Effects of dbcAMP on progesterone synthesis by cultured goat luteal cell subpopulations isolated from early and late luteal stage corpora lutea

S Arikan,H Kalender,O Simsek

doi:10.21451/1984-3143-ar774

Abstract

This research aimed to investigate the effects of dbcAMP on steroid accumulation by culturing two distinct luteal cell subpopulations isolated from early and late luteal stage corpora lutea. Cells were isolated from corpora lutea collected from eight Angora goats on either the 5th or 15th days of their estrous cycles. Cell isolation was performed by enzymatic digestion using collagenase and DNase. Isolated cells were separated into two distinct subpopulations enriched with small and large luteal cells by percoll density-gradient centrifugation. Isolated cells were stained in order to detect 3β-hydroxysteroid dehydrogenase (3β-HSD). Cells stained positively for 3β-HSD activity (5 x 104 cell⁄well) were incubated with dbcAMP in the absence or presence of 22(R)-hydroxycholesterol (22R-HC) for periods of up to 7 days. Large luteal cell enriched subpopulations produced more basal progesterone (P < 0.05) than did the small luteal cell enriched subpopulations. Treatment of cells with 22R-HC alone induced 4.00 to 11.60 times increase in steroid synthesis depending on type of cells incubated, luteal age and days of incubation. Incubation of cells with 1 mM dbcAMP in the absence or presence of 22R-HC induced in a significant increase (P < 0.01) in steroid accumulation in all treated groups. In contrast, when cells are treated with low dose dbcAMP (0.1 mM), treatment induced stimulation failed to reach significant level in most treated groups. In conclusion, although treatment of goat luteal cells with dbcAMP induces an increase in steroid accumulation, a high dose is necessary to reach significant levels. Stimulatory effect of dbcAMP on steroidogenesis maintains during long life culturing.

Highlights

The corpus luteum is known to be a temporary endocrine gland that secretes progesterone during the luteal stage of the estrous cycle (Band et al, 1987; Arikan et al, 2010) and pregnancy (Arikan and Yigit, 2003) in goats
Since response of luteal cells to the exogenous treatments might be affected by age and type of luteal cells, we used small and large luteal enriched subpopulations from early and late luteal corpora lutea as cell sources in this study
Progesterone production was stable in the cholesterol treated group during the 7 days of culture period

Summary

Introduction

The corpus luteum is known to be a temporary endocrine gland that secretes progesterone during the luteal stage of the estrous cycle (Band et al, 1987; Arikan et al, 2010) and pregnancy (Arikan and Yigit, 2003) in goats. The cells having steroidogenic activity can be determined by testing 3βHSD activity. This is due to the fact that the conversion of pregnenolone to progesterone requires 3β-HSD located largely in the endoplasmic reticulum of the luteal cells (Bao et al, 1995)

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