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Garofano K, Mariani V, Rashid K, Suwunnakorn S, Sidahmed A, Horvath A, Maggirwar SB, O’Brien TJ, Perera MA, Whalen M, Lee NH. Transcriptomic and functional characterization of megakaryocytic-derived platelet-like particles: impaired aggregation and prominent anti-tumor effects. Platelets 2025; 36:2449344. [PMID: 39812346 PMCID: PMC11890189 DOI: 10.1080/09537104.2024.2449344] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/14/2024] [Revised: 12/22/2024] [Accepted: 12/24/2024] [Indexed: 01/16/2025]
Abstract
Platelet-like particles (PLPs), derived from megakaryocytic cell lines MEG-01 and K-562, are widely used as a surrogate to study platelet formation and function. We demonstrate by RNA-Seq that PLPs are transcriptionally distinct from platelets. Expression of key genes in signaling pathways promoting platelet activation/aggregation, such as the PI3K/AKT, protein kinase A, phospholipase C, and α-adrenergic and GP6 receptor pathways, was missing or under-expressed in PLPs. Functionally, PLPs do not aggregate following epinephrine, collagen, or ADP stimulation. While PLPs aggregated in response to thrombin, they did not display enhanced expression of surface markers P-selectin and activated α2bβ3, in contrast to platelets. We have previously demonstrated that platelets physically couple to MDA-PCa-2b and RC77T/E prostate cancer (PCa) cells via specific ligand-receptor interactions, leading to platelet-stimulated cell invasiveness and apoptotic resistance, and reciprocal cell-induced platelet aggregation. In contrast, PLP interactions with PCa cells inhibited both cell invasion and apoptotic resistance while failing to promote PLP aggregation. Moreover, PLPs reduced platelet-PCa cell interactions and antagonized platelet-stimulated oncogenic effects in PCa cells. RNA-Seq analysis identified candidate ligand-transmembrane protein combinations involved in anti-tumorigenic signaling of PLPs to PCa cells. Antibody neutralization of the TIMP3-MMP15 and VEGFB-FGFR1 signaling axes reversed PLP-mediated anti-invasion and apoptotic sensitization, respectively. In summary, PLPs lack many transcriptomic, molecular and functional features of platelets and possess novel anti-tumorigenic properties. These findings indicate that PLPs may have a potential therapeutic role in targeting and disrupting the oncogenic signaling between platelets and cancer cells, offering a new avenue for anti-cancer strategies.
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Affiliation(s)
- Kaitlin Garofano
- Department of Pharmacology and Physiology, George Washington University, Washington, DC, 20037
| | - Vera Mariani
- Department of Pharmacology and Physiology, George Washington University, Washington, DC, 20037
| | - Kameron Rashid
- Department of Pharmacology and Physiology, George Washington University, Washington, DC, 20037
| | - Sumanun Suwunnakorn
- Department of Microbiology Immunology and Tropical Medicine, The George Washington University, Washington, DC, 20037
| | - Alfateh Sidahmed
- Department of Medicine, George Washington University, Washington, DC, 20037
| | - Anelia Horvath
- Department of Biochemistry and Molecular Medicine, George Washington University, Washington, DC, 20037
| | - Sanjay B. Maggirwar
- Department of Microbiology Immunology and Tropical Medicine, The George Washington University, Washington, DC, 20037
| | - Travis J. O’Brien
- Department of Pharmacology and Physiology, George Washington University, Washington, DC, 20037
| | - Minoli A. Perera
- Department of Pharmacology and Center for Pharmacogenomics, Northwestern University, Chicago, IL, 60611
| | - Michael Whalen
- GW Cancer Center, George Washington University, Washington, DC, 20037
| | - Norman H Lee
- Department of Pharmacology and Physiology, George Washington University, Washington, DC, 20037
- GW Cancer Center, George Washington University, Washington, DC, 20037
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Wang L, Zhang T, Yang X, Mo Q, Ran M, Li R, Yang B, Shen H, Li Q, Li Z, Jiang N, Zeng J, Xie X, He S, Huang F, Zhang C, Luo J, Wu J. Multimodal discovery of bavachinin A: A natural FLT3 agonist for treating thrombocytopenia. PHYTOMEDICINE : INTERNATIONAL JOURNAL OF PHYTOTHERAPY AND PHYTOPHARMACOLOGY 2025; 140:156597. [PMID: 40058315 DOI: 10.1016/j.phymed.2025.156597] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 04/04/2024] [Revised: 11/25/2024] [Accepted: 02/28/2025] [Indexed: 03/25/2025]
Abstract
BACKGROUND Radiation-induced thrombocytopenia (RIT) poses a serious risk to patients with cancer undergoing radiotherapy and leads to hemorrhage and mortality. Unfortunately, effective treatment options for RIT are currently limited. PURPOSE This study aimed to discover active compound from Fructus Psoraleae, a traditional Chinese medicine recognized for its hemostatic properties, and to elucidate its mechanism of action in the treatment of RIT. METHODS The efficacy of Fructus Psoraleae in treating thrombocytopenia was assessed using network pharmacology. A drug-screening model was built using a naive Bayes algorithm to determine the effective compounds in Fructus Psoraleae. Giemsa staining and flow cytometry were used to evaluate the effects of bavachinin A on megakaryocytes (MK) differentiation. RIT and thrombopoietin (TPO) receptor (c-MPL) knockout (c-MPL-/-) mice were used to assess the therapeutic efficacy of bavachinin A in mitigating thrombocytopenia. Tg (cd41:eGFP) zebrafish were used to investigate the effect of bavachinin A on thrombopoiesis. RNA sequencing (RNA-seq), molecular docking simulations, molecular dynamics simulations, drug affinity responsive target stability assay (DARTS), and biolayer interferometry (BLI) were used to elucidate the molecular mechanisms of action of bavachinin A against thrombocytopenia. RESULTS In silico analysis using a drug screening model identified bavachinin A as promising candidate compound derived from Fructus Psoraleae. In vitro experiments demonstrated that Bavachinin A induced MK differentiation. In vivo experiments revealed that bavachinin A augmented platelet levels and improved coagulation in RIT mice, facilitated megakaryopoiesis and platelet levels in c-MPL-/- mice, and accelerated thrombopoiesis in zebrafish. Furthermore, RNA-seq, molecular docking simulations, molecular dynamics simulations, DARTS, and BLI demonstrated that bavachinin A bound directly to fms-like tyrosine kinase 3 (FLT3). Notably, blocking FLT3 or phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) pathway hindered bavachinin-A-induced MK differentiation. However, repressing the TPO/c-MPL signaling pathway had no significant effect. CONCLUSION Bavachinin A promotes MK differentiation and thrombopoiesis by directly binding to FLT3 and activating PI3K/Akt signaling. Importantly, this effect was not dependent on the conventional TPO/c-MPL signaling pathway. This study underscores the translational potential of bavachinin A as a promising novel therapeutic for thrombocytopenia, offering novel insights into TPO-independent mechanisms of thrombopoiesis and establishing a robust multimodal approach for drug discovery.
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Affiliation(s)
- Long Wang
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, Sichuan, 646000, China
| | - Ting Zhang
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, Sichuan, 646000, China
| | - Xin Yang
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, Sichuan, 646000, China
| | - Qi Mo
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, Sichuan, 646000, China
| | - Mei Ran
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, Sichuan, 646000, China
| | - Rong Li
- Drug Discovery Research Center, Southwest Medical University, Luzhou, Sichuan, 646000, China; Laboratory for Cardiovascular Pharmacology of Department of Pharmacology, The School of Pharmacy, Southwest Medical University, Luzhou, Sichuan, 646000, China
| | - Bo Yang
- Department of Oncology, The Affiliated Hospital of Southwest Medical University, Luzhou, 646000, China
| | - Hongping Shen
- Clinical Trial Center, The Affiliated Traditional Chinese Medicine Hospital of Southwest Medical University, Luzhou, Sichuan, 646000, China
| | - Qinyao Li
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, Sichuan, 646000, China
| | - Zhichao Li
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, Sichuan, 646000, China
| | - Nan Jiang
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, Sichuan, 646000, China
| | - Jing Zeng
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, Sichuan, 646000, China
| | - Xiang Xie
- School of Basic Medical Sciences, Southwest Medical University, Luzhou, Sichuan, 646000, China
| | - Siyu He
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, Sichuan, 646000, China
| | - Feihong Huang
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, Sichuan, 646000, China
| | - Chunxiang Zhang
- Education Ministry Key Laboratory of Medical Electrophysiology, Sichuan Key Medical Laboratory of New Drug Discovery and Druggability Evaluation, Luzhou Key Laboratory of Activity Screening and Druggability Evaluation for Chinese Materia Medica, Southwest Medical University, Luzhou, Sichuan, 646000, China.
| | - Jiesi Luo
- School of Basic Medical Sciences, Southwest Medical University, Luzhou, Sichuan, 646000, China.
| | - Jianming Wu
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, Sichuan, 646000, China; School of Basic Medical Sciences, Southwest Medical University, Luzhou, Sichuan, 646000, China; Education Ministry Key Laboratory of Medical Electrophysiology, Sichuan Key Medical Laboratory of New Drug Discovery and Druggability Evaluation, Luzhou Key Laboratory of Activity Screening and Druggability Evaluation for Chinese Materia Medica, Southwest Medical University, Luzhou, Sichuan, 646000, China.
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Swain IX, Kresak AM. Proteins Involved in Focal Cell Adhesion and Podosome Formation Are Differentially Expressed during Colorectal Tumorigenesis in AOM-Treated Rats. Cancers (Basel) 2024; 16:1678. [PMID: 38730628 PMCID: PMC11083089 DOI: 10.3390/cancers16091678] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/10/2024] [Revised: 04/25/2024] [Accepted: 04/25/2024] [Indexed: 05/13/2024] Open
Abstract
Colorectal tumorigenesis involves the development of aberrant crypt foci (ACF) or preneoplastic lesions, representing the earliest morphological lesion visible in colon cancer. The purpose of this study was to determine changes in protein expression in carcinogen-induced ACF as they mature and transform into adenomas. Protein expression profiles of azoxymethane (AOM)-induced F344 rat colon ACF and adenomas were compared at four time points, 4 (control), 8, 16, and 24 weeks post AOM administration (n = 9/group), with time points correlating with induction and transformation events. At each time point, micro-dissected ACF and/or adenoma tissues were analyzed across multiple quantitative two-dimensional (2D-DIGE) gels using a Cy-dye labeling technique and a pooled internal standard to quantify expression changes with statistical confidence. Western blot and subsequent network pathway mapping were used to confirm and elucidate differentially expressed (p ≤ 0.05) proteins, including changes in vinculin (Vcl; p = 0.007), scinderin (Scin; p = 0.02), and profilin (Pfn1; p = 0.01), By determining protein expression changes in ACF as they mature and transform into adenomas, a "baseline" of altered regulatory proteins associated with adenocarcinoma development in this model has been elucidated. These data will enable future studies aimed at biomarker identification and understanding the molecular biology of intestinal tumorigenesis and adenocarcinoma maturation under varying intestinal conditions.
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Affiliation(s)
- Ian X. Swain
- Department of Pathology, School of Medicine, Case Western Reserve University, 2103 Cornell Road, Cleveland, OH 44106, USA;
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Lin X, Zhao Z, Sun SP, Liu W. Scinderin promotes glioma cell migration and invasion via remodeling actin cytoskeleton. World J Clin Oncol 2024; 15:32-44. [PMID: 38292665 PMCID: PMC10823943 DOI: 10.5306/wjco.v15.i1.32] [Citation(s) in RCA: 2] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/08/2023] [Revised: 11/20/2023] [Accepted: 12/19/2023] [Indexed: 01/23/2024] Open
Abstract
BACKGROUND Glioma is one of the most common intracranial tumors, characterized by invasive growth and poor prognosis. Actin cytoskeletal rearrangement is an essential event of tumor cell migration. The actin dynamics-related protein scinderin (SCIN) has been reported to be closely related to tumor cell migration and invasion in several cancers. AIM To investigate the role and mechanism of SCIN in glioma. METHODS The expression and clinical significance of SCIN in glioma were analyzed based on public databases. SCIN expression was examined using real-time quantitative polymerase chain reaction and Western blotting. Gene silencing was performed using short hairpin RNA transfection. Cell viability, migration, and invasion were assessed using cell counting kit 8 assay, wound healing, and Matrigel invasion assays, respectively. F-actin cytoskeleton organization was assessed using F-actin staining. RESULTS SCIN expression was significantly elevated in glioma, and high levels of SCIN were associated with advanced tumor grade and wild-type isocitrate dehydrogenase. Furthermore, SCIN-deficient cells exhibited decreased proliferation, migration, and invasion in U87 and U251 cells. Moreover, knockdown of SCIN inhibited the RhoA/focal adhesion kinase (FAK) signaling to promote F-actin depolymerization in U87 and U251 cells. CONCLUSION SCIN modulates the actin cytoskeleton via activating RhoA/FAK signaling, thereby promoting the migration and invasion of glioma cells. This study identified the cancer-promoting effect of SCIN and provided a potential therapeutic target for the treatment of glioma.
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Affiliation(s)
- Xin Lin
- Department of Neurosurgery, Tianjin Huanhu Hospital, Tianjin 300000, China
| | - Zhao Zhao
- Department of Neurosurgery, Tianjin Huanhu Hospital, Tianjin 300000, China
| | - Shu-Peng Sun
- Department of Neurosurgery, Tianjin Huanhu Hospital, Tianjin 300000, China
| | - Wei Liu
- Department of Neurosurgery, Tianjin Huanhu Hospital, Tianjin 300000, China
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Li Y, Tang L, Dang G, Ma M, Tang X. Scinderin Promotes Hydrogen Peroxide-induced Lens Epithelial Cell Injury in Age-related Cataract. Curr Mol Med 2024; 24:1426-1436. [PMID: 37936437 DOI: 10.2174/0115665240250050231030110542] [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: 02/20/2023] [Revised: 09/21/2023] [Accepted: 09/25/2023] [Indexed: 11/09/2023]
Abstract
BACKGROUND Scinderin (SCIN) is a calcium-dependent protein implicated in cell growth and apoptosis by regulating actin cleavage and capping. In this study, we investigated the role of SCIN in hydrogen peroxide-induced lens epithelial cell (LEC) injury related to age-related cataract (ARC). METHODS Anterior lens capsules from ARC patients were collected to examine SCIN expression levels. Immortalized human LEC cell line SRA01/04 and lens capsules freshly isolated from mice were induced by H2O2 to mimic the oxidative stress in ARC. The role of SCIN was investigated by gain-of-function (overexpression) and loss-offunction (knockdown) experiments. Flow cytometry (FCM) and Western-blot (WB) assays were performed to investigate the effect of SCIN on apoptosis. The oxidative stress (OS) was examined by detecting malondialdehyde (MDA) level, superoxide dismutase (SOD) and catalase (CAT) activity. The interaction between SCIN mRNA and miR-489-3p was predicted by StarBase and miRDB databases and validated by luciferase reporter activity assay. RESULTS SCIN was significantly elevated in cataract samples, and the expression levels were positively correlated with the nuclear sclerosis grades. SCIN overexpression promoted OS and apoptosis in H2O2-induced SRA01/04 cells, while SCIN silencing showed the opposite effect. We further showed that miR-489-3p was a negative regulator of SCIN. miR-489-3p overexpression suppressed apoptosis and OS in H2O2-induced SRA01/04 cells by targeting SCIN. CONCLUSION Our study identified SCIN as an upregulated gene in ARC, which is negatively regulated by miR-489-3p. Targeting miR-489-3p/SCIN axis could attenuate OS-induced apoptosis in LECs.
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Affiliation(s)
- Yan Li
- Shaanxi Eye Hospital, Xi'an People's Hospital (Xi'an Fourth Hospital), Affiliated People's Hospital Northwest University, Xi'an 710004 China
| | - Li Tang
- Shaanxi Eye Hospital, Xi'an People's Hospital (Xi'an Fourth Hospital), Affiliated People's Hospital Northwest University, Xi'an 710004 China
| | - Guanxing Dang
- Shaanxi Eye Hospital, Xi'an People's Hospital (Xi'an Fourth Hospital), Affiliated People's Hospital Northwest University, Xi'an 710004 China
| | - Mengyuan Ma
- Shaanxi Eye Hospital, Xi'an People's Hospital (Xi'an Fourth Hospital), Affiliated People's Hospital Northwest University, Xi'an 710004 China
| | - Xingfang Tang
- Shaanxi Eye Hospital, Xi'an People's Hospital (Xi'an Fourth Hospital), Affiliated People's Hospital Northwest University, Xi'an 710004 China
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Wang K, Kong F, Qiu Y, Chen T, Fu J, Jin X, Su Y, Gu Y, Hu Z, Li J. Autophagy regulation and protein kinase activity of PIK3C3 controls sertoli cell polarity through its negative regulation on SCIN (scinderin). Autophagy 2023; 19:2934-2957. [PMID: 37450577 PMCID: PMC10549198 DOI: 10.1080/15548627.2023.2235195] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/24/2022] [Revised: 06/25/2023] [Accepted: 07/06/2023] [Indexed: 07/18/2023] Open
Abstract
Sertoli cells are highly polarized testicular cells that provide a nurturing environment for germ cell development and maturation during spermatogenesis. The class III phosphatidylinositol 3-kinase (PtdIns3K) plays core roles in macroautophagy in various cell types; however, its role in Sertoli cells remains unclear. Here, we generated a mouse line in which the gene encoding the catalytic subunit, Pik3c3, was specifically deleted in Sertoli cells (cKO) and found that after one round of normal spermatogenesis, the cKO mice quickly became infertile and showed disruption of Sertoli cell polarity and impaired spermiogenesis. Subsequent proteomics and phosphoproteomics analyses enriched the F-actin cytoskeleton network involved in the disorganized Sertoli-cell structure in cKO testis which we identified a significant increase of the F-actin negative regulator SCIN (scinderin) and the reduced phosphorylation of HDAC6, an α-tubulin deacetylase. Our results further demonstrated that the accumulation of SCIN in cKO Sertoli cells caused the disorder and disassembly of the F-actin cytoskeleton, which was related to the failure of SCIN degradation through the autophagy-lysosome pathway. Additionally, we found that the phosphorylation of HDAC6 at site S59 by PIK3C3 was essential for its degradation through the ubiquitin-proteasome pathway. As a result, the HDAC6 that accumulated in cKO Sertoli cells deacetylated SCIN at site K189 and led to a disorganized F-actin cytoskeleton. Taken together, our findings elucidate a new mechanism for PIK3C3 in maintaining the polarity of Sertoli cells, in which both its autophagy regulation or protein kinase activities are required for the stabilization of the actin cytoskeleton.Abbreviations: ACTB: actin, beta; AR: androgen receptor; ATG14: autophagy related 14; BafA1: bafilomycin A1; BECN1: beclin 1, autophagy related; BTB: blood-testis barrier; CASP3: caspase 3; CDC42: cell division cycle 42; CDH2: cadherin 2; CHX: cycloheximide; CTNNA1: catenin (cadherin associated protein), alpha 1; CYP11A1: cytochrome P450, family 11, subfamily A, polypeptide 1; EBSS: Earle's balanced salt solution; ES: ectoplasmic specialization; FITC: fluorescein isothiocyanate; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GCNA: germ cell nuclear acidic protein; GJA1: gap junction protein, alpha 1; H2AX: H2A.X variant histone; HDAC6: histone deacetylase 6; KIT: KIT proto-oncogene, receptor tyrosine kinase; LAMP1: lysosomal associated membrane protein 1; MAP3K5: mitogen-activated protein kinase kinase kinase 5; MAP1LC3B: microtubule associated protein 1 light chain 3 beta; OCLN: occludin; PIK3C3: phosphatidylinositol 3-kinase catalytic subunit type 3; PIK3R4: phosphoinositide-3-kinase regulatory subunit 4; PNA: arachis hypogaea lectin; RAC1: Rac family small GTPase 1; SCIN: scinderin; SQSTM1/p62: sequestosome 1; SSC: spermatogonia stem cell; STK11: serine/threonine kinase 11; TJP1: tight junction protein 1; TubA: tubastatin A; TUBB3: tubulin beta 3 class III; TUNEL: TdT-mediated dUTP nick-end labeling; UB: ubiquitin; UVRAG: UV radiation resistance associated gene; VIM: vimentin; WT1: WT1 transcription factor; ZBTB16: zinc finger and BTB domain containing 16.
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Affiliation(s)
- Kehan Wang
- State Key Laboratory of Reproductive Medicine and Offspring Health, Nanjing Medical University, Nanjing, Jiangsu, China
| | - Feifei Kong
- Assisted Reproduction Unit, Department of Obstetrics and Gynecology, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, China
| | - Yuexin Qiu
- State Key Laboratory of Reproductive Medicine and Offspring Health, Nanjing Medical University, Nanjing, Jiangsu, China
| | - Tao Chen
- State Key Laboratory of Reproductive Medicine and Offspring Health, Nanjing Medical University, Nanjing, Jiangsu, China
| | - Jiayi Fu
- State Key Laboratory of Reproductive Medicine and Offspring Health, Nanjing Medical University, Nanjing, Jiangsu, China
| | - Xin Jin
- Department of Center of Reproductive Medicine, Wuxi Maternity and Child Health Care Hospital, Nanjing Medical University, Wuxi, Jiangsu, China
| | - Youqiang Su
- Shandong Provincial Key Laboratory of Animal Cells and Developmental Biology, School of Life Sciences, Shandong University, Qingdao, Shandong, China
| | - Yayun Gu
- State Key Laboratory of Reproductive Medicine and Offspring Health, Nanjing Medical University, Nanjing, Jiangsu, China
| | - Zhibin Hu
- State Key Laboratory of Reproductive Medicine and Offspring Health, Nanjing Medical University, Nanjing, Jiangsu, China
- Department of Epidemiology and Biostatistics, International Joint Research Center on Environment and Human Health, Center for Global Health, School of Public Health, Nanjing Medical University, Nanjing, Jiangsu, China
| | - Jing Li
- State Key Laboratory of Reproductive Medicine and Offspring Health, Nanjing Medical University, Nanjing, Jiangsu, China
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Nasimi Shad A, Fanoodi A, Maharati A, Akhlaghipour I, Moghbeli M. Molecular mechanisms of microRNA-301a during tumor progression and metastasis. Pathol Res Pract 2023; 247:154538. [PMID: 37209575 DOI: 10.1016/j.prp.2023.154538] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/26/2023] [Revised: 05/10/2023] [Accepted: 05/16/2023] [Indexed: 05/22/2023]
Abstract
Cancer is known as one of the leading causes of human deaths globally. Late diagnosis is considered as one of the main reasons for the high mortality rate among cancer patients. Therefore, the introduction of early diagnostic tumor markers can improve the efficiency of therapeutic modalities. MicroRNAs (miRNAs) have a key role in regulation of cell proliferation and apoptosis. MiRNAs deregulation has been frequently reported during tumor progressions. Since, miRNAs have a high stability in body fluids; they can be used as the reliable non-invasive tumor markers. Here, we discussed the role of miR-301a during tumor progressions. MiR-301a mainly functions as an oncogene via the modulation of transcription factors, autophagy, epithelial-mesenchymal transition (EMT), and signaling pathways. This review paves the way to suggest miR-301a as a non-invasive marker for the early tumor diagnosis. MiR-301a can also be suggested as an effective target in cancer therapy.
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Affiliation(s)
- Arya Nasimi Shad
- Student Research Committee, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran
| | - Ali Fanoodi
- Student Research Committee, Faculty of Medicine, Birjand University of Medical Sciences, Mashhad, Iran
| | - Amirhosein Maharati
- Student Research Committee, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran
| | - Iman Akhlaghipour
- Student Research Committee, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran
| | - Meysam Moghbeli
- Department of Medical Genetics and Molecular Medicine, School of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran.
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Perez-Castro L, Venkateswaran N, Garcia R, Hao YH, Lafita-Navarro MC, Kim J, Segal D, Saponzik E, Chang BJ, Fiolka R, Danuser G, Xu L, Brabletz T, Conacci-Sorrell M. The AHR target gene scinderin activates the WNT pathway by facilitating the nuclear translocation of β-catenin. J Cell Sci 2022; 135:jcs260028. [PMID: 36148682 PMCID: PMC10658791 DOI: 10.1242/jcs.260028] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/22/2022] [Accepted: 09/12/2022] [Indexed: 01/12/2023] Open
Abstract
The ligand-activated transcription factor aryl hydrocarbon receptor (AHR) regulates cellular detoxification, proliferation and immune evasion in a range of cell types and tissues, including cancer cells. In this study, we used RNA-sequencing to identify the signature of the AHR target genes regulated by the pollutant 2,3,7,8-tetrachlorodibenzodioxin (TCDD) and the endogenous ligand kynurenine (Kyn), a tryptophan-derived metabolite. This approach identified a signature of six genes (CYP1A1, ALDH1A3, ABCG2, ADGRF1 and SCIN) as commonly activated by endogenous or exogenous ligands of AHR in multiple colon cancer cell lines. Among these, the actin-severing protein scinderin (SCIN) was necessary for cell proliferation; SCIN downregulation limited cell proliferation and its expression increased it. SCIN expression was elevated in a subset of colon cancer patient samples, which also contained elevated β-catenin levels. Remarkably, SCIN expression promoted nuclear translocation of β-catenin and activates the WNT pathway. Our study identifies a new mechanism for adhesion-mediated signaling in which SCIN, likely via its ability to alter the actin cytoskeleton, facilitates the nuclear translocation of β-catenin. This article has an associated First Person interview with the first authors of the paper.
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Affiliation(s)
- Lizbeth Perez-Castro
- Department of Cell Biology, UT Southwestern Medical Center, Dallas, TX 75390, USA
| | | | - Roy Garcia
- Department of Cell Biology, UT Southwestern Medical Center, Dallas, TX 75390, USA
| | - Yi-Heng Hao
- Department of Cell Biology, UT Southwestern Medical Center, Dallas, TX 75390, USA
| | - M. C. Lafita-Navarro
- Department of Cell Biology, UT Southwestern Medical Center, Dallas, TX 75390, USA
| | - Jiwoong Kim
- Quantitative Biomedical Research Center, Department of Population & Data Sciences, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA
| | - Dagan Segal
- Department of Cell Biology, UT Southwestern Medical Center, Dallas, TX 75390, USA
- Lyda Hill Department of Bioinformatics, UT Southwestern Medical Center, Dallas, TX 75390, USA
| | - Etai Saponzik
- Lyda Hill Department of Bioinformatics, UT Southwestern Medical Center, Dallas, TX 75390, USA
| | - Bo-Jui Chang
- Lyda Hill Department of Bioinformatics, UT Southwestern Medical Center, Dallas, TX 75390, USA
| | - Reto Fiolka
- Lyda Hill Department of Bioinformatics, UT Southwestern Medical Center, Dallas, TX 75390, USA
| | - Gaudenz Danuser
- Department of Cell Biology, UT Southwestern Medical Center, Dallas, TX 75390, USA
- Lyda Hill Department of Bioinformatics, UT Southwestern Medical Center, Dallas, TX 75390, USA
| | - Lin Xu
- Quantitative Biomedical Research Center, Department of Population & Data Sciences, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA
- Harold C. Simmons Comprehensive Cancer Center, UT Southwestern Medical Center, Dallas, TX 75390, USA
- Department of Pediatrics, Division of Hematology/Oncology, UT Southwestern Medical Center, Dallas, TX 75390, USA
| | - Thomas Brabletz
- Nikolaus-Fiebiger Center for Molecular Medicine, University Erlangen-Nurnberg, Erlangen 91054, Germany
| | - Maralice Conacci-Sorrell
- Department of Cell Biology, UT Southwestern Medical Center, Dallas, TX 75390, USA
- Harold C. Simmons Comprehensive Cancer Center, UT Southwestern Medical Center, Dallas, TX 75390, USA
- Hamon Center for Regenerative Science and Medicine, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA
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Kang Y, Lin J, Wang L, Shen X, Li J, Wu A, Yue L, Wei L, Ye Y, Yang J, Wu J. Hirsutine, a novel megakaryopoiesis inducer, promotes thrombopoiesis via MEK/ERK/FOG1/TAL1 signaling. PHYTOMEDICINE : INTERNATIONAL JOURNAL OF PHYTOTHERAPY AND PHYTOPHARMACOLOGY 2022; 102:154150. [PMID: 35569185 DOI: 10.1016/j.phymed.2022.154150] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 10/18/2021] [Revised: 04/11/2022] [Accepted: 05/02/2022] [Indexed: 06/15/2023]
Abstract
BACKGROUND Thrombocytopenia (TP) remains a challenge in clinical hematology. TP may have serious consequences, such as recurrent skin and mucosal bleeding and increased risk of intracranial and internal organ hemorrhage. However, effective and safe therapeutic drugs for the long-term management of TP are still lacking. PURPOSE This study aimed to identify more effective active compounds for TP therapy. METHODS Liquid chromatography-mass spectrometry-nuclear magnetic resonance analysis was used to confirm the medicinal species and chemical structure of Hirsutine (HS). The proliferation of HS was examined by Cell Counting Kit (CCK-8) assay on cells lines. The effect of HS on megakaryocyte differentiation was analyzed by evaluating the expression of CD41, CD42b, and DNA ploidy via flow cytometry (FCM). The morphology of megakaryocytes and intermediate cells was observed using an optical microscope. K562 cells were then stained with Giemsa and benzidine. qRT-PCR was used to examine the mRNA expression of GATA-1, GATA-2, FOG-1, TAL-1, RUNX-1, NF-E2, and KLF-1 in K562 cells. Protein levels of the transcription factors were analyzed by western blotting. An MEK inhibitor was used to verify the relationship between the MEK/ERK signaling pathway and CD41/CD42b (FCM), FOG-1, and TAL-1. The Kunming thrombocytopenia mouse model was established by X-ray irradiation (4 Gy) and used to test HS activity and related hematopoietic organ index in vivo. Finally, computer simulations of molecular docking were used to predict the binding energies between HS-MEK and HS-ERK. RESULTS We preliminarily identified HS by screening a plant-sourced compound library for natural compounds with megakaryocytic differentiation and maturation (MKD/MKM)-promoting activity. We found that HS not only enhanced MKD/MKM of K562 and Meg01 cells, but also suppressed the decline of peripheral platelet levels in X-ray-induced myelosuppressive mice. In addition, HS promoted MKD via activation of MEK-ERK-FOG1/TAL1 signaling, which may be the key molecular mechanism of HS action in TP treatment. Molecular docking simulations further verified that HS targets the signaling protein MEK with high-affinity. CONCLUSION In this study, we report for the first time that hirsutine boosts MKD/MKM through the MEK/ERK/FOG1/TAL1 signaling pathway and thus represents a promising treatment option for TP.
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Affiliation(s)
- Yaqi Kang
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, China; State Key Laboratory of Experimental Hematology, National Clinical Research Center for Blood Diseases, Institute of Hematology and Blood Diseases Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Tianjin, China
| | - Jing Lin
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, China
| | - Long Wang
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, China
| | - Xin Shen
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, China
| | - Jingyan Li
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, China; Department of Medical Technology, Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai, Thailand
| | - Anguo Wu
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, China
| | - Liang Yue
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, China
| | - Liuping Wei
- Department of Pharmacy, Children's Hospital of Chongqing Medical University, Chongqing, China
| | - Yun Ye
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, China; Department of Pharmacy, Affiliated Hospital of Southwest Medical University, Luzhou, China
| | - Jing Yang
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, China
| | - Jianming Wu
- Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou, China; Department of Pharmacy, Affiliated Hospital of Southwest Medical University, Luzhou, China.
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10
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MEX3A promotes nasopharyngeal carcinoma progression via the miR-3163/SCIN axis by regulating NF-κB signaling pathway. Cell Death Dis 2022; 13:420. [PMID: 35490173 PMCID: PMC9056523 DOI: 10.1038/s41419-022-04871-0] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/16/2021] [Revised: 04/14/2022] [Accepted: 04/20/2022] [Indexed: 11/08/2022]
Abstract
AbstractMex-3 RNA Binding Family Member A (MEX3A) is an RNA-binding protein that plays complex and diverse roles in the development of various malignancies. However, its role and mechanism in nasopharyngeal carcinoma (NPC) remain undefined and were therefore evaluated in this study. By analyzing Gene Expression Omnibus data and using tissue microarrays, we found that MEX3A is significantly upregulated in NPC and negatively associated with prognosis. Notably, MEX3A depletion led to decreased cell proliferation, invasion, and migration, but increased apoptosis in NPC cells in vitro, while inhibiting tumor growth in vivo. Using whole-transcript expression arrays and bioinformatic analysis, we identified scinderin (SCIN) and miR-3163 as potential downstream targets of MEX3A in NPC. The regulatory mechanisms of MEX3A, SCIN and miR-3163 were further investigated using rescue experiments. Importantly, SCIN depletion and miR-3163 inhibition reversed and rescued the oncogenic effects of MEX3A, respectively. Moreover, NF-κB signaling inhibition reversed the oncogenic effects of both SCIN and MEX3A. In summary, our results demonstrate that MEX3A may promote NPC development and progression via the miR-3163/SCIN axis by regulating NF-κB signaling, thus providing a potential target for NPC treatment.
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11
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Huang Y, Du X, Chen X, Chen C, Wang H, Yang Y, Teng L. MiR-301a-5p/SCIN promotes gastric cancer progression via regulating STAT3 and NF-κB signaling. J Cancer 2021; 12:5394-5403. [PMID: 34405002 PMCID: PMC8364655 DOI: 10.7150/jca.59747] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/24/2021] [Accepted: 06/29/2021] [Indexed: 01/12/2023] Open
Abstract
Objective: Gastric cancer (GC) is a type of highly malignant cancer. Although the diagnostic and therapeutic methods are innovating, the outcome of GC patients is still poor. Therefore, our research was carried out to explore potential molecular mechanism in the diagnosis of GC. Materials and methods: Bioinformatics analyses were used to obtain microRNA and target mRNA of interest. The expression level of miR-301a-5p and Scinderin (SCIN) mRNA were detected by quantitative real-time PCR (qRT-PCR). Western blot assay was used to investigate SCIN protein level. Cell Counting Kit-8 assay (CCK-8) and colony formation assay were used to investigate cell proliferation ability. Transwell assay was employed to examine cell motility. The interaction between miR-301a-5p and SCIN mRNA was verified by dual-luciferase reporter assay. Results: The qRT-PCR analysis revealed that the expression of miR-301a-5p was higher in gastric cancer tissues than para-cancer tissues (P<0.05). Cox regression analysis showed upregulated miR-301a-5p was associated with larger tumor size (P=0.036) and more advanced TNM stage (P=0.048). The Kaplan-Meier analysis showed a correlation between increased miR-301a-5p expression and shorter overall survival (OS)(P=0.018). By using bioinformatic analysis, SCIN was predicted as one of the targets of miR-301a-5p. Overexpressing miR-301a-5p promoted proliferation and motility of GC cells while knockdown of SCIN exhibited the same performance. Further, we verified the alteration of miR-301a-5p and SCIN expression level could affect the epithelial-mesenchymal transition (EMT) progression on GC cells via STAT3 and NF-κB signaling. Conclusion: Highly expressed miR-301a-5p was associated with aggressiveness of GC. Upregulation of miR-301a-5p promoted malignant phenotype of GC by targeting SCIN. The present results indicated miR-301a-5p might be a promising molecule in the prognosis of GC.
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Affiliation(s)
- Yingying Huang
- Department of oncological surgery, The First Affiliated Hospital, College of Medicine, Zhejiang University.,Cancer Institute (Key Laboratory for Cancer Intervention and Prevention, China National Ministry of Education, Zhejiang Provincial Key Laboratory of Molecular Biology in Medical Sciences), The Second Affiliated Hospital, Zhejiang University School of Medicine, China
| | - Xiaoxiao Du
- Department of oncological surgery, The First Affiliated Hospital, College of Medicine, Zhejiang University
| | - Xiangliu Chen
- Department of oncological surgery, The First Affiliated Hospital, College of Medicine, Zhejiang University
| | - Chuanzhi Chen
- Department of oncological surgery, The First Affiliated Hospital, College of Medicine, Zhejiang University
| | - Haiyong Wang
- Department of oncological surgery, The First Affiliated Hospital, College of Medicine, Zhejiang University
| | - Yan Yang
- Department of oncological surgery, The First Affiliated Hospital, College of Medicine, Zhejiang University
| | - Lisong Teng
- Department of oncological surgery, The First Affiliated Hospital, College of Medicine, Zhejiang University
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12
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Fernández Muñoz B, Lopez-Navas L, Gonzalez Bermejo M, Lomas Romero IM, Montiel Aguilera MÁ, Campos Cuerva R, Arribas Arribas B, Nogueras S, Carmona Sánchez G, Santos González M. A PROPRIETARY GMP HUMAN PLATELET LYSATE FOR THE EXPANSION OF DERMAL FIBROBLASTS FOR CLINICAL APPLICATIONS. Platelets 2021; 33:98-109. [PMID: 33393414 DOI: 10.1080/09537104.2020.1856356] [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] [Indexed: 01/01/2023]
Abstract
Recent years have witnessed the introduction of ex vivo expanded dermal fibroblasts for several cell therapy and tissue-engineering applications, including the treatment of facial scars and burns, representing a promising cell type for regenerative medicine. We tested different in-house produced human platelet lysate (HPL) solutions against fetal bovine serum as supplements for in vitro fibroblast expansion by comparing cell yield, molecular marker expression, extracellular matrix (ECM) generation, genomic stability and global gene expression. Our in-house produced HPL supported fibroblast growth at levels similar to those for FBS and commercial HPL products and was superior to AB human serum. Cells grown in HPL maintained a fibroblast phenotype (VIM+, CD44+, CD13+, CD90+), ECM generation capacity (FN+, COL1+) and a normal karyotype, although gene expression profiling revealed changes related to cell metabolism, adhesion and cellular senescence. The HPL manufacturing process was validated within a GMP compliant system and the solution was stable at -80ºC and -20ºC for 2 years. Dermal fibroblasts expanded in vitro with HPL maintain a normal karyotype and expression of fibroblast markers, with only minor changes in their global gene expression profile. Our in-house produced GMP-HPL is an efficient, safe and economical cell culture supplement that can help increase the healthcare activity of blood transfusion centers through the re-use of transfusional plasma and platelets approaching their expiration date. Currently, our HPL solution is approved by the Spanish Agency of Medicines and Medical Devices and is being used in the manufacture of cell therapy products.
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Affiliation(s)
- Beatriz Fernández Muñoz
- Unidad de Producción y Reprogramación Celular de Sevilla (UPRC), Red Andaluza de Diseño y Traslación de Terapias Avanzadas (RADyTTA), Seville, Spain.,Departamento de Neurociencia Aplicada, Instituto de Investigaciones Biomédicas de Sevilla (IBIS), Seville, Spain
| | - Luis Lopez-Navas
- Unidad de Coordinación, Red Andaluza de Diseño y Traslación de Terapias Avanzadas (RADyTTA), Seville, Spain
| | - María Gonzalez Bermejo
- Unidad de Producción y Reprogramación Celular de Sevilla (UPRC), Red Andaluza de Diseño y Traslación de Terapias Avanzadas (RADyTTA), Seville, Spain.,Program in Biología Molecular, Biomedicina e Investigación Clínica, University of Seville, Seville, Spain
| | - Isabel María Lomas Romero
- Unidad de Producción y Reprogramación Celular de Sevilla (UPRC), Red Andaluza de Diseño y Traslación de Terapias Avanzadas (RADyTTA), Seville, Spain
| | - Miguel Ángel Montiel Aguilera
- Unidad de Producción y Reprogramación Celular de Sevilla (UPRC), Red Andaluza de Diseño y Traslación de Terapias Avanzadas (RADyTTA), Seville, Spain
| | - Rafael Campos Cuerva
- Unidad de Producción y Reprogramación Celular de Sevilla (UPRC), Red Andaluza de Diseño y Traslación de Terapias Avanzadas (RADyTTA), Seville, Spain.,Program in Biología Molecular, Biomedicina e Investigación Clínica, University of Seville, Seville, Spain.,Centro de Transfusiones, Tejidos y Células de Sevilla (CTTS), Fundación Pública Andaluza para la Gestión de la Investigación en Salud en Sevilla (FISEVI), Seville, Spain
| | - Blanca Arribas Arribas
- Unidad de Producción y Reprogramación Celular de Sevilla (UPRC), Red Andaluza de Diseño y Traslación de Terapias Avanzadas (RADyTTA), Seville, Spain.,Program in Pharmaceutical Technology and Medicine Sciences (Pharmacy), University of Seville, Seville, Spain
| | - Sonia Nogueras
- Departamento de Terapia Celular, Instituto Maimónides de Investigación Biomédica of Córdoba (IMIBIC), Córdoba, Spain.,Unidad de Terapia Celular, Hospital Universitario Reina Sofía, Cordoba, Spain
| | - Gloria Carmona Sánchez
- Unidad de Producción y Reprogramación Celular de Sevilla (UPRC), Red Andaluza de Diseño y Traslación de Terapias Avanzadas (RADyTTA), Seville, Spain.,Unidad de Coordinación, Red Andaluza de Diseño y Traslación de Terapias Avanzadas (RADyTTA), Seville, Spain.,Program in Biomedicine, University of Granada, Granada, Spain
| | - Mónica Santos González
- Unidad de Producción y Reprogramación Celular de Sevilla (UPRC), Red Andaluza de Diseño y Traslación de Terapias Avanzadas (RADyTTA), Seville, Spain.,Centro de Transfusiones, Tejidos y Células de Sevilla (CTTS), Fundación Pública Andaluza para la Gestión de la Investigación en Salud en Sevilla (FISEVI), Seville, Spain
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13
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Dengue virus infection impedes megakaryopoiesis in MEG-01 cells where the virus envelope protein interacts with the transcription factor TAL-1. Sci Rep 2020; 10:19587. [PMID: 33177556 PMCID: PMC7658202 DOI: 10.1038/s41598-020-76350-5] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/16/2020] [Accepted: 10/08/2020] [Indexed: 12/30/2022] Open
Abstract
Dengue virus (DENV) infection causes dengue fever in humans, which can lead to thrombocytopenia showing a marked reduction in platelet counts, and dengue hemorrhagic fever. The virus may cause thrombocytopenia either by destroying the platelets or by interfering with their generation via the process of megakaryopoiesis. MEG-01 is the human megakaryoblastic leukemia cell line that can be differentiated in vitro by phorbol-12-myristate-13-acetate (PMA) treatment to produce platelet-like-particles (PLPs). We have studied DENV infection of MEG-01 cells to understand its effect on megakaryopoiesis and the generation of PLPs. We observed that DENV could infect only naive MEG-01 cells, and differentiated cells were refractory to virus infection/replication. However, DENV-infected MEG-01 cells, when induced for differentiation with PMA, supported an enhanced viral replication. Following the virus infection, the MEG-01 cells showed a marked reduction in the surface expression of platelet markers (CD41, CD42a, and CD61), a decreased polyploidy, and significantly reduced PLP counts. DENV infection caused an enhanced Notch signaling in MEG-01 cells where the virus envelope protein was shown to interact with TAL-1, a host protein important for megakaryopoiesis. These observations provide new insight into the role of DENV in modulating the megakaryopoiesis and platelet production process.
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14
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Hearn JI, Green TN, Chopra M, Nursalim YNS, Ladvanszky L, Knowlton N, Blenkiron C, Poulsen RC, Singleton DC, Bohlander SK, Kalev-Zylinska ML. N-Methyl-D-Aspartate Receptor Hypofunction in Meg-01 Cells Reveals a Role for Intracellular Calcium Homeostasis in Balancing Megakaryocytic-Erythroid Differentiation. Thromb Haemost 2020; 120:671-686. [PMID: 32289863 PMCID: PMC7286128 DOI: 10.1055/s-0040-1708483] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/18/2023]
Abstract
The release of calcium ions (Ca
2+
) from the endoplasmic reticulum (ER) and related store-operated calcium entry (SOCE) regulate maturation of normal megakaryocytes. The
N
-methyl-D-aspartate (NMDA) receptor (NMDAR) provides an additional mechanism for Ca
2+
influx in megakaryocytic cells, but its role remains unclear. We created a model of NMDAR hypofunction in Meg-01 cells using CRISPR-Cas9 mediated knockout of the
GRIN1
gene, which encodes an obligate, GluN1 subunit of the NMDAR. We found that compared with unmodified Meg-01 cells, Meg-01-
GRIN1−/−
cells underwent atypical differentiation biased toward erythropoiesis, associated with increased basal ER stress and cell death. Resting cytoplasmic Ca
2+
levels were higher in Meg-01-
GRIN1−/−
cells, but ER Ca
2+
release and SOCE were lower after activation. Lysosome-related organelles accumulated including immature dense granules that may have contributed an alternative source of intracellular Ca
2+
. Microarray analysis revealed that Meg-01-
GRIN1−/−
cells had deregulated expression of transcripts involved in Ca
2+
metabolism, together with a shift in the pattern of hematopoietic transcription factors toward erythropoiesis. In keeping with the observed pro-cell death phenotype induced by
GRIN1
deletion, memantine (NMDAR inhibitor) increased cytotoxic effects of cytarabine in unmodified Meg-01 cells. In conclusion, NMDARs comprise an integral component of the Ca
2+
regulatory network in Meg-01 cells that help balance ER stress and megakaryocytic-erythroid differentiation. We also provide the first evidence that megakaryocytic NMDARs regulate biogenesis of lysosome-related organelles, including dense granules. Our results argue that intracellular Ca
2+
homeostasis may be more important for normal megakaryocytic and erythroid differentiation than currently recognized; thus, modulation may offer therapeutic opportunities.
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Affiliation(s)
- James I Hearn
- Department of Molecular Medicine and Pathology, School of Medical Sciences, University of Auckland, Auckland, New Zealand
| | - Taryn N Green
- Department of Molecular Medicine and Pathology, School of Medical Sciences, University of Auckland, Auckland, New Zealand
| | - Martin Chopra
- Department of Molecular Medicine and Pathology, School of Medical Sciences, University of Auckland, Auckland, New Zealand
| | - Yohanes N S Nursalim
- Department of Molecular Medicine and Pathology, School of Medical Sciences, University of Auckland, Auckland, New Zealand
| | - Leandro Ladvanszky
- Department of Molecular Medicine and Pathology, School of Medical Sciences, University of Auckland, Auckland, New Zealand
| | - Nicholas Knowlton
- Department of Molecular Medicine and Pathology, School of Medical Sciences, University of Auckland, Auckland, New Zealand
| | - Cherie Blenkiron
- Department of Molecular Medicine and Pathology, School of Medical Sciences, University of Auckland, Auckland, New Zealand
| | - Raewyn C Poulsen
- Department of Medicine, School of Medicine, University of Auckland, Auckland, New Zealand.,Department of Pharmacology and Clinical Pharmacology, School of Medical Sciences, University of Auckland, Auckland, New Zealand
| | - Dean C Singleton
- Auckland Cancer Society Research Centre, University of Auckland, Auckland, New Zealand
| | - Stefan K Bohlander
- Department of Molecular Medicine and Pathology, School of Medical Sciences, University of Auckland, Auckland, New Zealand
| | - Maggie L Kalev-Zylinska
- Department of Molecular Medicine and Pathology, School of Medical Sciences, University of Auckland, Auckland, New Zealand.,LabPlus Haematology, Auckland City Hospital, Auckland, New Zealand
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15
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Zhou B, Chen TW, Jiang YB, Wei XB, Lu CD, Li JJ, Xie D, Cheng SQ. Scinderin suppresses cell proliferation and predicts the poor prognosis of hepatocellular carcinoma. Oncol Lett 2020; 19:2011-2020. [PMID: 32194697 DOI: 10.3892/ol.2020.11262] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/19/2019] [Accepted: 11/26/2019] [Indexed: 12/20/2022] Open
Abstract
Hepatocellular carcinoma (HCC) remains an intractable disease despite numerous advancements made in the available treatments over recent decades. Therefore, investigation of the underlying pathogenesis of HCC is urgently required. Our previous microarray result showed that SCIN was generally downregulated in 23 paired tumor/normal tissues. Reverse transcription-quantitative PCR, western blotting and immunohistochemistry were performed in the present study in order to detect the expression of scinderin (SCIN). Lentivirus-mediated gene delivery was used in order to produce SCIN-manipulated cell lines. MTT and crystal violet assays were performed in order to investigate cell growth, and fluorescence-activated cell sorting analysis was used in order to determine cell cycle distribution. SCIN was downregulated in HCC samples, and low SCIN expression predicted the poor prognosis of patients with HCC. Notably, SCIN may have the potential to serve as an independent risk factor for overall survival (3-year overall survival rate of 28.6 and 10.3% in high SCIN expression and low SCIN expression groups, respectively) and disease-free survival (3-year recurrence rate of 71.4 and 84.6% in high SCIN expression and low SCIN expression groups, respectively) in HCC. SCIN inhibited HCC cell proliferation both in vitro and in subcutaneous tumor formation assay. Furthermore, SCIN decreased the levels of phosphorylated STAT3, thereby downregulating cyclin A1 levels in HCC cells. The results of the present study demonstrate the tumor suppressive role of SCIN in HCC, providing a candidate strategy to treat this disease.
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Affiliation(s)
- Bin Zhou
- Department of Hepatic Surgery VI, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai 200438, P.R. China
| | - Tian-Wei Chen
- Laboratory of Molecular Oncology, Institute for Nutritional Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, P.R. China
| | - Ya-Bo Jiang
- Department of Hepatic Surgery VI, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai 200438, P.R. China
| | - Xu-Biao Wei
- Department of Hepatic Surgery VI, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai 200438, P.R. China
| | - Chong-De Lu
- Department of Hepatic Surgery VI, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai 200438, P.R. China
| | - Jing-Jing Li
- Laboratory of Molecular Oncology, Institute for Nutritional Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, P.R. China
| | - Dong Xie
- Laboratory of Molecular Oncology, Institute for Nutritional Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, P.R. China
| | - Shu-Qun Cheng
- Department of Hepatic Surgery VI, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai 200438, P.R. China
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16
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Lin Q, Li J, Zhu D, Niu Z, Pan X, Xu P, Ji M, Wei Y, Xu J. Aberrant Scinderin Expression Correlates With Liver Metastasis and Poor Prognosis in Colorectal Cancer. Front Pharmacol 2019; 10:1183. [PMID: 31736743 PMCID: PMC6836707 DOI: 10.3389/fphar.2019.01183] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/25/2019] [Accepted: 09/13/2019] [Indexed: 12/22/2022] Open
Abstract
Many genes and mutations have been reported for colorectal cancer (CRC); however, very few have been associated with colorectal cancer liver metastasis (CRLM). We performed gene expression profiling experiments to identify genetic markers for CRLM and elucidate the molecular mechanisms. Microarray experiments were performed on CRC primary tumor samples with or without liver metastasis (LM) using the Affymetrix U133 plus 2.0 GeneChip Array. A new identified gene-scinderin (SCIN) was overexpressed with synchronous LM at both the RNA level evaluated with quantitative real-time PCR and protein level evaluated with immunohistochemistry and also with short overall survival analyzed with Kaplan-Meier method. With multivariate analysis indicated that SCIN served as an independent poor prognostic predictor for CRC patients. Disease-free survival was also significantly lower in SCIN overexpressing CRC patients with metachronous LM. In addition, SCIN knockdown significantly reduced cell proliferation, induced cell cycle arrest, and promoted the expression of some cell cycle apoptosis-related protein. Moreover, the DIAPH1, STAT3, CDK2, CDK4, and EGFR levels were downregulated, whereas CDKN2B and COL4A1 were upregulated in DLD-1-shSCIN cells by microarray analysis compared with DLD-1 shCon cells. These findings revealed that SCIN may serve as an important predictor of CRLM and poor outcome for CRC patients. SCIN may be a potential therapeutic target in human CRC. However, translation of its roles into clinical practice will require further investigation and additional experimental validation.
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Affiliation(s)
- Qi Lin
- Department of General Surgery, Zhongshan Hospital, Fudan University, Shanghai, China
| | - Jun Li
- Department of General Surgery, Zhongshan Hospital, Fudan University, Shanghai, China
| | - Dexiang Zhu
- Department of General Surgery, Zhongshan Hospital, Fudan University, Shanghai, China
| | - Zhengchuan Niu
- Department of General Surgery, Zhongshan Hospital, Fudan University, Shanghai, China
| | - Xiangou Pan
- Department of Radiology, Zhongshan Hospital, Fudan University, Shanghai, China
| | - Pingping Xu
- Department of General Surgery, Zhongshan Hospital, Fudan University, Shanghai, China
| | - Meiling Ji
- Department of General Surgery, Zhongshan Hospital, Fudan University, Shanghai, China
| | - Ye Wei
- Department of General Surgery, Zhongshan Hospital, Fudan University, Shanghai, China
| | - Jianmin Xu
- Department of General Surgery, Zhongshan Hospital, Fudan University, Shanghai, China
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17
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Jian W, Zhang X, Wang J, Liu Y, Hu C, Wang X, Liu R. Scinderin-knockdown inhibits proliferation and promotes apoptosis in human breast carcinoma cells. Oncol Lett 2018; 16:3207-3214. [PMID: 30127916 DOI: 10.3892/ol.2018.9009] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/16/2017] [Accepted: 05/11/2018] [Indexed: 01/16/2023] Open
Abstract
Previous studies have reported that scinderin (SCIN) affects multiple cellular processes, including proliferation, migration and differentiation in cancer. However, the specific role of SCIN in breast cancer (BC) cells is unknown. Immunohistochemistry was used to investigate SCIN expression in 46 BC and 21 mammary fibroadenoma or fibroadenomatoid hyperplasia tissue samples. SCIN expression was ablated in MDA-MB-231 and T-47D cells using lentivirus-mediated small interfering RNA technology. Cell proliferation was tested using Celigo and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays. Cell apoptosis was analyzed by measuring Caspase 3/7 activity and annexin-V staining. The results of the present study demonstrated that SCIN expression was elevated in BC tissues compared with mammary fibroadenoma or fibroadenomatoid hyperplasia tissues. Specifically, higher SCIN expression was observed in Ki-67-positive BC tissues (78.6%) compared with Ki-67-negative BC tissues. Furthermore, knockdown of SCIN expression in the BC cell lines significantly suppressed cell proliferation and induced apoptosis. The data presented in the present study indicate that SCIN serves an important role in the development of breast cancer.
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Affiliation(s)
- Wenjing Jian
- Department of Thyroid and Breast Surgery, The Third Affiliated Hospital, Sun Yat-Sen University, Guangzhou, Guangdong 510630, P.R. China.,Department of Thyroid and Breast Surgery, The Second People's Hospital of Shenzhen, Shenzhen, Guangdong 518035, P.R. China
| | - Xiaoli Zhang
- Central Laboratory, The Second People's Hospital of Shenzhen, Shenzhen, Guangdong 518035, P.R. China
| | - Jiguo Wang
- Department of Medical Oncology, Baoan District Traditional Chinese Medicine Hospital, Shenzhen, Guangdong 518133, P.R. China
| | - Yunlong Liu
- Department of Thyroid and Breast Surgery, The Second People's Hospital of Shenzhen, Shenzhen, Guangdong 518035, P.R. China
| | - Chuting Hu
- Department of Thyroid and Breast Surgery, The Second People's Hospital of Shenzhen, Shenzhen, Guangdong 518035, P.R. China
| | - Xianming Wang
- Department of Thyroid and Breast Surgery, The Second People's Hospital of Shenzhen, Shenzhen, Guangdong 518035, P.R. China
| | - Renbin Liu
- Department of Thyroid and Breast Surgery, The Third Affiliated Hospital, Sun Yat-Sen University, Guangzhou, Guangdong 510630, P.R. China
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18
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Zhang ZH, Zhang W, Zhou JD, Zhang TJ, Ma JC, Xu ZJ, Lian XY, Wu DH, Wen XM, Deng ZQ, Lin J, Qian J. Decreased SCIN expression, associated with promoter methylation, is a valuable predictor for prognosis in acute myeloid leukemia. Mol Carcinog 2018; 57:735-744. [PMID: 29457658 DOI: 10.1002/mc.22794] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/07/2017] [Revised: 01/29/2018] [Accepted: 02/12/2018] [Indexed: 12/19/2022]
Abstract
The present study was aimed to investigate SCIN expression as well as promoter methylation and further explore their clinical relevance in acute myeloid leukemia (AML) patients. Real-time quantitative PCR was carried out to detect the expression level of SCIN in 119 AML patients and 37 healthy controls. Real-time quantitative methylation-specific PCR and bisulfite sequencing PCR were carried out to detect SCIN promoter methylation levels in 103 AML patients and 29 controls. As compared with controls, the level of SCIN transcript was significantly down-regulated in AML patients (P = 0.001), and the level of methylated SCIN promoter was significantly higher in AML patients (P = 0.001). Moreover, the level of promoter methylation was weakly negatively correlated with SCIN expression in AML patients (R = -0.265, P = 0.027). Demethylation of SCIN promoter by 5-aza-2'-deoxycytidine could restore its expression in leukemic cell line THP1. The age of SCINlow patients was significantly higher and C/EBPA mutation was significantly less than SCINhigh patients (P = 0.039 and 0.038, respectively). Moreover, the rate of complete remission (CR) of SCINlow patients was significantly lower than SCINhigh patients (P = 0.009). Kaplan-Meier analysis showed that low SCIN expression was associated with shorter overall survival (P = 0.036). Cox regression analysis demonstrated low SCIN expression was an independent poor prognostic factor (P = 0.047). Furthermore, SCIN expression was restored in those patients who achieved CR after induction therapy (P = 0.003). These findings indicate that decreased SCIN expression associated with its promoter methylation is a valuable biomarker for predicting adverse prognosis in AML patients.
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Affiliation(s)
- Zhi-Hui Zhang
- Department of Hematology, Affiliated People's Hospital of Jiangsu University, Zhenjiang, Jiangsu, People's Republic of China.,The Key Lab of Precision Diagnosis and Treatment of Zhenjiang City, Zhenjiang, Jiangsu, People's Republic of China
| | - Wei Zhang
- Department of Hematology, Affiliated People's Hospital of Jiangsu University, Zhenjiang, Jiangsu, People's Republic of China.,The Key Lab of Precision Diagnosis and Treatment of Zhenjiang City, Zhenjiang, Jiangsu, People's Republic of China
| | - Jing-Dong Zhou
- Department of Hematology, Affiliated People's Hospital of Jiangsu University, Zhenjiang, Jiangsu, People's Republic of China.,The Key Lab of Precision Diagnosis and Treatment of Zhenjiang City, Zhenjiang, Jiangsu, People's Republic of China
| | - Ting-Juan Zhang
- Department of Hematology, Affiliated People's Hospital of Jiangsu University, Zhenjiang, Jiangsu, People's Republic of China.,The Key Lab of Precision Diagnosis and Treatment of Zhenjiang City, Zhenjiang, Jiangsu, People's Republic of China
| | - Ji-Chun Ma
- Department of Hematology, Affiliated People's Hospital of Jiangsu University, Zhenjiang, Jiangsu, People's Republic of China.,The Key Lab of Precision Diagnosis and Treatment of Zhenjiang City, Zhenjiang, Jiangsu, People's Republic of China
| | - Zi-Jun Xu
- The Key Lab of Precision Diagnosis and Treatment of Zhenjiang City, Zhenjiang, Jiangsu, People's Republic of China.,Laboratory Center, Affiliated People's Hospital of Jiangsu University, Zhenjiang, Jiangsu, People's Republic of China
| | - Xin-Yue Lian
- Department of Hematology, Affiliated People's Hospital of Jiangsu University, Zhenjiang, Jiangsu, People's Republic of China.,The Key Lab of Precision Diagnosis and Treatment of Zhenjiang City, Zhenjiang, Jiangsu, People's Republic of China
| | - De-Hong Wu
- Department of Hematology, The Third People's Hospital of KunShan City, Kunshan, Jiangsu, People's Republic of China
| | - Xiang-Mei Wen
- The Key Lab of Precision Diagnosis and Treatment of Zhenjiang City, Zhenjiang, Jiangsu, People's Republic of China.,Laboratory Center, Affiliated People's Hospital of Jiangsu University, Zhenjiang, Jiangsu, People's Republic of China
| | - Zhao-Qun Deng
- The Key Lab of Precision Diagnosis and Treatment of Zhenjiang City, Zhenjiang, Jiangsu, People's Republic of China.,Laboratory Center, Affiliated People's Hospital of Jiangsu University, Zhenjiang, Jiangsu, People's Republic of China
| | - Jiang Lin
- The Key Lab of Precision Diagnosis and Treatment of Zhenjiang City, Zhenjiang, Jiangsu, People's Republic of China.,Laboratory Center, Affiliated People's Hospital of Jiangsu University, Zhenjiang, Jiangsu, People's Republic of China
| | - Jun Qian
- Department of Hematology, Affiliated People's Hospital of Jiangsu University, Zhenjiang, Jiangsu, People's Republic of China.,The Key Lab of Precision Diagnosis and Treatment of Zhenjiang City, Zhenjiang, Jiangsu, People's Republic of China
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19
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Lomnytska M, Pinto R, Becker S, Engström U, Gustafsson S, Björklund C, Templin M, Bergstrand J, Xu L, Widengren J, Epstein E, Franzén B, Auer G. Platelet protein biomarker panel for ovarian cancer diagnosis. Biomark Res 2018; 6:2. [PMID: 29344361 PMCID: PMC5767003 DOI: 10.1186/s40364-018-0118-y] [Citation(s) in RCA: 35] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/05/2017] [Accepted: 01/03/2018] [Indexed: 01/17/2023] Open
Abstract
BACKGROUND Platelets support cancer growth and spread making platelet proteins candidates in the search for biomarkers. METHODS Two-dimensional (2D) gel electrophoresis, Partial Least Squares Discriminant Analysis (PLS-DA), Western blot, DigiWest. RESULTS PLS-DA of platelet protein expression in 2D gels suggested differences between the International Federation of Gynaecology and Obstetrics (FIGO) stages III-IV of ovarian cancer, compared to benign adnexal lesions with a sensitivity of 96% and a specificity of 88%. A PLS-DA-based model correctly predicted 7 out of 8 cases of FIGO stages I-II of ovarian cancer after verification by western blot. Receiver-operator curve (ROC) analysis indicated a sensitivity of 83% and specificity of 76% at cut-off >0.5 (area under the curve (AUC) = 0.831, p < 0.0001) for detecting these cases. Validation on an independent set of samples by DigiWest with PLS-DA differentiated benign adnexal lesions and ovarian cancer, FIGO stages III-IV, with a sensitivity of 70% and a specificity of 83%. CONCLUSION We identified a group of platelet protein biomarker candidates that can quantify the differential expression between ovarian cancer cases as compared to benign adnexal lesions.
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Affiliation(s)
- Marta Lomnytska
- Department of Obstetrics and Gynaecology, Academical Uppsala University Hospital, Uppsala University, SE-751 85 Uppsala, Sweden
- Institute of Women’s and Children’s Health, Karolinska Institute, SE-171 76 Stockholm, Sweden
- Department of Oncology and Pathology, Cancer Centre Karolinska, Karolinska Institute, SE-171 76 Stockholm, Sweden
| | - Rui Pinto
- Department of Epidemiology and Biostatistics, MRC-PHE Centre for Environment and Health, School of Public Health, Imperial College London, St. Mary’s Campus, Norfolk Place, W2 1PG, London, England UK
| | - Susanne Becker
- Department of Oncology and Pathology, Cancer Centre Karolinska, Karolinska Institute, SE-171 76 Stockholm, Sweden
| | - Ulla Engström
- Ludwig Institute for Cancer Research Ltd, Box 595, SE-751 24 Uppsala, Sweden
| | - Sonja Gustafsson
- NeoProteomics AB, Cancer Centre Karolinska, SE-17176 Stockholm, Sweden
| | | | - Markus Templin
- NMI Natural and Medical Sciences Institute at the University of Tübingen, 72770 Reutlingen, Germany
| | - Jan Bergstrand
- Experimental Biomolecular Physics, Department of Applied Physics, Royal Institute of Technology, AlbaNova University Center, SE-106 91 Stockholm, Sweden
| | - Lei Xu
- Experimental Biomolecular Physics, Department of Applied Physics, Royal Institute of Technology, AlbaNova University Center, SE-106 91 Stockholm, Sweden
| | - Jerker Widengren
- Experimental Biomolecular Physics, Department of Applied Physics, Royal Institute of Technology, AlbaNova University Center, SE-106 91 Stockholm, Sweden
| | - Elisabeth Epstein
- Institute of Women’s and Children’s Health, Karolinska Institute, SE-171 76 Stockholm, Sweden
- Department of Obstetrics and Gynaecology, Department of Clinical Science and Education, Södersjukhuset, SE-118 83 Stockholm, Sweden
| | - Bo Franzén
- Department of Oncology and Pathology, Cancer Centre Karolinska, Karolinska Institute, SE-171 76 Stockholm, Sweden
- NeoProteomics AB, Cancer Centre Karolinska, SE-17176 Stockholm, Sweden
| | - Gert Auer
- Department of Oncology and Pathology, Cancer Centre Karolinska, Karolinska Institute, SE-171 76 Stockholm, Sweden
- NeoProteomics AB, Cancer Centre Karolinska, SE-17176 Stockholm, Sweden
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20
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Cao Y, Wang Y, Sprangers S, Picavet DI, Glogauer M, McCulloch CA, Everts V. Deletion of Adseverin in Osteoclasts Affects Cell Structure But Not Bone Metabolism. Calcif Tissue Int 2017; 101:207-216. [PMID: 28389691 PMCID: PMC5498625 DOI: 10.1007/s00223-017-0271-6] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/09/2017] [Accepted: 03/20/2017] [Indexed: 11/30/2022]
Abstract
Adseverin is an actin-severing/capping protein that may contribute to osteoclast differentiation in vitro but its role in bone remodeling of healthy animals is not defined. We analyzed bone and osteoclast structure in adseverin conditional null mice at alveolar and long bone sites. In wild-type and adseverin null mice, as measured by dual-energy X-ray absorptiometry, there were no differences of bone mineral content or bone mineral density, indicating no change of bone metabolism. In tibiae, TRAcP+ osteoclasts were formed in comparable numbers in adseverin null and wild-type mice. Ultrastructural analysis showed normal and similar abundance of ruffled borders, sealing zones, and mitochondria, and with no difference of osteoclast nuclear numbers. In contrast, analyses of long bone showed that in the absence of adseverin osteoclasts were smaller (120 ± 13 vs. 274 ± 19 µm2; p < 0.05), as were nuclear size and the surface area of cytoplasm. The nuclei of adseverin null osteoclasts exhibited more heterochromatin (31 ± 3%) than wild-type cells (8 ± 1%), suggesting that adseverin affects cell differentiation. The data indicate that in healthy, developing tissues, adseverin contributes to the regulation of osteoclast structure but not to bone metabolism in vivo.
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Affiliation(s)
- Yixuan Cao
- Department of Oral Cell Biology and Functional Anatomy, Academic Centre for Dentistry Amsterdam (ACTA), Research Institute MOVE, University of Amsterdam and VU University Amsterdam, 11N-43, Gustav Mahlerlaan 3004, 1081 LA, Amsterdam, The Netherlands.
| | - Yongqiang Wang
- Matrix Dynamics Group, Faculty of Dentistry, University of Toronto, Ontario, Canada
| | - Sara Sprangers
- Department of Oral Cell Biology and Functional Anatomy, Academic Centre for Dentistry Amsterdam (ACTA), Research Institute MOVE, University of Amsterdam and VU University Amsterdam, 11N-43, Gustav Mahlerlaan 3004, 1081 LA, Amsterdam, The Netherlands
| | - Daisy I Picavet
- Department of Cell Biology and Histology, Core Facility Cellular Imaging, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands
| | - Michael Glogauer
- Matrix Dynamics Group, Faculty of Dentistry, University of Toronto, Ontario, Canada
| | | | - Vincent Everts
- Department of Oral Cell Biology and Functional Anatomy, Academic Centre for Dentistry Amsterdam (ACTA), Research Institute MOVE, University of Amsterdam and VU University Amsterdam, 11N-43, Gustav Mahlerlaan 3004, 1081 LA, Amsterdam, The Netherlands
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21
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FocusHeuristics - expression-data-driven network optimization and disease gene prediction. Sci Rep 2017; 7:42638. [PMID: 28205611 PMCID: PMC5311990 DOI: 10.1038/srep42638] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/05/2016] [Accepted: 01/10/2017] [Indexed: 12/27/2022] Open
Abstract
To identify genes contributing to disease phenotypes remains a challenge for bioinformatics. Static knowledge on biological networks is often combined with the dynamics observed in gene expression levels over disease development, to find markers for diagnostics and therapy, and also putative disease-modulatory drug targets and drugs. The basis of current methods ranges from a focus on expression-levels (Limma) to concentrating on network characteristics (PageRank, HITS/Authority Score), and both (DeMAND, Local Radiality). We present an integrative approach (the FocusHeuristics) that is thoroughly evaluated based on public expression data and molecular disease characteristics provided by DisGeNet. The FocusHeuristics combines three scores, i.e. the log fold change and another two, based on the sum and difference of log fold changes of genes/proteins linked in a network. A gene is kept when one of the scores to which it contributes is above a threshold. Our FocusHeuristics is both, a predictor for gene-disease-association and a bioinformatics method to reduce biological networks to their disease-relevant parts, by highlighting the dynamics observed in expression data. The FocusHeuristics is slightly, but significantly better than other methods by its more successful identification of disease-associated genes measured by AUC, and it delivers mechanistic explanations for its choice of genes.
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22
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Oittinen M, Popp A, Kurppa K, Lindfors K, Mäki M, Kaikkonen MU, Viiri K. Polycomb Repressive Complex 2 Enacts Wnt Signaling in Intestinal Homeostasis and Contributes to the Instigation of Stemness in Diseases Entailing Epithelial Hyperplasia or Neoplasia. Stem Cells 2016; 35:445-457. [PMID: 27570105 DOI: 10.1002/stem.2479] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/07/2016] [Revised: 07/11/2016] [Accepted: 07/29/2016] [Indexed: 12/15/2022]
Abstract
Canonical Wnt/β-catenin signaling regulates the homeostasis of intestinal epithelium by controlling the balance between intestinal stem cell self-renewal and differentiation but epigenetic mechanisms enacting the process are not known. We hypothesized that epigenetic regulator, Polycomb Repressive Complex-2 (PRC2), is involved in Wnt-mediated epithelial homeostasis on the crypt-villus axis and aberrancies therein are implicated both in celiac disease and in intestinal malignancies. We found that PRC2 establishes repressive crypt and villus specific trimethylation of histone H3 lysine 27 (H3K27me3) signature on genes responsible for, for example, nutrient transport and cell killing in crypts and, for example, proliferation and differentiation in mature villi, suggesting that PRC2 facilitates the Wnt-governed intestinal homeostasis. When celiac patients are on gluten-containing diet PRC2 is out-of-bounds active and consequently its target genes were found affected in intestinal epithelium. Significant set of effective intestinal PRC2 targets are also differentially expressed in colorectal adenoma and carcinomas. Our results suggest that PRC2 gives rise and maintains polar crypt and villus specific H3K27me3 signatures. As H3K27me3 is a mark enriched in developmentally important genes, identified intestinal PRC2 targets are possibly imperative drivers for enterocyte differentiation and intestinal stem cell maintenance downstream to Wnt-signaling. Our work also elucidates the mechanism sustaining the crypt hyperplasia in celiac disease and suggest that PRC2-dependent fostering of epithelial stemness is a common attribute in intestinal diseases in which epithelial hyperplasia or neoplasia prevails. Finally, this work demonstrates that in intestine PRC2 represses genes having both pro-stemness and pro-differentiation functions, fact need to be considered when designing epigenetic therapies including PRC2 as a drug target. Stem Cells 2017;35:445-457.
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Affiliation(s)
- Mikko Oittinen
- Tampere Centre for Child Health Research, University of Tampere, Department of Pediatrics and Tampere University Hospital, Tampere, Finland
| | - Alina Popp
- Tampere Centre for Child Health Research, University of Tampere, Department of Pediatrics and Tampere University Hospital, Tampere, Finland.,University of Medicine and Pharmacy "Carol Davila", Department of Pediatrics and Institute for Mother and Child Care, Bucharest, Romania
| | - Kalle Kurppa
- Tampere Centre for Child Health Research, University of Tampere, Department of Pediatrics and Tampere University Hospital, Tampere, Finland
| | - Katri Lindfors
- Tampere Centre for Child Health Research, University of Tampere, Department of Pediatrics and Tampere University Hospital, Tampere, Finland
| | - Markku Mäki
- Tampere Centre for Child Health Research, University of Tampere, Department of Pediatrics and Tampere University Hospital, Tampere, Finland
| | - Minna U Kaikkonen
- Department of Biotechnology and Molecular Medicine, A.I. Virtanen Institute for Molecular Sciences, University of Eastern Finland, Kuopio, Finland
| | - Keijo Viiri
- Tampere Centre for Child Health Research, University of Tampere, Department of Pediatrics and Tampere University Hospital, Tampere, Finland
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23
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Scinderin promotes the invasion and metastasis of gastric cancer cells and predicts the outcome of patients. Cancer Lett 2016; 376:110-7. [PMID: 27033455 DOI: 10.1016/j.canlet.2016.03.035] [Citation(s) in RCA: 41] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/14/2016] [Revised: 03/20/2016] [Accepted: 03/21/2016] [Indexed: 12/31/2022]
Abstract
Invasion and metastasis are major malignant characteristics of human gastric cancer (GC), but the underlying molecular mechanisms are poorly understood. Recent studies have shown that scinderin (SCIN), an actin severing and capping protein that regulates the actin cytoskeleton, is involved in the proliferation and migration of certain cancer cells. Accordingly, this study aimed to investigate the potential role of SCIN in the invasion and metastasis of human GC cells and to evaluate its prognostic value for GC patients. We found that high levels of SCIN expression in GC tumors were correlated with poor overall survival of patients. Silencing of SCIN effectively suppressed the migratory and invasive capabilities of human GC cells in vitro and tumorigenicity and metastasis in vivo. Furthermore, knockdown of SCIN markedly inhibited the formation of filopodia, decreasing GC cell migration and the expression of Cdc42, an important regulator of filopodia by GC cells. These findings suggest that SCIN may be a novel prognostic marker and a potential therapeutic target in human GC.
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24
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Adseverin mediates RANKL-induced osteoclastogenesis by regulating NFATc1. Exp Mol Med 2015; 47:e199. [PMID: 26642432 PMCID: PMC4686697 DOI: 10.1038/emm.2015.94] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/23/2015] [Revised: 09/07/2015] [Accepted: 09/17/2015] [Indexed: 11/08/2022] Open
Abstract
Adseverin is a Ca2+-dependent actin filament-severing protein that has been reported to regulate exocytosis via rearrangements of the actin cytoskeleton in secretory cells. However, the role of adseverin in bone cells has not yet been well characterized. Here, we investigated the role of adseverin in osteoclastogenesis using primary osteoclast precursor cells. Adseverin expression was upregulated during RANKL (receptor activator of nuclear factor-κB ligand)-induced osteoclast differentiation. Moreover, genetic silencing of adseverin decreased the number of osteoclasts generated by RANKL. Adseverin knockdown also suppressed the RANKL-mediated induction of nuclear factor of activated T-cell c1 (NFATc1), which is a key transcription factor in osteoclastogenesis. In addition, adseverin knockdown impaired bone resorption and the secretion of bone-degrading enzymes from osteoclasts. These effects were accompanied by decreased NFATc1 expression and the activation of nuclear factor-κB. Collectively, our results indicate that adseverin has a crucial role in osteoclastogenesis by regulating NFATc1.
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25
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An RNA interference screen identifies new avenues for nephroprotection. Cell Death Differ 2015; 23:608-15. [PMID: 26564400 DOI: 10.1038/cdd.2015.128] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/18/2015] [Revised: 08/03/2015] [Accepted: 08/20/2015] [Indexed: 01/28/2023] Open
Abstract
Acute kidney injury is a major public health problem, which is commonly caused by renal ischemia and is associated with a high risk of mortality and long-term disability. Efforts to develop a treatment for this condition have met with very limited success. We used an RNA interference screen to identify genes (BCL2L14, BLOC1S2, C2ORF42, CPT1A, FBP1, GCNT3, RHOB, SCIN, TACR1, and TNFAIP6) whose suppression improves survival of kidney epithelial cells in in vitro models of oxygen and glucose deprivation. Some of the genes also modulate the toxicity of cisplatin, an anticancer agent whose use is currently limited by nephrotoxicity. Furthermore, pharmacological inhibition of TACR1 product NK1R was protective in a model of mouse renal ischemia, attesting to the in vivo relevance of our findings. These data shed new light on the mechanisms of stress response in mammalian cells, and open new avenues to reduce the morbidity and mortality associated with renal injury.
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26
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Calcium-controlled conformational choreography in the N-terminal half of adseverin. Nat Commun 2015; 6:8254. [PMID: 26365202 PMCID: PMC4647846 DOI: 10.1038/ncomms9254] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/10/2015] [Accepted: 08/03/2015] [Indexed: 01/23/2023] Open
Abstract
Adseverin is a member of the calcium-regulated gelsolin superfamily of actin-binding proteins. Here we report the crystal structure of the calcium-free N-terminal half of adseverin (iA1-A3) and the Ca(2+)-bound structure of A3, which reveal structural similarities and differences with gelsolin. Solution small-angle X-ray scattering combined with ensemble optimization revealed a dynamic Ca(2+)-dependent equilibrium between inactive, intermediate and active conformations. Increasing calcium concentrations progressively shift this equilibrium from a main population of inactive conformation to the active form. Molecular dynamics simulations of iA1-A3 provided insights into Ca(2+)-induced destabilization, implicating a critical role for the A2 type II calcium-binding site and the A2A3 linker in the activation process. Finally, mutations that disrupt the A1/A3 interface increase Ca(2+)-independent F-actin severing by A1-A3, albeit at a lower efficiency than observed for gelsolin domains G1-G3. Together, these data address the calcium dependency of A1-A3 activity in relation to the calcium-independent activity of G1-G3.
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27
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Li X, Jiang H, Huang Y, Gong Q, Wang J, Ling J. Expression and Function of the Actin-severing Protein Adseverin in the Proliferation, Migration, and Differentiation of Dental Pulp Cells. J Endod 2015; 41:493-500. [DOI: 10.1016/j.joen.2014.11.030] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/06/2014] [Revised: 11/29/2014] [Accepted: 11/30/2014] [Indexed: 12/18/2022]
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28
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Liu H, Shi D, Liu T, Yu Z, Zhou C. Lentivirus-mediated silencing of SCIN inhibits proliferation of human lung carcinoma cells. Gene 2014; 554:32-9. [PMID: 25303873 DOI: 10.1016/j.gene.2014.10.013] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/28/2014] [Revised: 09/11/2014] [Accepted: 10/06/2014] [Indexed: 10/25/2022]
Abstract
SCIN (scinderin) is a calcium-dependent actin severing and capping protein. Homologue in zebrafish has been found to be related with cell death. In the present study, we found that SCIN is highly expressed in human lung cancer specimens. However, the role of SCIN in lung cancer has not yet been determined. To investigate the function of SCIN in lung carcinoma cells, we took advantage of lentivirus-mediated RNA interference (RNAi) to knockdown SCIN expression in two lung carcinoma cell lines A549 and H1299. Silencing of SCIN significantly inhibited the proliferation and colony formation ability of both cell lines in vitro. Moreover, flow cytometry analysis showed that knockdown of SCIN led to G0/G1 phase cell cycle arrest as well as an excess accumulation of cells in the sub-G1 phase. Furthermore, depletion of SCIN resulted in a significant increase in Cyclin B1, p21 and PARP expression, and a little decrease in Cyclin D1 expression. These results suggest that SCIN plays an important role in lung carcinoma cell proliferation, and lentivirus-mediated silencing of SCIN might be a potential therapeutic approach for the treatment of lung cancer.
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Affiliation(s)
- Hongxu Liu
- Department of Thoracic Surgery, The First Hospital, China Medical University, Shenyang 110001, China.
| | - Daiwang Shi
- Department of Thoracic Surgery, The First Hospital, China Medical University, Shenyang 110001, China
| | - Tieqin Liu
- Department of Thoracic Surgery, The First Hospital, China Medical University, Shenyang 110001, China
| | - Zhanwu Yu
- Department of Thoracic Surgery, Liaoning Cancer Hospital & Institute, Shenyang 110042, China
| | - Chuanjiang Zhou
- Department of Thoracic Surgery, Benxi Central Hospital, Benxi 117000, China
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29
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Hassanpour S, Jiang H, Wang Y, Kuiper JWP, Glogauer M. The actin binding protein adseverin regulates osteoclastogenesis. PLoS One 2014; 9:e109078. [PMID: 25275604 PMCID: PMC4183545 DOI: 10.1371/journal.pone.0109078] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/05/2013] [Accepted: 09/07/2014] [Indexed: 11/29/2022] Open
Abstract
Adseverin (Ads), a member of the Gelsolin superfamily of actin binding proteins, regulates the actin cytoskeleton architecture by severing and capping existing filamentous actin (F-actin) strands and nucleating the assembly of new F-actin filaments. Ads has been implicated in cellular secretion, exocytosis and has also been shown to regulate chondrogenesis and megakaryoblastic leukemia cell differentiation. Here we report for the first time that Ads is involved in regulating osteoclastogenesis (OCG). Ads is induced during OCG downstream of RANK-ligand (RANKL) stimulation and is highly expressed in mature osteoclasts. The D5 isoform of Ads is not involved in regulating OCG, as its expression is not induced in response to RANKL. Three clonal Ads knockdown RAW264.7 (RAW) macrophage cell lines with varying degrees of Ads expression and OCG deficiency were generated. The most drastic OCG defect was noted in the clonal cell line with the greatest degree of Ads knockdown as indicated by a lack of TRAcP staining and multinucleation. RNAi mediated knockdown of Ads in osteoclast precursors resulted in distinct morphological changes characterized by altered F-actin distribution and increased filopodia formation. Ads knockdown precursor cells experienced enhanced migration while fusion of knockdown precursors cells was limited. Transient reintroduction of de novo Ads back into the knockdown system was capable of rescuing TRAcP expression but not osteoclast multinucleation most likely due to the transient nature of Ads expression. This preliminary study allows us to conclude that Ads is a RANKL induced early regulator of OCG with a potential role in pre-osteoclast differentiation and fusion.
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Affiliation(s)
- Siavash Hassanpour
- Matrix Dynamics Group, Faculty of Dentistry, University of Toronto, Toronto, Ontario, Canada
| | - Hongwei Jiang
- Matrix Dynamics Group, Faculty of Dentistry, University of Toronto, Toronto, Ontario, Canada
- Department of Operative Dentistry and Endodontics, Guanghua School of Stomatology, Sun Yat-sen University, Guangdong Provincial Key Laboratory of Stomatology, Guangzhou, P. R. China
| | - Yongqiang Wang
- Matrix Dynamics Group, Faculty of Dentistry, University of Toronto, Toronto, Ontario, Canada
| | - Johannes W. P. Kuiper
- Matrix Dynamics Group, Faculty of Dentistry, University of Toronto, Toronto, Ontario, Canada
| | - Michael Glogauer
- Matrix Dynamics Group, Faculty of Dentistry, University of Toronto, Toronto, Ontario, Canada
- * E-mail:
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30
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CHEN XIAOMIN, GUO JUNMING, CHEN PING, MAO LIANGANG, FENG WEIYUN, LE DONGHAI, LI KEQIANG. Suppression of scinderin modulates epithelial-mesenchymal transition markers in highly metastatic gastric cancer cell line SGC-7901. Mol Med Rep 2014; 10:2327-33. [DOI: 10.3892/mmr.2014.2523] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/30/2013] [Accepted: 06/17/2014] [Indexed: 11/05/2022] Open
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Lim MN, Hussin NH, Othman A, Umapathy T, Gurbind S, Baharuddin P, Jamal R, Zakaria Z. Comparative global gene expression profile of human limbal stromal cells, bone marrow mesenchymal stromal cells, adipose-derived mesenchymal stromal cells and foreskin fibroblasts. ACTA ACUST UNITED AC 2014. [DOI: 10.7243/2054-717x-1-1] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/15/2022]
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Hasmim M, Badoual C, Vielh P, Drusch F, Marty V, Laplanche A, de Oliveira Diniz M, Roussel H, De Guillebon E, Oudard S, Hans S, Tartour E, Chouaib S. Expression of EPHRIN-A1, SCINDERIN and MHC class I molecules in head and neck cancers and relationship with the prognostic value of intratumoral CD8+ T cells. BMC Cancer 2013; 13:592. [PMID: 24330498 PMCID: PMC3867221 DOI: 10.1186/1471-2407-13-592] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/13/2013] [Accepted: 12/02/2013] [Indexed: 03/17/2023] Open
Abstract
Background Our group has previously shown that EPHRIN-A1 and SCINDERIN expression by tumor cells rendered them resistant to cytotoxic T lymphocyte-mediated lysis. Whereas the prognostic value of EPHRIN-A1 expression in cancer has already been studied, the role of SCINDERIN presence remains to be established. In the present work, we investigated the prognosis value of EPHRIN-A1 and SCINDERIN expression in head and neck carcinomas. In addition, we monitored the HLA-class I expression by tumor cells and the presence of tumor-infiltrating CD8+ T cells to evaluate a putative correlation between these factors and the survival prognosis by themselves or related to EPHRIN-A1 and SCINDERIN expression. Methods Tumor tissue sections of 83 patients with head and neck cancer were assessed by immunohistochemistry for the expression of EPHRIN-A1, SCINDERIN, HLA class I molecules and the presence of CD8+ T cells. Results No significant prognosis value could be attributed to these factors independently, despite a tendency of association between EPHRIN-A1 and a worse clinical outcome. No prognostic value could be observed when CD8+ T cell tumor infiltration was analyzed combined with EPHRIN-A1, SCINDERIN or HLA class I expression. Conclusion These results highlight that molecules involved in cancer cell resistance to cytotoxic T lymphocytes by themselves are not a sufficient criteria for prognosis determination in cancer patients. Other intrinsic or tumor microenvironmental features should be considered in prognostic evaluation.
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Affiliation(s)
| | | | | | | | | | | | | | | | | | | | | | | | - Salem Chouaib
- U753-INSERM, Institut Gustave Roussy, 114 rue Edouard Vaillant, 94800 Villejuif, France.
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Wang D, Sun SQ, Yu YH, Wu WZ, Yang SL, Tan JM. Suppression of SCIN inhibits human prostate cancer cell proliferation and induces G0/G1 phase arrest. Int J Oncol 2013; 44:161-6. [PMID: 24212916 DOI: 10.3892/ijo.2013.2170] [Citation(s) in RCA: 38] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/08/2013] [Accepted: 10/08/2013] [Indexed: 11/05/2022] Open
Abstract
SCIN is a calcium regulated actin severing and capping protein. Its homologue in zebrafish is found to be related with cell death. In the present study, we found that SCIN is highly expressed in human prostate cancer specimens. However, the functions of SCIN in human prostate carcinoma cells are largely unknown. To address the function of SCIN in prostate carcinoma cells, we used lentivirus-mediated RNAi to knock down SCIN expression in PC3 cells, a prostate carcinoma cell line. We found that in vitro silencing of SCIN could inhibit the proliferation and colony formation ability of PC3 cells. Furthermore, cell cycle analysis showed that reduced SCIN expression lead to G0/G1 cell cycle arrest through the regulation of cell cycle-related genes, such as p21Waf1/Cip1, cyclin-dependent kinase inhibitor 2A (CDKN2A, p16Ink4A) and cyclin A2. These results suggest that SCIN plays an important role in the proliferation of prostate cancer cells and lentivirus-mediated inhibition of SCIN expression may be a potential therapeutic method for the treatment of prostate cancer.
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Affiliation(s)
- Dong Wang
- Department of Urology, Fuzhou General Hospital, Fuzhou 350025, P.R. China
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Estrada JC, Torres Y, Benguría A, Dopazo A, Roche E, Carrera-Quintanar L, Pérez RA, Enríquez JA, Torres R, Ramírez JC, Samper E, Bernad A. Human mesenchymal stem cell-replicative senescence and oxidative stress are closely linked to aneuploidy. Cell Death Dis 2013; 4:e691. [PMID: 23807220 PMCID: PMC3702285 DOI: 10.1038/cddis.2013.211] [Citation(s) in RCA: 176] [Impact Index Per Article: 14.7] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/24/2023]
Abstract
In most clinical trials, human mesenchymal stem cells (hMSCs) are expanded in vitro before implantation. The genetic stability of human stem cells is critical for their clinical use. However, the relationship between stem-cell expansion and genetic stability is poorly understood. Here, we demonstrate that within the normal expansion period, hMSC cultures show a high percentage of aneuploid cells that progressively increases until senescence. Despite this accumulation, we show that in a heterogeneous culture the senescence-prone hMSC subpopulation has a lower proliferation potential and a higher incidence of aneuploidy than the non-senescent subpopulation. We further show that senescence is linked to a novel transcriptional signature that includes a set of genes implicated in ploidy control. Overexpression of the telomerase catalytic subunit (human telomerase reverse transcriptase, hTERT) inhibited senescence, markedly reducing the levels of aneuploidy and preventing the dysregulation of ploidy-controlling genes. hMSC-replicative senescence was accompanied by an increase in oxygen consumption rate (OCR) and oxidative stress, but in long-term cultures that overexpress hTERT, these parameters were maintained at basal levels, comparable to unmodified hMSCs at initial passages. We therefore propose that hTERT contributes to genetic stability through its classical telomere maintenance function and also by reducing the levels of oxidative stress, possibly, by controlling mitochondrial physiology. Finally, we propose that aneuploidy is a relevant factor in the induction of senescence and should be assessed in hMSCs before their clinical use.
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Affiliation(s)
- J C Estrada
- Department of Cardiovascular Development and Repair, Fundación Centro Nacional de Investigaciones Cardiovasculares Carlos III (CNIC), Melchor Fernández Almagro 3, Madrid, Spain
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Hur D, Hong S. Cloning and characterization of a fish specific gelsolin family gene, ScinL, in olive flounder (Paralichthys olivaceus). Comp Biochem Physiol B Biochem Mol Biol 2012; 164:89-98. [PMID: 23159325 DOI: 10.1016/j.cbpb.2012.11.002] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/04/2012] [Revised: 11/07/2012] [Accepted: 11/07/2012] [Indexed: 12/23/2022]
Abstract
Scinderin like (ScinL) gene is a unique gelsolin family gene found only in fish. In this study ScinL gene was cloned in olive flounder for the first time and characterized its expression and function. Flounder ScinL cDNA consists of 2911 nucleotides encoding a putative protein of 720 amino acids (79.4 kDa). In phylogenetic analysis, flounder ScinL is closely related to ScinL of zebra fish, anableps, and fugu with the similarity of 51-72%. Fish ScinLs are positioned between gelsolin and scinderin of other species. Flounder ScinL protein has the highly conserved actin and PIP2 binding sites, Ca(2+) coordination site, and a C-terminal latch helix preventing the activation of ScinL protein in the absence of Ca(2+). Putative binding sites for NFAT and AP-1 were found in 5' flanking region. Constitutive ScinL expression was found in most organs and the expression level was higher in gill, head kidney, trunk kidney, spleen and skin than muscle, stomach, intestine and brain. In Q-PCR analysis ScinL and CYP1A1 gene expression were significantly upregulated by BaP in head kidney in vivo and in vitro, and in macrophage cells. Upregulated ScinL expression by BaP was blocked by EGTA, indicating a calcium dependent regulation of ScinL expression.
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Affiliation(s)
- Deokhwe Hur
- Department of Marine Biotechnology, Gangneung Wonju National University, Gangneung 210-702, South Korea
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Abstract
The role of platelets in hemostasis and thrombosis is clearly established; however, the mechanisms by which platelets mediate inflammatory and immune pathways are less well understood. Platelets interact and modulate the function of blood and vascular cells by releasing bioactive molecules. Although the platelet is anucleate, it contains transcripts that may mirror disease. Platelet mRNA is only associated with low-level protein translation; however, platelets have a unique membrane structure allowing for the passage of small molecules, leading to the possibility that its cytoplasmic RNA may be passed to nucleated cells. To examine this question, platelet-like particles with labeled RNA were cocultured with vascular cells. Coculture of platelet-like particles with activated THP-1, monocytic, and endothelial cells led to visual and functional RNA transfer. Posttransfer microarray gene expression analysis of THP-1 cells showed an increase in HBG1/HBG2 and HBA1/HBA2 expression that was directly related to the transfer. Infusion of wild-type platelets into a TLR2-deficient mouse model established in vivo confirmation of select platelet RNA transfer to leukocytes. By specifically transferring green fluorescent protein, we also observed external RNA was functional in the recipient cells. The observation that platelets possess the capacity to transfer cytosolic RNA suggests a new function for platelets in the regulation of vascular homeostasis.
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Sigismund S, Confalonieri S, Ciliberto A, Polo S, Scita G, Di Fiore PP. Endocytosis and signaling: cell logistics shape the eukaryotic cell plan. Physiol Rev 2012; 92:273-366. [PMID: 22298658 DOI: 10.1152/physrev.00005.2011] [Citation(s) in RCA: 243] [Impact Index Per Article: 18.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022] Open
Abstract
Our understanding of endocytosis has evolved remarkably in little more than a decade. This is the result not only of advances in our knowledge of its molecular and biological workings, but also of a true paradigm shift in our understanding of what really constitutes endocytosis and of its role in homeostasis. Although endocytosis was initially discovered and studied as a relatively simple process to transport molecules across the plasma membrane, it was subsequently found to be inextricably linked with almost all aspects of cellular signaling. This led to the notion that endocytosis is actually the master organizer of cellular signaling, providing the cell with understandable messages that have been resolved in space and time. In essence, endocytosis provides the communications and supply routes (the logistics) of the cell. Although this may seem revolutionary, it is still likely to be only a small part of the entire story. A wealth of new evidence is uncovering the surprisingly pervasive nature of endocytosis in essentially all aspects of cellular regulation. In addition, many newly discovered functions of endocytic proteins are not immediately interpretable within the classical view of endocytosis. A possible framework, to rationalize all this new knowledge, requires us to "upgrade" our vision of endocytosis. By combining the analysis of biochemical, biological, and evolutionary evidence, we propose herein that endocytosis constitutes one of the major enabling conditions that in the history of life permitted the development of a higher level of organization, leading to the actuation of the eukaryotic cell plan.
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Affiliation(s)
- Sara Sigismund
- IFOM, Fondazione Istituto FIRC di Oncologia Molecolare, Milan, Italy
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Kim H, Gillis LC, Jarvis JD, Yang S, Huang K, Der S, Barber DL. Tyrosine kinase chromosomal translocations mediate distinct and overlapping gene regulation events. BMC Cancer 2011; 11:528. [PMID: 22204395 PMCID: PMC3295743 DOI: 10.1186/1471-2407-11-528] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/11/2011] [Accepted: 12/28/2011] [Indexed: 12/19/2022] Open
Abstract
Background Leukemia is a heterogeneous disease commonly associated with recurrent chromosomal translocations that involve tyrosine kinases including BCR-ABL, TEL-PDGFRB and TEL-JAK2. Most studies on the activated tyrosine kinases have focused on proximal signaling events, but little is known about gene transcription regulated by these fusions. Methods Oligonucleotide microarray was performed to compare mRNA changes attributable to BCR-ABL, TEL-PDGFRB and TEL-JAK2 after 1 week of activation of each fusion in Ba/F3 cell lines. Imatinib was used to control the activation of BCR-ABL and TEL-PDGFRB, and TEL-JAK2-mediated gene expression was examined 1 week after Ba/F3-TEL-JAK2 cells were switched to factor-independent conditions. Results Microarray analysis revealed between 800 to 2000 genes induced or suppressed by two-fold or greater by each tyrosine kinase, with a subset of these genes commonly induced or suppressed among the three fusions. Validation by Quantitative PCR confirmed that eight genes (Dok2, Mrvi1, Isg20, Id1, gp49b, Cxcl10, Scinderin, and collagen Vα1(Col5a1)) displayed an overlapping regulation among the three tested fusion proteins. Stat1 and Gbp1 were induced uniquely by TEL-PDGFRB. Conclusions Our results suggest that BCR-ABL, TEL-PDGFRB and TEL-JAK2 regulate distinct and overlapping gene transcription profiles. Many of the genes identified are known to be involved in processes associated with leukemogenesis, including cell migration, proliferation and differentiation. This study offers the basis for further work that could lead to an understanding of the specificity of diseases caused by these three chromosomal translocations.
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Affiliation(s)
- Hani Kim
- Campbell Family Cancer Research Institute, Ontario Cancer Institute, University Health Network, 610 University Avenue, Toronto, Ontario M5G 2M9, Canada
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Laiosa MD, Mills JH, Lai ZW, Singh KP, Middleton FA, Gasiewicz TA, Silverstone AE. Identification of stage-specific gene modulation during early thymocyte development by whole-genome profiling analysis after aryl hydrocarbon receptor activation. Mol Pharmacol 2010; 77:773-83. [PMID: 20159946 PMCID: PMC2872972 DOI: 10.1124/mol.109.062497] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/17/2009] [Accepted: 02/12/2010] [Indexed: 11/22/2022] Open
Abstract
The aryl hydrocarbon receptor (AHR) is a basic helix-loop-helix transcription factor, implicated as an important modulator of the immune system and of early thymocyte development. We have shown previously that AHR activation by the environmental contaminant and potent AHR agonist 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) leads to a significant decline in the percentage of S-phase cells in the CD3(-)CD4(-)CD8(-) triple-negative stage (TN) 3 and TN4 T-cell committed thymocytes 9 to 12 h after exposure. In the more immature TN1- or TN2-stage cells, no effect on cell cycle was observed. To identify early molecular targets, which could provide insight into how the AHR acts as a modulator of thymocyte development and cell cycle regulation, we performed gene-profiling experiments using RNA isolated from four intrathymic progenitor populations in which the AHR was activated for 6 or 12 h. This microarray analysis of AHR activation identified 108 distinct gene probes that were significantly modulated in the TN1-4 thymocyte progenitor stages. Although most of the genes identified have specific AHR recognition sequences, only seven genes were altered exclusively in the two T-cell committed stages of early thymocyte development (TN3 and TN4) in which the decline of S-phase cells is seen. Moreover, all seven of these genes were reduced in expression, and five of the seven are associated with cell cycle regulatory processes. These seven genes are novel targets for modulation by the TCDD-activated AHR and may be involved in the observed cell-cycle arrest and suppression of early thymocyte development.
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Affiliation(s)
- Michael D Laiosa
- Department of Environmental Medicine, University of Rochester, Rochester, New York, USA.
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Furness SGB, Whelan F. The pleiotropy of dioxin toxicity--xenobiotic misappropriation of the aryl hydrocarbon receptor's alternative physiological roles. Pharmacol Ther 2009; 124:336-53. [PMID: 19781569 DOI: 10.1016/j.pharmthera.2009.09.004] [Citation(s) in RCA: 44] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/31/2009] [Accepted: 09/01/2009] [Indexed: 10/20/2022]
Abstract
The aryl hydrocarbon receptor is a signal regulated transcription factor that has best been characterised as regulating the xenobiotic response to a variety of planar aromatic hydrocarbons. There is compelling evidence that it mediates most, if not all, of the toxic effects of dioxin (2,3,7,8-tetrachlorodibenzo-p-dioxin). Dioxin exposure results in a wide variety of toxic outcomes including severe wasting syndrome, chloracne, thymic involution, severe immune suppression, reduced fertility, hepatotoxicity, teratogenicity, tumour promotion and death. The pleiotropy of toxic outcomes implies the disruption of a wide range of normal physiological functions. The aryl hydrocarbon receptor has developmentally restricted expression as well as developmental defects in gene-targeted mice. It has a wide range of target genes that do not fit into the classical xenobiotic metabolising gene battery and has recently been shown to interact with NF-kappa B and the estrogen receptor. There is also evidence for its activation in the absence of exogenous ligand, all of which point to various roles outside xenobiotic metabolism. Ligands so far identified display differential activation potential with respect to receptor activity. This article addresses activities of the aryl hydrocarbon receptor that are outside the xenobiotic response. Known physiological roles are discussed as well as how their disruption contributes to the pleiotropic toxicity of TCDD.
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Affiliation(s)
- Sebastian G B Furness
- Drug Discovery Biology Laboratory, Monash Institute of Pharmaceutical Sciences, Monash University, Parkville, Victoria 3052, Australia.
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41
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Tada M, Nagasima T, Udagawa T, Tachikawa M, Sugawara H. Ab initio fragment molecular orbital (FMO) analysis of the structure of the phosphoinositide-binding peptide from gelsolin. ACTA ACUST UNITED AC 2009. [DOI: 10.1016/j.theochem.2008.12.003] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/21/2022]
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Isakari Y, Sogo S, Ishida T, Kawakami T, Ono T, Taki T, Kiwada H. Gene Expression Analysis during Platelet-Like Particle Production in Phorbol Myristate Acetate-Treated MEG-01 Cells. Biol Pharm Bull 2009; 32:354-8. [DOI: 10.1248/bpb.32.354] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Affiliation(s)
- Yoshimasa Isakari
- Department of Pharmacokinetics and Biopharmaceutics, Subdivision of Biopharmaceutical Sciences, Institute of Health Biosciences, The University of Tokushima
- Molecular Medical Science Institute, Otsuka Pharmaceutical Co., Ltd
| | - Shinji Sogo
- Molecular Medical Science Institute, Otsuka Pharmaceutical Co., Ltd
| | - Tatsuhiro Ishida
- Department of Pharmacokinetics and Biopharmaceutics, Subdivision of Biopharmaceutical Sciences, Institute of Health Biosciences, The University of Tokushima
| | - Takuma Kawakami
- Molecular Medical Science Institute, Otsuka Pharmaceutical Co., Ltd
| | - Toshihide Ono
- BioInfomatics Institute, Otsuka Pharmaceutical Co., Ltd
| | - Takao Taki
- Molecular Medical Science Institute, Otsuka Pharmaceutical Co., Ltd
| | - Hiroshi Kiwada
- Department of Pharmacokinetics and Biopharmaceutics, Subdivision of Biopharmaceutical Sciences, Institute of Health Biosciences, The University of Tokushima
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Abstract
Thrombocytopenia is a critical problem that occurs in many hematologic diseases, as well as after cancer therapy and radiation exposure. Platelet transfusion is the most commonly used therapy but has limitations of alloimmunization, availability, and expense. Thus, the development of safe, small, molecules to enhance platelet production would be advantageous for the treatment of thrombocytopenia. Herein, we report that an important lipid mediator and a peroxisome proliferator-activated receptor gamma (PPARgamma) ligand called 15-deoxy-Delta(12,14) prostaglandin J(2) (15d-PGJ(2)), increases Meg-01 maturation and platelet production. 15d-PGJ(2) also promotes platelet formation from culture-derived mouse and human megakaryocytes and accelerates platelet recovery after in vivo radiation-induced bone marrow injury. Interestingly, the platelet-enhancing effects of 15d-PGJ(2) in Meg-01 cells are independent of PPARgamma, but dependent on reactive oxygen species (ROS) accumulation; treatment with antioxidants such as glutathione ethyl ester (GSH-EE); or N-acetylcysteine (NAC) attenuate 15d-PGJ(2)-induced platelet production. Collectively, these data support the concept that megakaryocyte redox status plays an important role in platelet generation and that small electrophilic molecules may have clinical efficacy for improving platelet numbers in thrombocytopenic patients.
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Jia S, Omelchenko M, Garland D, Vasiliou V, Kanungo J, Spencer M, Wolf Y, Koonin E, Piatigorsky J. Duplicated gelsolin family genes in zebrafish: a novel scinderin-like gene (scinla) encodes the major corneal crystallin. FASEB J 2007; 21:3318-28. [PMID: 17548429 PMCID: PMC6007973 DOI: 10.1096/fj.07-8172com] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/06/2023]
Abstract
We have previously identified a gelsolin-like protein (C/L-gelsolin) as a corneal crystallin in zebrafish. Here we show by phylogenetic analysis that there are at least six genes encoding gelsolin-like proteins based on their gelsolin domains in zebrafish: gsna and gsnb group with the vertebrate gelsolin gene, scina and scinb group with the scinderin (adseverin) gene, and scinla (C/L-gelsolin) and scinlb are novel scinderin-like genes. RT-PCR showed that scinla, scinlb, and gsnb are preferentially expressed in the adult cornea whereas gsna is expressed to a similar extent in cornea, lens, brain, and heart; scina and scinb expression were detectable only in whole zebrafish and not in these adult tissues. Quantitative RT-PCR and 2-dimensional polyacrylamide gel electrophoresis followed by MALDI/TOF mass spectroscopy confirmed high expression of beta-actin and scinla, moderate expression of scinlb, and very low expression of gsna and gsnb in the cornea. Finally, transgenic zebrafish carrying a green fluorescent protein reporter transgene driven by a 4 kb scinla promoter fragment showed expression in the cornea, snout, dorsal fin, and tail fin of 3-day-old zebrafish larvae. Our data suggest that scinla and scinlb are diverged paralogs of the vertebrate scinderin gene and show that scinla encodes the zebrafish corneal crystallin previously called C/L-gelsolin.
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Affiliation(s)
- Sujuan Jia
- Laboratory of Molecular and Developmental Biology, Bethesda, Maryland, USA
| | - Marina Omelchenko
- National Library of Medicine, National Institutes of Health, Bethesda, Maryland, USA
| | - Donita Garland
- Laboratory of Retinal Cellular and Molecular Biology, National Eye Institute, Bethesda, Maryland, USA
| | - Vasilis Vasiliou
- University of Colorado Health Sciences Center, School of Pharmacy, University of Colorado, Denver, Colorado, USA
| | | | - Michael Spencer
- Laboratory of Molecular and Developmental Biology, Bethesda, Maryland, USA
| | - Yuri Wolf
- National Library of Medicine, National Institutes of Health, Bethesda, Maryland, USA
| | - Eugene Koonin
- National Library of Medicine, National Institutes of Health, Bethesda, Maryland, USA
| | - Joram Piatigorsky
- Laboratory of Molecular and Developmental Biology, Bethesda, Maryland, USA
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Nurminsky D, Magee C, Faverman L, Nurminskaya M. Regulation of chondrocyte differentiation by actin-severing protein adseverin. Dev Biol 2007; 302:427-37. [PMID: 17097081 PMCID: PMC3387683 DOI: 10.1016/j.ydbio.2006.09.052] [Citation(s) in RCA: 53] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/11/2006] [Revised: 09/25/2006] [Accepted: 09/29/2006] [Indexed: 11/28/2022]
Abstract
The importance of actin organization in controlling the chondrocyte phenotype is well established, but little is known about the cytoskeletal components regulating chondrocyte differentiation. Previously, we have observed up-regulation of an actin-binding gelsolin-like protein in hypertrophic chondrocytes. We have now identified it as adseverin (scinderin). Adseverin is drastically up-regulated during chondrocyte maturation, as shown by Northern blot analysis, in situ hybridization, and real-time RT-PCR. Its expression is positively regulated by PKC and MEK signaling as shown by inhibitory analyses. Over-expression of adseverin in non-hypertrophic chondrocytes causes rearrangement of the actin cytoskeleton, a change in cell morphology, a dramatic (3.5-fold) increase in cell volume, and up-regulation of Indian hedgehog (Ihh) and of collagen type X--all indicative of chondrocyte differentiation. These changes are mediated by ERK1/2 and p38 kinase pathways. Thus, adseverin-induced rearrangements of the actin cytoskeleton may mediate the PKC-dependent activation of p38 and Erk1/2 signaling pathways necessary for chondrocyte hypertrophy, as evidenced by changes in cell morphology, increase in cell size and expression of the chondrocyte maturation markers. These results demonstrate that interdependence of cytoskeletal organization and chondrogenic gene expression is regulated, at least in part, by actin-binding proteins such as adseverin.
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Affiliation(s)
- Dmitry Nurminsky
- Tufts University School of Medicine, Department of Anatomy and Cellular Biology, 136 Harrison Avenue Boston, MA 02111, USA
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Isakari Y, Harada Y, Ishikawa D, Matsumura-Takeda K, Sogo S, Ishida T, Taki T, Kiwada H. Efficient gene expression in megakaryocytic cell line using nucleofection. Int J Pharm 2007; 338:157-64. [PMID: 17331684 DOI: 10.1016/j.ijpharm.2007.01.042] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/22/2006] [Revised: 01/09/2007] [Accepted: 01/28/2007] [Indexed: 11/25/2022]
Abstract
To clarify the mechanism of platelet production from megakaryocytes, expression of target proteins by gene transfection was examined using various gene delivery techniques. Transfection into hematopoietic cells, including megakaryocytes, by conventional gene delivery techniques such as electroporation and lipofection are known to be difficult. In this study, in addition to electroporation and lipofection, we tested other gene-transfer methods (nucleofection, transfection using inactivated virus envelope, and transferrin-linked cationic polymer) with the green fluorescent protein (GFP) gene into the human megakaryocytic cell line MEG-01. We found that nucleofection, which uses a combination of special electrical parameters and specific solutions, was the best, judging from the expression ratio of GFP-positive cells (approximately 70% of cells) and low toxicity. The efficiency of GFP expression was not related to the amount of pDNA delivered into the MEG-01 cells. To verify the utility of nucleofection, the thrombopoietin (TPO) receptor c-mpl was transfected into MEG-01 cells. Transfected cells showed a higher responsiveness to TPO than mock-transfected MEG-01 cells. We propose that nucleofection is a useful method for transfecting target genes to megakaryocytic cells when addressing the mechanism of platelet production.
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Affiliation(s)
- Yoshimasa Isakari
- Department of Pharmacokinetics and Biopharmaceutics, Subdivision of Biopharmaceutical Sciences, Institute of Health Biosciences, The University of Tokushima, 1-78-1, Sho-machi, Tokushima 770-8505, Japan.
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Archer SK, Claudianos C, Campbell HD. Evolution of the gelsolin family of actin-binding proteins as novel transcriptional coactivators. Bioessays 2005; 27:388-96. [PMID: 15770676 DOI: 10.1002/bies.20200] [Citation(s) in RCA: 50] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/07/2022]
Abstract
The gelsolin gene family encodes a number of higher eukaryotic actin-binding proteins that are thought to function in the cytoplasm by severing, capping, nucleating or bundling actin filaments. Recent evidence, however, suggests that several members of the gelsolin family may have adopted unexpected nuclear functions including a role in regulating transcription. In particular, flightless I, supervillin and gelsolin itself have roles as coactivators for nuclear receptors, despite the fact that their divergence appears to predate the evolutionary appearance of nuclear receptors. Flightless I has been shown to bind both actin and the actin-related BAF53a protein, which are subunits of SWI/SNF-like chromatin remodelling complexes. The primary sequences of some actin-related proteins such as BAF53a exhibit conservation of residues that, in actin itself, are known to interact with gelsolin-related proteins. In summary, there is a growing body of evidence supporting a biological role in the nucleus for actin, Arps and actin-binding proteins and, in particular, the gelsolin family of actin-binding proteins.
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Affiliation(s)
- Stuart K Archer
- Molecular Genetics and Evolution Group and Centre for the Molecular Genetics of Development, Research School of Biological Sciences, Australian National University, Canberra, ACT 2601, Australia
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Unwin RD, Sternberg DW, Lu Y, Pierce A, Gilliland DG, Whetton AD. Global effects of BCR/ABL and TEL/PDGFRbeta expression on the proteome and phosphoproteome: identification of the Rho pathway as a target of BCR/ABL. J Biol Chem 2004; 280:6316-26. [PMID: 15569670 DOI: 10.1074/jbc.m410598200] [Citation(s) in RCA: 37] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/04/2023] Open
Abstract
Many leukemic oncogenes form as a consequence of gene fusions or mutation that result in the activation or overexpression of a tyrosine kinase. To identify commonalities and differences in the action of two such kinases, breakpoint cluster region (BCR)/ABL and TEL/PDGFRbeta, two-dimensional gel electrophoresis was employed to characterize their effects on the proteome. While both oncogenes affected expression of specific proteins, few common effects were observed. A number of proteins whose expression is altered by BCR/ABL, including gelsolin and stathmin, are related to cytoskeletal function whereas no such changes were seen in TEL/PDGFRbeta-transfected cells. Treatment of cells with the kinase inhibitor STI571 for 4-h reversed changes in expression of some of these cytoskeletal proteins. Correspondingly, BCR/ABL-transfected cells were less responsive to chemotactic and chemokinetic stimuli than non-transfected cells and TEL/PDGFRbeta-transfected Ba/F3 cells. Decreased motile response was reversed by a 16-h treatment with STI571. A phosphoprotein-specific gel stain was used to identify TEL/PDGFRbeta and BCR/ABL-mediated changes in the phosphoproteome. These included changes on Crkl, Ras-GAP-binding protein 1, and for BCR/ABL, cytoskeletal proteins such as tubulin, and Nedd5. Decreased phosphorylation of Rho-GTPase dissociation inhibitor (Rho GDI) was also observed in BCR/ABL-transfected cells. This results in the activation of the Rho pathway, and treatment of cells with Y27632, an inhibitor of Rho kinase, inhibited DNA synthesis in BCR/ABL-transfected Ba/F3 cells but not TEL/PDGFRbeta-expressing cells. Expression of a dominant-negative RhoA inhibited both DNA synthesis and transwell migration, demonstrating the significance of this pathway in BCR/ABL-mediated transformation.
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Affiliation(s)
- Richard D Unwin
- Faculty of Medical and Human Sciences, University of Manchester
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Abstract
Platelets are anucleate cells that fragment from mature megakaryocytes and play an essential role in thrombosis and hemostasis. Platelets are among the first cell types to be recruited to an injured blood vessel, assisting in endothelial repair. Platelet hyperactivation contributes to the development of atherosclerosis, myocardial infarction, and ischemia of peripheral limbs. A fall in platelet counts, due to a variety of conditions, including disseminated intravascular coagulation, chemotherapy or genetic disorders, may lead, in most severe cases, to death from hemorrhage. This review focuses on the late stages of megakaryocyte differentiation and platelet fragmentation, including associated cytoskeletal changes, and on the importance of apoptotic events for these processes. Studies point to a unique biological system in which programmed cell death may be linked with biogenesis of new cells.
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Affiliation(s)
- Yulia Kaluzhny
- Department of Biochemistry and Whitaker Cardiovascular Institute, Boston University School of Medicine, Boston, MA 02118, USA
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Abstract
Apoptosis or programmed cell death was discovered in nucleate cells 30 years ago and has been well documented. In contrast, apoptosis in anucleate platelets has only a five-year research history and as yet but few publications related to it. In this review, we will present the data on platelet apoptosis in several models. These include in vitro models where platelet apoptosis was induced by calcium ionophores, natural platelet agonists, storage in capped tubes at 37 degrees C and storage at room temperature under standard blood banking conditions, and in vivo models where apoptosis was provoked by suppression of thrombopoiesis, malaria infection and injection of tumor necrosis factor or anti-platelet antibodies. Understanding of platelet apoptosis and its role in the platelet storage lesion is an exciting challenge; future research is likely to provide us with further insight into this field.
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Affiliation(s)
- Valery Leytin
- Department of Transfusion Medicine, St. Michael's Hospital, Room 2003, Shuter Wing, 30 Bond Street, Toronto, Ont., Canada M5B 1W8.
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