title: Differential gene expression profiling of enriched human spermatogonia after short- and long-term culture.
abstract: This study aimed to provide a molecular signature for enriched adult human stem/progenitor spermatogonia during short-term (<2 weeks) and long-term culture (up to more than 14 months) in comparison to human testicular fibroblasts and human embryonic stem cells. Human spermatogonia were isolated by CD49f magnetic activated cell sorting and collagen(-)/laminin(+) matrix binding from primary testis cultures obtained from ten adult men. For transcriptomic analysis, single spermatogonia-like cells were collected based on their morphology and dimensions using a micromanipulation system from the enriched germ cell cultures. Immunocytochemical, RT-PCR and microarray analyses revealed that the analyzed populations of cells were distinct at the molecular level. The germ- and pluripotency-associated genes and genes of differentiation/spermatogenesis pathway were highly expressed in enriched short-term cultured spermatogonia. After long-term culture, a proportion of cells retained and aggravated the "spermatogonial" gene expression profile with the expression of germ and pluripotency-associated genes, while in the majority of long-term cultured cells this molecular profile, typical for the differentiation pathway, was reduced and more genes related to the extracellular matrix production and attachment were expressed. The approach we provide here to study the molecular status of in vitro cultured spermatogonia may be important to optimize the culture conditions and to evaluate the germ cell plasticity in the future.
authors.affiliation: Institute of Anatomy, University of Tübingen, Österbergstraße 3, 72074 Tübingen, Germany.
authors.forename: Sabine
authors.initials: S
authors.lastname: Conrad
authors.affiliation: Institute for Anatomy and Cell Biology, Medical Faculty, University of Heidelberg, Im Neuenheimer Feld 307, 69120 Heidelberg, Germany ; Amol University of Special Modern Technologies, Special Modern Technologies, P.O. Box 46168-49767, Amol, Iran ; Department of Stem Cells and Developmental Biology, Royan Institute, P.O. Box 19395-4644, Tehran, Iran.
authors.forename: Hossein
authors.initials: H
authors.lastname: Azizi
authors.affiliation: Institute for Anatomy and Cell Biology, Medical Faculty, University of Heidelberg, Im Neuenheimer Feld 307, 69120 Heidelberg, Germany.
authors.forename: Maryam
authors.initials: M
authors.lastname: Hatami
authors.affiliation: TATAA Biocenter AB, Odinsgatan 28, 41103 Göteborg, Sweden and Institute of Biotechnology at the Czech Academy of Sciences, Vídenská 1083, 14220 Prague 4, Czech Republic.
authors.forename: Mikael
authors.initials: M
authors.lastname: Kubista
authors.affiliation: Institute of Anthropology and Human Genetics, Microarray Facility, University Clinic, Calwerstraße 7, 72076 Tübingen, Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Bonin
authors.affiliation: Department of Urology, University Clinic Tübingen, Hoppe-Seyler-Straße 3, 72076 Tübingen, Germany.
authors.forename: Jörg
authors.initials: J
authors.lastname: Hennenlotter
authors.affiliation: Department of Urology, University Clinic Tübingen, Hoppe-Seyler-Straße 3, 72076 Tübingen, Germany.
authors.forename: Markus
authors.initials: M
authors.lastname: Renninger
authors.affiliation: Institute for Anatomy and Cell Biology, Medical Faculty, University of Heidelberg, Im Neuenheimer Feld 307, 69120 Heidelberg, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Skutella
mesh_terms: D000328:Adult; D018929:Cell Culture Techniques / Q000379:methods; D002454:Cell Differentiation / Q000235:genetics; D002469:Cell Separation / Q000379:methods; D002478:Cells, Cultured; D053595:Embryonic Stem Cells / Q000502:physiology; D005347:Fibroblasts / Q000502:physiology; D020869:Gene Expression Profiling / Q000379:methods; D006801:Humans; D008297:Male; D008875:Middle Aged; D013091:Spermatogenesis / Q000235:genetics*; D013093:Spermatogonia / Q000502:physiology*; D013737:Testis / Q000502:physiology; D059467:Transcriptome / Q000235:genetics*; D055815:Young Adult
references.citation: JAMA. 2009 Nov 18;302(19):2127-34
references.pmid: 19920237
references.citation: J Cell Physiol. 2013 Feb;228(2):298-305
references.pmid: 22740373
references.citation: Cancer Res. 2002 Sep 15;62(18):5358-64
references.pmid: 12235007
references.citation: Proc Natl Acad Sci U S A. 1999 Jun 8;96(12):6734-8
references.pmid: 10359781
references.citation: Oncogene. 2002 May 30;21(24):3879-88
references.pmid: 12032826
references.citation: Mol Reprod Dev. 2006 Apr;73(4):437-45
references.pmid: 16425234
references.citation: Asian J Androl. 2009 Sep;11(5):557-65
references.pmid: 19734913
references.citation: APMIS. 2003 Jan;111(1):197-210; discussion 210-1
references.pmid: 12760378
references.citation: Hum Mol Genet. 2002 Jul 1;11(14):1647-58
references.pmid: 12075009
references.citation: Oncogene. 2002 Apr 25;21(18):2880-9
references.pmid: 11973648
references.citation: Hum Mol Genet. 2007 May 1;16(9):1058-71
references.pmid: 17341483
references.citation: Hum Reprod. 2011 Jun;26(6):1296-306
references.pmid: 21349855
references.citation: Asian J Androl. 2008 Jul;10(4):569-76
references.pmid: 18478160
references.citation: Hum Reprod. 2010 Jan;25(1):158-67
references.pmid: 19815622
references.citation: Hum Reprod. 2012 Jan;27(1):210-21
references.pmid: 22095788
references.citation: Exp Cell Res. 2007 Apr 15;313(7):1373-92
references.pmid: 17359973
references.citation: Biol Reprod. 2006 Jan;74(1):102-8
references.pmid: 16162871
references.citation: Ren Fail. 2013;35(10):1387-91
references.pmid: 23991628
references.citation: Biomed Res Int. 2013;2013:143028
references.pmid: 23484080
references.citation: J Cell Sci. 2003 Aug 15;116(Pt 16):3363-72
references.pmid: 12857787
references.citation: Fertil Steril. 2009 Oct;92(4):1347-1350
references.pmid: 19463995
references.citation: Am J Hum Genet. 1997 Jan;60(1):237-41
references.pmid: 8981970
references.citation: Nature. 2011 Mar 24;471(7339):504-7
references.pmid: 21430778
references.citation: Dev Dyn. 2008 Jan;237(1):97-111
references.pmid: 18069690
references.citation: Nat Genet. 2004 Jun;36(6):647-52
references.pmid: 15156142
references.citation: J Biomed Biotechnol. 2012;2012:291038
references.pmid: 22431916
references.citation: Andrologia. 2012 May;44 Suppl 1:41-55
references.pmid: 21806653
references.citation: Nature. 2010 Jun 24;465(7301):E1; discussion E3
references.pmid: 20577160
references.citation: Blood. 1995 Dec 15;86(12):4544-52
references.pmid: 8541544
references.citation: Hum Reprod. 2013 Nov;28(11):3012-25
references.pmid: 24001715
references.citation: Stem Cells Dev. 2009 Oct;18(8):1115-26
references.pmid: 19281326
references.citation: Am J Anat. 1966 Mar;118(2):509-24
references.pmid: 5917196
references.citation: Cytogenet Genome Res. 2003;101(3-4):261-5
references.pmid: 14684992
references.citation: JAMA. 2011 Jun 15;305(23):2416-8
references.pmid: 21673293
references.citation: Acta Histochem. 2012 May;114(3):237-43
references.pmid: 21764106
references.citation: J Biol Chem. 1999 Jun 4;274(23):16180-7
references.pmid: 10347172
references.citation: Stem Cells. 2009 Jan;27(1):138-49
references.pmid: 18927477
references.citation: Exp Cell Res. 2008 Jan 1;314(1):213-26
references.pmid: 17964570
references.citation: Cell Prolif. 2010 Aug;43(4):405-17
references.pmid: 20590666
references.citation: Proc Natl Acad Sci U S A. 2000 Aug 15;97(17):9585-90
references.pmid: 10920202
references.citation: Clin Chem. 2009 Oct;55(10):1816-23
references.pmid: 19643838
references.citation: Cell Res. 2011 Aug;21(8):1164-71
references.pmid: 21747415
references.citation: Folia Histochem Cytobiol. 2004;42(2):119-21
references.pmid: 15253135
references.citation: Biochem Biophys Res Commun. 2009 Aug 7;385(4):570-5
references.pmid: 19482010
references.citation: Mech Dev. 2001 Jun;104(1-2):61-7
references.pmid: 11404080
references.citation: J Natl Cancer Inst. 1994 Jul 20;86(14):1070-5
references.pmid: 8021956
references.citation: Nature. 2009 Aug 13;460(7257):909-13
references.pmid: 19578360
references.citation: Biol Reprod. 2010 Feb;82(2):363-72
references.pmid: 19846602
references.citation: Nat Genet. 2002 Dec;32 Suppl:496-501
references.pmid: 12454644
references.citation: Nature. 2008 Nov 20;456(7220):344-9
references.pmid: 18849962