Follistatin (FST)， a local regulator of gonadal functions is a powerful inhibitor of follicle stimulating hormone (FSH) secretion. In the present study， the expression of FST was partially silenced at both transcriptional and translational levels by RNAi-Ready pSIREN-RetroQ-ZsGreen Vector mediated recombinant pshRNA vectors in bovine granulosa cells (bGCs). The results showed that transfection with FST-1 and FST-2 vectors significantly down-regulated mRNA and protein expressions of follistatin by 51% (P = 0.0093) and 72% (P = 0.0078) respectively. After down-regulation of FST in bGCs， cell cycle was arrested at S-phase (9.2 ± 0.6 vs 12.5 ± 0.2， P = 0.0055)， and apoptosis was significantly (21.3 ± 2... More
Follistatin (FST)， a local regulator of gonadal functions is a powerful inhibitor of follicle stimulating hormone (FSH) secretion. In the present study， the expression of FST was partially silenced at both transcriptional and translational levels by RNAi-Ready pSIREN-RetroQ-ZsGreen Vector mediated recombinant pshRNA vectors in bovine granulosa cells (bGCs). The results showed that transfection with FST-1 and FST-2 vectors significantly down-regulated mRNA and protein expressions of follistatin by 51% (P = 0.0093) and 72% (P = 0.0078) respectively. After down-regulation of FST in bGCs， cell cycle was arrested at S-phase (9.2 ± 0.6 vs 12.5 ± 0.2， P = 0.0055)， and apoptosis was significantly (21.3 ± 2.7 vs 13.9 ± 2.5， P = 0.0051) increased. These findings were further verified by down-regulation of protein level of B-cell leukemia/lymphoma 2 (Bcl2， P = 0.0423)， and up-regulation of caspase-3 (P = 0.0362)， p21 (P = 0.0067) and mRNA levels of Bcl2-associated X protein (Bax， P = 0.041). Knockdown of FST in bGCs significantly increased activin A concentration in culture medium， while level of estradiol (E2) was suppressed without affecting progesterone production. In addition， mRNA levels of all activin receptor subtypes [activin receptor types I (ACRI) and II (ACRIIA and ACRIIB)] and inhibin α-subunit were augmented (P < 0.05) without altering both inhibin β-subunits. These findings suggest that follistatin may participate in caspase3-dependent apoptosis through Bcl2/Bax gene family in bovine GCs， whereas， activin and its receptors are associated with its regulation. Activin-induced up-regulation of inhibin-α subunit in bGCs seems to be involved in the regulation of steroidogenesis.
