Histochemistry and Cell Biology

, Volume 135, Issue 1, pp 73–81 | Cite as

Immunohistochemical expression analysis of Cx43, Cx26, c-KIT and PlAP in contralateral testis biopsies of patients with non-seminomatous testicular germ cell tumor

  • Marianne Steiner
  • Klara Weipoltshammer
  • Gerhard Viehberger
  • Eva-Maria Meixner
  • Gerhard Lunglmayr
  • Christian Schöfer
Original Paper


Non seminomatous testicular germ cell tumors (NSTGCTs) express fetal stem cell markers and display dysregulation of connexin 43 expression. Persistence of fetal spermatogonial characteristics was implicated in the emergence of testicular germ cell tumors. The objective of this study was to analyze the tubular architecture in contralateral testes of patients with NSTGCT. We studied morphologic alterations, expression patterns of markers for the integrity of the germinal epithelium (gap junction proteins connexin 43 and 26), as well as of the embryonic markers c-KIT and placental alkaline phosphatase (PlAP), both established markers to detect carcinoma in situ (CIS). In all samples, tubules showing maturation of germ cells up to spermatozoa were observed. In addition, tubules with alterations in tubular architecture and with impaired spermatogenesis occurred. In tubules showing aberrant spermatogenesis, connexin 43 (Cx43) signal was down-regulated and a shift of signal from gap junctions to the cytoplasm occurred. Concomitantly, Cx26 was found highly up-regulated in tubules with incomplete and aberrant germ cell maturation. All testes exhibited single spermatogonia with positive reaction for c-KIT and a significant positive correlation was found between the mean number of c-KIT positive spermatogonia per tubule and the percentage of tubules presenting severely impaired spermatogenesis. Our data show alterations of the normal architecture of the germinal epithelium and disturbances of spermatogenesis in the contralateral testes of patients with NSTGCT in all cases evaluated. The concomitant occurrence of c-KIT positive spermatogonia and defects in tubular architecture is in line with the hypothesis that patients with NSTGCT suffer from disturbed germ cell development.


CIS (carcinoma in situ) Gap junction Immunohistochemistry Dysgenesis Testicular architecture 



The expert technical assistance of Ines Rammel is greatly acknowledged. We would like to thank Dr. Christa Freibauer, Institute of Clinical Pathology, Mistelbach General Hospital, for providing archival material and Michaela Leisser for assistance with PlAP and c-KIT staining. We are grateful to Mag. Christian Huber, Bernhard Gottlieb University for Dental Medicine, for his assistance with statistics.

Supplementary material

418_2010_769_MOESM1_ESM.pdf (100 kb)
Supplementary material 1 (PDF 99 kb)
418_2010_769_MOESM2_ESM.pdf (39 kb)
Supplementary material 2 (PDF 39.4 kb)
418_2010_769_MOESM3_ESM.pdf (10 kb)
Supplementary material 3 (PDF 10.1 kb)


  1. Brehm R, Marks A, Rey R, Kliesch S, Bergmann M, Steger K (2002) Altered expression of connexins 26 and 43 in Sertoli cells in seminiferous tubules infiltrated with carcinoma-in situ or seminoma. J Pathol 197:647–653CrossRefPubMedGoogle Scholar
  2. Brehm R, Ruttinger C, Fischer P, Gashaw I, Winterhager E, Kliesch S, Bohle RM, Steger K, Bergmann M (2006) Transition from preinvasive carcinoma in situ to seminoma is accompanied by a reduction of connexin 43 expression in Sertoli cells and germ cells. Neoplasia 8:499–509CrossRefPubMedGoogle Scholar
  3. Cronier L, Crespin S, Strale PO, Defamie N, Mesnil M (2009) Gap junctions and cancer: new functions for an old story. Antioxid Redox Signal 11:323–338CrossRefPubMedGoogle Scholar
  4. Dieckmann KP, Pichlmeier U (2004) Clinical epidemiology of testicular germ cell tumors. World J Urol 22:2–14CrossRefPubMedGoogle Scholar
  5. Dieckmann KP, Linke J, Pichlmeier U, Kulejewski M, Loy V (2007) Spermatogenesis in the contralateral testis of patients with testicular germ cell cancer: histological evaluation of testicular biopsies and a comparison with healthy males. BJU Int 99:1079–1085CrossRefPubMedGoogle Scholar
  6. Eble JN SG, Epstein JI, Sesterhenn IAE (2004) World health organization of tumours: pathology and genetics of tumours of the urinary system and male genital organs. IARC Press, LyonGoogle Scholar
  7. Fiorini C, Gilleron J, Carette D, Valette A, Tilloy A, Chevalier S, Segretain D, Pointis G (2008) Accelerated internalization of junctional membrane proteins (connexin 43, N-cadherin and ZO-1) within endocytic vacuoles: an early event of DDT carcinogenicity. Biochim Biophys Acta 1778:56–67CrossRefPubMedGoogle Scholar
  8. Fossa SD, Chen J, Schonfeld SJ, McGlynn KA, McMaster ML, Gail MH, Travis LB (2005) Risk of contralateral testicular cancer: a population-based study of 29, 515 U.S. men. J Natl Cancer Inst 97:1056–1066CrossRefPubMedGoogle Scholar
  9. Geczi L, Gomez F, Bak M, Bodrogi I (2003) The incidence, prognosis, clinical and histological characteristics, treatment, and outcome of patients with bilateral germ cell testicular cancer in Hungary. J Cancer Res Clin Oncol 129:309–315PubMedGoogle Scholar
  10. Gilleron J, Carette D, Carpentier F, Segretain D, Pointis G (2009) Three-dimensional analysis of connexin 43 gap junction in the ex vivo rat seminiferous tubules: short-term effects of hormonal effectors. Microsc Res Tech 72:845–855CrossRefPubMedGoogle Scholar
  11. Hayashi T, Arai G, Hyochi N, Suzuki M, Masuda H, Kawakami S, Okuno T, Ishizaka K, Kageyama Y, Kihara K (2001) Suppression of spermatogenesis in ipsilateral and contralateral testicular tissues in patients with seminoma by human chorionic gonadotropin beta subunit. Urology 58:251–257CrossRefPubMedGoogle Scholar
  12. Hoei-Hansen CE, Holm M, Rajpert-De Meyts E, Skakkebaek NE (2003) Histological evidence of testicular dysgenesis in contralateral biopsies from 218 patients with testicular germ cell cancer. J Pathol 200:370–374CrossRefPubMedGoogle Scholar
  13. Hoei-Hansen CE, Olesen IA, Jorgensen N, Carlsen E, Holm M, Almstrup K, Leffers H, Rajpert-De Meyts E (2007) Current approaches for detection of carcinoma in situ testis. Int J Androl 30:398–404 (discussion 404-395)CrossRefPubMedGoogle Scholar
  14. Honecker F, Stoop H, de Krijger RR, Chris Lau YF, Bokemeyer C, Looijenga LH (2004) Pathobiological implications of the expression of markers of testicular carcinoma in situ by fetal germ cells. J Pathol 203:849–857CrossRefPubMedGoogle Scholar
  15. Jorgensen N, Rajpert-De Meyts E, Graem N, Muller J, Giwercman A, Skakkebaek NE (1995) Expression of immunohistochemical markers for testicular carcinoma in situ by normal human fetal germ cells. Lab Invest 72:223–231PubMedGoogle Scholar
  16. Krege S, Beyer J, Souchon R et al (2008) European consensus conference on diagnosis and treatment of germ cell cancer: a report of the second meeting of the European Germ Cell Cancer Consensus group (EGCCCG): part I. Eur Urol 53:478–496CrossRefPubMedGoogle Scholar
  17. Laird DW (2005) Connexin phosphorylation as a regulatory event linked to gap junction internalization and degradation. Biochim Biophys Acta 1711:172–182CrossRefPubMedGoogle Scholar
  18. Oosterhuis JW, Looijenga LH (2005) Testicular germ-cell tumours in a broader perspective. Nat Rev Cancer 5:210–222CrossRefPubMedGoogle Scholar
  19. Pauls K, Schorle H, Jeske W, Brehm R, Steger K, Wernert N, Buttner R, Zhou H (2006) Spatial expression of germ cell markers during maturation of human fetal male gonads: an immunohistochemical study. Hum Reprod 21:397–404CrossRefPubMedGoogle Scholar
  20. Rajpert-De Meyts E, Skakkebaek NE (1994) Expression of the c-kit protein product in carcinoma-in situ and invasive testicular germ cell tumours. Int J Androl 17:85–92CrossRefPubMedGoogle Scholar
  21. Rajpert-De Meyts E, Bartkova J, Samson M, Hoei-Hansen CE, Frydelund-Larsen L, Bartek J, Skakkebaek NE (2003) The emerging phenotype of the testicular carcinoma in situ germ cell. APMIS 111:267–278 (discussion 278-269)PubMedGoogle Scholar
  22. Sette C, Dolci S, Geremia R, Rossi P (2000) The role of stem cell factor and of alternative c-kit gene products in the establishment, maintenance and function of germ cells. Int J Dev Biol 44:599–608PubMedGoogle Scholar
  23. Steger K, Tetens F, Bergmann M (1999) Expression of connexin 43 in human testis. Histochem Cell Biol 112:215–220CrossRefPubMedGoogle Scholar
  24. Stoop H, Honecker F, van de Geijn GJ, Gillis AJ, Cools MC, de Boer M, Bokemeyer C, Wolffenbuttel KP, Drop SL, de Krijger RR, Dennis N, Summersgill B, McIntyre A, Shipley J, Oosterhuis JW, Looijenga LH (2008) Stem cell factor as a novel diagnostic marker for early malignant germ cells. J Pathol 216:43–54CrossRefPubMedGoogle Scholar
  25. UICC (2009) Classification of malignant tumours. Wiley & Sons, New YorkGoogle Scholar
  26. Winterhager E, Pielensticker N, Freyer J, Ghanem A, Schrickel JW, Kim JS, Behr R, Grummer R, Maass K, Urschel S, Lewalter T, Tiemann K, Simoni M, Willecke K (2007) Replacement of connexin43 by connexin26 in transgenic mice leads to dysfunctional reproductive organs and slowed ventricular conduction in the heart. BMC Dev Biol 7:26CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  • Marianne Steiner
    • 1
  • Klara Weipoltshammer
    • 1
  • Gerhard Viehberger
    • 1
  • Eva-Maria Meixner
    • 2
  • Gerhard Lunglmayr
    • 3
  • Christian Schöfer
    • 1
  1. 1.Department for Cell- and Developmental Biology, Center for Anatomy and Cell BiologyMedical University of ViennaViennaAustria
  2. 2.Department of Clinical PathologyMistelbach General HospitalMistelbachAustria
  3. 3.Karl Landsteiner Institute for Andrology and Prostate ResearchMistelbachAustria

Personalised recommendations