APOPTOSIS MODULATION ON PORCINE GRANULOSA CELLS BY A GnRH ANALOG

FIORITO, C; CAROU C; REVILLA, M; LOMBARDO, DM

INTERNATIONAL JOURNAL OF MORPHOLOGY

universidad nacional de la frontera, temuco, chile

Año: 2008 vol. 26 p. 726 - 727

0717-9367

Granulosa cells (GC) contribute on ovocit maturationand most of the growth factors that it needs act through them.This makes that GC viability and integrity to be an importantfactor throughout follicle genesis. It?s known that GC deathduring follicular atresia and luteolisys is apoptotic, although,the regulation process is not yet well understood.Gonadotropin releasing hormone receptor (GnRHr) wasfound on GC and luteal cells of various species. There wasobserved a decrease on GCs proliferate activity on porcineimmature follicles and an increase on apoptosis levelsindependently on follicle genesisstage. Our objective is todetermine if a GnRHa (Leuprolide Acetate) is an apoptoticmodulator on GC from adult porcine?s ovaries and also precisethe best experimental conditions to observe LAmodulation on this model. This study was carried on a primaryGC porcine culture (CPGP) model obtained from punction of5-8 nm follicles. CPGPs were grown in DMEM + F12supplemented with SFB 5% and antibiotics. We assay threeLA concentrations (1, 10 and 100 nM) and two incubationtimes (12 and 24 Hs). Three experiments by triplicate werepurchased and were measured for apoptosis with DAPI,TUNEL and by a citometric analysis (FACS) using Anexina Vand PI label techniques. Results were statistical analyzedwith a two way ANOVA and Bonferroni comparison of means.TUNEL and DAPI techniques show that 100 nM LA for 24 Hstreatment an 27 % augment of apoptosis over control (p =0.058). This effect was corroborated by FACS with AnexinaV and PI label, being, the differences, significant for LA 1 nMfor 24 Hs treatment, with an apoptosis increment of 74 %over control (p = 0.05). FACS analysis shows a characteristicapoptotic temporal dynamic on CPGPs exposed to LA forthis model on this experimental conditions.