SCF Improves In Vitro Differentiation of SSCs Through Transcriptionally Up-regulating PRTM1, STRA8, c-KIT, PIWIL2, and OCT4 Genes

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Abstract

Several lines of evidence strongly suggest that retinoic acid (RA) and stem cell factor (SCF)/c-Kit signal transduction pathways are involved in the differentiation of spermatogonial stem cells (SSCs). This study was aimed to investigate the effect of RA and SCF on in vitro differentiation of SSCs via evaluation of the mRNA expression of meiosis-specific genes in cultured testicular tissues. Testicular tissue samples were obtained from bilaterally vasectomized rats and also healthy adult rats and then were cultured for 25, 30, and 35 days on different conditions. The cultured testicular pieces were sectioned and stained with PAS to histological analysis. The total RNA was extracted from cultured testicular samples, and the expression of ACR, PRTM1, SYCP3, STRA8, c-KIT, PIWIL2, and OCT4 genes at mRNA level was quantified using real-time polymerase chain reaction (qPCR) procedure. After 1-month surgery, bilateral testicular weight showed a significant decrease in vasectomized adult rats compared with healthy adult rats (P < 0.05). Reduction in the diameter of the seminiferous tubules and depletion of advanced germinal elements in vasectomized rats compared with healthy adult rats were also observed. Our findings also demonstrated that the mRNA expression level of PRTM1, STRA8, c-KIT, PIWIL2, and OCT4 genes in cultured testicular tissues significantly up-regulated in experimental group II compared with the control group (P < 0.001). Our findings lead us to conclude that SCF improves in vitro differentiation of SSCs in the OA rats, at least partially, by transcriptionally upregulating PRTM1, STRA8, c-KIT, PIWIL2, and OCT4 genes.

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References

  1. 1.

    Krausz C, Riera-Escamilla A. Genetics of male infertility. Nat Rev Urol. 2018;15:369–84.

    CAS  PubMed  PubMed Central  Google Scholar 

  2. 2.

    Aston KI. Genetic susceptibility to male infertility: news from genome-wide association studies. Andrology. 2014;2:315–21.

    CAS  PubMed  PubMed Central  Google Scholar 

  3. 3.

    Miyamoto T, Minase G, Shin T, Ueda H, Okada H, Sengoku K. Human male infertility and its genetic causes. Reprod Med Biol. 2017;16:81–8.

    CAS  PubMed  PubMed Central  Google Scholar 

  4. 4.

    Leaver RB. Male infertility: an overview of causes and treatment options. Br J Nurs. 2016;25:35–40.

    Google Scholar 

  5. 5.

    Lotti F, Maggi M. Sexual dysfunction and male infertility. Nat Rev Urol. 2018;15:287–307.

    PubMed  PubMed Central  Google Scholar 

  6. 6.

    Bracke A, Peeters K, Punjabi U, Hoogewijs D, Dewilde S. A search for molecular mechanisms underlying male idiopathic infertility. Reprod BioMed Online. 2018;36:327–39.

    CAS  PubMed  PubMed Central  Google Scholar 

  7. 7.

    Oatley JM, Brinster RL. Regulation of spermatogonial stem cell self-renewal in mammals. Annu Rev Cell Dev Biol. 2008;24:263–86.

    CAS  PubMed  PubMed Central  Google Scholar 

  8. 8.

    Griswold MD. Spermatogenesis: the commitment to meiosis. Physiol Rev. 2016;96:1–17.

    CAS  PubMed  PubMed Central  Google Scholar 

  9. 9.

    Mecklenburg JM, Hermann BP. Mechanisms regulating spermatogonial differentiation. Results Probl Cell Differ. 2016;58:253–87.

    CAS  PubMed  PubMed Central  Google Scholar 

  10. 10.

    de Rooij DG. Proliferation and differentiation of spermatogonial stem cells. Reproduction. 2001;121:347–54.

    PubMed  PubMed Central  Google Scholar 

  11. 11.

    Sun M, Yuan Q, Niu M, Wang H, Wen L, Yao C, et al. Efficient generation of functional haploid spermatids from human germline stem cells by three-dimensional-induced system. Cell Death Differ. 2018;25:749–66.

    PubMed  PubMed Central  Google Scholar 

  12. 12.

    Song HW, Wilkinson MF. In vitro spermatogenesis: a long journey to get tails. Spermatogenesis. 2012;2:238–44.

    PubMed  PubMed Central  Google Scholar 

  13. 13.

    Ibtisham F, Wu J, Xiao M, An L, Banker Z, Nawab A, et al. Progress and future prospect of in vitro spermatogenesis. Oncotarget. 2017;8:66709–27.

    PubMed  PubMed Central  Google Scholar 

  14. 14.

    Sato T, Katagiri K, Yokonishi T, Kubota Y, Inoue K, Ogonuki N, et al. In vitro production of fertile sperm from murine spermatogonial stem cell lines. Nat Commun. 2011;2:472.

    PubMed  PubMed Central  Google Scholar 

  15. 15.

    Sato T, Katagiri K, Kubota Y, Ogawa T. In vitro sperm production from mouse spermatogonial stem cell lines using an organ culture method. Nat Protoc. 2013;8:2098–104.

    CAS  PubMed  PubMed Central  Google Scholar 

  16. 16.

    Sato T, Katagiri K, Kojima K, Komeya M, Yao M, Ogawa T. In vitro spermatogenesis in explanted adult mouse testis tissues. PLoS One. 2015;10:e0130171.

    PubMed  PubMed Central  Google Scholar 

  17. 17.

    Reda A, Hou M, Winton TR, Chapin RE, Söder O, Stukenborg JB. In vitro differentiation of rat spermatogonia into round spermatids in tissue culture. Mol Hum Reprod. 2016;22:601–12.

    CAS  PubMed  PubMed Central  Google Scholar 

  18. 18.

    Gohbara A, Katagiri K, Sato T, Kubota Y, Kagechika H, Araki Y, et al. In vitro murine spermatogenesis in an organ culture system. Biol Reprod. 2010;83:261–7.

    CAS  Google Scholar 

  19. 19.

    Sanjo H, Komeya M, Sato T, Abe T, Katagiri K, Yamanaka H, et al. In vitro mouse spermatogenesis with an organ culture method in chemically defined medium. PLoS One. 2018;13:e0192884.

    PubMed  PubMed Central  Google Scholar 

  20. 20.

    Mohammadzadeh E, Mirzapour T, Nowroozi MR, Nazarian H, Piryaei A, Alipour F, et al. Differentiation of spermatogonial stem cells by soft agar three-dimensional culture system. Artif Cells Nanomed Biotechnol. 2019;47:1772–81.

    CAS  PubMed  PubMed Central  Google Scholar 

  21. 21.

    Ziloochi Kashani M, Bagher Z, Asgari HR, Najafi M, Koruji M, Mehraein F. Differentiation of neonate mouse spermatogonial stem cells on three-dimensional agar/polyvinyl alcohol nanofiber scaffold. Syst Biol Reprod Med. 2020;6(1):1–14.

    Google Scholar 

  22. 22.

    AbuMadighem A, Solomon R, Stepanovsky A, Kapelushnik J, Shi Q, Meese E, et al. Development of spermatogenesis in vitro in three-dimensional culture from spermatogonial cells of busulfan-treated immature mice. Int J Mol Sci. 2018;19:E3804.

    PubMed  PubMed Central  Google Scholar 

  23. 23.

    Roser JF. Regulation of testicular function in the stallion: an intricate network of endocrine, paracrine and autocrine systems. Anim Reprod Sci. 2008;107:179–96.

    CAS  PubMed  PubMed Central  Google Scholar 

  24. 24.

    Huleihel M, Lunenfeld E. Regulation of spermatogenesis by paracrine/autocrine testicular factors. Asian J Androl. 2004;6:259–68.

    CAS  PubMed  PubMed Central  Google Scholar 

  25. 25.

    Li X, Long XY, Xie YJ, Zeng X, Chen X, Mo ZC. The roles of retinoic acid in the differentiation of spermatogonia and spermatogenic disorders. Clin Chim Acta. 2019;497:54–60.

    CAS  PubMed  PubMed Central  Google Scholar 

  26. 26.

    Busada JT, Geyer CB. The role of retinoic acid (RA) in spermatogonial differentiation. Biol Reprod. 2016;94:10.

    PubMed  PubMed Central  Google Scholar 

  27. 27.

    Feng LX, Ravindranath N, Dym M. Stem cell factor/c-kit up-regulates cyclin D3 and promotes cell cycle progression via the phosphoinositide 3-kinase/p70 S6 kinase pathway in spermatogonia. J Biol Chem. 2000;275:25572–6.

    CAS  PubMed  PubMed Central  Google Scholar 

  28. 28.

    Koli S, Mukherjee A, Reddy KVR. Retinoic acid triggers c-kit gene expression in spermatogonial stem cells through an enhanceosome constituted between transcription factor binding sites for retinoic acid response element (RARE), spleen focus forming virus proviral integration oncogene (SPFI1) (PU.1) and E26 transformation-specific (ETS). Reprod Fertil Dev. 2017;29:521–43.

    CAS  PubMed  PubMed Central  Google Scholar 

  29. 29.

    Amann PM, Eichmüller SB, Schmidt J, Bazhin AV. Regulation of gene expression by retinoids. Curr Med Chem. 2011;18:1405–12.

    CAS  PubMed  PubMed Central  Google Scholar 

  30. 30.

    Ijiri TW, Mahbub Hasan AK, Sato K. Protein-tyrosine kinase signaling in the biological functions associated with sperm. J Signal Transduct. 2012;2012:181560.

    PubMed  PubMed Central  Google Scholar 

  31. 31.

    Sato T, Katagiri K, Gohbara A, Inoue K, Ogonuki N, Ogura A, et al. In vitro production of functional sperm in cultured neonatal mouse testes. Nature. 2011;471:504–7.

    CAS  PubMed  PubMed Central  Google Scholar 

  32. 32.

    Russell LD, Ettlin RA, Hikim APS, Clegg ED. Histological and histopathological evaluation of the testis. Int J Androl. 1993;16:83.

    Google Scholar 

  33. 33.

    Travers A, Arkoun B, Safsaf A, Milazzo JP, Absyte A, Bironneau A, et al. Effects of vitamin a on in vitro maturation of pre-pubertal mouse spermatogonial stem cells. PLoS One. 2013;8:e82819.

    PubMed  PubMed Central  Google Scholar 

  34. 34.

    Arkoun B, Dumont L, Milazzo JP, Rondanino C, Bironneau A, Wils J, et al. Does soaking temperature during controlled slow freezing of pre-pubertal mouse testes influence course of in vitro spermatogenesis? Cell Tissue Res. 2016;364:661–74.

    CAS  PubMed  PubMed Central  Google Scholar 

  35. 35.

    Dumont L, Oblette A, Rondanino C, Jumeau F, Bironneau A, Liot D, et al. Vitamin a prevents round spermatid nuclear damage and promotes the production of motile sperm during in vitro maturation of vitrified pre-pubertal mouse testicular tissue. Mol Hum Reprod. 2016;22:819–32.

    CAS  PubMed  PubMed Central  Google Scholar 

  36. 36.

    Di Lorenzo A, Bedford MT. Histone arginine methylation. FEBS Lett. 2011;585:2024–31.

    PubMed  PubMed Central  Google Scholar 

  37. 37.

    Li J, Zhou F, Zhan D, Gao Q, Cui N, Li J, et al. A novel histone H4 arginine 3 methylation-sensitive histone H4 binding activity and transcriptional regulatory function for signal recognition particle subunits SRP68 and SRP72. J Biol Chem. 2012;287:40641–51.

    CAS  PubMed  PubMed Central  Google Scholar 

  38. 38.

    Jeong HS, Bhin J, Joon Kim H, Hwang D, Ryul Lee D, Kim KS. Transcriptional regulatory networks underlying the reprogramming of spermatogonial stem cells to multipotent stem cells. Exp Mol Med. 2017;49:e315.

    CAS  PubMed  PubMed Central  Google Scholar 

  39. 39.

    Ma HT, Niu CM, Xia J, Shen XY, Xia MM, Hu YQ, et al. Stimulated by retinoic acid gene 8 (Stra8) plays important roles in many stages of spermatogenesis. Asian J Androl. 2018;20:479–87.

    CAS  PubMed  PubMed Central  Google Scholar 

  40. 40.

    Anderson EL, Baltus AE, Roepers-Gajadien HL, Hassold TJ, de Rooij DG, van Pelt AM, et al. Stra8 and its inducer, retinoic acid, regulate meiotic initiation in both spermatogenesis and oogenesis in mice. Proc Natl Acad Sci U S A. 2008;105:14976–80.

    CAS  PubMed  PubMed Central  Google Scholar 

  41. 41.

    Raverdeau M, Gely-Pernot A, Féret B, Dennefeld C, Benoit G, Davidson I, et al. Retinoic acid induces Sertoli cell paracrine signals for spermatogonia differentiation but cell autonomously drives spermatocyte meiosis. Proc Natl Acad Sci U S A. 2012;109:16582–7.

    CAS  PubMed  PubMed Central  Google Scholar 

  42. 42.

    Zhou Q, Nie R, Li Y, Friel P, Mitchell D, Hess RA, et al. Expression of stimulated by retinoic acid gene 8 (Stra8) in spermatogenic cells induced by retinoic acid: an in vivo study in vitamin A-sufficient postnatal murine testes. Biol Reprod. 2008;79:35–42.

    CAS  PubMed  PubMed Central  Google Scholar 

  43. 43.

    Cardoso HJ, Figueira MI, Socorro S. The stem cell factor (SCF)/c-KIT signalling in testis and prostate cancer. J Cell Commun Signal. 2017;11:297–307.

    PubMed  PubMed Central  Google Scholar 

  44. 44.

    Zhang L, Tang J, Haines CJ, Feng H, Lai L, Teng X, et al. c-kit expression profile and regulatory factors during spermatogonial stem cell differentiation. BMC Dev Biol. 2013;13:38.

    PubMed  PubMed Central  Google Scholar 

  45. 45.

    Lee JH, Engel W, Nayernia K. Stem cell protein Piwil2 modulates expression of murine spermatogonial stem cell expressed genes. Mol Reprod Dev. 2006;73:173–9.

    CAS  PubMed  PubMed Central  Google Scholar 

  46. 46.

    Esch D, Vahokoski J, Groves MR, Pogenberg V, Cojocaru V, Vom Bruch H, et al. A unique Oct4 interface is crucial for reprogramming to pluripotency. Nat Cell Biol. 2013;15:295–301.

    CAS  PubMed  PubMed Central  Google Scholar 

  47. 47.

    Niwa H, Miyazaki J, Smith AG. Quantitative expression of Oct-3/4 defines differentiation, dedifferentiation or self-renewal of ES cells. Nat Genet. 2000;24:372–6.

    CAS  PubMed  PubMed Central  Google Scholar 

  48. 48.

    Dann CT, Alvarado AL, Molyneux LA, Denard BS, Garbers DL, Porteus MH. Spermatogonial stem cell self-renewal requires OCT4, a factor downregulated during retinoic acid-induced differentiation. Stem Cells. 2008;26:2928–37.

    CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgments

The authors would like to express their sincere gratitude to Vice-Chancellor of Research and Technology at Babol University of Medical Sciences, Cellular and Molecular Biology Research Center at Babol University of Medical Sciences, the staff of Stem Cell Laboratory of the Reproductive Health and Infertility Research Center of Fatemeh Al-Zahra Hospital and Islamic Azad University of Ayatollah Amoli.

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Mahnaz Nasimi, Seyed Gholam Ali Jorsaraei, and Esmail Fattahi designed the experiments, performed the statistical analysis, contributed to the interpretation of the results, and wrote the manuscript; Mahnaz Nasimi, Maryam Gholamitabar Tabari, and Ebrahim Zabihi Neyshaburi carried out the experiments; Seyed Gholam Ali Jorsaraei and Esmail Fattahi supervised the project.

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Correspondence to Seyed Gholam Ali Jorsaraei.

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The current study was approved by the ethics committee for experimental research on animals at Babol University of Medical Sciences, Babol, Iran.

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This article was updated to correct the spelling of Esmail Fattahi’s name in the author listing.

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Nasimi, M., Jorsaraei, S.G.A., Fattahi, E. et al. SCF Improves In Vitro Differentiation of SSCs Through Transcriptionally Up-regulating PRTM1, STRA8, c-KIT, PIWIL2, and OCT4 Genes. Reprod. Sci. (2021). https://doi.org/10.1007/s43032-020-00326-z

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Keywords

  • Male infertility
  • Retinoid
  • SCF
  • Spermatogonial differentiation
  • Meiotic marker