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Serin N, Dihazi GH, Tayyeb A, Lenz C, Müller GA, Zeisberg M, Dihazi H. Calreticulin Deficiency Disturbs Ribosome Biogenesis and Results in Retardation in Embryonic Kidney Development. Int J Mol Sci 2021; 22:5858. [PMID: 34070742 PMCID: PMC8198291 DOI: 10.3390/ijms22115858] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/09/2021] [Revised: 05/18/2021] [Accepted: 05/26/2021] [Indexed: 11/27/2022] Open
Abstract
Nephrogenesis is driven by complex signaling pathways that control cell growth and differentiation. The endoplasmic reticulum chaperone calreticulin (Calr) is well known for its function in calcium storage and in the folding of glycoproteins. Its role in kidney development is still not understood. We provide evidence for a pivotal role of Calr in nephrogenesis in this investigation. We show that Calr deficiency results in the disrupted formation of an intact nephrogenic zone and in retardation of nephrogenesis, as evidenced by the disturbance in the formation of comma-shaped and s-shaped bodies. Using proteomics and transcriptomics approaches, we demonstrated that in addition to an alteration in Wnt-signaling key proteins, embryonic kidneys from Calr-/- showed an overall impairment in expression of ribosomal proteins which reveals disturbances in protein synthesis and nephrogenesis. CRISPR/cas9 mediated knockout confirmed that Calr deficiency is associated with a deficiency of several ribosomal proteins and key proteins in ribosome biogenesis. Our data highlights a direct link between Calr expression and the ribosome biogenesis.
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Affiliation(s)
- Nazli Serin
- Clinic for Nephrology and Rheumatology, University Medical Center Göttingen, Robert-Koch-Strasse 40, 37075 Göttingen, Germany; (N.S.); (G.A.M.); (M.Z.)
- Department of Hematology and Oncology, University of Medical Center Göttingen, Robert-Koch-Strasse 40, 37075 Göttingen, Germany
| | - Gry H. Dihazi
- Institute of Clinical Chemistry/UMG-Laboratories, University Medical Center Göttingen, Robert-Koch-Strasse 40, 37075 Göttingen, Germany; (G.H.D.); (C.L.)
| | - Asima Tayyeb
- School of Biological Sciences, University of the Punjab, Lahore 54590, Pakistan;
| | - Christof Lenz
- Institute of Clinical Chemistry/UMG-Laboratories, University Medical Center Göttingen, Robert-Koch-Strasse 40, 37075 Göttingen, Germany; (G.H.D.); (C.L.)
- Bioanalytical Mass Spectrometry, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany
| | - Gerhard A. Müller
- Clinic for Nephrology and Rheumatology, University Medical Center Göttingen, Robert-Koch-Strasse 40, 37075 Göttingen, Germany; (N.S.); (G.A.M.); (M.Z.)
| | - Michael Zeisberg
- Clinic for Nephrology and Rheumatology, University Medical Center Göttingen, Robert-Koch-Strasse 40, 37075 Göttingen, Germany; (N.S.); (G.A.M.); (M.Z.)
| | - Hassan Dihazi
- Clinic for Nephrology and Rheumatology, University Medical Center Göttingen, Robert-Koch-Strasse 40, 37075 Göttingen, Germany; (N.S.); (G.A.M.); (M.Z.)
- Center for Biostructural Imaging of Neurodegeneration (BIN), University Medical Center Göttingen, 37075 Göttingen, Germany
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Aldahhan RA, Stanton PG. Heat stress response of somatic cells in the testis. Mol Cell Endocrinol 2021; 527:111216. [PMID: 33639219 DOI: 10.1016/j.mce.2021.111216] [Citation(s) in RCA: 43] [Impact Index Per Article: 10.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/05/2020] [Revised: 11/30/2020] [Accepted: 02/15/2021] [Indexed: 02/06/2023]
Abstract
The testis is a temperature-sensitive organ that needs to be maintained 2-7 °C below core body temperature to ensure the production of normal sperm. Failure to maintain testicular temperature in mammals impairs spermatogenesis and leads to low sperm counts, poor sperm motility and abnormal sperm morphology in the ejaculate. This review discusses the recent knowledge on the response of testicular somatic cells to heat stress and, specifically, regarding the relevant contributions of heat, germ cell depletion and inflammatory reactions on the functions of Sertoli and Leydig cells. It also outlines mechanisms of testicular thermoregulation, as well as the thermogenic factors that impact testicular function.
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Affiliation(s)
- Rashid A Aldahhan
- Department of Anatomy, College of Medicine, Imam Abdulrahman Bin Faisal University, P.O. Box 2114, Dammam, 31541, Saudi Arabia.
| | - Peter G Stanton
- Centre for Reproductive Health, Hudson Institute of Medical Research, Clayton, Victoria, Australia; Department of Molecular and Translational Sciences, Monash University, Clayton, Victoria, Australia
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Guttula PK, Gupta MK. Examining the co-expression, transcriptome clustering and variation using fuzzy cluster network of testicular stem cells and pluripotent stem cells compared with other cell types. Comput Biol Chem 2020; 85:107227. [PMID: 32044562 DOI: 10.1016/j.compbiolchem.2020.107227] [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: 05/03/2019] [Revised: 10/10/2019] [Accepted: 01/31/2020] [Indexed: 10/25/2022]
Abstract
Stem cells are crucial in the field of tissue regeneration and developmental biology. Embryonic stem cells (ESCs) which are pluripotent in nature are derived from the inner cell mass of blastocyst. The gene expression profiles of ESCs and Induced pluripotent stem cells (iPSCs) were compared to identify the differences. Spermatogonial stem cells (SSCs) are also known as Germ-line stem cells (GSCs) present in testis is having the capability of producing the sperm in their whole lifetime. Therefore can be reprogrammed into pluripotent cells called male germline pluripotent cells (gPSCs). It is very difficult to interpret the larger genomic data sets which are available in public databases without high computational facilities. In order to identify the similar groups We studied the co-expression, clustering of the transcriptome and variation of the transcriptome of the GSCs, gPSCs, ESCs and other cell types using fuzzy clustering using AutoSOME. The series matrix file with GSE ID GSE11274 was retrieved and subjected to the various normalization methods, corresponding rows and columns were clustered using p values, ensemble runs, and different running modes. Transcriptome analysis using the proposed approach intuitively and consistently characterized the variation in cell-cell significantly. Collectively, our results suggest that the GSCs and the ESCs displayed differential gene expression profiles, and the GSCs possessed the potential to acquire pluripotency based on the high expression of epigenetic factors and transcription factors. These data may provide novel insights into the reprogramming mechanism of GSCs.
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Affiliation(s)
- Praveen Kumar Guttula
- Gene Manipulation Laboratory, Department of Biotechnology and Medical Engineering, National Institute of Technology, Rourkela, 769008, India
| | - Mukesh Kumar Gupta
- Gene Manipulation Laboratory, Department of Biotechnology and Medical Engineering, National Institute of Technology, Rourkela, 769008, India.
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Nejad-Moghaddam A, Tahmasbpour E, Sohrabiyan M, Jafari H, Ghanei M. Stem cells therapy: a review on approaches that can be used for treatment of respiratory failures in sulfur mustard-injured patients. Immunopharmacol Immunotoxicol 2018; 40:359-367. [PMID: 30488735 DOI: 10.1080/08923973.2018.1510961] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/25/2018] [Accepted: 07/06/2018] [Indexed: 12/28/2022]
Abstract
Sulfur mustard (SM) is a toxic agent which causes severe abnormalities in an airway system such as necrosis and inflammation, oxidative stress, chronic bronchitis, shortness of breath, and chronic obstructive pulmonary disease. Although possible mechanisms of SM toxicity have been extensively considered, there is still need to find an appropriate clinical treatment to decrease chronic lung injuries caused by SM. Due to extensive progresses and achievement in tissue repairing through stem cells therapy, the importance of cell therapy for the treatment of lung injuries has been increased. However, several factors such as types of stem cells, necessary conditions for growth and proliferation of stem cells, and their homing into the target tissues are considered as the most important problems in this issue. Mesenchymal stem cells (MSCs) are a class of multipotent stem cells with proliferative and self-renewal capacity which are able to differentiate into different cell lines such as lung epithelial cells. They have a potential repairing and immune modulatory properties which make them as a good candidate for the regeneration of bronchioles tract in SM-exposed patients. Unlike chemical drugs, the differentiation and high-level safety properties of MSCs can be considered as a new strategy for the treatment of SM-injured patients with pulmonary complications. This review aims to consider the therapeutic effects of MSCs in the treatment of SM-induced pulmonary injuries in both animals and humans.
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Affiliation(s)
- Amir Nejad-Moghaddam
- a Marine Medicine Research Center , Baqiyatallah University of Medical Sciences , Tehran , Iran
| | - Eisa Tahmasbpour
- b Laboratory of Regenerative Medicine & Biomedical Innovations , Pasteur Institute of Iran , Tehran , Iran
| | - Milad Sohrabiyan
- c Chemical Injuries Research Center, Systems Biology and Poisonings Institute , Baqiyatallah University of Medical Sciences , Tehran , Iran
| | - Hosein Jafari
- a Marine Medicine Research Center , Baqiyatallah University of Medical Sciences , Tehran , Iran
| | - Mostafa Ghanei
- c Chemical Injuries Research Center, Systems Biology and Poisonings Institute , Baqiyatallah University of Medical Sciences , Tehran , Iran
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Abstract
Compared to genomics or transcriptomics, proteomics is often regarded as an "emerging technology," i.e., as not having reached the same level of maturity. While the successful implementation of proteomics workflows and technology still requires significant levels of expertise and specialization, great strides have been made to make the technology more powerful, streamlined and accessible. In 2014, two landmark studies published the first draft versions of the human proteome.We aim to provide an introduction specifically into the background of mass spectrometry (MS)-based proteomics. Within the field, mass spectrometry has emerged as a core technology. Coupled to increasingly powerful separations and data processing and bioinformatics solution, it allows the quantitative analysis of whole proteomes within a matter of days, a timescale that has made global comparative proteome studies feasible at last. We present and discuss the basic concepts behind proteomics mass spectrometry and the accompanying topic of protein and peptide separations, with a focus on the properties of datasets emerging from such studies.
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Hillmer M, Wagner D, Summerer A, Daiber M, Mautner VF, Messiaen L, Cooper DN, Kehrer-Sawatzki H. Fine mapping of meiotic NAHR-associated crossovers causing large NF1 deletions. Hum Mol Genet 2015; 25:484-96. [PMID: 26614388 DOI: 10.1093/hmg/ddv487] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/12/2015] [Accepted: 11/19/2015] [Indexed: 02/06/2023] Open
Abstract
Large deletions encompassing the NF1 gene and its flanking regions belong to the group of genomic disorders caused by copy number changes that are mediated by the local genomic architecture. Although nonallelic homologous recombination (NAHR) is known to be a major mutational mechanism underlying such genomic copy number changes, the sequence determinants of NAHR location and frequency are still poorly understood since few high-resolution mapping studies of NAHR hotspots have been performed to date. Here, we have characterized two NAHR hotspots, PRS1 and PRS2, separated by 20 kb and located within the low-copy repeats NF1-REPa and NF1-REPc, which flank the human NF1 gene region. High-resolution mapping of the crossover sites identified in 78 type 1 NF1 deletions mediated by NAHR indicated that PRS2 is a much stronger NAHR hotspot than PRS1 since 80% of these deletions exhibited crossovers within PRS2, whereas 20% had crossovers within PRS1. The identification of the most common strand exchange regions of these 78 deletions served to demarcate the cores of the PRS1 and PRS2 hotspots encompassing 1026 and 1976 bp, respectively. Several sequence features were identified that may influence hotspot intensity and direct the positional preference of NAHR to the hotspot cores. These features include regions of perfect sequence identity encompassing 700 bp at the hotspot core, the presence of PRDM9 binding sites perfectly matching the consensus motif for the most common PRDM9 variant, specific pre-existing patterns of histone modification and open chromatin conformations that are likely to facilitate PRDM9 binding.
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Affiliation(s)
- Morten Hillmer
- Institute of Human Genetics, University of Ulm, 89081 Ulm, Germany
| | - David Wagner
- Institute of Human Genetics, University of Ulm, 89081 Ulm, Germany
| | - Anna Summerer
- Institute of Human Genetics, University of Ulm, 89081 Ulm, Germany
| | - Michaela Daiber
- Institute of Human Genetics, University of Ulm, 89081 Ulm, Germany
| | - Victor-Felix Mautner
- Department of Neurology, University Hospital Hamburg Eppendorf, 20246 Hamburg, Germany
| | - Ludwine Messiaen
- Medical Genomics Laboratory, Department of Genetics, University of Alabama at Birmingham, Birmingham, AL 35242, USA and
| | - David N Cooper
- Institute of Medical Genetics, School of Medicine, Cardiff University, Cardiff CF14 4XN, UK
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Dihazi GH, Jahn O, Tampe B, Zeisberg M, Müller C, Müller GA, Dihazi H. Proteomic analysis of embryonic kidney development: Heterochromatin proteins as epigenetic regulators of nephrogenesis. Sci Rep 2015; 5:13951. [PMID: 26359909 PMCID: PMC4566080 DOI: 10.1038/srep13951] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/04/2014] [Accepted: 07/10/2015] [Indexed: 01/18/2023] Open
Abstract
Elucidation of the mechanisms underlying the nephrogenesis will boost enormously the regenerative medicine. Here we performed 2-D gel-based comparative proteome analyses of rat embryonic kidney from different developmental stages. Out of 288 non-redundant identified proteins, 102 were common in all developmental stages. 86% of the proteins found in E14 and E16 were identical, in contrast only 37% of the identified proteins overlap between E14 and P1. Bioinformatics analysis suggests developmental stage-specific pathway activation and highlighted heterochromatin protein 1 (Cbx1, Cbx3, Cbx5) and Trim28 as potential key players in nephrogenesis. These are involved in the epigenetic regulation of gene silencing and were down-regulated in the course of kidney development. Trim28 is a potential epigenetic regulator of the branching inhibitor Bmp4. Silencing of Trim28 in cultured kidneys resulted in branching arrest. In contrast knockdown of Cbx5 was associated with abnormal ureteric bud growth and slight impairment of branching. ChIP analysis showed that the H3K9me3 distribution on Bmp4 promoters at E14 and E19 inversely correlate with mRNA expression levels. The concentrated expression-pattern of heterochromatin proteins and the negative impact of their silencing on kidney development, suggest an important role in reciprocal and inductive signaling between the ureteric bud and the metanephric mesenchyme.
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Affiliation(s)
- Gry H Dihazi
- Department of Nephrology and Rheumatology, Georg-August University Göttingen, Robert-Koch-Strasse 40, D-37075 Göttingen, Germany
| | - Olaf Jahn
- Proteomics Group, Max-Planck-Institute of Experimental Medicine, Hermann-Rein-Strasse 3, D-37075 Göttingen, Germany.,Deutsche Forschungsgemeinschaft Research Center for Molecular Physiology of the Brain, Humboldtallee 23, D-37073 Göttingen, Germany
| | - Björn Tampe
- Department of Nephrology and Rheumatology, Georg-August University Göttingen, Robert-Koch-Strasse 40, D-37075 Göttingen, Germany
| | - Michael Zeisberg
- Department of Nephrology and Rheumatology, Georg-August University Göttingen, Robert-Koch-Strasse 40, D-37075 Göttingen, Germany
| | - Claudia Müller
- Department of Nephrology and Rheumatology, Georg-August University Göttingen, Robert-Koch-Strasse 40, D-37075 Göttingen, Germany.,Section for Transplantation- Immunology and Immunohematology, ZMF, Eberhard-Karls-University Tübingen, Germany
| | - Gerhard A Müller
- Department of Nephrology and Rheumatology, Georg-August University Göttingen, Robert-Koch-Strasse 40, D-37075 Göttingen, Germany
| | - Hassan Dihazi
- Department of Nephrology and Rheumatology, Georg-August University Göttingen, Robert-Koch-Strasse 40, D-37075 Göttingen, Germany
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Blaschke S, Rinke K, Maring M, Flad T, Patschan S, Jahn O, Mueller CA, Mueller GA, Dihazi H. Haptoglobin-α1, -α2, vitamin D-binding protein and apolipoprotein C-III as predictors of etanercept drug response in rheumatoid arthritis. Arthritis Res Ther 2015; 17:45. [PMID: 25884688 PMCID: PMC4383078 DOI: 10.1186/s13075-015-0553-1] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/26/2014] [Accepted: 02/11/2015] [Indexed: 01/03/2023] Open
Abstract
Introduction The introduction of tumor necrosis factor-alpha (TNF-α) antagonists has substantially improved patient’s clinical outcome in rheumatoid arthritis (RA). However, nearly 20% to 40% of RA patients do not respond to anti-TNF-α treatment strategies. To identify valid predictors of TNF-α antagonist response in RA, serum proteome profiles from responders (R) and non-responders (NR) to etanercept, a soluble recombinant TNF-α receptor/IgG Fc fusion protein receptor, were compared in a prospective cohort study. Methods In this clinical study 50 RA patients with inadequate response to conventional DMARDs were included and treated with etanercept. The primary efficacy endpoint was response according to the European League against Rheumatism (EULAR) improvement criteria. Serum samples collected prior to initiation and after six months of etanercept therapy were cleared of the most abundant major proteins by immunoaffinity chromatography. After separation by two-dimensional differential gel electrophoresis (2D-DIGE) and identification by mass spectrometry (MS) data were validated by Western blot analysis. Results After six months of etanercept treatment 62% (n = 31) of RA patients achieved response. Haptoglobin-α1 (Hp-α1) and -α2 (Hp-α2) and vitamin D-binding protein (VDBP) were found to be significantly upregulated in responder sera (P ≤0.02) at study entry. In contrast, apolipoprotein C-III (ApoC-III) showed significantly higher levels in non-responders (P = 0.0162). At study end ApoA-II, Hp-α1, Hp-α2 and VDBP were identified to be expressed at significantly higher levels (P <0.05) in responder sera. Conclusions By application of clinical proteomics in immunodepleted sera we could identify and validate for the first time Hp-α1, -α2, VDBP and ApoC-III as potential biomarkers for prediction of etanercept drug response in RA. Electronic supplementary material The online version of this article (doi:10.1186/s13075-015-0553-1) contains supplementary material, which is available to authorized users.
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Affiliation(s)
- Sabine Blaschke
- Department of Nephrology and Rheumatology, University Medical Center of Göttingen, Robert-Koch Straße 40, 37075, Göttingen, Germany.
| | - Kathinka Rinke
- Department of Nephrology and Rheumatology, University Medical Center of Göttingen, Robert-Koch Straße 40, 37075, Göttingen, Germany.
| | - Michael Maring
- Department of Nephrology and Rheumatology, University Medical Center of Göttingen, Robert-Koch Straße 40, 37075, Göttingen, Germany.
| | - Thomas Flad
- PANATecs GmbH, Inselwiesenstr. 10, 74076, Heilbronn, Germany.
| | - Susann Patschan
- Department of Nephrology and Rheumatology, University Medical Center of Göttingen, Robert-Koch Straße 40, 37075, Göttingen, Germany.
| | - Olaf Jahn
- Proteomics Group, Max-Planck-Institute of Experimental Medicine, Hermann-Rein Str. 3, 37075, Göttingen, Germany.
| | - Claudia A Mueller
- Section for Transplantation Immunology and Immunohematology, Waldhörnlestr. 22, 72072, Tübingen, Germany.
| | - Gerhard A Mueller
- Department of Nephrology and Rheumatology, University Medical Center of Göttingen, Robert-Koch Straße 40, 37075, Göttingen, Germany.
| | - Hassan Dihazi
- Department of Nephrology and Rheumatology, University Medical Center of Göttingen, Robert-Koch Straße 40, 37075, Göttingen, Germany.
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Mukherjee A, Koli S, Reddy KVR. Regulatory non-coding transcripts in spermatogenesis: shedding light on ‘dark matter’. Andrology 2014; 2:360-9. [DOI: 10.1111/j.2047-2927.2014.00183.x] [Citation(s) in RCA: 37] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/28/2013] [Revised: 12/26/2013] [Accepted: 12/26/2013] [Indexed: 11/29/2022]
Affiliation(s)
- A. Mukherjee
- Division of Molecular Immunology and Microbiology; National Institute for Research in Reproductive Health; Indian Council of Medical Research; Mumbai India
| | - S. Koli
- Division of Molecular Immunology and Microbiology; National Institute for Research in Reproductive Health; Indian Council of Medical Research; Mumbai India
| | - K. V. R. Reddy
- Division of Molecular Immunology and Microbiology; National Institute for Research in Reproductive Health; Indian Council of Medical Research; Mumbai India
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MacLeod G, Varmuza S. The application of proteomic approaches to the study of mammalian spermatogenesis and sperm function. FEBS J 2013; 280:5635-51. [DOI: 10.1111/febs.12461] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/22/2013] [Revised: 07/04/2013] [Accepted: 07/26/2013] [Indexed: 12/22/2022]
Affiliation(s)
- Graham MacLeod
- Department of Cell & Systems Biology; University of Toronto; ON Canada
| | - Susannah Varmuza
- Department of Cell & Systems Biology; University of Toronto; ON Canada
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Pešić I, Müller GA, Baumann C, Dihazi GH, Koziolek MJ, Eltoweissy M, Bramlage C, Asif AR, Dihazi H. Cellulose membranes are more effective in holding back vital proteins and exhibit less interaction with plasma proteins during hemodialysis. BIOCHIMICA ET BIOPHYSICA ACTA-PROTEINS AND PROTEOMICS 2013; 1834:754-62. [PMID: 23369790 DOI: 10.1016/j.bbapap.2013.01.021] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 10/01/2012] [Revised: 01/15/2013] [Accepted: 01/18/2013] [Indexed: 11/27/2022]
Abstract
The vast majority of patients with end-stage renal disease are treated with intermittent hemodialysis as a form of renal replacement therapy. To investigate the impact of hemodialysis membrane material on vital protein removal, dialysates from 26 well-characterized hemodialysis patients were collected 5 min after beginning, during 5h of treatment, as well as 5 min before ending of the dialysis sessions. Dialysis sessions were performed using either modified cellulose (n=12) (low-flux and high flux) or synthetic Polyflux (n=14) (low-flux and high-flux) dialyzer. Protein removal during hemodialysis was quantified and the dialysate proteome patterns were analyzed by 2-DE, MS and Western blot. There was a clear correlation between the type of membrane material and the amount of protein removed. Synthetic Polyflux membranes exhibit strong interaction with plasma proteins resulting in a significantly higher protein loss compared to modified cellulosic membrane. Moreover, the proteomics analysis showed that the removed proteins represented different molecular weight range and different functional groups: transport proteins, protease inhibitors, proteins with role in immune response and regulations, constructive proteins and as a part of HLA immune complex. The effect of this protein removal on hemodialysis treatment outcome should be investigated in further studies.
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Affiliation(s)
- Ivana Pešić
- Department of Nephrology and Rheumatology, Georg-August University Goettingen, Goettingen, Germany
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Dihazi GH, Bibi A, Jahn O, Nolte J, Mueller GA, Engel W, Dihazi H. Impact of the antiproliferative agent ciclopirox olamine treatment on stem cells proteome. World J Stem Cells 2013; 5:9-25. [PMID: 23362436 PMCID: PMC3557350 DOI: 10.4252/wjsc.v5.i1.9] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/07/2012] [Revised: 09/19/2012] [Accepted: 12/20/2012] [Indexed: 02/06/2023] Open
Abstract
AIM To investigate the proteome changes of stem cells due to ciclopirox olamine (CPX) treatment compared to control and retinoic acid treated cells. METHODS Stem cells (SCs) are cells, which have the ability to continuously divide and differentiate into various other kinds of cells. Murine embryonic stem cells (ESCs) and multipotent adult germline stem cells (maGSCs) were treated with CPX, which has been shown to have an antiproliferative effect on stem cells, and compared to stem cells treated with retinoic acid (RA), which is known to have a differentiating effect on stem cells. Classical proteomic techniques like 2-D gel electrophoresis and differential in-gel electrophoresis (DIGE) were used to generate 2D protein maps from stem cells treated with RA or CPX as well as from non-treated stem cells. The resulting 2D gels were scanned and the digitalized images were collated with the help of Delta 2D software. The differentially expressed proteins were analyzed by a MALDI-TOF-TOF mass spectrometer, and the identified proteins were investigated and categorized using bioinformatics. RESULTS Treatment of stem cells with CPX, a synthetic antifungal clinically used to treat superficial mycoses, resulted in an antiproliferative effect in vitro, without impairment of pluripotency. To understand the mechanisms induced by CPX treatments which results in arrest of cell cycle without any marked effect on pluripotency, a comparative proteomics study was conducted. The obtained data revealed that the CPX impact on cell proliferation was accompanied with a significant alteration in stem cell proteome. By peptide mass fingerprinting and tandem mass spectrometry combined with searches of protein sequence databases, a set of 316 proteins was identified, corresponding to a library of 125 non-redundant proteins. With proteomic analysis of ESCs and maGSCs treated with CPX and RA, we could identify more than 90 single proteins, which were differently expressed in both cell lines. We could highlight, that CPX treatment of stem cells, with subsequent proliferation inhibition, resulted in an alteration of the expression of 56 proteins compared to non-treated cells, and 54 proteins compared to RA treated cells. Bioinformatics analysis of the regulated proteins demonstrated their involvement in various biological processes. To our interest, a number of proteins have potential roles in the regulation of cell proliferation either directly or indirectly. Furthermore the classification of the altered polypeptides according to their main known/postulated functions revealed that the majority of these proteins are involved in molecular functions like nucleotide binding and metal ion binding, and biological processes like nucleotide biosynthetic processes, gene expression, embryonic development, regulation of transcription, cell cycle processes, RNA and mRNA processing. Proteins, which are involved in nucleotide biosynthetic process and proteolysis, were downregulated in CPX treated cells compared to control, as well as in RA treated cells, which may explain the cell cycle arrest. Moreover, proteins which were involved in cell death, positive regulation of biosynthetic process, response to organic substance, glycolysis, anti-apoptosis, and phosphorylation were downregulated in RA treated cells compared to control and CPX treated cells. CONCLUSION The CPX treatment of SCs results in downregulation of nucleotide binding proteins and leads to cell cycle stop without impairment of pluripotency.
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Affiliation(s)
- Gry H Dihazi
- Gry H Dihazi, Asima Bibi, Gerhard A Mueller, Hassan Dihazi, Department of Nephrology and Rheumatology, Georg-August University Goettingen, D-37075 Goettingen, Germany
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Khromov T, Dressel R, Siamishi I, Nolte J, Opitz L, Engel W, Pantakani DVK. Apoptosis-related gene expression profiles of mouse ESCs and maGSCs: role of Fgf4 and Mnda in pluripotent cell responses to genotoxicity. PLoS One 2012; 7:e48869. [PMID: 23145002 PMCID: PMC3492253 DOI: 10.1371/journal.pone.0048869] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/07/2012] [Accepted: 10/02/2012] [Indexed: 01/27/2023] Open
Abstract
Stem cells in the developing embryo proliferate and differentiate while maintaining genomic integrity, failure of which may lead to accumulation of mutations and subsequent damage to the embryo. Embryonic stem cells (ESCs), the in vitro counterpart of embryo stem cells are highly sensitive to genotoxic stress. Defective ESCs undergo either efficient DNA damage repair or apoptosis, thus maintaining genomic integrity. However, the genotoxicity- and apoptosis-related processes in germ-line derived pluripotent cells, multipotent adult germ-line stem cells (maGSCs), are currently unknown. Here, we analyzed the expression of apoptosis-related genes using OligoGEArray in undifferentiated maGSCs and ESCs and identified a similar set of genes expressed in both cell types. We detected the expression of intrinsic, but not extrinsic, apoptotic pathway genes in both cell types. Further, we found that apoptosis-related gene expression patterns of differentiated ESCs and maGSCs are identical to each other. Comparative analysis revealed that several pro- and anti-apoptotic genes are expressed specifically in pluripotent cells, but markedly downregulated in the differentiated counterparts of these cells. Activation of the intrinsic apoptotic pathway cause approximately ∼35% of both ESCs and maGSCs to adopt an early-apoptotic phenotype. Moreover, we performed transcriptome studies using early-apoptotic cells to identify novel pluripotency- and apoptosis-related genes. From these transcriptome studies, we selected Fgf4 (Fibroblast growth factor 4) and Mnda (Myeloid cell nuclear differentiating antigen), which are highly downregulated in early-apoptotic cells, as novel candidates and analyzed their roles in apoptosis and genotoxicity responses in ESCs. Collectively, our results show the existence of common molecular mechanisms for maintaining the pristine stem cell pool of both ESCs and maGSCs.
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Affiliation(s)
- Tatjana Khromov
- Institute of Human Genetics, University of Goettingen, Goettingen, Germany
| | - Ralf Dressel
- Department of Cellular and Molecular Immunology, University of Goettingen, Goettingen, Germany
| | - Iliana Siamishi
- Institute of Human Genetics, University of Goettingen, Goettingen, Germany
| | - Jessica Nolte
- Institute of Human Genetics, University of Goettingen, Goettingen, Germany
| | - Lennart Opitz
- DNA Microarray Facility, University of Goettingen, Goettingen, Germany
| | - Wolfgang Engel
- Institute of Human Genetics, University of Goettingen, Goettingen, Germany
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14
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Abstract
Spermatogenesis is a highly sophisticated process involved in the transmission of genetic heritage. It includes halving ploidy, repackaging of the chromatin for transport, and the equipment of developing spermatids and eventually spermatozoa with the advanced apparatus (e.g., tightly packed mitochondrial sheat in the mid piece, elongating of the tail, reduction of cytoplasmic volume) to elicit motility once they reach the epididymis. Mammalian spermatogenesis is divided into three phases. In the first the primitive germ cells or spermatogonia undergo a series of mitotic divisions. In the second the spermatocytes undergo two consecutive divisions in meiosis to produce haploid spermatids. In the third the spermatids differentiate into spermatozoa in a process called spermiogenesis. Paracrine, autocrine, juxtacrine, and endocrine pathways all contribute to the regulation of the process. The array of structural elements and chemical factors modulating somatic and germ cell activity is such that the network linking the various cellular activities during spermatogenesis is unimaginably complex. Over the past two decades, advances in genomics have greatly improved our knowledge of spermatogenesis, by identifying numerous genes essential for the development of functional male gametes. Large-scale analyses of testicular function have deepened our insight into normal and pathological spermatogenesis. Progress in genome sequencing and microarray technology have been exploited for genome-wide expression studies, leading to the identification of hundreds of genes differentially expressed within the testis. However, although proteomics has now come of age, the proteomics-based investigation of spermatogenesis remains in its infancy. Here, we review the state-of-the-art of large-scale proteomic analyses of spermatogenesis, from germ cell development during sex determination to spermatogenesis in the adult. Indeed, a few laboratories have undertaken differential protein profiling expression studies and/or systematic analyses of testicular proteomes in entire organs or isolated cells from various species. We consider the pros and cons of proteomics for studying the testicular germ cell gene expression program. Finally, we address the use of protein datasets, through integrative genomics (i.e., combining genomics, transcriptomics, and proteomics), bioinformatics, and modelling.
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Affiliation(s)
- Sophie Chocu
- Inserm, U1085, IRSET, University of Rennes I, Campus de Beaulieu, Rennes, France
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15
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Imamura M, Lin ZYC, Okano H. Cell-intrinsic reprogramming capability: gain or loss of pluripotency in germ cells. Reprod Med Biol 2012; 12:1-14. [PMID: 29699125 DOI: 10.1007/s12522-012-0131-z] [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: 04/02/2012] [Accepted: 05/30/2012] [Indexed: 12/23/2022] Open
Abstract
In multicellular organisms, germ cells are an extremely specialized cell type with the vital function of transmitting genetic information across generations. In this respect, they are responsible for the perpetuity of species, and are separated from somatic lineages at each generation. Interestingly, in the past two decades research has shown that germ cells have the potential to proceed along two distinct pathways: gametogenesis or pluripotency. Unequivocally, the primary role of germ cells is to produce gametes, the sperm or oocyte, to produce offspring. However, under specific conditions germ cells can become pluripotent, as shown by teratoma formation in vivo or cell culture-induced reprogramming in vitro. This phenomenon seems to be a general propensity of germ cells, irrespective of developmental phase. Recent attempts at cellular reprogramming have resulted in the generation of induced pluripotent stem cells (iPSCs). In iPSCs, the intracellular molecular networks instructing pluripotency have been activated and override the exclusively somatic cell programs that existed. Because the generation of iPSCs is highly artificial and depends on gene transduction, whether the resulting machinery reflects any physiological cell-intrinsic programs is open to question. In contrast, germ cells can spontaneously shift their fate to pluripotency during in-vitro culture. Here, we review the two fates of germ cells, i.e., differentiation and reprogramming. Understanding the molecular mechanisms regulating differentiation versus reprogramming would provide invaluable insight into understanding the mechanisms of cellular reprogramming that generate iPSCs.
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Affiliation(s)
- Masanori Imamura
- Department of Physiology, School of Medicine Keio University 35 Shinanomachi 160-8582 Shinjuku-ku Tokyo Japan
| | - Zachary Yu-Ching Lin
- Department of Physiology, School of Medicine Keio University 35 Shinanomachi 160-8582 Shinjuku-ku Tokyo Japan
| | - Hideyuki Okano
- Department of Physiology, School of Medicine Keio University 35 Shinanomachi 160-8582 Shinjuku-ku Tokyo Japan
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16
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Kim SY, Kim MJ, Jung H, Kim WK, Kwon SO, Son MJ, Jang IS, Choi JS, Park SG, Park BC, Han YM, Lee SC, Cho YS, Bae KH. Comparative Proteomic Analysis of Human Somatic Cells, Induced Pluripotent Stem Cells, and Embryonic Stem Cells. Stem Cells Dev 2012; 21:1272-86. [DOI: 10.1089/scd.2011.0243] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/03/2023] Open
Affiliation(s)
- Sun Young Kim
- Medical Proteomics Research Center, KRIBB, Daejeon, South Korea
- Department of Biological Sciences, KAIST, Daejeon, South Korea
| | - Min-Jeong Kim
- Development and Differentiation Research Center, KRIBB, Daejeon, South Korea
| | - Hyeyun Jung
- Medical Proteomics Research Center, KRIBB, Daejeon, South Korea
| | - Won Kon Kim
- Medical Proteomics Research Center, KRIBB, Daejeon, South Korea
| | - Sang Oh Kwon
- Proteome Research Team, Korea Basic Science Institute, Daejeon, South Korea
| | - Myung Jin Son
- Development and Differentiation Research Center, KRIBB, Daejeon, South Korea
| | - Ik-Soon Jang
- Proteome Research Team, Korea Basic Science Institute, Daejeon, South Korea
| | - Jong-Soon Choi
- Proteome Research Team, Korea Basic Science Institute, Daejeon, South Korea
| | - Sung Goo Park
- Medical Proteomics Research Center, KRIBB, Daejeon, South Korea
| | | | - Yong-Mahn Han
- Department of Biological Sciences, KAIST, Daejeon, South Korea
| | - Sang Chul Lee
- Medical Proteomics Research Center, KRIBB, Daejeon, South Korea
| | - Yee Sook Cho
- Development and Differentiation Research Center, KRIBB, Daejeon, South Korea
| | - Kwang-Hee Bae
- Medical Proteomics Research Center, KRIBB, Daejeon, South Korea
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17
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Pešić I, Stefanović V, Müller GA, Müller CA, Čukuranović R, Jahn O, Bojanić V, Koziolek M, Dihazi H. Identification and validation of six proteins as marker for endemic nephropathy. J Proteomics 2011; 74:1994-2007. [DOI: 10.1016/j.jprot.2011.05.020] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/24/2011] [Revised: 05/07/2011] [Accepted: 05/10/2011] [Indexed: 01/09/2023]
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18
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19
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Gu B, Zhang J, Wu Y, Zhang X, Tan Z, Lin Y, Huang X, Chen L, Yao K, Zhang M. Proteomic analyses reveal common promiscuous patterns of cell surface proteins on human embryonic stem cells and sperms. PLoS One 2011; 6:e19386. [PMID: 21559292 PMCID: PMC3086920 DOI: 10.1371/journal.pone.0019386] [Citation(s) in RCA: 38] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/20/2011] [Accepted: 03/28/2011] [Indexed: 12/30/2022] Open
Abstract
Background It has long been proposed that early embryos and reproductive organs exhibit
similar gene expression profiles. However, whether this similarity is
propagated to the protein level remains largely unknown. We have previously
characterised the promiscuous expression pattern of cell surface proteins on
mouse embryonic stem (mES) cells. As cell surface proteins also play
critical functions in human embryonic stem (hES) cells and germ cells, it is
important to reveal whether a promiscuous pattern of cell surface proteins
also exists for these cells. Methods and Principal Findings Surface proteins of hES cells and human mature sperms (hSperms) were purified
by biotin labelling and subjected to proteomic analyses. More than 1000
transmembrane or secreted cell surface proteins were identified on the two
cell types, respectively. Proteins from both cell types covered a large
variety of functional categories including signal transduction, adhesion and
transporting. Moreover, both cell types promiscuously expressed a wide
variety of tissue specific surface proteins, and some surface proteins were
heterogeneously expressed. Conclusions/Significance Our findings indicate that the promiscuous expression of functional and
tissue specific cell surface proteins may be a common pattern in embryonic
stem cells and germ cells. The conservation of gene expression patterns
between early embryonic cells and reproductive cells is propagated to the
protein level. These results have deep implications for the cell surface
signature characterisation of pluripotent stem cells and germ cells and may
lead the way to a new area of study, i.e., the functional significance of
promiscuous gene expression in pluripotent and germ cells.
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Affiliation(s)
- Bin Gu
- The Institute of Genetics, College of Life
Sciences, Zhejiang University, Hangzhou, China
| | - Jiarong Zhang
- The Institute of Genetics, College of Life
Sciences, Zhejiang University, Hangzhou, China
| | - Ying Wu
- Zhejiang Institute of Planned Parenthood
Research and Zhejiang Human Sperm Bank, Hangzhou, China
| | - Xinzong Zhang
- Zhejiang Institute of Planned Parenthood
Research and Zhejiang Human Sperm Bank, Hangzhou, China
| | - Zhou Tan
- The Institute of Genetics, College of Life
Sciences, Zhejiang University, Hangzhou, China
| | - Yuanji Lin
- The Institute of Genetics, College of Life
Sciences, Zhejiang University, Hangzhou, China
| | - Xiao Huang
- The Institute of Cell and Developmental
Biology, College of Life Sciences, Zhejiang University, Hangzhou,
China
| | - Liangbiao Chen
- The Institute of Genetics and Developmental
Biology, Chinese Academy of Sciences, Beijing, China
| | - Kangshou Yao
- Zhejiang Institute of Planned Parenthood
Research and Zhejiang Human Sperm Bank, Hangzhou, China
- * E-mail: (MZ); (KY)
| | - Ming Zhang
- The Institute of Genetics, College of Life
Sciences, Zhejiang University, Hangzhou, China
- * E-mail: (MZ); (KY)
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20
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Condic ML, Rao M. Alternative sources of pluripotent stem cells: ethical and scientific issues revisited. Stem Cells Dev 2011; 19:1121-9. [PMID: 20397928 DOI: 10.1089/scd.2009.0482] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022] Open
Abstract
Stem cell researchers in the United States continue to face an uncertain future, because of the changing federal guidelines governing this research, the restrictive patent situation surrounding the generation of new human embryonic stem cell lines, and the ethical divide over the use of embryos for research. In this commentary, we describe how recent advances in the derivation of induced pluripotent stem cells and the isolation of germ-line-derived pluripotent stem cells resolve a number of these uncertainties. The availability of patient-matched, pluripotent stem cells that can be obtained by ethically acceptable means provides important advantages for stem cell researchers, by both avoiding protracted ethical debates and giving U.S. researchers full access to federal funding. Thus, ethically uncompromised stem cells, such as those derived by direct reprogramming or from germ-cell precursors, are likely to yield important advances in stem cell research and move the field rapidly toward clinical applications.
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Affiliation(s)
- Maureen L Condic
- Department of Neurobiology and Anatomy, University of Utah School of Medicine, Salt Lake City, Utah 84132-3401, USA.
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21
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Oliva R, Castillo J. Proteomics and the genetics of sperm chromatin condensation. Asian J Androl 2010; 13:24-30. [PMID: 21042303 DOI: 10.1038/aja.2010.65] [Citation(s) in RCA: 56] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022] Open
Abstract
Spermatogenesis involves extremely marked cellular, genetic and chromatin changes resulting in the generation of the highly specialized sperm cell. Proteomics allows the identification of the proteins that compose the spermatogenic cells and the study of their function. The recent developments in mass spectrometry (MS) have markedly increased the throughput to identify and to study the sperm proteins. Catalogs of thousands of testis and spermatozoan proteins in human and different model species are becoming available, setting up the basis for subsequent research, diagnostic applications and possibly the future development of specific treatments. The present review intends to summarize the key genetic and chromatin changes at the different stages of spermatogenesis and in the mature sperm cell and to comment on the presently available proteomic studies.
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Affiliation(s)
- Rafael Oliva
- Human Genetics Research Group, IDIBAPS, Department of Ciencias Fisiológicas I, Faculty of Medicine, University of Barcelona, Barcelona 08036, Spain.
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22
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Khromov T, Pantakani DVK, Nolte J, Wolf M, Dressel R, Engel W, Zechner U. Global and gene-specific histone modification profiles of mouse multipotent adult germline stem cells. Mol Hum Reprod 2010; 17:166-74. [DOI: 10.1093/molehr/gaq085] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022] Open
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23
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Meyer S, Nolte J, Opitz L, Salinas-Riester G, Engel W. Pluripotent embryonic stem cells and multipotent adult germline stem cells reveal similar transcriptomes including pluripotency-related genes. Mol Hum Reprod 2010; 16:846-55. [PMID: 20624824 DOI: 10.1093/molehr/gaq060] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022] Open
Abstract
DNA microarray analysis was performed with mouse multipotent adult germline stem cells (maGSCs) and embryonic stem cells (ESCs) from different genetic backgrounds cultured under standard ESC-culture conditions and under differentiation-promoting conditions by the withdrawal of the leukemia inhibitory factor (LIF) and treatment with retinoic acid (RA). The analyzed undifferentiated cell lines are very similar based on their global gene expression pattern and show 97-99% identity dependent on the analyzed background. Only 621 genes are differentially expressed in cells derived from mouse 129SV-background and 72 genes show differences in expression in cells generated from transgenic Stra8-EGFP/Rosa26-LacZ-background. Both maGSCs and ESCs express the same genes involved in the regulation of pluripotency and even show no differences in the expression level of these genes. When comparing maGSCs with previously published signature genes of other pluripotent cell lines, we found that maGSCs shared a very similar gene expression pattern with embryonic germ cells (EGCs). Also after differentiation of maGSCs and ESCs the transcriptomes of the cell lines are nearly identical which suggests that both cell types differentiate spontaneously in a very similar way. This is the first study, at transcriptome level, to compare ESCs and a pluripotent cell line derived from an adult organism (maGSCs).
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Affiliation(s)
- S Meyer
- Institute of Human Genetics, Georg-August-University Göttingen, Heinrich-Düker-Weg 12, D-37073 Göttingen, Germany
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