Effect of vitamin C on colony formation of ovine spermatogonial stem cells in vitro

The technology of using spermatogonial stem cells (SSCs) has been limited due to lack of an ideal culture system for growth and proliferation. The aim of this study was to investigate the effects of different doses of vitamin C on SSCs colony formation in vitro.
The cells were isolated from testes of prepubertal lambs by two enzymatic digestions, purified by differential platting, and then treated for 10 days by using 4 Simple culture including SSCs in DMEM containing 1% antibiotic and 5% FBS as our control group and for the three other cultures we used the same culture medium as that in control group plus 20, 40 and 60 µg/ml of vitamin C respectively. Culture media were refreshed every 72h and colony numbers and diameters were determined on the 4th , 7th and 10th days after the beginning of culture by using inverted microscope. Spermatogonial cells were identified by immunocytochemistry staining against PGP9.5. Using R software, the results obtained from 5 repeats were evaluated by ANOVA. P On the 7th day, we found a significant difference between the culture No. 2 (0.41 mm2) and culture No. 3 (0.08 mm2) in regard to spermatogonial colonies surface areas (P The results of this study showed that vitamin C with a dose of 40 (μg/ml) was effective in increasing the surface area of spermatogonial colonies. But it had no effect on spermatogonial cell number.