Apoptosis of bovine granulosa cells: Intracellular pathways and differentiation

CAROU MA. CLARA; CRUZANS PAULA; FARINA MARIANA; FIORITO CARLA; MARURI ALEJANDRO; OLEA GABRIELA; LOMBARDO DANIEL

ACTA HISTOCHEMICA

ELSEVIER GMBH, URBAN & FISCHER VERLAG

Año: 2017

0065-1281

GnRH receptors have been identified in ovarian granulosa and luteal cells. In vivo and in vitro hormonal regulation experiences with GnRH agonists on rat ovaries demonstrated an increase of follicular atresia. Our previous experimental results showed increment of apoptosis on BGC-1established bovine granulosa cell line induced for 24 and 48 hours with GnRH agonist (leuprolide acetate100 nM). This induction was partially inhibited by the preincubation of 100 nM Antide, a GnRH antagonist. In the present paper we aim to describe the mechanisms of intracellular pathway regulation of apoptosis involved in BGC-1. Besides, we purpose and discussed a dependence on differentiation stage in the response of bovine granulosa cells treated with different doses of this analogue. Intrinsic apoptosis activation pathway was explored determining BAX and BCL-2 expression by western blot and extrinsic protection pathway, through phospholipase D activity assay. The data obtained by caspase 3 activity substantiated previous observations for agonist behaviour on BGC-1 but intriguingly showed Antide per se activity. Western blot for BAX and BCL-2 showed an increase on BAX over BCL-2 protein expression with 100 nM agonist but Antide did not, showing antagonism. All treatments showed inhibition of PLD activity compared to control, implying agonist behaviour of Antide. We conclude that leuprolide acetate induced apoptosis on BGC-1 by activation of mitochondrial pathway and by inhibition of PLD activity. We purpose Antide might work both, as an antagonist of intrinsic pathway and as an agonist of extrinsic protection pathway by inhibiting PLD activity.