Journal of Reproduction & Infertility

Journal of Reproduction & Infertility

Decreased Expression of CDC25A in Azoospermia as the Etiology of Spermatogenesis Failure

Authors
Dwi Suryandari 1
Yurnadi Midoen 1
Luluk Yunaini 1
Sari Setyaningsih 2
Hans-Joachim Freisleben 3
1 Department of Medical Biology, Faculty of Medicine, Universitas Indonesia, Jakarta Pusat, Indonesia
2 Master Program Biomedical Sciences, Faculty of Medicine, Universitas Indonesia, Jakarta Pusat, Indonesia
3 Medical Research Unit, Faculty of Medicine, Universitas Indonesia, Jakarta Pusat, Indonesia
Abstract
Background: Spermatogenesis is a tightly regulated developmental process of male germ cells. The stages in spermatogenesis are mitosis, meiosis and spermiogenesis. One of the genes playing a role in meiosis is Cell Division Cycle 25A (CDC25A). Decreased expression of CDC25A is associated with failure of spermatogenesis and sperm retrieval. Infertility examination for azoospermia has been limited on histological examination. Hence, molecular research to find marker genes for infertility will improve the examination of testis biopsies. Methods: This research is a cross sectional study of 50 testicular biopsies with Johnsen scoring categories from scoring 2 to 8. Analysis of mRNA expression used qPCR and protein expression using immunohistochemistry. Statistical analysis with Spearman correlation was considered significant at p<0.05. Results: The result showed that transcript level and protein expression of CDC25A decreased in score 5 of Johnsen scoring categories. Moderate Spearman rho correlation (r=0.546) between mRNA relative expression and protein expression of CDC25A was significant at p<0.01. Conclusion: Decreased expression of CDC25A is associated with meiotic arrest as the etiology of spermatogenic failure in many azoospermic men.
Keywords
CDC25A
Infertility
Johnsen scoring
Spermatogenic failure
  1. Mahanta R, Gogoi A, Roy S, Bhattacharyya IK, Sharma P. Prevalence of azoospermia factor (AZF) deletions in idiopathic infertile males in North-East India. Int J Hum Genet. 2011;11(2):99-104.
  2. Massart A, Lissens W, Tournaye H, Stouffs K. Genetic causes of spermatogenic failure. Asian J Androl. 2012;14(1):40-8.
  3. Parikh UR, Goswami HM, Deliwala KJ, Shah AM, Barot HP. Testicular biopsy in male infertility: study of 80 cases. Int J Pathol 2012;25(2):75-7.
  4. Khabour OF, Fararjeh AS, Alfaouri AA. Genetic screening for AZF Y chromosome microdeletions in Jordanian azoospermic infertile men. Int J Mol Epidemiol Genet. 2014;5(1):47-50.
  5. Nourashrafeddin S, Ebrahimzadeh-Vesal R, Modaressi MH, Zekri A, Nouri M. Identification of Spata-19 new variant with exspression beyond meiotic phase of mouse testis development. Rep Biochem Mol Biol. 2014;2(2):89-93.
  6. Dohle GR, Weidner W, Jungwirth A, Colpi G, Papp G, Pomerol J, et al. Guidelines on male infertility. Netherlands: Europ Assc Urol; 2004 Mar.
  7. Don J, Stelzer G. The expanding family of CREB/CREM transcription factors that are involved with spermatogenesis. Mol Cell Endocrinol. 2002;187(1-2):115-24.
  8. Carrel DT, Emery BR, Hammoud S. Altered protamine expression and diminished spermatogenesis: what is the link? Hum Reprod Update. 2007;13(3):313-27.
  9. Luetjens CM, Xu EY, Rejo Pera RA, Kamischke A, Nieschlag E, Gromoll J. Association of meiotic arrest with lack of BOULE protein expression in infertile men. J Clin Endocrinol Metab. 2004;89(4):1926-33.
  10. Johnsen SG. Testicular biopsy score count-a method for registration of spermatogenesis in human testis: normal values and results in 335 hypogonadal males. Horm Res Paediatr. 1970;1(1):2-5.
  11. Gray P. Handbook of basic microtechnique. 1st ed. New York; Mc Graw Company; 1952. 252 p.
  12. Bibikova M, Yeakley JM, Chudin E, Chen J, Wickham E, Wang-Rodriguez J, et al. Gene expression profile in formalin-fixed, paraffin-embedded tissue obtained with a novel assay for microarray analysis. Clin Chem. 2004;50(12):2384-6.
  13. Brooke-Powell ET, Mandal TN, Ajioka JW. Use of transcriptor reverse transcriptase in microarray analysis. Biochemica. 2004;1:27-30.
  14. Untergasser A, Cutcutache I, Koressaar T, Ye J, Faircloth BC, Remm M, Rozen SG. Primer3--new capabilities and interfaces. Nucleic Acids Res. 2012;40(15):e115.
  15. Koressaar T, Remm M. Enhancements and modifications of primer design program Primer3. Bioinformatics. 2007;23(10):1289-91.
  16. Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods. 2001;25(4):402-8.
  17. Dorak M. Real Time PCR. 1st ed. USA: Taylor and Francais Group; 2006. 333 p.
  18. Bustin SA, Benes V, Garson JA, Hellemans J, Huggett J, Kubista M, et al. The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiment. Clin Chem. 2009;55(4):611-22.
  19. Life Technologies Corporation. Real Time PCR Handbook [Internet]. USA: Life technologies; 2012 [cited 2015 Dec 4]. 70 p. Available from: www.gene-quantification.com/real-time-pcr-handbook-life-technologies-update-flr.pdf.
  20. Kumar GL, Rudbeck L. Education guide: immunohistochemical (IHC) staining methods. 5th ed. California: Dako North America; 2009. 172 p.
  21. Hauke J, Kossowski T. Comparison of values of Pearson’s and Spearman’s correlation coefficients on the same sets of data. Quaestiones Geogr. 2011;30(2):87-93.
  22. Dahlan MS. Statistik untuk kedokteran dan kesehatan. 3rd ed. Jakarta: PT Arkans; 2011. 85 p.
  23. Green MR, Sambrook J. Molecular cloning: A laboratory manual. 4th ed. New York: Cold Spring Harbor Laboratory Press; 2012. 2028 p.
  24. Wistuba J, Luetjens CM, Wesselmann R, Nieschlag E, Simoni M, Schlatt S. Meiosis in autologous ectopic transplants of immature testicular tissue grafted to Callithrix jacchus. Biol Reprod. 2006;74(4):706-13.
  25. Cheng YS, Kuo PL, Teng YN, Kuo TY, Chung CL, Lin YH, et al. Association of spermatogenic failure with decreased CDC25A expression in infertile men. Hum Reprod. 2006;21(9):2346-52.
  26. Jiao X, Trifillis P, Kiledjian M. Identification of target messenger RNA substrates for murine deleted in azoospermia-like RNA-binding protein. Biol Reprod. 2002;66(2):475-85.
  27. Novitasari. Analysis of CREMτ and protamine expression in male azoospermic testicular tissue: the pathobiological review of spermatogenic arrest [master’s thesis]. [Indonesia]: Universitas Indonesia; 2015. 83 p.
  28. Ritz C, Spiess AN. qPCR: an R package for sigmoidal model selection in quantitative real-time polymerase chain reaction analysis. Bioinformatics. 2008;24(13):1549-51.
  29. Banard L, Kenneth AI. Spermatogenesis: methods and protocols. 1st ed. New York: Humana Press; 2013. Immunohistochemical approaches for the study of spermatogenesis; p. 309-20.
  30. Saleh R, Mahfouz RZ, Agarwal A, Farouk H. Histopathologic pattern of testicular biopsies in infertile azoospermic men with varicocele. Fertil Steril. 2010;94(6):2482-5.
  31. Lin ML, Chung CL, Cheng YS. Posttranscriptional regulation of CDC25A by BOLL is a conserved fertility mechanism essensial for human spermatogenesis. J Clin Endocrinol Metab. 2009;94(7):2650-7.
  32. Mizoguchi S, Kim KH. Expression of cdc25 phosphatase in germ cells of rat testis. Bio Reprod. 1997;56(6):1474-81.
  33. Bernardi R, Liebermann DA, Hoffman B. Cdc25A stability is controlled by the ubiquitin-proteasome pathway during cell cycle progression and terminal differentiation. Oncogene. 2000;19(20):2447-54.
  34. Tang Z, Coleman TR, Dunphy WG. Two distinct mechanisms for negative regulation of the Wee 1 protein kinase. EMBO J. 1993;12(9):3427-36.
  35. Lin YM, Teng YN, Chung CL, Tsai WC, Lin YH, Lin JS, et al. Decreased mRNA transcripts of M-phase promoting factor and its regulators in the testes of infertile men. Hum Reprod. 2006;21(1):138-44.
  36. Wang H, Wang Q, Pape UJ, Shen B, Huang J, Wu B, et al. Systemic investigation of global coordination among mRNA and protein in cellular society. BMC Genomics. 2010;11:364.
  37. Garcia E. A tutorial on correlation coefficients [Internet]. 2011 Jan 8 [cited 2015 Dec 3]. 13 p. Available from: https://pdfs.semanticscholar.org/c3e1/095209d3f72ff66e07b8f3b152fab099edea.pdf
  38. Schwanhäusser B, Busse D, Li N, Dittmar G, Schuchhardt J, Wolf J, et al. Corrigendum: Global quantification of mammalian gene expression control. Nature. 2013;495(7439):126-7.
  39. Vogel C, de Sousa Abreu R, Ko D, Le SY, Shapiro BA, Burns SC, et al. Sequence signature and mRNA concentration can explain two-thirds of protein abundance variation in human cell line. Mol Syst Biol. 2010;6(1):400.
  40. Moore KJ. microRNAs: small regulators with a big impact on lipid metabolism. J lipid Res. 2013;54(5):1159-60.
  41. Kendrick N. A gene’s mRNA level does not usually predict its protein level [Internet]. Madison, Wisconsin USA: Kendrick Laboratories, Inc. [cited 2015 Dec 20]; Available from: http://www.kendricklabs.com.
  42. Forbes K, Westwood M. The IGF axis and placental function. a mini review. Horm Res. 2008;69(3):129-37.
Journal of Reproduction & Infertility
Volume 19, Issue 2
Spring 2018
Pages 100-108