Osteocalcin improved Spermatogenesis in Azoospermic Mouse Model

Infertility is one of the major health concerns in the world whose globally incidence rate is on the rise. Several hormones are effective in this process, such as testosterone, estrogen and Osteocalcin (OCN). The Osteocalcin has Gprc6a receptors on leydig cells, which promotes the production of testosterone when mounted on these receptors. Therefore, the aim of this study was to investigate the role of Osteocalcin on the improvement of spermatogenesis and expression of Gprc6a receptors on leydig cells in the azoospermic mouse model.Twenty-five mice (4 to 6 weeks) were randomly divided into five groups: control, sham I Group that was initially injected by busulfan solvent (DMSO) at 5 weeks old and then by Osteocalcin solvent (PBS) after 5 weeks for one month, azoospermia Group that received busulfan (40 mg/kg/ip) at 5 weeks old. Sham II Group received busulfan 40mg/kg/ipandt hen after 5 weeks received PBS and experimental group, including azoospermic mice, was administered by Osteocalcin (3ng/g/d) for one month. After the last injection, the tests were dissected and then exposed to the tissue passage. To measure morphological changes, H & E staining was performed on a number of sections to measure the diameter of the seminiferous tubule, thickness of the germinal layer, count the spermatogonial cells, spermatocyte, round spermatid, long spermatid, Sertoli, leydigandmyoid cells. Image J software was used to conduct quantitative studies. Immunohistochemical method was employed to examine the expression of the specific receptor of Osteocalcin, Gprc6α, in the Leydig cell among the groups.The H & E staining showed a significant difference in the count the spermatogonial cells, spermatocytes, round spermatids, long spermatids, the thickness of the germ layer, the seminiferous tubule diameter between the studied groups (P<0.05).There was also no significant difference in the count of Sertoli, leydig, myoid cells and seminiferous tubule diameter between the groups (P<0.05). In the immune histochemistry, no significant difference was found in the count of GpRC6α-positive leydig cells between the groups (P< 0.05).According to the current results, the OCN plays an important role in spermatogenesis, which has a positive effect on the count of spermatocytes and spermatids, and can be further explored as an appropriate therapeutic strategy proposed for the infertility.