A Regulatory Mechanism Between Progesterone and Notch Signaling Pathway Maintains the Functionality of the Corpus Luteum from Pregnant Rats.

Abstract

The Notch signaling pathway includes a family of transmembrane receptors (Notch1-4) that interact with specific ligands (Delta-like family, Jagged1 and Jagged2). This system regulates cell fate decisions, including proliferation, differentiation and apoptosis. The transmembrane receptors are cleaved upon binding of their ligands, leading to the release of the intracellular domain (NICD), which translocates to the nucleus where it functions as a transcriptional coactivator of regulatory genes of cellular fate. This processing requires the activity of two proteases, namely tumour necrosis factor a-converting enzyme and presenilin/gamma-secretase. We have previously shown that Notch1-4 receptors and the ligand Dll4 are expressed in the corpus luteum (CL) of pregnant rats and that the administration of prostaglandin F2 alpha decreases the levels of mRNA and protein expression of these proteins. Furthermore, blocking ovarian Notch system with the gamma-secretase inhibitor DAPT decreases serum levels of progesterone (P4) and increases pro-apoptotic protein expression in the CL. However, the role of P4 in regulating the Notch pathway in the CL of pregnant rats remains unclear. In this study CL′s were isolated from ovaries of rats on day 16 of gestation and cultured for 4 hours in the presence of aminoglutethimide (inhibitor of cytochrome P450scc 0.15 mM), DAPT (gamma secretase inhibitor, 20 uM), both inhibitors and the inhibitors with the addition of P4 (500ng/ml). Luteal function was assessed by measuring P4 levels in the medium by RIA. The CL levels of Notch1-4 and members of the ERK and PI3K/AKT pathways were measured by Western blot. The treatment of CL′s with each inhibitor significantly decreased P4 levels (P < 0.05, n = 5) in the culture medium, effect which was similar in the presence of both compounds. Furthermore, these inhibitors decreased protein levels of Notch1 intracellular fragment (NICD) compared to the control group (P < 0.05, n = 5). However, no significant difference was observed for NICD of Notch4. In addition, ERK and AKT phosphorylation decreased with the addition of aminoglutethimide or/and DAPT in the CL culture medium. These results suggest the existence of a regulatory mechanism between progesterone and the Notch signaling pathway to maintain the functionality of the CL in pregnancy. Supported by: CONICET (PIP1223), Universidad de Buenos Aires (X-218) and ANPCYT (PICT, 2010-0248).