (presumably innocuous to humans). Our results highlight specific miRNA-based immunity pathways and substantial differences between the strains beyond their clinical manifestation and pathogenicity. These analyses shed new light into the molecular signature of liver cells upon EBOV infection and reveal new insights into miRNA-based virus attack and host defense strategy.
Collapse
Affiliation(s)
- Idrissa Diallo
- CHU de Québec Research Center, Department of Microbiology, Infectious Diseases and Immunology, Faculty of Medicine, Université Laval, Quebec, QC G1V 4G2, Canada; (I.D.); (J.H.); (B.L.); (J.L.); (A.B.); (M.L.); (Z.H.); (G.P.K.)
| | - Jeffrey Ho
- CHU de Québec Research Center, Department of Microbiology, Infectious Diseases and Immunology, Faculty of Medicine, Université Laval, Quebec, QC G1V 4G2, Canada; (I.D.); (J.H.); (B.L.); (J.L.); (A.B.); (M.L.); (Z.H.); (G.P.K.)
| | - Benoit Laffont
- CHU de Québec Research Center, Department of Microbiology, Infectious Diseases and Immunology, Faculty of Medicine, Université Laval, Quebec, QC G1V 4G2, Canada; (I.D.); (J.H.); (B.L.); (J.L.); (A.B.); (M.L.); (Z.H.); (G.P.K.)
| | - Jonathan Laugier
- CHU de Québec Research Center, Department of Microbiology, Infectious Diseases and Immunology, Faculty of Medicine, Université Laval, Quebec, QC G1V 4G2, Canada; (I.D.); (J.H.); (B.L.); (J.L.); (A.B.); (M.L.); (Z.H.); (G.P.K.)
| | - Abderrahim Benmoussa
- CHU de Québec Research Center, Department of Microbiology, Infectious Diseases and Immunology, Faculty of Medicine, Université Laval, Quebec, QC G1V 4G2, Canada; (I.D.); (J.H.); (B.L.); (J.L.); (A.B.); (M.L.); (Z.H.); (G.P.K.)
| | - Marine Lambert
- CHU de Québec Research Center, Department of Microbiology, Infectious Diseases and Immunology, Faculty of Medicine, Université Laval, Quebec, QC G1V 4G2, Canada; (I.D.); (J.H.); (B.L.); (J.L.); (A.B.); (M.L.); (Z.H.); (G.P.K.)
| | - Zeinab Husseini
- CHU de Québec Research Center, Department of Microbiology, Infectious Diseases and Immunology, Faculty of Medicine, Université Laval, Quebec, QC G1V 4G2, Canada; (I.D.); (J.H.); (B.L.); (J.L.); (A.B.); (M.L.); (Z.H.); (G.P.K.)
| | - Geoff Soule
- Special Pathogens Program, National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg, MB R3B 3M9, Canada; (G.S.); (R.K.)
| | - Robert Kozak
- Special Pathogens Program, National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg, MB R3B 3M9, Canada; (G.S.); (R.K.)
- Division of Microbiology, Department of Laboratory Medicine & Molecular Diagnostics, Sunnybrook Health Sciences Centre, Toronto, ON M4N 3M5, Canada
| | - Gary P. Kobinger
- CHU de Québec Research Center, Department of Microbiology, Infectious Diseases and Immunology, Faculty of Medicine, Université Laval, Quebec, QC G1V 4G2, Canada; (I.D.); (J.H.); (B.L.); (J.L.); (A.B.); (M.L.); (Z.H.); (G.P.K.)
- Special Pathogens Program, National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg, MB R3B 3M9, Canada; (G.S.); (R.K.)
- Département de Microbiologie Médicale, Université du Manitoba, Winnipeg, MB R3E 0J9, Canada
| | - Patrick Provost
- CHU de Québec Research Center, Department of Microbiology, Infectious Diseases and Immunology, Faculty of Medicine, Université Laval, Quebec, QC G1V 4G2, Canada; (I.D.); (J.H.); (B.L.); (J.L.); (A.B.); (M.L.); (Z.H.); (G.P.K.)
- CHUQ Research Center/CHUL Pavilion, 2705 Blvd Laurier, Room T1-65, Quebec, QC G1V 4G2, Canada
- Correspondence: ; Tel.: +1-418-525-4444 (ext. 48842)
| |
Collapse
|
15
|
Rajendiran S, Maji S, Haddad A, Lotan Y, Nandy RR, Vishwanatha JK, Chaudhary P. MicroRNA-940 as a Potential Serum Biomarker for Prostate Cancer. Front Oncol 2021; 11:628094. [PMID: 33816263 PMCID: PMC8017318 DOI: 10.3389/fonc.2021.628094] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/11/2020] [Accepted: 02/23/2021] [Indexed: 12/22/2022] Open
Abstract
Prostate cancer is one of the leading causes of death despite an astoundingly high survival rate for localized tumors. Though prostate specific antigen (PSA) test, performed in conjunction with digital rectal examinations, is reasonably accurate, there are major caveats requiring a thorough assessment of risks and benefits prior to conducting the test. MicroRNAs, a class of small non-coding RNAs, are stable molecules that can be detected in circulation by non-invasive methods and have gained importance in cancer prognosis and diagnosis in the recent years. Here, we investigate circulating miR-940, a miRNA known to play a role in prostate cancer progression, in both cell culture supernatants as well as patient serum and urine samples to determine the utility of miR-940 as a new molecular marker for prostate cancer detection. We found that miR-940 was significantly higher in serum from cancer patients, specifically those with clinically significant tumors (GS ≥ 7). Analysis of receiver operating characteristic curve demonstrated that miR-940 in combination with PSA had a higher area under curve value (AUC: 0.818) than the miR-940 alone (AUC: 0.75) for the diagnosis of prostate cancer. This study provides promising results suggesting the use of miR-940 for prostate cancer diagnosis.
Collapse
Affiliation(s)
- Smrithi Rajendiran
- Department of Microbiology, Immunology, and Genetics, University of North Texas Health Science Center, Fort Worth, TX, United States
| | - Sayantan Maji
- Department of Microbiology, Immunology, and Genetics, University of North Texas Health Science Center, Fort Worth, TX, United States
| | - Ahmed Haddad
- Department of Urology, University of Texas Southwestern Medical Center, Dallas, TX, United States
| | - Yair Lotan
- Department of Urology, University of Texas Southwestern Medical Center, Dallas, TX, United States
| | - Rajesh R Nandy
- School of Public Health, University of North Texas Health Science Center, Fort Worth, TX, United States
| | - Jamboor K Vishwanatha
- Department of Microbiology, Immunology, and Genetics, University of North Texas Health Science Center, Fort Worth, TX, United States.,Texas Center for Health Disparities, University of North Texas Health Science Center, Fort Worth, TX, United States
| | - Pankaj Chaudhary
- Department of Microbiology, Immunology, and Genetics, University of North Texas Health Science Center, Fort Worth, TX, United States.,Texas Center for Health Disparities, University of North Texas Health Science Center, Fort Worth, TX, United States
| |
Collapse
|
16
|
Tran CM, Lee H, Lee B, Ra JS, Kim KT. Effects of the chorion on the developmental toxicity of organophosphate esters in zebrafish embryos. JOURNAL OF HAZARDOUS MATERIALS 2021; 401:123389. [PMID: 32763690 DOI: 10.1016/j.jhazmat.2020.123389] [Citation(s) in RCA: 60] [Impact Index Per Article: 15.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 03/29/2020] [Revised: 05/13/2020] [Accepted: 07/02/2020] [Indexed: 05/21/2023]
Abstract
Many toxicological studies have utilized zebrafish embryos to investigate the developmental toxicity of organophosphate esters (OPEs). However, in respect of the presence or absence of the chorion, a consistent experimental methodology has yet to be developed. In this study, we used a fixed exposure scheme to compare the developmental toxicities of six major OPEs in chorionated and dechorionated zebrafish embryos. Removal of the chorion increased sensitivity to OPEs: we found higher incidence of mortality and malformation in dechorionated embryos. In a behavioral assay, the locomotive activity of zebrafish larvae was consistently inhibited by OPEs except tris (1-chloropropyl) phosphate regardless of chorion presence. However, at the molecular level, the expression of ZHE1 and mmp9 was affected by the presence of the chorion in zebrafish embryos exposed to tributyl phosphate and triphenyl phosphate (TPHP), respectively. Furthermore, in zebrafish embryos exposed to TPHP, the increased expression of miR-137 and miR-141 was abolished by the presence of the chorion. Our results demonstrate for the first time that the presence of the chorion influences phenotypic morbidity, organismal behavior, and gene expression in zebrafish embryos exposed to chemicals; thus, we suggest that dechorionation is desirable for exploring the toxicity mechanisms that underlie effects of chemical exposure.
Collapse
Affiliation(s)
- Cong M Tran
- Department of Environmental Engineering, Seoul 01811 Republic of Korea; Department of Environmental Energy Engineering, Seoul National University of Science and Technology, Seoul 01811, Republic of Korea
| | - Hyojin Lee
- Department of Environmental Energy Engineering, Seoul National University of Science and Technology, Seoul 01811, Republic of Korea
| | - Byoungcheun Lee
- Risk Assessment Division, National Institute of Environmental Research, Incheon 22689, Republic of Korea
| | - Jin-Sung Ra
- Eco-testing and Risk Assessment Center, Korea Institute of Industrial Technology (KITECH), Ansan 15588, Republic of Korea
| | - Ki-Tae Kim
- Department of Environmental Engineering, Seoul 01811 Republic of Korea; Department of Environmental Energy Engineering, Seoul National University of Science and Technology, Seoul 01811, Republic of Korea.
| |
Collapse
|
17
|
Qin D, Zhao Y, Guo Q, Zhu S, Zhang S, Min L. Detection of Pancreatic Ductal Adenocarcinoma by A qPCR-based Normalizer-free Circulating Extracellular Vesicles RNA Signature. J Cancer 2021; 12:1445-1454. [PMID: 33531989 PMCID: PMC7847660 DOI: 10.7150/jca.50716] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/15/2020] [Accepted: 10/25/2020] [Indexed: 12/17/2022] Open
Abstract
Background: Pancreatic ductal adenocarcinoma (PDAC) is difficult to diagnose and many efforts have been made to evaluate EVs-derived RNAs as biomarkers to predict PDAC. However, lack of robust internal references largely limited their clinical application. Here we proposed an RNA ratio-based, normalizer-free algorithm to quantitate EVs-derived RNAs in PDAC. Methods: Differentially expressed RNAs in the training group were identified using "limma" package. The ratio of any two candidate RNAs in the same sample was calculated and used as a new biomarker. LASSO regression was performed to build prediction models based on those RNA ratios. RNA-seq data of 116 plasma samples and RT-qPCR data of 111 plasma samples were used for internal and external validation, separately. Three algorithms (lasso regression, logistic regression, and SVM) were compared to improve the performance of this RNA signature. Results: We developed an RNA-ratio based prediction model which comprised eight EVs-derived RNAs, including FBXO7, MORF4L1, DDX17, TALDO1, AHNAK, TUBA1B, CD44, and SETD3. This model could well differentiate PDAC patients with a minimal AUC of 0.86 in internal verification using testing group. External validation using RT-qPCR data also exhibited a good classifier ability with an AUC of 0.89 when distinguishing PDAC from healthy controls. Conclusion: We've developed a qPCR-based, normalizer-free circulating EVs RNA classifier, which could well distinguish PDAC patients from noncancerous controls.
Collapse
Affiliation(s)
- Da Qin
- Department of Gastroenterology, Beijing Friendship Hospital, Capital Medical University, National Clinical Research Center for Digestive Disease, Beijing Digestive Disease Center, Beijing Key Laboratory for Precancerous Lesion of Digestive Disease, Beijing, 100050, P. R. China
| | - Yu Zhao
- Department of Gastroenterology, Beijing Friendship Hospital, Capital Medical University, National Clinical Research Center for Digestive Disease, Beijing Digestive Disease Center, Beijing Key Laboratory for Precancerous Lesion of Digestive Disease, Beijing, 100050, P. R. China
| | - Qingdong Guo
- Department of Gastroenterology, Beijing Friendship Hospital, Capital Medical University, National Clinical Research Center for Digestive Disease, Beijing Digestive Disease Center, Beijing Key Laboratory for Precancerous Lesion of Digestive Disease, Beijing, 100050, P. R. China
| | - Shengtao Zhu
- Department of Gastroenterology, Beijing Friendship Hospital, Capital Medical University, National Clinical Research Center for Digestive Disease, Beijing Digestive Disease Center, Beijing Key Laboratory for Precancerous Lesion of Digestive Disease, Beijing, 100050, P. R. China
| | - Shutian Zhang
- Department of Gastroenterology, Beijing Friendship Hospital, Capital Medical University, National Clinical Research Center for Digestive Disease, Beijing Digestive Disease Center, Beijing Key Laboratory for Precancerous Lesion of Digestive Disease, Beijing, 100050, P. R. China
| | - Li Min
- Department of Gastroenterology, Beijing Friendship Hospital, Capital Medical University, National Clinical Research Center for Digestive Disease, Beijing Digestive Disease Center, Beijing Key Laboratory for Precancerous Lesion of Digestive Disease, Beijing, 100050, P. R. China
| |
Collapse
|
18
|
Elshafie NO, Nascimento NCD, Lichti NI, Kasinski AL, Childress MO, Santos APD. MicroRNA Biomarkers in Canine Diffuse Large B-Cell Lymphoma. Vet Pathol 2020; 58:34-41. [PMID: 33287683 DOI: 10.1177/0300985820967902] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/25/2022]
Abstract
Lymphoma is among the most common cancer in dogs. Diffuse large B-cell lymphoma (DLBCL) is the predominant type, accounting for up to half of all cases. Definitive diagnosis of DLBCL relies on cytologic evaluation with immunophenotyping, or histopathology and immunohistochemistry when needed. A rapid and specific molecular test aiding in the diagnosis could be beneficial. Noncoding microRNAs (miRNAs) are regulators of gene expression involved in a variety of cellular processes, including cell differentiation, cell cycle progression, and apoptosis. Not surprisingly, miRNA expression is aberrant in diseases such as cancers. Their high stability and abundance in tissues make them promising biomarkers for diagnosing and monitoring diseases. This study aimed to identify miRNA signatures of DLBCL to develop ancillary molecular diagnostic tools. miRNA was isolated from formalin-fixed, paraffin-embedded lymph node tissue from 22 DLBCL and 14 nonneoplastic controls. Relative gene expression of 8 tumor-regulating miRNAs was achieved by RT-qPCR (reverse transcriptase quantitative polymerase chain reaction). The results showed downregulation of the let-7 family of miRNAs and miR-155, whereas miR-34a was upregulated in DLBCL compared to the controls. We demonstrated that the combination of expression levels of miR-34a and let-7f or of let-7b and let-7f achieved 100% differentiation between DLBCL and controls. Furthermore, let-7f alone discriminated DLBCL from nonneoplastic tissue in 97% of cases. Our results represent one step forward in search of a rapid and accurate ancillary diagnostic test for DLBCL in dogs.
Collapse
|
19
|
Identification of miR-29c-3p as a Robust Normalizer for Urine microRNA Studies in Bladder Cancer. Biomedicines 2020; 8:biomedicines8110447. [PMID: 33105660 PMCID: PMC7690381 DOI: 10.3390/biomedicines8110447] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/17/2020] [Revised: 10/09/2020] [Accepted: 10/19/2020] [Indexed: 12/24/2022] Open
Abstract
Bladder cancer (BC) is among the most frequent malignancies worldwide, being the most expensive cancer to treat and monitor and the most lethal urological cancer. Urine microRNAs (miRNAs) have been proposed as novel non-invasive biomarkers to early diagnose and monitor BC patients in order to avoid the performance of current aggressive diagnostic techniques. However, huge discrepancies arise among studies mainly due to the lack of standardization in the normalization, a crucial step in all miRNA studies. Our aim was to identify the best miRNA normalizer for miRNA studies in urine of BC patients. We evaluated the performance of 110 candidate miRNAs in urine of 35 BC patients and 15 healthy controls by Real Time quantitative Polymerase Chain Reaction (RT-qPCR) followed by a stability analysis with RefFinder. In this screening stage, miR-29c-3p arose as the most stably expressed miRNA in BC and controls, with a good expression level. Stability of miR-29c-3p expression was validated in an independent cohort of 153 BC patients and 57 controls. Finally, we evaluated the robustness of miR-29c-3p as normalizer in the expression study of miR-200c-3p, a potential diagnostic marker for BC. We propose miR-29c-3p as a normalizer for miRNA studies in BC urine. This is the first study that characterizes a reliable normalizer that may allow the comparison of future urine miRNA studies as non-invasive biomarkers for BC diagnosis and monitoring.
Collapse
|
20
|
Abstract
MicroRNAs (miRNAs) are endogenous noncoding RNAs, which regulate gene expression on the post-transcriptional level. Since miRNAs are involved in the regulation of apoptosis, cellular proliferation, differentiation, and other important cellular processes, their deregulation is important for the development of a wide range of diseases including cancer. Apart from tissue, specific disease-related miRNA signatures can be found in body fluids as well. Especially for urologic diseases or injuries, urine miRNAs represent a promising group of biomarkers. Despite a large number of studies describing the importance of urinary miRNAs, there is a lack of recommendations for urine management and subsequent miRNA analysis. Thus, in this chapter, we aim to describe the origin and functions of urinary miRNAs and discuss the technical aspects of their detection including the pre-analytical phase principles and new directions in quantification, which could forward urine miRNA into clinical practice.
Collapse
Affiliation(s)
- Jaroslav Juracek
- Central European Institute of Technology, Masaryk University, Brno, Czech Republic
| | - Ondrej Slaby
- Central European Institute of Technology, Masaryk University, Brno, Czech Republic.
| |
Collapse
|
21
|
Geeurickx E, Hendrix A. Targets, pitfalls and reference materials for liquid biopsy tests in cancer diagnostics. Mol Aspects Med 2019; 72:100828. [PMID: 31711714 DOI: 10.1016/j.mam.2019.10.005] [Citation(s) in RCA: 93] [Impact Index Per Article: 15.5] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/31/2019] [Revised: 10/14/2019] [Accepted: 10/18/2019] [Indexed: 12/12/2022]
Abstract
Assessment of cell free DNA (cfDNA) and RNA (cfRNA), circulating tumor cells (CTC) and extracellular vesicles (EV) in blood or other bodily fluids can enable early cancer detection, tumor dynamics assessment, minimal residual disease detection and therapy monitoring. However, few liquid biopsy tests progress towards clinical application because results are often discordant and challenging to reproduce. Reproducibility can be enhanced by the development and implementation of standard operating procedures and reference materials to identify and correct for pre-analytical variables. In this review we elaborate on the technological considerations, pre-analytical variables and the use and availability of reference materials for the assessment of liquid biopsy targets in blood and highlight initiatives towards the standardization of liquid biopsy testing.
Collapse
Affiliation(s)
- Edward Geeurickx
- Laboratory of Experimental Cancer Research, Department of Human Structure and Repair, Ghent University, 9000, Ghent, Belgium; Cancer Research Institute Ghent, 9000, Ghent, Belgium
| | - An Hendrix
- Laboratory of Experimental Cancer Research, Department of Human Structure and Repair, Ghent University, 9000, Ghent, Belgium; Cancer Research Institute Ghent, 9000, Ghent, Belgium.
| |
Collapse
|
22
|
A Novel Predictor Tool of Biochemical Recurrence after Radical Prostatectomy Based on a Five-MicroRNA Tissue Signature. Cancers (Basel) 2019; 11:cancers11101603. [PMID: 31640261 PMCID: PMC6826532 DOI: 10.3390/cancers11101603] [Citation(s) in RCA: 22] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/11/2019] [Accepted: 10/17/2019] [Indexed: 12/24/2022] Open
Abstract
Within five to ten years after radical prostatectomy (RP), approximately 15–34% of prostate cancer (PCa) patients experience biochemical recurrence (BCR), which is defined as recurrence of serum levels of prostate-specific antigen >0.2 µg/L, indicating probable cancer recurrence. Models using clinicopathological variables for predicting this risk for patients lack accuracy. There is hope that new molecular biomarkers, like microRNAs (miRNAs), could be potential candidates to improve risk prediction. Therefore, we evaluated the BCR prognostic capability of 20 miRNAs, which were selected by a systematic literature review. MiRNA expressions were measured in formalin-fixed, paraffin-embedded (FFPE) tissue RP samples of 206 PCa patients by RT-qPCR. Univariate and multivariate Cox regression analyses were performed, to assess the independent prognostic potential of miRNAs. Internal validation was performed, using bootstrapping and the split-sample method. Five miRNAs (miR-30c-5p/31-5p/141-3p/148a-3p/miR-221-3p) were finally validated as independent prognostic biomarkers. Their prognostic ability and accuracy were evaluated using C-statistics of the obtained prognostic indices in the Cox regression, time-dependent receiver-operating characteristics, and decision curve analyses. Models of miRNAs, combined with relevant clinicopathological factors, were built. The five-miRNA-panel outperformed clinically established BCR scoring systems, while their combination significantly improved predictive power, based on clinicopathological factors alone. We conclude that this miRNA-based-predictor panel will be worth to be including in future studies.
Collapse
|
23
|
Liu J, Jia E, Shi H, Li X, Jiang G, Chi C, Liu W, Zhang D. Selection of reference genes for miRNA quantitative PCR and its application in miR-34a/Sirtuin-1 mediated energy metabolism in Megalobrama amblycephala. FISH PHYSIOLOGY AND BIOCHEMISTRY 2019; 45:1663-1681. [PMID: 31127447 DOI: 10.1007/s10695-019-00658-x] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 11/26/2018] [Accepted: 05/06/2019] [Indexed: 05/19/2023]
Abstract
MiRNAs are small, non-coding RNAs that downregulate gene expression at post-transcriptional levels. They have emerged as important regulators involved in metabolism, immunity, and cancer. Real-time quantitative PCR (RT-qPCR) is an effective and main method for quantifying target miRNA. For robust RT-qPCR method, suitable reference genes play crucial roles in data normalization. Blunt snout bream (Megalobrama amblycephala) is an economically important aquaculture species; however, no reference genes dedicated for qPCR method has been identified for this species so far. The objective of this study was to screen stable reference genes for miRNA RT-qPCR and demonstrated its application in energy metabolism in blunt snout bream. The stabilities of ten potential reference genes (miR-21-1-5p, miR-107a-3p, miR-222a-3p, miR-146a-5p, miR-101a-3p, miR-22a-3p, miR-103-3p, miR-456-3p, miR-221-3p, and U6 (RNU6A)) were evaluated in nine tissues (brain, muscle, liver, skin, spleen, heart, gill, intestine, and eye) under normal condition and in three tissues (liver, intestine, and spleen) under four stresses (heat stress, ammonia stress, bacterial challenge, and glycolipid stress). Using GeNorm, NormFinder, and RefFinder softwares, we discovered that different tissues and stresses are both important variability factors for the expression stability of miRNAs. After verifying miR-34a/Sirtuin-1 expressions in high-carbohydrate diet-induced blunt snout bream, we eventually identified that the most stable reference gene in this species was miR-221-3p, and the best combination of reference genes were miR-221-3p and miR-103-3p.
Collapse
Affiliation(s)
- Jie Liu
- Key Laboratory of Aquatic Nutrition and Feed Science of Jiangsu Province, College of Animal Science and Technology, Nanjing Agricultural University, No. 1 Weigang Road, Nanjing, 210095, People's Republic of China
| | - Erteng Jia
- Key Laboratory of Aquatic Nutrition and Feed Science of Jiangsu Province, College of Animal Science and Technology, Nanjing Agricultural University, No. 1 Weigang Road, Nanjing, 210095, People's Republic of China
| | - Huajuan Shi
- Key Laboratory of Aquatic Nutrition and Feed Science of Jiangsu Province, College of Animal Science and Technology, Nanjing Agricultural University, No. 1 Weigang Road, Nanjing, 210095, People's Republic of China
| | - Xiangfei Li
- Key Laboratory of Aquatic Nutrition and Feed Science of Jiangsu Province, College of Animal Science and Technology, Nanjing Agricultural University, No. 1 Weigang Road, Nanjing, 210095, People's Republic of China
| | - Guangzhen Jiang
- Key Laboratory of Aquatic Nutrition and Feed Science of Jiangsu Province, College of Animal Science and Technology, Nanjing Agricultural University, No. 1 Weigang Road, Nanjing, 210095, People's Republic of China
| | - Cheng Chi
- Key Laboratory of Aquatic Nutrition and Feed Science of Jiangsu Province, College of Animal Science and Technology, Nanjing Agricultural University, No. 1 Weigang Road, Nanjing, 210095, People's Republic of China
| | - Wenbin Liu
- Key Laboratory of Aquatic Nutrition and Feed Science of Jiangsu Province, College of Animal Science and Technology, Nanjing Agricultural University, No. 1 Weigang Road, Nanjing, 210095, People's Republic of China
| | - Dingdong Zhang
- Key Laboratory of Aquatic Nutrition and Feed Science of Jiangsu Province, College of Animal Science and Technology, Nanjing Agricultural University, No. 1 Weigang Road, Nanjing, 210095, People's Republic of China.
| |
Collapse
|
24
|
Madadi S, Schwarzenbach H, Lorenzen J, Soleimani M. MicroRNA expression studies: challenge of selecting reliable reference controls for data normalization. Cell Mol Life Sci 2019; 76:3497-3514. [PMID: 31089747 PMCID: PMC11105490 DOI: 10.1007/s00018-019-03136-y] [Citation(s) in RCA: 23] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/02/2019] [Revised: 04/13/2019] [Accepted: 05/06/2019] [Indexed: 02/06/2023]
Abstract
Accurate determination of microRNA expression levels is a prerequisite in using these small non-coding RNA molecules as novel biomarkers in disease diagnosis and prognosis. Quantitative PCR is the method of choice for measuring the expression levels of microRNAs. However, a major obstacle that affects the reliability of results is the lack of validated reference controls for data normalization. Various non-coding RNAs have previously been used as reference controls, but their use may lead to variations and lack of comparability of microRNA data among the studies. Despite the growing number of studies investigating microRNA profiles to discriminate between healthy and disease stages, robust reference controls for data normalization have so far not been established. In the present article, we provide an overview of different reference controls used in various diseases, and highlight the urgent need for the identification of suitable reference controls to produce reliable data. Our analysis shows, among others, that RNU6 is not an ideal normalizer in studies using patient material from different diseases. Finally, our article tries to disclose the challenges to find a reference control which is uniformly and stably expressed across all body tissues, fluids, and diseases.
Collapse
Affiliation(s)
- Soheil Madadi
- Department of Pharmaceutical Biotechnology, School of Pharmacy, Hamadan University of Medical Sciences, Hamadan, Iran
| | - Heidi Schwarzenbach
- Department of Tumor Biology, University Medical Center Hamburg-Eppendorf, 20246, Hamburg, Germany
| | - Johan Lorenzen
- Department of Nephrology, University Hospital Zürich, Zurich, Switzerland
| | - Meysam Soleimani
- Department of Pharmaceutical Biotechnology, School of Pharmacy, Hamadan University of Medical Sciences, Hamadan, Iran.
| |
Collapse
|
25
|
Variance component analysis of circulating miR-122 in serum from healthy human volunteers. PLoS One 2019; 14:e0220406. [PMID: 31348817 PMCID: PMC6660082 DOI: 10.1371/journal.pone.0220406] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/23/2019] [Accepted: 07/15/2019] [Indexed: 12/25/2022] Open
Abstract
Micro-RNA (miR)-122 is a promising exploratory biomarker for detecting liver injury in preclinical and clinical studies. Elevations in serum or plasma have been associated with viral and autoimmune hepatitis, non-alcoholic steatohepatitis (NASH), hepatocellular carcinoma, and drug-induced liver injury (DILI). However, these associations were primarily based upon population differences between the disease state and the controls. Thus, little is known about the variability and subsequent variance components of circulating miR-122 in healthy humans, which has implications for the practical use of the biomarker clinically. To address this, we set out to perform variance components analysis of miR-122 in a cohort of 40 healthy volunteers. Employing a quantitative real-time polymerase chain reaction (qRT-PCR) assay to detect miR-122 and other circulating miRNAs in human serum, the relative expression of miR-122 was determined using two different normalization approaches: to the mean expression of a panel of several endogenous miRNAs identified using an adaptive algorithm (miRA-Norm) and to the expression of an exogenous miRNA control (Caenorhabditis elegans miR-39). Results from a longitudinal study in healthy volunteers (N = 40) demonstrated high variability with 117- and 111-fold 95% confidence reference interval, respectively. This high variability of miR-122 in serum appeared to be due in part to ethnicity, as 95% confidence reference intervals were approximately three-fold lower in volunteers that identified as Caucasian relative to those that identified as Non-Caucasian. Variance analysis revealed equivalent contributions of intra- and inter-donor variability to miR-122. Surprisingly, miR-122 exhibited the highest variability compared to other 36 abundant miRNAs in circulation; the next variable miRNA, miR-133a, demonstrated a 45- to 62-fold reference interval depending on normalization approaches. In contrast, alanine aminotransferase (ALT) activity levels in this population exhibited a 5-fold total variance, with 80% of this variance due to inter-donor sources. In conclusion, miR-122 demonstrated higher than expected variability in serum from healthy volunteers, which has implications for its potential utility as a prospective biomarker of liver damage or injury.
Collapse
|
26
|
Wang F, Yang QW, Zhao WJ, Du QY, Chang ZJ. Selection of suitable candidate genes for miRNA expression normalization in Yellow River Carp (Cyprinus carpio. var). Sci Rep 2019; 9:8691. [PMID: 31213623 PMCID: PMC6581906 DOI: 10.1038/s41598-019-44982-x] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/24/2018] [Accepted: 05/27/2019] [Indexed: 12/19/2022] Open
Abstract
Yellow River carp is widely cultivated in the world due to its economic value in aquaculture, and the faster growth of females compared to males. It is believed that microRNAs (miRNA) are involved in gonadal differentiation and development. qPCR is the most preferred method for miRNA functional analysis. Reliable reference genes for normalization in qRT-PCR are the key to ensuring the accuracy of this method. The aim of present research was to evaluate as well as identify the efficacy of reference genes for miRNA expression using qRT-PCR in Yellow River carp. Nine ncRNAs (miR-101, miR-23a, let7a, miR-26a, miR-146a, miR-451, U6, 5S, and 18S) were chosen and tested in four sample sets: (1) different tissues in adult carp, (2) different tissues in juvenile carp, (3) different early developmental stages of carp, and (4) different developmental stages of carp gonads. The stability and suitability values were calculated using NormFinder, geNorm, and BestKeeper software. The results showed that 5S was a suitable reference gene in different tissues of adult and juvenile carp. The genes 5S, 18S, and U6 were the most stable reference genes in the early developmental stages of carp. Let-7a and miR-23a were considered as the suitable reference genes in the development of gonads. All these reference genes were subsequently validated using miR-430. The results showed that genes 5S and 18S were the most suitable reference genes to normalize miRNA expression under normal growth conditions in early different developmental stages. The genes Let-7a, and miR-23a were the most suitable in different developmental stages. The present study is the first comprehensive study of the stability of miRNA reference genes in Yellow River carp, providing valuable as well as basic data for investigating more accurate miRNA expression during gonadal differentiation and development of carp.
Collapse
Affiliation(s)
- Fang Wang
- College of Life Science, Henan Normal University, Xinxiang, Henan, 453007, People's Republic of China
| | - Qian-Wen Yang
- College of Life Science, Henan Normal University, Xinxiang, Henan, 453007, People's Republic of China
| | - Wen-Jie Zhao
- College of Life Science, Henan Normal University, Xinxiang, Henan, 453007, People's Republic of China
| | - Qi-Yan Du
- College of Life Science, Henan Normal University, Xinxiang, Henan, 453007, People's Republic of China
| | - Zhong-Jie Chang
- College of Life Science, Henan Normal University, Xinxiang, Henan, 453007, People's Republic of China.
| |
Collapse
|
27
|
Buonpane C, Ares G, Benyamen B, Yuan C, Hunter CJ. Identification of suitable reference microRNA for qPCR analysis in pediatric inflammatory bowel disease. Physiol Genomics 2019; 51:169-175. [PMID: 30978148 DOI: 10.1152/physiolgenomics.00126.2018] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022] Open
Abstract
Pediatric inflammatory bowel disease (IBD) accounts for 10-15% of IBD and is associated with considerable morbidity for patients. Dysregulated microRNAs (miRNA, miR), small noncoding RNA molecules that modulate gene expression, have been the target of research in IBD diagnosis, surveillance, and therapy. Proper selection of reference genes, which are a prerequisite for accurate measurement of miRNA expression, is currently lacking. We hypothesize that appropriate normalization requires unique reference genes for different tissue and disease types. Through the study of 28 pediatric intestinal samples, we sought to create a protocol for selection of suitable endogenous reference genes. Candidate reference genes (miR-16, 193a, 27a, 103a, 191) were analyzed by RT-quantitative (q)PCR. Criteria used for designation of suitable reference genes were as follows: 1) ubiquitous: present in all tissue samples with quantification cycle value 15-35; 2) uniform expression: no differential expression between control and disease samples (P > 0.05); 3) stability: stability value <0.5 by NormFinder. Our results suggest the use of miR-27a/191 for Crohn's disease small bowel, none of the five candidate genes for Crohn's disease colon, and miR-16/27a for ulcerative colitis. Additionally, target miR-874 had differential expression when normalized with different reference genes. Our results demonstrate that reference gene choice for qPCR analysis has a significant effect on study results and that proper data normalization is imperative.
Collapse
Affiliation(s)
- Christie Buonpane
- Department of Pediatrics, Feinberg School of Medicine, Northwestern University , Chicago, Illinois.,Ann and Robert H. Lurie Children's Hospital of Chicago , Chicago, Illinois
| | - Guillermo Ares
- Ann and Robert H. Lurie Children's Hospital of Chicago , Chicago, Illinois.,Department of Surgery, University of Illinois at Chicago , Chicago, Illinois
| | - Beshoy Benyamen
- Department of Pediatrics, Feinberg School of Medicine, Northwestern University , Chicago, Illinois.,Ann and Robert H. Lurie Children's Hospital of Chicago , Chicago, Illinois
| | - Carrie Yuan
- Department of Pediatrics, Feinberg School of Medicine, Northwestern University , Chicago, Illinois
| | - Catherine J Hunter
- Department of Pediatrics, Feinberg School of Medicine, Northwestern University , Chicago, Illinois.,Ann and Robert H. Lurie Children's Hospital of Chicago , Chicago, Illinois
| |
Collapse
|
28
|
Chipont A, Esposito B, Challier I, Montabord M, Tedgui A, Mallat Z, Loyer X, Potteaux S. MicroRNA-21 Deficiency Alters the Survival of Ly-6C
lo
Monocytes in
ApoE
−/−
Mice and Reduces Early-Stage Atherosclerosis—Brief Report. Arterioscler Thromb Vasc Biol 2019; 39:170-177. [DOI: 10.1161/atvbaha.118.311942] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/24/2022]
Abstract
Objective—
To determine the role of microRNA-21 (miR-21) on the homeostasis of monocyte subsets and on atherosclerosis development in
ApoE
−/−
(apolipoprotein E) mice.
Approach and Results—
In
ApoE
−/−
mice, miR-21 expression was increased in circulating Ly-6C
lo
nonclassical monocytes in comparison to Ly-6C
hi
monocytes. The absence of miR-21 significantly altered the survival and number of circulating Ly-6C
lo
nonclassical monocytes in
ApoE
−/−
mice. In the early stages of atherosclerosis, the absence of miR-21 limited lesion development both in the aortic sinus (by almost 30%) and in the aorta (by almost 50%). This was associated with less monocyte availability in circulation and increased apoptosis of local macrophages in plaques. At later stages of atherosclerosis, lesion size in the aortic root was similar in
ApoE
−/−
and
ApoE
−/−
miR-21
−/−
mice, but plaques showed a less stable phenotype (larger necrotic cores) in the latter. The loss of protection in advanced stages was most likely because of excessive inflammatory apoptosis related to an impairment of local efficient efferocytosis.
Conclusions—
Gene deletion of miR-21 in
ApoE
−/−
mice alters Ly-6C
lo
nonclassical monocytes homeostasis and contribute to limit early-stage atherosclerosis.
Collapse
Affiliation(s)
- Anna Chipont
- From the Inserm U970, Paris Cardiovascular Research Center (PARCC), Université René Descartes Paris 5, France
| | - Bruno Esposito
- From the Inserm U970, Paris Cardiovascular Research Center (PARCC), Université René Descartes Paris 5, France
| | - Inès Challier
- From the Inserm U970, Paris Cardiovascular Research Center (PARCC), Université René Descartes Paris 5, France
| | - Mélanie Montabord
- From the Inserm U970, Paris Cardiovascular Research Center (PARCC), Université René Descartes Paris 5, France
| | - Alain Tedgui
- From the Inserm U970, Paris Cardiovascular Research Center (PARCC), Université René Descartes Paris 5, France
| | - Ziad Mallat
- From the Inserm U970, Paris Cardiovascular Research Center (PARCC), Université René Descartes Paris 5, France
| | - Xavier Loyer
- From the Inserm U970, Paris Cardiovascular Research Center (PARCC), Université René Descartes Paris 5, France
| | - Stephane Potteaux
- From the Inserm U970, Paris Cardiovascular Research Center (PARCC), Université René Descartes Paris 5, France
| |
Collapse
|
29
|
Khorasani M, Teimoori-Toolabi L, Farivar TN, Asgari M, Abolhasani M, Shahrokh H, Afgar A, Kalantari E, Peymani A, Mahdian R. Aberrant expression of miR-141 and nuclear receptor small heterodimer partner in clinical samples of prostate cancer. Cancer Biomark 2018; 22:19-28. [DOI: 10.3233/cbm-170696] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022]
Affiliation(s)
- Maryam Khorasani
- Department of Molecular Medicine, School of Medicine, Qazvin University of Medical Sciences, Qazvin, Iran
| | | | | | - Mojgan Asgari
- Oncopathology Research Center, Iran University of Medical Sciences, Tehran, Iran
- Department of Pathology, Hasheminejad Kidney Center, Iran University of Medical Sciences, Tehran, Iran
| | - Maryam Abolhasani
- Oncopathology Research Center, Iran University of Medical Sciences, Tehran, Iran
- Department of Pathology, Hasheminejad Kidney Center, Iran University of Medical Sciences, Tehran, Iran
| | - Hossein Shahrokh
- Department of Uro-oncology, Hasheminejad Kidney Center, Iran University of Medical Sciences, Tehran, Iran
| | - Ali Afgar
- Molecular Medicine Department, Pasteur Institute of Iran, Tehran, Iran
| | - Elham Kalantari
- Oncopathology Research Center, Iran University of Medical Sciences, Tehran, Iran
| | - Amir Peymani
- Medical Microbiology Research Center, Qazvin University of Medical Sciences, Qazvin, Iran
| | - Reza Mahdian
- Molecular Medicine Department, Pasteur Institute of Iran, Tehran, Iran
| |
Collapse
|
30
|
Identification of valid reference genes for mRNA and microRNA normalisation in prostate cancer cell lines. Sci Rep 2018; 8:1949. [PMID: 29386530 PMCID: PMC5792445 DOI: 10.1038/s41598-018-19458-z] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/26/2016] [Accepted: 12/14/2017] [Indexed: 01/10/2023] Open
Abstract
RT-qPCR offers high sensitivity, for accurate interpretations of qPCR results however, normalisation using suitable reference genes is fundamental. Androgens can regulate transcriptional expression including reference gene expression in prostate cancer. In this study, we evaluated ten mRNA and six non-protein coding RNA reference genes in five prostate cell lines under varied dihydrotestosterone (DHT) treatments. We validated the effects of DHT-treatments using media containing charcoal-stripped serum prior to DHT stimulation on the test samples by Western blot experiments. Reference gene expression stability was analysed using three programs (geNorm, NormFinder and BestKeeper), and the recommended comprehensive ranking is provided. Our results reveal that ACTB and GAPDH, and miR-16 and miR-1228-3p are the most suitable mRNA and miRNA reference genes across all cell lines, respectively. Considering prostate cancer cell types, ACTB/GAPDH and ACTB/HPRT1 are the most suitable reference gene combinations for mRNA analysis, and miR-16/miR-1228-3p and RNU6-2/RNU43 for miRNA analysis in AR+, and AR− and normal cell lines, respectively. Comparison of relative target gene (PCA3 and miR-141) expression reveals different patterns depending on reference genes used for normalisation. To our knowledge, this is the first report on validation of reference genes under different DHT treatments in prostate cancer cells. This study provides insights for discovery of reliable DHT-regulated genes in prostate cells.
Collapse
|
31
|
miR-331-3p and Aurora Kinase inhibitor II co-treatment suppresses prostate cancer tumorigenesis and progression. Oncotarget 2017; 8:55116-55134. [PMID: 28903407 PMCID: PMC5589646 DOI: 10.18632/oncotarget.18664] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/26/2017] [Accepted: 05/22/2017] [Indexed: 01/10/2023] Open
Abstract
RNA-based therapeutics could represent a new avenue of cancer treatment. miRNA 331-3p (miR-331-3p) is implicated in prostate cancer (PCa) as a putative tumor suppressor, but its functional activity and synergy with other anti-tumor agents is largely unknown. We found miR-331-3p expression in PCa tumors was significantly decreased compared to non-malignant matched tissue. Analysis of publicly available PCa gene expression data sets showed miR-331-3p expression negatively correlated with Gleason Score, tumor stage, lymph node involvement and PSA value, and was significantly down regulated in tumor tissue relative to normal prostate tissue. Overexpression of miR-331-3p reduced PCa cell growth, migration and colony formation, as well as xenograft tumor initiation, proliferation and survival of mice. Microarray analysis identified seven novel targets of miR-331-3p in PCa. The 3’-untranslated regions of PLCγ1 and RALA were confirmed as targets of miR-331-3p, with mutation analyses confirming RALA as a direct target. Expression of miR-331-3p or RALA siRNA in PCa cells reduced RALA expression, proliferation, migration and colony formation in vitro. RALA expression positively correlated with Gleason grade in two separate studies, as well as in a PCa tissue microarray. Co-treatment using siRALA with an Aurora Kinase inhibitor (AKi-II) decreased colony formation of PCa cells while the combination of AKi-II with miR-331-3p resulted in significant reduction of PCa cell proliferation in vitro and PCa xenograft growth in vivo. Thus, miR-331-3p directly targets the RALA pathway and the addition of the AKi-II has a synergistic effect on tumor growth inhibition, suggesting a potential role as combination therapy in PCa.
Collapse
|
32
|
Nabavi N, Saidy NRN, Venalainen E, Haegert A, Parolia A, Xue H, Wang Y, Wu R, Dong X, Collins C, Crea F, Wang Y. miR-100-5p inhibition induces apoptosis in dormant prostate cancer cells and prevents the emergence of castration-resistant prostate cancer. Sci Rep 2017; 7:4079. [PMID: 28642484 PMCID: PMC5481412 DOI: 10.1038/s41598-017-03731-8] [Citation(s) in RCA: 38] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/03/2017] [Accepted: 05/11/2017] [Indexed: 12/16/2022] Open
Abstract
Carcinoma of the prostate is the most common cancer in men. Treatment of aggressive prostate cancer involves a regiment of radical prostectomy, radiation therapy, chemotherapy and hormonal therapy. Despite significant improvements in the last decade, the treatment of prostate cancer remains unsatisfactory, because a significant fraction of prostate cancers develop resistance to multiple treatments and become incurable. This prompts an urgent need to investigate the molecular mechanisms underlying the evolution of therapy-induced resistance of prostate cancer either in the form of castration-resistant prostate cancer (CRPC) or transdifferentiated neuroendocrine prostate cancer (NEPC). By analyzing micro-RNA expression profiles in a set of patient-derived prostate cancer xenograft tumor lines, we identified miR-100-5p as one of the key molecular components in the initiation and evolution of androgen ablation therapy resistance in prostate cancer. In vitro results showed that miR-100-5p is required for hormone-independent survival and proliferation of prostate cancer cells post androgen ablation. In Silico target predictions revealed that miR-100-5p target genes are involved in key aspects of cancer progression, and are associated with clinical outcome. Our results suggest that mir-100-5p is a possible therapeutic target involved in prostate cancer progression and relapse post androgen ablation therapy.
Collapse
Affiliation(s)
- Noushin Nabavi
- Experimental Therapeutics, BC Cancer Research Centre, Vancouver, BC, Canada
- The Vancouver Prostate Centre, Vancouver General Hospital, Vancouver, BC, Canada
- Department of Urologic Sciences, University of British Columbia, Vancouver, BC, Canada
| | - Nur Ridzwan Nur Saidy
- Honors Biotechnology Program, Department of Microbiology and Immunology, University of British Columbia, 2329 West Mall, Vancouver, BC, V6T 1Z4, Canada
- Experimental Therapeutics, BC Cancer Research Centre, Vancouver, BC, Canada
| | - Erik Venalainen
- Honors Biotechnology Program, Department of Microbiology and Immunology, University of British Columbia, 2329 West Mall, Vancouver, BC, V6T 1Z4, Canada
- Experimental Therapeutics, BC Cancer Research Centre, Vancouver, BC, Canada
| | - Anne Haegert
- The Vancouver Prostate Centre, Vancouver General Hospital, Vancouver, BC, Canada
| | - Abhijit Parolia
- Honors Biotechnology Program, Department of Microbiology and Immunology, University of British Columbia, 2329 West Mall, Vancouver, BC, V6T 1Z4, Canada
- Experimental Therapeutics, BC Cancer Research Centre, Vancouver, BC, Canada
| | - Hui Xue
- Experimental Therapeutics, BC Cancer Research Centre, Vancouver, BC, Canada
- The Vancouver Prostate Centre, Vancouver General Hospital, Vancouver, BC, Canada
| | - Yuwei Wang
- Experimental Therapeutics, BC Cancer Research Centre, Vancouver, BC, Canada
- The Vancouver Prostate Centre, Vancouver General Hospital, Vancouver, BC, Canada
| | - Rebecca Wu
- Experimental Therapeutics, BC Cancer Research Centre, Vancouver, BC, Canada
- The Vancouver Prostate Centre, Vancouver General Hospital, Vancouver, BC, Canada
| | - Xin Dong
- Experimental Therapeutics, BC Cancer Research Centre, Vancouver, BC, Canada
| | - Colin Collins
- The Vancouver Prostate Centre, Vancouver General Hospital, Vancouver, BC, Canada
| | - Francesco Crea
- Experimental Therapeutics, BC Cancer Research Centre, Vancouver, BC, Canada.
- School of Life, Health and Chemical Sciences, The Open University, Milton Keynes, UK.
| | - Yuzhuo Wang
- Experimental Therapeutics, BC Cancer Research Centre, Vancouver, BC, Canada.
- The Vancouver Prostate Centre, Vancouver General Hospital, Vancouver, BC, Canada.
- Department of Urologic Sciences, University of British Columbia, Vancouver, BC, Canada.
| |
Collapse
|
33
|
Zavala E, Reyes D, Deerenberg R, Vidal R. Selection of reference genes for microRNA analysis associated to early stress response to handling and confinement in Salmo salar. Sci Rep 2017; 7:1756. [PMID: 28496155 PMCID: PMC5431957 DOI: 10.1038/s41598-017-01970-3] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/24/2017] [Accepted: 04/05/2017] [Indexed: 12/31/2022] Open
Abstract
MicroRNAs are key non-coding RNA molecules that play a relevant role in the regulation of gene expression through translational repression and/or transcript cleavage during normal development and physiological adaptation processes like stress. Quantitative reverse transcription polymerase chain reaction (RT-qPCR) has become the approach normally used to determine the levels of microRNAs. However, this approach needs the use of endogenous reference. An improper selection of endogenous references can result in confusing interpretation of data. The aim of this study was to identify and validate appropriate endogenous reference miRNA genes for normalizing RT-qPCR survey of miRNAs expression in four different tissues of Atlantic salmon, under handling and confinement stress conditions associated to early or primary stress response. Nine candidate reference normalizers, including microRNAs and nuclear genes, normally used in vertebrate microRNA expression studies were selected from literature, validated by RT-qPCR and analyzed by the algorithms geNorm and NormFinder. The results revealed that the ssa-miR-99-5p gene was the most stable overall and that ssa-miR-99-5p and ssa-miR-23a-5p genes were the best combination. Moreover, the suitability of ssa-miR-99-5p and ssa-miR-23a-5p as endogeneuos reference genes was demostrated by the expression analysis of ssa-miR-193-5p gene.
Collapse
Affiliation(s)
- Eduardo Zavala
- Department of Biology, Universidad de Santiago de Chile. Av. Libertador Bernardo O'Higgins 3363, Santiago, Chile
| | - Daniela Reyes
- Department of Biology, Universidad de Santiago de Chile. Av. Libertador Bernardo O'Higgins 3363, Santiago, Chile
| | | | - Rodrigo Vidal
- Department of Biology, Universidad de Santiago de Chile. Av. Libertador Bernardo O'Higgins 3363, Santiago, Chile.
| |
Collapse
|
34
|
Anauate AC, Leal MF, Wisnieski F, Santos LC, Gigek CO, Chen ES, Geraldis JC, Calcagno DQ, Assumpção PP, Demachki S, Arasaki CH, Lourenço LG, Artigiani R, Burbano RR, Smith MAC. Identification of suitable reference genes for miRNA expression normalization in gastric cancer. Gene 2017; 621:59-68. [PMID: 28411081 DOI: 10.1016/j.gene.2017.04.016] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/15/2017] [Revised: 03/28/2017] [Accepted: 04/10/2017] [Indexed: 12/13/2022]
Affiliation(s)
- Ana Carolina Anauate
- Disciplina de Genética, Departamento de Morfologia e Genética, Universidade Federal de São Paulo, São Paulo, Brazil
| | - Mariana Ferreira Leal
- Disciplina de Genética, Departamento de Morfologia e Genética, Universidade Federal de São Paulo, São Paulo, Brazil; Departamento de Ortopedia e Traumatologia, Universidade Federal de São Paulo, São Paulo, Brazil; Núcleo de Pesquisas em Oncologia, Hospital Universitário João de Barros Barreto, Universidade Federal do Pará, Belém, Brazil
| | - Fernanda Wisnieski
- Disciplina de Genética, Departamento de Morfologia e Genética, Universidade Federal de São Paulo, São Paulo, Brazil
| | - Leonardo Caires Santos
- Disciplina de Genética, Departamento de Morfologia e Genética, Universidade Federal de São Paulo, São Paulo, Brazil
| | - Carolina Oliveira Gigek
- Disciplina de Genética, Departamento de Morfologia e Genética, Universidade Federal de São Paulo, São Paulo, Brazil; Disciplina de Gastroenterologia Cirúrgica, Departamento de Cirurgia, Universidade Federal de São Paulo, São Paulo, Brazil
| | - Elizabeth Suchi Chen
- Disciplina de Genética, Departamento de Morfologia e Genética, Universidade Federal de São Paulo, São Paulo, Brazil
| | - Jaqueline Cruz Geraldis
- Disciplina de Genética, Departamento de Morfologia e Genética, Universidade Federal de São Paulo, São Paulo, Brazil
| | - Danielle Queiroz Calcagno
- Núcleo de Pesquisas em Oncologia, Hospital Universitário João de Barros Barreto, Universidade Federal do Pará, Belém, Brazil
| | - Paulo Pimentel Assumpção
- Núcleo de Pesquisas em Oncologia, Hospital Universitário João de Barros Barreto, Universidade Federal do Pará, Belém, Brazil
| | - Samia Demachki
- Núcleo de Pesquisas em Oncologia, Hospital Universitário João de Barros Barreto, Universidade Federal do Pará, Belém, Brazil
| | - Carlos Haruo Arasaki
- Disciplina de Gastroenterologia Cirúrgica, Departamento de Cirurgia, Universidade Federal de São Paulo, São Paulo, Brazil
| | - Laércio Gomes Lourenço
- Disciplina de Gastroenterologia Cirúrgica, Departamento de Cirurgia, Universidade Federal de São Paulo, São Paulo, Brazil
| | - Ricardo Artigiani
- Departamento de Patologia, Universidade Federal de São Paulo, São Paulo, Brazil
| | - Rommel Rodríguez Burbano
- Núcleo de Pesquisas em Oncologia, Hospital Universitário João de Barros Barreto, Universidade Federal do Pará, Belém, Brazil; Laboratório de Citogenética Humana, Instituto de Ciências Biológicas, Universidade Federal do Pará, Belém, Brazil; Laboratório de Biologia Molecular, Hospital Ophir Loyola, Belém, Brazil
| | - Marília Arruda Cardoso Smith
- Disciplina de Genética, Departamento de Morfologia e Genética, Universidade Federal de São Paulo, São Paulo, Brazil.
| |
Collapse
|
35
|
Kao HW, Pan CY, Lai CH, Wu CW, Fang WL, Huang KH, Lin WC. Urine miR-21-5p as a potential non-invasive biomarker for gastric cancer. Oncotarget 2017; 8:56389-56397. [PMID: 28915598 PMCID: PMC5593569 DOI: 10.18632/oncotarget.16916] [Citation(s) in RCA: 52] [Impact Index Per Article: 6.5] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/08/2016] [Accepted: 03/14/2017] [Indexed: 12/12/2022] Open
Abstract
Many reports have implicated that microRNAs involve in cancer development and progression, such as miR-155 in breast cancers and miR-196 in gastric cancers. Furthermore, microRNAs are more stable than typical protein-coding gene mRNAs in varieties of clinical samples including body fluids. This suggests that they are potentially valuable biomarkers for cancer monitoring. In this study, we have used urine samples of gastric cancer patients to demonstrate the feasibility of urine microRNAs for gastric cancer detection. Urine samples of gastric cancer patients were extracted for total RNA, which were examined for the expression of miR-21-5p using quantitative stem-loop PCR. Our results demonstrated that miR-21-5p could be detected in small amounts of urine samples with good stability, and the expression levels of miR-21-5p were reduced following surgical removal of gastric cancer tissues. These results implicate that urine miR-21-5p could be utilized as a novel non-invasive biomarker of gastric cancer detection and monitoring.
Collapse
Affiliation(s)
- Hsiao-Wei Kao
- Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan
| | - Chao-Yu Pan
- Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan.,Institute of Biomedical Informatics, National Yang-Ming University, Taipei, Taiwan
| | - Chun-Hung Lai
- Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan
| | - Chew-Wun Wu
- Department of Surgery, Veterans General Hospital and National Yang-Ming University, Taipei, Taiwan
| | - Wen-Liang Fang
- Department of Surgery, Veterans General Hospital and National Yang-Ming University, Taipei, Taiwan
| | - Kuo-Hung Huang
- Department of Surgery, Veterans General Hospital and National Yang-Ming University, Taipei, Taiwan
| | - Wen-Chang Lin
- Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan.,Institute of Biomedical Informatics, National Yang-Ming University, Taipei, Taiwan
| |
Collapse
|
36
|
Malla B, Zaugg K, Vassella E, Aebersold DM, Dal Pra A. Exosomes and Exosomal MicroRNAs in Prostate Cancer Radiation Therapy. Int J Radiat Oncol Biol Phys 2017; 98:982-995. [PMID: 28721912 DOI: 10.1016/j.ijrobp.2017.03.031] [Citation(s) in RCA: 55] [Impact Index Per Article: 6.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/19/2016] [Revised: 03/14/2017] [Accepted: 03/20/2017] [Indexed: 12/11/2022]
Abstract
Despite current risk stratification systems using traditional clinicopathologic factors, many localized and locally advanced prostate cancers fail radical treatment (ie, radical prostatectomy, radiation therapy with or without androgen deprivation therapy). Therefore, a pressing need exists for enhanced methods of disease stratification through novel prognostic and predictive tools that can reliably be applied in clinical practice. Exosomes are 50- to 150-nm small vesicles released by cancer cells that reflect the genetic and nongenetic materials of parent cancer cells. Cancer cells can contain distinct sets of microRNA profiles, the expression of which can change owing to stress such as radiation therapy. These alterations or distinctions in contents allow exosomes to be used as prognostic and/or predictive biomarkers and to monitor the treatment response. Additionally, microRNAs have been shown to influence multiple processes in prostate tumorigenesis, including cell proliferation, induction of apoptosis, migration, oncogene inhibition, and radioresistance. Thus, comparative exosomal microRNA profiling at different levels could help portray tumor aggressiveness and response to radiation therapy. Although technical challenges persist in exosome isolation and characterization, recent improvements in microRNA profiling have evolved toward in-depth analyses of the exosomal cargo and its functions. We have reviewed the role of exosomes and exosomal microRNAs in biologic processes of prostate cancer progression and radiation therapy response, with a particular focus on the development of clinical assays for treatment personalization.
Collapse
Affiliation(s)
- Bijaya Malla
- Department of Radiation Oncology, Bern University Hospital, Inselspital, Bern, Switzerland
| | - Kathrin Zaugg
- Department of Radiation Oncology, Bern University Hospital, Inselspital, Bern, Switzerland
| | - Erik Vassella
- Institute of Pathology, University of Bern, Bern, Switzerland
| | - Daniel M Aebersold
- Department of Radiation Oncology, Bern University Hospital, Inselspital, Bern, Switzerland
| | - Alan Dal Pra
- Department of Radiation Oncology, Bern University Hospital, Inselspital, Bern, Switzerland.
| |
Collapse
|
37
|
Masè M, Grasso M, Avogaro L, D’Amato E, Tessarolo F, Graffigna A, Denti MA, Ravelli F. Selection of reference genes is critical for miRNA expression analysis in human cardiac tissue. A focus on atrial fibrillation. Sci Rep 2017; 7:41127. [PMID: 28117343 PMCID: PMC5259703 DOI: 10.1038/srep41127] [Citation(s) in RCA: 69] [Impact Index Per Article: 8.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/25/2016] [Accepted: 12/15/2016] [Indexed: 12/23/2022] Open
Abstract
MicroRNAs (miRNAs) are emerging as key regulators of complex biological processes in several cardiovascular diseases, including atrial fibrillation (AF). Reverse transcription-quantitative polymerase chain reaction is a powerful technique to quantitatively assess miRNA expression profile, but reliable results depend on proper data normalization by suitable reference genes. Despite the increasing number of studies assessing miRNAs in cardiac disease, no consensus on the best reference genes has been reached. This work aims to assess reference genes stability in human cardiac tissue with a focus on AF investigation. We evaluated the stability of five reference genes (U6, SNORD48, SNORD44, miR-16, and 5S) in atrial tissue samples from eighteen cardiac-surgery patients in sinus rhythm and AF. Stability was quantified by combining BestKeeper, delta-Cq, GeNorm, and NormFinder statistical tools. All methods assessed SNORD48 as the best and U6 as the worst reference gene. Applications of different normalization strategies significantly impacted miRNA expression profiles in the study population. Our results point out the necessity of a consensus on data normalization in AF studies to avoid the emergence of divergent biological conclusions.
Collapse
Affiliation(s)
- Michela Masè
- Department of Physics, University of Trento, Trento, Italy
| | | | - Laura Avogaro
- Department of Physics, University of Trento, Trento, Italy
- Centre for Integrative Biology, University of Trento, Trento, Italy
| | - Elvira D’Amato
- Department of Physics, University of Trento, Trento, Italy
| | - Francesco Tessarolo
- Healthcare Research and Innovation Program (IRCS-PAT), Bruno Kessler Foundation, Trento, Italy
- Department of Industrial Engineering, University of Trento, Trento, Italy
| | - Angelo Graffigna
- Division of Cardiac Surgery, Santa Chiara Hospital, Trento, Italy
| | | | - Flavia Ravelli
- Department of Physics, University of Trento, Trento, Italy
| |
Collapse
|
38
|
Akbayır S, Muşlu N, Erden S, Bozlu M. Diagnostic value of microRNAs in prostate cancer patients with prostate specific antigen (PSA) levels between 2, and 10 ng/mL. Turk J Urol 2016; 42:247-255. [PMID: 27909617 DOI: 10.5152/tud.2016.52463] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022]
Abstract
OBJECTIVE Prostate specific antigen (PSA), used for the early diagnosis of prostate cancer (PCa), is one of the best tumour markers known so far. However, in situations when PSA is between 2-10 ng/mL, which is named as grey zone, PSA falls short of distinguishing benign prostate diseases from PCa. On the other hand, it was demonstrated in many previous studies that microRNA (miRNA) could be a marker for cancer. Therefore, in this study, it was aimed to enhance the diagnostic power of PSA, especially with grey zone patients, by the use of miRNA. MATERIAL AND METHODS Ninety-four patients included in the study were divided into three groups as "control group" (n=44, PSA=2-10 ng/mL and benign), "PCa 1 group" (n=37, PSA=2-10 ng/mL), and "PCa 2 group" (n=13, PSA >10 ng/mL), according to their pathological results and PSA levels. Free PSA (fPSA) and total PSA (T-PSA) levels were measured with chemiluminometric sandwich immunoassay method. Expressions of miRNAs were analyzed using quantitative reverse transcription-polymerase chain reaction (qRT-PCR) method. The most appropriate specificity, sensitivity and prediction values were found by drawing the receiver operating characteristic (ROC) curves of total PSA, free/total PSA (f/T PSA) ratio, and miRNAs, and the diagnostic powers were compared with each other. RESULTS Diagnostic powers of the f/T PSA ratio and miRNA were compared in PCa 1 and the control groups to determine the marker with higher area under the curve (AUC). It was shown that the diagnostic power of the combination of miR-16-5p and f/T PSA was higher than that obtained when they were used separately. CONCLUSION As a result, while making the the discrimination between benign and malignant prostate in patients with grey zone, it was determined that the combination of miR-16-5p and f/T PSA was more valuable than T-PSA or f/T PSA alone. It was thought that diagnostic role of miRNAs in the early diagnosis of the different stages of PCa needed to be examined in further studies with larger groups.
Collapse
Affiliation(s)
- Serin Akbayır
- Karaman State Hospital, Biochemistry Laboratory, Karaman, Turkey
| | - Necati Muşlu
- Department of Biochemistry, Mersin University School of Medicine, Mersin, Turkey
| | - Sema Erden
- Mersin University, Vocational School of Health Services, Mersin, Turkey
| | - Murat Bozlu
- Department of Urology, Mersin University School of Medicine, Mersin, Turkey
| |
Collapse
|
39
|
Loyer X, Paradis V, Hénique C, Vion AC, Colnot N, Guerin CL, Devue C, On S, Scetbun J, Romain M, Paul JL, Rothenberg ME, Marcellin P, Durand F, Bedossa P, Prip-Buus C, Baugé E, Staels B, Boulanger CM, Tedgui A, Rautou PE. Liver microRNA-21 is overexpressed in non-alcoholic steatohepatitis and contributes to the disease in experimental models by inhibiting PPARα expression. Gut 2016; 65:1882-1894. [PMID: 26338827 PMCID: PMC5099209 DOI: 10.1136/gutjnl-2014-308883] [Citation(s) in RCA: 130] [Impact Index Per Article: 14.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/24/2014] [Revised: 08/09/2015] [Accepted: 08/13/2015] [Indexed: 12/13/2022]
Abstract
OBJECTIVE Previous studies suggested that microRNA-21 may be upregulated in the liver in non-alcoholic steatohepatitis (NASH), but its role in the development of this disease remains unknown. This study aimed to determine the role of microRNA-21 in NASH. DESIGN We inhibited or suppressed microRNA-21 in different mouse models of NASH: (a) low-density lipoprotein receptor-deficient (Ldlr-/-) mice fed a high-fat diet and treated with antagomir-21 or antagomir control; (b) microRNA-21-deficient and wild-type mice fed a methionine-choline-deficient (MCD) diet; (c) peroxisome proliferation-activator receptor α (PPARα)-deficient mice fed an MCD diet and treated with antagomir-21 or antagomir control. We assessed features of NASH and determined liver microRNA-21 levels and cell localisation. MicroRNA-21 levels were also quantified in the liver of patients with NASH, bland steatosis or normal liver and localisation was determined. RESULTS Inhibiting or suppressing liver microRNA-21 expression reduced liver cell injury, inflammation and fibrogenesis without affecting liver lipid accumulation in Ldlr-/- fed a high-fat diet and in wild-type mice fed an MCD diet. Liver microRNA-21 was overexpressed, primarily in biliary and inflammatory cells, in mouse models as well as in patients with NASH, but not in patients with bland steatosis. PPARα, a known microRNA-21 target, implicated in NASH, was decreased in the liver of mice with NASH and restored following microRNA-21 inhibition or suppression. The effect of antagomir-21 was lost in PPARα-deficient mice. CONCLUSIONS MicroRNA-21 inhibition or suppression decreases liver injury, inflammation and fibrosis, by restoring PPARα expression. Antagomir-21 might be a future therapeutic strategy for NASH.
Collapse
Affiliation(s)
- Xavier Loyer
- INSERM, U970, Paris Cardiovascular Research Center—PARCC, Paris, France,Université Paris Descartes, Sorbonne Paris Cité, Paris, France
| | - Valérie Paradis
- Service d'Anatomie Pathologique, Hôpital Beaujon, Assistance Publique-Hôpitaux de Paris, Clichy, France,INSERM, U773, Centre de Recherche Biomédicale Bichat-Beaujon CRB3, Clichy, France,Université Denis Diderot-Paris 7, Sorbonne Paris Cité, Paris, France
| | - Carole Hénique
- INSERM, U970, Paris Cardiovascular Research Center—PARCC, Paris, France,Université Paris Descartes, Sorbonne Paris Cité, Paris, France
| | - Anne-Clémence Vion
- INSERM, U970, Paris Cardiovascular Research Center—PARCC, Paris, France,Université Paris Descartes, Sorbonne Paris Cité, Paris, France
| | - Nathalie Colnot
- Service d'Anatomie Pathologique, Hôpital Beaujon, Assistance Publique-Hôpitaux de Paris, Clichy, France
| | - Coralie L Guerin
- INSERM, U970, Paris Cardiovascular Research Center—PARCC, Paris, France,Université Paris Descartes, Sorbonne Paris Cité, Paris, France
| | - Cécile Devue
- INSERM, U970, Paris Cardiovascular Research Center—PARCC, Paris, France,Université Paris Descartes, Sorbonne Paris Cité, Paris, France
| | - Sissi On
- INSERM, U970, Paris Cardiovascular Research Center—PARCC, Paris, France,Université Paris Descartes, Sorbonne Paris Cité, Paris, France
| | - Jérémy Scetbun
- INSERM, U970, Paris Cardiovascular Research Center—PARCC, Paris, France,Université Paris Descartes, Sorbonne Paris Cité, Paris, France
| | - Mélissa Romain
- INSERM, U970, Paris Cardiovascular Research Center—PARCC, Paris, France,Université Paris Descartes, Sorbonne Paris Cité, Paris, France
| | - Jean-Louis Paul
- Service de Biochimie, Hôpital Européen Georges Pompidou, AP-HP (Assistance Publique-Hôpitaux de Paris), Paris, France
| | - Marc E Rothenberg
- Division of Allergy and Immunology, Department of Pediatrics, Cincinnati Children's Hospital Medical Center, University of Cincinnati College of Medicine, Cincinnati, Ohio, USA
| | - Patrick Marcellin
- INSERM, U773, Centre de Recherche Biomédicale Bichat-Beaujon CRB3, Clichy, France,Université Denis Diderot-Paris 7, Sorbonne Paris Cité, Paris, France,Service d'Hépatologie, Hôpital Beaujon, Assistance Publique-Hôpitaux de Paris, Clichy, France
| | - François Durand
- INSERM, U773, Centre de Recherche Biomédicale Bichat-Beaujon CRB3, Clichy, France,Université Denis Diderot-Paris 7, Sorbonne Paris Cité, Paris, France,Service d'Hépatologie, Hôpital Beaujon, Assistance Publique-Hôpitaux de Paris, Clichy, France
| | - Pierre Bedossa
- Service d'Anatomie Pathologique, Hôpital Beaujon, Assistance Publique-Hôpitaux de Paris, Clichy, France,INSERM, U773, Centre de Recherche Biomédicale Bichat-Beaujon CRB3, Clichy, France,Université Denis Diderot-Paris 7, Sorbonne Paris Cité, Paris, France
| | - Carina Prip-Buus
- Université Paris Descartes, Sorbonne Paris Cité, Paris, France,INSERM, U1016, Institut Cochin, Paris, France,CNRS, UMR8104, Paris, France
| | - Eric Baugé
- Univ. Lille, Inserm, Institut Pasteur de Lille, U1011—EGID, Lille, France
| | - Bart Staels
- Univ. Lille, Inserm, Institut Pasteur de Lille, U1011—EGID, Lille, France
| | - Chantal M Boulanger
- INSERM, U970, Paris Cardiovascular Research Center—PARCC, Paris, France,Université Paris Descartes, Sorbonne Paris Cité, Paris, France
| | - Alain Tedgui
- INSERM, U970, Paris Cardiovascular Research Center—PARCC, Paris, France,Université Paris Descartes, Sorbonne Paris Cité, Paris, France
| | - Pierre-Emmanuel Rautou
- INSERM, U970, Paris Cardiovascular Research Center—PARCC, Paris, France,Université Paris Descartes, Sorbonne Paris Cité, Paris, France,Université Denis Diderot-Paris 7, Sorbonne Paris Cité, Paris, France,Service d'Hépatologie, Hôpital Beaujon, Assistance Publique-Hôpitaux de Paris, Clichy, France
| |
Collapse
|
40
|
Koenig EM, Fisher C, Bernard H, Wolenski FS, Gerrein J, Carsillo M, Gallacher M, Tse A, Peters R, Smith A, Meehan A, Tirrell S, Kirby P. The beagle dog MicroRNA tissue atlas: identifying translatable biomarkers of organ toxicity. BMC Genomics 2016; 17:649. [PMID: 27535741 PMCID: PMC4989286 DOI: 10.1186/s12864-016-2958-x] [Citation(s) in RCA: 47] [Impact Index Per Article: 5.2] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/06/2015] [Accepted: 07/21/2016] [Indexed: 01/28/2023] Open
Abstract
Background MicroRNAs (miRNA) are varied in length, under 25 nucleotides, single-stranded noncoding RNA that regulate post-transcriptional gene expression via translational repression or mRNA degradation. Elevated levels of miRNAs can be detected in systemic circulation after tissue injury, suggesting that miRNAs are released following cellular damage. Because of their remarkable stability, ease of detection in biofluids, and tissue specific expression patterns, miRNAs have the potential to be specific biomarkers of organ injury. The identification of miRNA biomarkers requires a systematic approach: 1) determine the miRNA tissue expression profiles within a mammalian species via next generation sequencing; 2) identify enriched and/or specific miRNA expression within organs of toxicologic interest, and 3) in vivo validation with tissue-specific toxicants. While miRNA tissue expression has been reported in rodents and humans, little data exists on miRNA tissue expression in the dog, a relevant toxicology species. The generation and evaluation of the first dog miRNA tissue atlas is described here. Results Analysis of 16 tissues from five male beagle dogs identified 106 tissue enriched miRNAs, 60 of which were highly enriched in a single organ, and thus may serve as biomarkers of organ injury. A proof of concept study in dogs dosed with hepatotoxicants evaluated a qPCR panel of 15 tissue enriched miRNAs specific to liver, heart, skeletal muscle, pancreas, testes, and brain. Dogs with elevated serum levels of miR-122 and miR-885 had a correlative increase of alanine aminotransferase, and microscopic analysis confirmed liver damage. Other non-liver enriched miRNAs included in the screening panel were unaffected. Eli Lilly authors created a complimentary Sprague Dawely rat miRNA tissue atlas and demonstrated increased pancreas enriched miRNA levels in circulation, following caerulein administration in rat and dog. Conclusion The dog miRNA tissue atlas provides a resource for biomarker discovery and can be further mined with refinement of dog genome annotation. The 60 highly enriched tissue miRNAs identified within the dog miRNA tissue atlas could serve as diagnostic biomarkers and will require further validation by in vivo correlation to histopathology. Once validated, these tissue enriched miRNAs could be combined into a powerful qPCR screening panel to identify organ toxicity during early drug development. Electronic supplementary material The online version of this article (doi:10.1186/s12864-016-2958-x) contains supplementary material, which is available to authorized users.
Collapse
Affiliation(s)
- Erik M Koenig
- Takeda Pharmaceuticals International Co., 40 Landsdowne Street, Cambridge, MA, 02139, USA.
| | - Craig Fisher
- Takeda Pharmaceuticals International Co., 40 Landsdowne Street, Cambridge, MA, 02139, USA
| | - Hugues Bernard
- Takeda Pharmaceuticals International Co., 40 Landsdowne Street, Cambridge, MA, 02139, USA
| | - Francis S Wolenski
- Takeda Pharmaceuticals International Co., 40 Landsdowne Street, Cambridge, MA, 02139, USA
| | - Joseph Gerrein
- Takeda Pharmaceuticals International Co., 40 Landsdowne Street, Cambridge, MA, 02139, USA
| | - Mary Carsillo
- Takeda Pharmaceuticals International Co., 40 Landsdowne Street, Cambridge, MA, 02139, USA
| | - Matt Gallacher
- Takeda Pharmaceuticals International Co., 40 Landsdowne Street, Cambridge, MA, 02139, USA
| | - Aimy Tse
- Takeda Pharmaceuticals International Co., 40 Landsdowne Street, Cambridge, MA, 02139, USA
| | - Rachel Peters
- Takeda Pharmaceuticals International Co., 40 Landsdowne Street, Cambridge, MA, 02139, USA
| | - Aaron Smith
- Eli Lilly and Company, 893 S. Delaware, Indianapolis, IN, 46285, USA
| | - Alexa Meehan
- Takeda Pharmaceuticals International Co., 40 Landsdowne Street, Cambridge, MA, 02139, USA
| | - Stephen Tirrell
- Takeda Pharmaceuticals International Co., 40 Landsdowne Street, Cambridge, MA, 02139, USA
| | - Patrick Kirby
- Takeda Pharmaceuticals International Co., 40 Landsdowne Street, Cambridge, MA, 02139, USA
| |
Collapse
|
41
|
Liu Y, Lu Q. Extracellular vesicle microRNAs: biomarker discovery in various diseases based on RT-qPCR. Biomark Med 2016; 9:791-805. [PMID: 26287938 DOI: 10.2217/bmm.15.45] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/28/2022] Open
Abstract
In recent years, biomarker discovery based on extracellular microRNAs (miRNAs), especially exosome miRNAs, has drawn wide attention. While exosome isolation and identification technologies are increasingly sophisticated, the preanalytical process of exosome miRNAs seems to be no longer a crucial problem. Though next-generation sequencing, microarray and digital PCR have been recommended as good downstream analytical platforms for exosome miRNA quantification, they are still more constrained in clinical utility compared with RT-qPCR method at present. In this review, we will trace back to the origin and summarize current studies of biomarker discovery based on extracellular vesicle miRNAs, and provide an overview and latest developments of RT-qPCR-based data normalization, in order to further assist the development of translational medicine.
Collapse
Affiliation(s)
- Ying Liu
- Hunan Key Laboratory of Medical Epigenomics, Department of Dermatology, Second Xiangya Hospital, Central South University, #139 Renmin Middle Rd, Changsha, Hunan 410011, PR China
| | - Qianjin Lu
- Hunan Key Laboratory of Medical Epigenomics, Department of Dermatology, Second Xiangya Hospital, Central South University, #139 Renmin Middle Rd, Changsha, Hunan 410011, PR China
| |
Collapse
|
42
|
Stephan C, Jung M, Rabenhorst S, Kilic E, Jung K. Urinary miR-183 and miR-205 do not surpass PCA3 in urine as predictive markers for prostate biopsy outcome despite their highly dysregulated expression in prostate cancer tissue. Clin Chem Lab Med 2016; 53:1109-18. [PMID: 25720086 DOI: 10.1515/cclm-2014-1000] [Citation(s) in RCA: 18] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/10/2014] [Accepted: 11/30/2014] [Indexed: 12/24/2022]
Abstract
BACKGROUND MicroRNAs (miRNAs) have shown to be promising novel biomarkers in various cancers. We aimed to translate the results of an own previous tissue-based miRNA profile of prostate carcinoma (PCa) with upregulated miR-183 and downregulated miR-205 into a urine-based testing procedure for diagnosis of PCa. METHODS Urine sediments were prepared from urine samples collected after a standardized digital-rectal examination (DRE) of patients undergoing prostate biopsy with PSA (prostate-specific antigen) values <20 μg/L in consecutive order. According to the sample-size calculation (α=0.05, power=0.95), 38 patients each with PCa and without PCa were randomly enrolled in this study. PCA3 (prostate cancer associated 3) in urine as Food and Drug Administration-approved assay was determined as reference standard for comparison. The miRNAs were measured by RT-qPCR using TaqMan assays and normalized using different approaches. RESULTS Both miRNAs were correlated to the mRNA PSA concentrations in the sediments indicating a relationship to the released prostate cells after DRE. However, they had no discriminating capacity between patients with and without PCa. In contrast, PCA3 clearly differentiated between these two patients groups. There was also no significant correlation between miRNAs and standard clinicopathologic variables like Gleason score and serum PSA. CONCLUSIONS The data of our study show that miR-183 and miR-205 failed to detect early and aggressive PCa despite their highly dysregulated expression in cancer tissue. Our results and the critical evaluation of the few data of other studies raise serious doubts concerning the capability of urinary miRNAs to replace or improve PCA3 as predictive marker for prostate biopsy outcome.
Collapse
|
43
|
Plasma extracellular RNA profiles in healthy and cancer patients. Sci Rep 2016; 6:19413. [PMID: 26786760 PMCID: PMC4726401 DOI: 10.1038/srep19413] [Citation(s) in RCA: 209] [Impact Index Per Article: 23.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/27/2015] [Accepted: 12/11/2015] [Indexed: 12/15/2022] Open
Abstract
Extracellular vesicles are selectively enriched in RNA that has potential as disease biomarkers. To systemically characterize circulating extracellular RNA (exRNA) profiles, we performed RNA sequencing analysis on plasma extracellular vesicles derived from 50 healthy individuals and 142 cancer patients. Of ~12.6 million raw reads for each individual, the number of mappable reads aligned to RNA references was ~5.4 million including miRNAs (~40.4%), piwiRNAs (~40.0%), pseudo-genes (~3.7%), lncRNAs (~2.4%), tRNAs (~2.1%), and mRNAs (~2.1%). By expression stability testing, we identified a set of miRNAs showing relatively consistent expression, which may serve as reference control for exRNA quantification. By performing multivariate analysis of covariance, we identified significant associations of these exRNAs with age, sex and different types of cancers. In particular, down-regulation of miR-125a-5p and miR-1343-3p showed an association with all cancer types tested (false discovery rate <0.05). We developed multivariate statistical models to predict cancer status with an area under the curve from 0.68 to 0.92 depending cancer type and staging. This is the largest RNA-seq study to date for profiling exRNA species, which has not only provided a baseline reference profile for circulating exRNA, but also revealed a set of RNA candidates for reference controls and disease biomarkers.
Collapse
|
44
|
Circulating cell-free microRNAs as biomarkers for colorectal cancer. Surg Today 2016; 46:13-24. [PMID: 25712224 DOI: 10.1007/s00595-015-1138-y] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/14/2014] [Accepted: 02/11/2015] [Indexed: 02/07/2023]
Abstract
MicroRNAs (miRNAs) are a class of small, endogenous, non-coding, single-stranded RNAs that act as post-transcriptional regulators. Their discovery has provided new avenues for the diagnosis and treatment of cancer. The expression of both oncogenic and tumor suppressor miRNAs can be aberrantly either up- or down-regulated in cancer cells. These miRNAs target mRNAs of genes that either promote or inhibit tumor growth, and are one of several epigenetic factors that control the initiation and progression of colorectal cancer (CRC) and other cancers. Investigations of miRNAs as CRC biomarkers have employed the expression profiling of traditional tissue samples and, more recently, non-invasive samples, such as feces and body fluids, have been analyzed. MiRNAs may also be able to predict responses to chemo- and radiotherapy, and may be manipulated to modify CRC characteristics. We herein discuss the use of circulating miRNAs as possible non-invasive biomarkers of early CRC onset, relapse, or response to treatment. We also discuss the obstacles that currently limit the routine use of epigenetic biomarkers in clinical settings.
Collapse
|
45
|
Robinson JM. MicroRNA expression during demosponge dissociation, reaggregation, and differentiation and a evolutionarily conserved demosponge miRNA expression profile. Dev Genes Evol 2015; 225:341-51. [PMID: 26553380 DOI: 10.1007/s00427-015-0520-5] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/24/2015] [Accepted: 10/30/2015] [Indexed: 10/22/2022]
Abstract
Demosponges share eight orthologous microRNAs (miRNAs), with none in common with Bilateria. Biological functions of these demosponge miRNAs are unknown. Bilaterian miRNAs are key regulators of cellular processes including cell cycle, differentiation, and metabolism. Resolving if demosponge miRNAs participate in such biological functions will provide clues whether these functions are convergent, evidence on the mode of evolution of cellular developmental processes. Here, a quantitative PCR (qPCR) assay was developed and used to test for differential miRNA expression during dissociation and reaggregation in Spongosorites, compare expression profiles between choanosome and cortex in Spongosorites, and compare undifferentiated gemmules to differentiated juveniles in Ephydatia. During Spongosorites dissociation and reaggregation, miRNA expression showed a global decrease in expression across a range of reaggregating cell densities. miRNA differential response could be related to various general cellular responses, potentially related to nutrient-poor conditions of the minimal artificial seawater media, stress response from tissue dissociation, or loss of cell-cell or cell-matrix contact. In Ephydatia, overall increase in miRNA expression in gemmule-hatched stage 4/5 juveniles relative to gemmules is observed, indicating that increased miRNA expression may be related to increased cellular activity such as migration, cell cycle, and/or differentiation. Observed differential miRNA expression of miRNA during dissociation in Spongosorites (lowered global expression), and during activation, and differentiation of Ephydatia gemmules (increased global expression) could indicate that miRNA expression is associated with cell cycle, differentiation, or metabolism pathways. Interspecies comparison was performed, results indicating that orthologous miRNAs share similar relative expression pattern between the four species tested (Spongosorites, Cinachyrella, Haliclona, and Ephydatia), demonstrating and evolutionarily conserved miRNA expression profile across Demospongia. While these results do not elucidate specific molecular and cellular pathways, together they provide a broad survey of miRNA expression in demosponge systems.
Collapse
Affiliation(s)
- Jeffrey M Robinson
- Department of Biological Sciences, Molecular and Cellular Biology Program, Dartmouth College, Hanover, USA.
- Anatomy Department, College of Medicine, Howard University, 520 W Street NW, Washington, DC, 20059, USA.
| |
Collapse
|
46
|
Gharbi S, Shamsara M, Khateri S, Soroush MR, Ghorbanmehr N, Tavallaei M, Nourani MR, Mowla SJ. Identification of Reliable Reference Genes for Quantification of MicroRNAs in Serum Samples of Sulfur Mustard-Exposed Veterans. CELL JOURNAL 2015; 17:494-501. [PMID: 26464821 PMCID: PMC4601870 DOI: 10.22074/cellj.2015.9] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 11/20/2013] [Accepted: 03/11/2014] [Indexed: 12/22/2022]
Abstract
Objective In spite of accumulating information about pathological aspects of sulfur
mustard (SM), the precise mechanism responsible for its effects is not well understood.
Circulating microRNAs (miRNAs) are promising biomarkers for disease diagnosis and
prognosis. Accurate normalization using appropriate reference genes, is a critical step in
miRNA expression studies. In this study, we aimed to identify appropriate reference gene
for microRNA quantification in serum samples of SM victims.
Materials and Methods In this case and control experimental study, using quantitative
real-time polymerase chain reaction (qRT-PCR), we evaluated the suitability of a panel
of small RNAs including SNORD38B, SNORD49A, U6, 5S rRNA, miR-423-3p, miR-191,
miR-16 and miR-103 in sera of 28 SM-exposed veterans of Iran-Iraq war (1980-1988)
and 15 matched control volunteers. Different statistical algorithms including geNorm,
Normfinder, best-keeper and comparative delta-quantification cycle (Cq) method were
employed to find the least variable reference gene.
Results miR-423-3p was identified as the most stably expressed reference gene, and miR-
103 and miR-16 ranked after that.
Conclusion We demonstrate that non-miRNA reference genes have the least stabil-
ity in serum samples and that some house-keeping miRNAs may be used as more
reliable reference genes for miRNAs in serum. In addition, using the geometric mean
of two reference genes could increase the reliability of the normalizers.
Collapse
Affiliation(s)
- Sedigheh Gharbi
- Department of Molecular Genetics, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran ; Department of Biology, Faculty of Sciences, Shahid Bahonar University of Kerman, Kerman, Iran
| | - Mehdi Shamsara
- National Institute of Genetic Engineering and Biotechnology, Tehran, Iran
| | - Shahriar Khateri
- Janbazan Medical and Engineering Research Center (JMERC), Tehran, Iran
| | | | - Nassim Ghorbanmehr
- Departmen of Biotechnology, Faculty of Biological Sciences, Alzahra University, Tehran, Iran
| | - Mahmood Tavallaei
- Genetic Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran
| | - Mohammad Reza Nourani
- Chemical Injury Research Center (CIRC), Baqiyatallah University of Medical Sciences, Tehran, Iran
| | - Seyed Javad Mowla
- Department of Molecular Genetics, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran
| |
Collapse
|
47
|
Schwarzenbach H, da Silva AM, Calin G, Pantel K. Data Normalization Strategies for MicroRNA Quantification. Clin Chem 2015; 61:1333-42. [PMID: 26408530 DOI: 10.1373/clinchem.2015.239459] [Citation(s) in RCA: 362] [Impact Index Per Article: 36.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/27/2015] [Accepted: 08/25/2015] [Indexed: 12/15/2022]
Abstract
BACKGROUND Different technologies, such as quantitative real-time PCR or microarrays, have been developed to measure microRNA (miRNA) expression levels. Quantification of miRNA transcripts implicates data normalization using endogenous and exogenous reference genes for data correction. However, there is no consensus about an optimal normalization strategy. The choice of a reference gene remains problematic and can have a serious impact on the actual available transcript levels and, consequently, on the biological interpretation of data. CONTENT In this review article we discuss the reliability of the use of small RNAs, commonly reported in the literature as miRNA expression normalizers, and compare different strategies used for data normalization. SUMMARY A workflow strategy is proposed for normalization of miRNA expression data in an attempt to provide a basis for the establishment of a global standard procedure that will allow comparison across studies.
Collapse
Affiliation(s)
- Heidi Schwarzenbach
- Department of Tumour Biology, Center of Experimental Medicine, University Cancer Center Hamburg, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Andreia Machado da Silva
- Department of Experimental Therapeutics and The Center for RNA Interference and Non-Coding RNAs, The University of Texas MD Anderson Cancer Center, Houston, TX; Instituto de Investigação em Saúde, Universidade do Porto, Porto, Portugal; INEB, Institute of Biomedical Engineering, Universidade do Porto, Porto, Portugal
| | - George Calin
- Department of Experimental Therapeutics and The Center for RNA Interference and Non-Coding RNAs, The University of Texas MD Anderson Cancer Center, Houston, TX
| | - Klaus Pantel
- Department of Tumour Biology, Center of Experimental Medicine, University Cancer Center Hamburg, University Medical Center Hamburg-Eppendorf, Hamburg, Germany;
| |
Collapse
|
48
|
Simonson OE, Mougiakakos D, Heldring N, Bassi G, Johansson HJ, Dalén M, Jitschin R, Rodin S, Corbascio M, El Andaloussi S, Wiklander OPB, Nordin JZ, Skog J, Romain C, Koestler T, Hellgren-Johansson L, Schiller P, Joachimsson PO, Hägglund H, Mattsson M, Lehtiö J, Faridani OR, Sandberg R, Korsgren O, Krampera M, Weiss DJ, Grinnemo KH, Le Blanc K. In Vivo Effects of Mesenchymal Stromal Cells in Two Patients With Severe Acute Respiratory Distress Syndrome. Stem Cells Transl Med 2015; 4:1199-213. [PMID: 26285659 DOI: 10.5966/sctm.2015-0021] [Citation(s) in RCA: 109] [Impact Index Per Article: 10.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/10/2015] [Accepted: 04/13/2015] [Indexed: 12/17/2022] Open
Abstract
UNLABELLED Mesenchymal stromal cells (MSCs) have been investigated as a treatment for various inflammatory diseases because of their immunomodulatory and reparative properties. However, many basic questions concerning their mechanisms of action after systemic infusion remain unanswered. We performed a detailed analysis of the immunomodulatory properties and proteomic profile of MSCs systemically administered to two patients with severe refractory acute respiratory distress syndrome (ARDS) on a compassionate use basis and attempted to correlate these with in vivo anti-inflammatory actions. Both patients received 2×10(6) cells per kilogram, and each subsequently improved with resolution of respiratory, hemodynamic, and multiorgan failure. In parallel, a decrease was seen in multiple pulmonary and systemic markers of inflammation, including epithelial apoptosis, alveolar-capillary fluid leakage, and proinflammatory cytokines, microRNAs, and chemokines. In vitro studies of the MSCs demonstrated a broad anti-inflammatory capacity, including suppression of T-cell responses and induction of regulatory phenotypes in T cells, monocytes, and neutrophils. Some of these in vitro potency assessments correlated with, and were relevant to, the observed in vivo actions. These experiences highlight both the mechanistic information that can be gained from clinical experience and the value of correlating in vitro potency assessments with clinical effects. The findings also suggest, but do not prove, a beneficial effect of lung protective strategies using adoptively transferred MSCs in ARDS. Appropriate randomized clinical trials are required to further assess any potential clinical efficacy and investigate the effects on in vivo inflammation. SIGNIFICANCE This article describes the cases of two patients with severe refractory adult respiratory syndrome (ARDS) who failed to improve after both standard life support measures, including mechanical ventilation, and additional measures, including extracorporeal ventilation (i.e., in a heart-lung machine). Unlike acute forms of ARDS (such in the current NIH-sponsored study of mesenchymal stromal cells in ARDS), recovery does not generally occur in such patients.
Collapse
Affiliation(s)
- Oscar E Simonson
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Dimitrios Mougiakakos
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Nina Heldring
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Giulio Bassi
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Henrik J Johansson
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Magnus Dalén
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Regina Jitschin
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Sergey Rodin
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Matthias Corbascio
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Samir El Andaloussi
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Oscar P B Wiklander
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Joel Z Nordin
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Johan Skog
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Charlotte Romain
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Tina Koestler
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Laila Hellgren-Johansson
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Petter Schiller
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Per-Olof Joachimsson
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Hans Hägglund
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Mattias Mattsson
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Janne Lehtiö
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Omid R Faridani
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Rickard Sandberg
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Olle Korsgren
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Mauro Krampera
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Daniel J Weiss
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Karl-Henrik Grinnemo
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| | - Katarina Le Blanc
- Departments of Molecular Medicine and Surgery, Cardiothoracic Surgery and Anesthesia, and Laboratory Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; Department of Internal Medicine, Hematology and Oncology, University of Erlangen-Nuremberg, Erlangen, Germany; Stem Cell Research Laboratory, Section of Hematology, Department of Medicine, University of Verona, Verona, Italy; Cancer Proteomics Mass Spectrometry, Science for Life Laboratory, Department of Oncology-Pathology, Department of Medical Biochemistry and Biophysics, and Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; Center for Diseases of Aging, Vaccine and Gene Therapy Institute Florida, Port St. Lucie, Florida, USA; Exosome Diagnostics Inc., New York, New York, USA; Departments of Cardiothoracic Surgery, Hematology, and Immunology, Genetics and Pathology, Uppsala University Hospital, Uppsala, Sweden; Ludwig Institute for Cancer Research, Stockholm, Sweden; Health Sciences Research Facility, Department of Medicine, University of Vermont, Burlington, Vermont, USA
| |
Collapse
|
49
|
Mahdipour M, van Tol HTA, Stout TAE, Roelen BAJ. Validating reference microRNAs for normalizing qRT-PCR data in bovine oocytes and preimplantation embryos. BMC DEVELOPMENTAL BIOLOGY 2015; 15:25. [PMID: 26062615 PMCID: PMC4464232 DOI: 10.1186/s12861-015-0075-8] [Citation(s) in RCA: 25] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 02/03/2015] [Accepted: 06/03/2015] [Indexed: 01/09/2023]
Abstract
BACKGROUND MicroRNAs (miRNAs) are small noncoding RNAs that act as post-transcriptional regulators of gene targets. Accurate quantification of miRNA expression using validated internal controls should aid in the understanding of their role in epigenetic modification of genome function. To date, most studies that have examined miRNA expression levels have used the global mean expression of all expressed genes or the expression of reference mRNAs or nuclear RNAs for normalization. RESULTS We analyzed the suitability of a number of miRNAs as potential expression normalizers in bovine oocytes and early embryos, and porcine oocytes. The stages examined were bovine oocytes at the germinal vesicle (GV) and metaphase II stages, bovine zygotes, 2, 4 and 8 cell embryos, morulae and blastocysts, as well as porcine cumulus oocyte complexes, GV, metaphase I and II oocytes. qRT-PCR was performed to quantify expression of miR-93, miR-103, miR-26a, miR-191, miR-23b, Let-7a and U6 for bovine samples and miR-21, miR-26a, miR-93, miR-103, miR-148a, miR-182 and miR-191 for porcine oocytes. The average starting material for each sample was determined using specific standard curves for each primer set. Subsequently, geNorm and BestKeeper software were used to identify a set of stably expressed miRNAs. Stepwise removal to determine the optimum number of reference miRNAs identified miR-93 and miR-103 as the most stably expressed in bovine samples and miR-26a, miR-191 and miR-93 in porcine samples. CONCLUSIONS The combination of miR-93 and miR-103 is optimal for normalizing miRNA expression for qPCR experiments on bovine oocytes and preimplantation embryos; the preferred combination for porcine oocytes is miR-26a, miR-191 and miR-93.
Collapse
Affiliation(s)
- Mahdi Mahdipour
- Department of Farm Animal Health, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 104, 3584, CM, Utrecht, The Netherlands.
| | - Helena T A van Tol
- Department of Farm Animal Health, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 104, 3584, CM, Utrecht, The Netherlands.
| | - Tom A E Stout
- Department of Farm Animal Health, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 104, 3584, CM, Utrecht, The Netherlands.
- Department of Equine Sciences, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 104, 3584, CM, Utrecht, The Netherlands.
| | - Bernard A J Roelen
- Department of Farm Animal Health, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 104, 3584, CM, Utrecht, The Netherlands.
- Department of Equine Sciences, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 104, 3584, CM, Utrecht, The Netherlands.
| |
Collapse
|
50
|
Pasqualini L, Bu H, Puhr M, Narisu N, Rainer J, Schlick B, Schäfer G, Angelova M, Trajanoski Z, Börno ST, Schweiger MR, Fuchsberger C, Klocker H. miR-22 and miR-29a Are Members of the Androgen Receptor Cistrome Modulating LAMC1 and Mcl-1 in Prostate Cancer. Mol Endocrinol 2015; 29:1037-54. [PMID: 26052614 DOI: 10.1210/me.2014-1358] [Citation(s) in RCA: 68] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/24/2023] Open
Abstract
The normal prostate as well as early stages and advanced prostate cancer (PCa) require a functional androgen receptor (AR) for growth and survival. The recent discovery of microRNAs (miRNAs) as novel effector molecules of AR disclosed the existence of an intricate network between AR, miRNAs and downstream target genes. In this study DUCaP cells, characterized by high content of wild-type AR and robust AR transcriptional activity, were chosen as the main experimental model. By integrative analysis of chromatin immunoprecipitation-sequencing (ChIP-seq) and microarray expression profiling data, miRNAs putatively bound and significantly regulated by AR were identified. A direct AR regulation of miR-22, miR-29a, and miR-17-92 cluster along with their host genes was confirmed. Interestingly, endogenous levels of miR-22 and miR-29a were found to be reduced in PCa cells expressing AR. In primary tumor samples, miR-22 and miR-29a were less abundant in the cancerous tissue compared with the benign counterpart. This specific expression pattern was associated with a differential DNA methylation of the genomic AR binding sites. The identification of laminin gamma 1 (LAMC1) and myeloid cell leukemia 1 (MCL1) as direct targets of miR-22 and miR-29a, respectively, suggested a tumor-suppressive role of these miRNAs. Indeed, transfection of miRNA mimics in PCa cells induced apoptosis and diminished cell migration and viability. Collectively, these data provide additional information regarding the complex regulatory machinery that guides miRNAs activity in PCa, highlighting an important contribution of miRNAs in the AR signaling.
Collapse
Affiliation(s)
- Lorenza Pasqualini
- Department of Urology (L.P., H.B., M.P., B.S., G.S., H.K.), Division of Experimental Urology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Research Institute for Biomedical Aging Research (H.B.), University of Innsbruck, 6020 Innsbruck, Austria; Medical Genomics and Metabolic Genetics Branch (N.N.), National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland 20892; Biocenter Innsbruck (J.R.), Section for Molecular Pathophysiology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Center for Biomedicine (J.R., C.F.), EURAC Bolzano, 39100 Bolzano, Italy; Oncotyrol (B.S.), Center for Personalized Cancer Medicine, 6020 Innsbruck, Austria; Department of Pathology (G.S.), Medical University of Innsbruck, 6020 Innsbruck, Austria; Biocenter Innsbruck (M.A., Z.T.), Division of Bioinformatics, Medical University of Innsbruck, 6020 Innsbruck, Austria; Max Planck Institute for Molecular Genetics (S.T.B., M.R.S.), 14195 Berlin, Germany; Cologne Center for Genomics (M.R.S.), University of Cologne, 50931 Cologne, Germany; and Department of Biostatistic (C.F.), University of Michigan, Ann Arbor, Michigan 48109
| | - Huajie Bu
- Department of Urology (L.P., H.B., M.P., B.S., G.S., H.K.), Division of Experimental Urology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Research Institute for Biomedical Aging Research (H.B.), University of Innsbruck, 6020 Innsbruck, Austria; Medical Genomics and Metabolic Genetics Branch (N.N.), National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland 20892; Biocenter Innsbruck (J.R.), Section for Molecular Pathophysiology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Center for Biomedicine (J.R., C.F.), EURAC Bolzano, 39100 Bolzano, Italy; Oncotyrol (B.S.), Center for Personalized Cancer Medicine, 6020 Innsbruck, Austria; Department of Pathology (G.S.), Medical University of Innsbruck, 6020 Innsbruck, Austria; Biocenter Innsbruck (M.A., Z.T.), Division of Bioinformatics, Medical University of Innsbruck, 6020 Innsbruck, Austria; Max Planck Institute for Molecular Genetics (S.T.B., M.R.S.), 14195 Berlin, Germany; Cologne Center for Genomics (M.R.S.), University of Cologne, 50931 Cologne, Germany; and Department of Biostatistic (C.F.), University of Michigan, Ann Arbor, Michigan 48109
| | - Martin Puhr
- Department of Urology (L.P., H.B., M.P., B.S., G.S., H.K.), Division of Experimental Urology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Research Institute for Biomedical Aging Research (H.B.), University of Innsbruck, 6020 Innsbruck, Austria; Medical Genomics and Metabolic Genetics Branch (N.N.), National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland 20892; Biocenter Innsbruck (J.R.), Section for Molecular Pathophysiology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Center for Biomedicine (J.R., C.F.), EURAC Bolzano, 39100 Bolzano, Italy; Oncotyrol (B.S.), Center for Personalized Cancer Medicine, 6020 Innsbruck, Austria; Department of Pathology (G.S.), Medical University of Innsbruck, 6020 Innsbruck, Austria; Biocenter Innsbruck (M.A., Z.T.), Division of Bioinformatics, Medical University of Innsbruck, 6020 Innsbruck, Austria; Max Planck Institute for Molecular Genetics (S.T.B., M.R.S.), 14195 Berlin, Germany; Cologne Center for Genomics (M.R.S.), University of Cologne, 50931 Cologne, Germany; and Department of Biostatistic (C.F.), University of Michigan, Ann Arbor, Michigan 48109
| | - Narisu Narisu
- Department of Urology (L.P., H.B., M.P., B.S., G.S., H.K.), Division of Experimental Urology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Research Institute for Biomedical Aging Research (H.B.), University of Innsbruck, 6020 Innsbruck, Austria; Medical Genomics and Metabolic Genetics Branch (N.N.), National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland 20892; Biocenter Innsbruck (J.R.), Section for Molecular Pathophysiology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Center for Biomedicine (J.R., C.F.), EURAC Bolzano, 39100 Bolzano, Italy; Oncotyrol (B.S.), Center for Personalized Cancer Medicine, 6020 Innsbruck, Austria; Department of Pathology (G.S.), Medical University of Innsbruck, 6020 Innsbruck, Austria; Biocenter Innsbruck (M.A., Z.T.), Division of Bioinformatics, Medical University of Innsbruck, 6020 Innsbruck, Austria; Max Planck Institute for Molecular Genetics (S.T.B., M.R.S.), 14195 Berlin, Germany; Cologne Center for Genomics (M.R.S.), University of Cologne, 50931 Cologne, Germany; and Department of Biostatistic (C.F.), University of Michigan, Ann Arbor, Michigan 48109
| | - Johannes Rainer
- Department of Urology (L.P., H.B., M.P., B.S., G.S., H.K.), Division of Experimental Urology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Research Institute for Biomedical Aging Research (H.B.), University of Innsbruck, 6020 Innsbruck, Austria; Medical Genomics and Metabolic Genetics Branch (N.N.), National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland 20892; Biocenter Innsbruck (J.R.), Section for Molecular Pathophysiology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Center for Biomedicine (J.R., C.F.), EURAC Bolzano, 39100 Bolzano, Italy; Oncotyrol (B.S.), Center for Personalized Cancer Medicine, 6020 Innsbruck, Austria; Department of Pathology (G.S.), Medical University of Innsbruck, 6020 Innsbruck, Austria; Biocenter Innsbruck (M.A., Z.T.), Division of Bioinformatics, Medical University of Innsbruck, 6020 Innsbruck, Austria; Max Planck Institute for Molecular Genetics (S.T.B., M.R.S.), 14195 Berlin, Germany; Cologne Center for Genomics (M.R.S.), University of Cologne, 50931 Cologne, Germany; and Department of Biostatistic (C.F.), University of Michigan, Ann Arbor, Michigan 48109
| | - Bettina Schlick
- Department of Urology (L.P., H.B., M.P., B.S., G.S., H.K.), Division of Experimental Urology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Research Institute for Biomedical Aging Research (H.B.), University of Innsbruck, 6020 Innsbruck, Austria; Medical Genomics and Metabolic Genetics Branch (N.N.), National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland 20892; Biocenter Innsbruck (J.R.), Section for Molecular Pathophysiology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Center for Biomedicine (J.R., C.F.), EURAC Bolzano, 39100 Bolzano, Italy; Oncotyrol (B.S.), Center for Personalized Cancer Medicine, 6020 Innsbruck, Austria; Department of Pathology (G.S.), Medical University of Innsbruck, 6020 Innsbruck, Austria; Biocenter Innsbruck (M.A., Z.T.), Division of Bioinformatics, Medical University of Innsbruck, 6020 Innsbruck, Austria; Max Planck Institute for Molecular Genetics (S.T.B., M.R.S.), 14195 Berlin, Germany; Cologne Center for Genomics (M.R.S.), University of Cologne, 50931 Cologne, Germany; and Department of Biostatistic (C.F.), University of Michigan, Ann Arbor, Michigan 48109
| | - Georg Schäfer
- Department of Urology (L.P., H.B., M.P., B.S., G.S., H.K.), Division of Experimental Urology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Research Institute for Biomedical Aging Research (H.B.), University of Innsbruck, 6020 Innsbruck, Austria; Medical Genomics and Metabolic Genetics Branch (N.N.), National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland 20892; Biocenter Innsbruck (J.R.), Section for Molecular Pathophysiology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Center for Biomedicine (J.R., C.F.), EURAC Bolzano, 39100 Bolzano, Italy; Oncotyrol (B.S.), Center for Personalized Cancer Medicine, 6020 Innsbruck, Austria; Department of Pathology (G.S.), Medical University of Innsbruck, 6020 Innsbruck, Austria; Biocenter Innsbruck (M.A., Z.T.), Division of Bioinformatics, Medical University of Innsbruck, 6020 Innsbruck, Austria; Max Planck Institute for Molecular Genetics (S.T.B., M.R.S.), 14195 Berlin, Germany; Cologne Center for Genomics (M.R.S.), University of Cologne, 50931 Cologne, Germany; and Department of Biostatistic (C.F.), University of Michigan, Ann Arbor, Michigan 48109
| | - Mihaela Angelova
- Department of Urology (L.P., H.B., M.P., B.S., G.S., H.K.), Division of Experimental Urology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Research Institute for Biomedical Aging Research (H.B.), University of Innsbruck, 6020 Innsbruck, Austria; Medical Genomics and Metabolic Genetics Branch (N.N.), National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland 20892; Biocenter Innsbruck (J.R.), Section for Molecular Pathophysiology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Center for Biomedicine (J.R., C.F.), EURAC Bolzano, 39100 Bolzano, Italy; Oncotyrol (B.S.), Center for Personalized Cancer Medicine, 6020 Innsbruck, Austria; Department of Pathology (G.S.), Medical University of Innsbruck, 6020 Innsbruck, Austria; Biocenter Innsbruck (M.A., Z.T.), Division of Bioinformatics, Medical University of Innsbruck, 6020 Innsbruck, Austria; Max Planck Institute for Molecular Genetics (S.T.B., M.R.S.), 14195 Berlin, Germany; Cologne Center for Genomics (M.R.S.), University of Cologne, 50931 Cologne, Germany; and Department of Biostatistic (C.F.), University of Michigan, Ann Arbor, Michigan 48109
| | - Zlatko Trajanoski
- Department of Urology (L.P., H.B., M.P., B.S., G.S., H.K.), Division of Experimental Urology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Research Institute for Biomedical Aging Research (H.B.), University of Innsbruck, 6020 Innsbruck, Austria; Medical Genomics and Metabolic Genetics Branch (N.N.), National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland 20892; Biocenter Innsbruck (J.R.), Section for Molecular Pathophysiology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Center for Biomedicine (J.R., C.F.), EURAC Bolzano, 39100 Bolzano, Italy; Oncotyrol (B.S.), Center for Personalized Cancer Medicine, 6020 Innsbruck, Austria; Department of Pathology (G.S.), Medical University of Innsbruck, 6020 Innsbruck, Austria; Biocenter Innsbruck (M.A., Z.T.), Division of Bioinformatics, Medical University of Innsbruck, 6020 Innsbruck, Austria; Max Planck Institute for Molecular Genetics (S.T.B., M.R.S.), 14195 Berlin, Germany; Cologne Center for Genomics (M.R.S.), University of Cologne, 50931 Cologne, Germany; and Department of Biostatistic (C.F.), University of Michigan, Ann Arbor, Michigan 48109
| | - Stefan T Börno
- Department of Urology (L.P., H.B., M.P., B.S., G.S., H.K.), Division of Experimental Urology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Research Institute for Biomedical Aging Research (H.B.), University of Innsbruck, 6020 Innsbruck, Austria; Medical Genomics and Metabolic Genetics Branch (N.N.), National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland 20892; Biocenter Innsbruck (J.R.), Section for Molecular Pathophysiology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Center for Biomedicine (J.R., C.F.), EURAC Bolzano, 39100 Bolzano, Italy; Oncotyrol (B.S.), Center for Personalized Cancer Medicine, 6020 Innsbruck, Austria; Department of Pathology (G.S.), Medical University of Innsbruck, 6020 Innsbruck, Austria; Biocenter Innsbruck (M.A., Z.T.), Division of Bioinformatics, Medical University of Innsbruck, 6020 Innsbruck, Austria; Max Planck Institute for Molecular Genetics (S.T.B., M.R.S.), 14195 Berlin, Germany; Cologne Center for Genomics (M.R.S.), University of Cologne, 50931 Cologne, Germany; and Department of Biostatistic (C.F.), University of Michigan, Ann Arbor, Michigan 48109
| | - Michal R Schweiger
- Department of Urology (L.P., H.B., M.P., B.S., G.S., H.K.), Division of Experimental Urology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Research Institute for Biomedical Aging Research (H.B.), University of Innsbruck, 6020 Innsbruck, Austria; Medical Genomics and Metabolic Genetics Branch (N.N.), National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland 20892; Biocenter Innsbruck (J.R.), Section for Molecular Pathophysiology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Center for Biomedicine (J.R., C.F.), EURAC Bolzano, 39100 Bolzano, Italy; Oncotyrol (B.S.), Center for Personalized Cancer Medicine, 6020 Innsbruck, Austria; Department of Pathology (G.S.), Medical University of Innsbruck, 6020 Innsbruck, Austria; Biocenter Innsbruck (M.A., Z.T.), Division of Bioinformatics, Medical University of Innsbruck, 6020 Innsbruck, Austria; Max Planck Institute for Molecular Genetics (S.T.B., M.R.S.), 14195 Berlin, Germany; Cologne Center for Genomics (M.R.S.), University of Cologne, 50931 Cologne, Germany; and Department of Biostatistic (C.F.), University of Michigan, Ann Arbor, Michigan 48109
| | - Christian Fuchsberger
- Department of Urology (L.P., H.B., M.P., B.S., G.S., H.K.), Division of Experimental Urology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Research Institute for Biomedical Aging Research (H.B.), University of Innsbruck, 6020 Innsbruck, Austria; Medical Genomics and Metabolic Genetics Branch (N.N.), National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland 20892; Biocenter Innsbruck (J.R.), Section for Molecular Pathophysiology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Center for Biomedicine (J.R., C.F.), EURAC Bolzano, 39100 Bolzano, Italy; Oncotyrol (B.S.), Center for Personalized Cancer Medicine, 6020 Innsbruck, Austria; Department of Pathology (G.S.), Medical University of Innsbruck, 6020 Innsbruck, Austria; Biocenter Innsbruck (M.A., Z.T.), Division of Bioinformatics, Medical University of Innsbruck, 6020 Innsbruck, Austria; Max Planck Institute for Molecular Genetics (S.T.B., M.R.S.), 14195 Berlin, Germany; Cologne Center for Genomics (M.R.S.), University of Cologne, 50931 Cologne, Germany; and Department of Biostatistic (C.F.), University of Michigan, Ann Arbor, Michigan 48109
| | - Helmut Klocker
- Department of Urology (L.P., H.B., M.P., B.S., G.S., H.K.), Division of Experimental Urology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Research Institute for Biomedical Aging Research (H.B.), University of Innsbruck, 6020 Innsbruck, Austria; Medical Genomics and Metabolic Genetics Branch (N.N.), National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland 20892; Biocenter Innsbruck (J.R.), Section for Molecular Pathophysiology, Medical University of Innsbruck, 6020 Innsbruck, Austria; Center for Biomedicine (J.R., C.F.), EURAC Bolzano, 39100 Bolzano, Italy; Oncotyrol (B.S.), Center for Personalized Cancer Medicine, 6020 Innsbruck, Austria; Department of Pathology (G.S.), Medical University of Innsbruck, 6020 Innsbruck, Austria; Biocenter Innsbruck (M.A., Z.T.), Division of Bioinformatics, Medical University of Innsbruck, 6020 Innsbruck, Austria; Max Planck Institute for Molecular Genetics (S.T.B., M.R.S.), 14195 Berlin, Germany; Cologne Center for Genomics (M.R.S.), University of Cologne, 50931 Cologne, Germany; and Department of Biostatistic (C.F.), University of Michigan, Ann Arbor, Michigan 48109
| |
Collapse
|