Characterization of endothelin-1 and nitric oxide generating systems in corpus luteum-derived endothelial cells

Abstract

Endothelium-derived endothelin-1 (ET-1) and nitric oxide (NO) are pivotal regulators of corpus luteum (CL) function. To have a better insight into their synthesis and action, members of the ET system (ET-1, ET converting enzyme (ECE-1) isoforms a–d, ETAand ETBreceptors) along with NO synthase (NOS) isoforms – endothelial (e)NOS and inducible (i)NOS – were quantified in CL-derived endothelial cells (CLEC). The expression of these genes in microvascular CLEC, obtained by lectin-coated magnetic beads, was compared with cells removed from the luteal microenvironment and maintained in culture for different durations, and with endothelial cells (EC) derived from a large blood vessel (i.e. bovine aortic endothelial cells, BAEC). The profile of gene expression in the different EC types was determined by quantitative real-time PCR. Freshly isolated EC from mid-cycle CL exhibited high ET-1 receptor expression (both ETAand ETB), low ET-1 synthesizing ability (both prepro (pp) ET-1 and ECE-1), but elevated iNOS – the high throughput NOS isoform. The distinct phenotype of CLEC was lost soon after an overnight culture. ETAand ETBreceptor levels declined, ppET-1 levels increased while iNOS was reduced. These changes were extenuated during long-term culture of CLEC. The general pattern of gene expression in BAEC and long-term cultured CLEC was similar yet some differences, reminiscent of freshly isolated CLEC, remained: ECE-1c, ETBreceptor and NOS isoforms were expressed differently in BAEC as compared with lines of CLEC.This study suggests that the luteal microenvironment is necessary to sustain the selective phenotype of its resident endothelial cells. The inverse relationship between ppET-1 and iNOS observed in freshly isolated CLEC and in cultured cells is physiologically significant and suggests that ET-1 and NO may modulate the production of each other.