Expression of 3β-Hydroxysteroid Dehydrogenase in Rat Leydig Cells Cultured by Differential Adhesion In Vitro

Objective To investigate the constitution of cells population, changes in 3β-hydroxysteroid dehydrogenase (3β-HSD) expression and androgen productivity in rat Leydig cells cultured by differential adhesion during 4 days primary culture. Methods Testes from 7 days and 3 weeks old rats were harvested, then Leydig cells were isolated by using collagenase dispersion, stainless steel mesh infiltration and differential adhesion. 3β-HSD expression in cultured cells was observed by immunohistochemistry and flow Cytometry analysis immediately after cell isolation and 4 days later. Testosterone level in isolated Leydig cells with or without HCG stimulation was also tested. Results In 7 days group and 3 weeks old group 3β-HSD positive rates in cultured cells were 5.4%±1.2% and 59.2%±3.2% respectively after isolation immediately. 4 days after culture, most of the cultured cells (93.6%±1.2% in 7 days old group, 95.4%±3.2% in 3 weeks old group) became 3β-HSD positive. Testosterone level, increased by HCG stimulation, in cultured cells from both groups. Conclusion Leydig cells obtained by differential adhesion from 7 days and 3 weeks old rats consist Stem Leydig cells (SLCs). These SLCs were differentiated and gained 3β-HSD activity during cell culture.