Local effects of the sphingosine 1-phosphate on PGF2alpha-induced

MARINA C. PELUFFO; SILVIA F. HERNANDEZ; GRISELDA IRUSTA; RICHARD L. STOUFFER; MARTA TESONE

kailua-kona, Big Island,Hawaii

Congreso; 41st Annual Meeting of The Society for the Study of Reproduction; 2008

Since apoptosis is associated with regression of the corpus luteum (CL) inmany species, studies were designed to determine if local administrationof the antiapoptotic agent sphingosine-1-phosphate (S1P) effectivelyblocks the luteolytic action of PGF-2alpha. On day 19 of pregnancy, 2 hbefore systemic PGF-2alpha administration, rats were injected with eitherS1P or vehicle (control) into the bursa of both ovaries. The activity offour caspases, which contribute to the initial (caspase-2, -8 and -9) andfinal (caspase-3) events in apoptosis, was measured in pooled CL from 4individual ovaries at 0 h (pretreatment) and 4 h after PGF-2alphainjection in both control and S1P groups. The expression of thephosphorylated form of AKT was analyzed in CL using an ELISA. In addition,cell death was evaluated by electronic microscopy (EM) in CL 36 h afterPGF-2alpha injection. The activity of caspase-2, -3 and -8 wassignificantly greater by 4h after PGF-2alpha administration (0h vs 4h:8491±1406 vs 36702±3155; 45049±5679 vs 155053±7391; 1895±283 vs 9888±585RFU respectively, p<0.05), but not caspase-9 activity. In contrast,expression of the phosphorylated form of AKT decreased (0h vs 4h: 22.5±0.5vs 19.5±0.2 U/ml; p<0.05). Administration with S1P suppressed theseeffects, decreasing caspase activities and increasing the phosphorylatedform of AKT (p<0.05). EM studies revealed ultrastructural indices ofapoptotic andSince apoptosis is associated with regression of the corpus luteum (CL) inmany species, studies were designed to determine if local administrationof the antiapoptotic agent sphingosine-1-phosphate (S1P) effectivelyblocks the luteolytic action of PGF-2alpha. On day 19 of pregnancy, 2 hbefore systemic PGF-2alpha administration, rats were injected with eitherS1P or vehicle (control) into the bursa of both ovaries. The activity offour caspases, which contribute to the initial (caspase-2, -8 and -9) andfinal (caspase-3) events in apoptosis, was measured in pooled CL from 4individual ovaries at 0 h (pretreatment) and 4 h after PGF-2alphainjection in both control and S1P groups. The expression of thephosphorylated form of AKT was analyzed in CL using an ELISA. In addition,cell death was evaluated by electronic microscopy (EM) in CL 36 h afterPGF-2alpha injection. The activity of caspase-2, -3 and -8 wassignificantly greater by 4h after PGF-2alpha administration (0h vs 4h:8491±1406 vs 36702±3155; 45049±5679 vs 155053±7391; 1895±283 vs 9888±585RFU respectively, p<0.05), but not caspase-9 activity. In contrast,expression of the phosphorylated form of AKT decreased (0h vs 4h: 22.5±0.5vs 19.5±0.2 U/ml; p<0.05). Administration with S1P suppressed theseeffects, decreasing caspase activities and increasing the phosphorylatedform of AKT (p<0.05). EM studies revealed ultrastructural indices ofapoptotic and non-apoptotic cell death in the CL at day 19 of pregnancy.PGF-2alpha treatment increased luteal cells with advanced signs ofapoptosis (i.e. containing multiple nuclear fragments, chromatincondensation or apoptotic bodies) by 36h post-injection. S1P treatmentsuppressed these changes and increased the blood vessel density. Theseresults suggest that S1P can block the luteolytic effect of the PGF-2alphaby decreasing caspase-2, -3 and -8 activities and increasing thephosphorylation of AKT. Supported by: ANPCYT (BID 1201 OC-AR PICT99:05-06384), NIH-FIRCA RO3-TW007041 and NIH-NCCR RR00163. non-apoptotic cell death in the CL at day 19 of pregnancy.PGF-2alpha treatment increased luteal cells with advanced signs ofapoptosis (i.e. containing multiple nuclear fragments, chromatincondensation or apoptotic bodies) by 36h post-injection. S1P treatmentsuppressed these changes and increased the blood vessel density. Theseresults suggest that S1P can block the luteolytic effect of the PGF-2alphaby decreasing caspase-2, -3 and -8 activities and increasing thephosphorylation of AKT. Supported by: ANPCYT (BID 1201 OC-AR PICT99:05-06384), NIH-FIRCA RO3-TW007041 and NIH-NCCR RR00163.