Effect of age on expression of spermatogonial markers in bovine testis and isolated cells

Spermatogonial stem cells (SSC) are the most undifferentiated germ cell present in adult male testes and, it is responsible to maintain the spermatogenesis. Age has a negative effect over stem cell, but the aging effect on SSC is not elucidated for bovine. The present study aim to evaluate the effect of age on the expression of undifferentiated spermatogonial markers in testis and in enriched testicular cells from prepubertal calves and adult bulls. In this matter, testicular parenchyma from calves (3–5 months) (n = 5) and bulls with 3 years of age (n = 5) were minced and, isolated cells were obtained after two enzymatic digestions. Differential platting was performed for two hours onto BSA coated dish. Cell viability was assessed by Trypan Blue solution exclusion method and testicular cells enriched for SSC was evaluated by expression of specific molecular markers by qRT-PCR (POU5F1, GDNF, CXCR4, UCHL1, ST3GAL, SELP, ICAM1 and ITGA6) and flow cytometry (GFRA1, CXCR4 and ITGA6). CXCR4 and UCHL1 expression was evaluated in fixated testes by immunohistochemistry. We observed that age just affected the expression of selective genes [SELP (Fold Change = 5.61; p = 0.0023) and UCHL1 (Fold Change = 4.98; p = 0.0127)]. By flow cytometry, age affected only the proportion of ITGA6+ cells (P < 0.001), which was higher in prepubertal calves when compared to adult bulls. In situ, we observed an effect of age on the number of UCHL1+ (p = 0.0006) and CXCR4+ (p = 0.0139) cells per seminiferous tubule. At conclusion, age affects gene expression and the population of cells expressing specific spermatogonial markers in the bovine testis.