Influence of Nitric Oxide on Steroid Synthesis, Growth and Apoptosis of Buffalo (Bubalus bubalis) Granulosa Cells In vitro

Abstract

Objective of this study was to examine the effect of sodium nitroprusside (SNP), a nitric oxide (NO) donor on steroid synthesis, growth and apoptosis of buffalo granulosa cells (GCs) in vitro. Follicular fluid of antral follicles (3-5 mm diameter) was aspirated and GCs were cultured in 0 (control), , , , of SNP for 48 h. To evaluate whether this effect was reversible, GCs were cultured in presence of SNP+1.0 mM -nitro-L-arginine methyl ester (L-NAME) a NO synthase (NOS) inhibitor or hemoglobin (Hb, ) as NO scavenger. Nitrate/nitrite concentration was evaluated by Griess method, progesterone and estradiol concentrations by RIA and apoptosis by TUNEL assay. SNP (, , ) significantly (p SNP induced the progesterone synthesis and stimulated GCs to develop into a uniform monolayer. Combination of SNP M+L-NAME strengthened the inhibitory effect while, SNP+Hb together reversed these inhibitory effects. In conclusion, SNP at greater concentrations (, and ) has a cytotoxic effect and it may lead to cell death whereas, at a lower concentration () induced progesterone synthesis and growth of GCs. These findings have important implications that NOS derived NO are involved at physiological level during growth and development of buffalo GCs which regulates the steroidogenesis, growth and apoptosis.