Three-dimensional image analysis to quantify the temporo-spacial expression of cellular receptors

Abstract

Ovarian folliculo genesis is primarily controlled by the action of gonadotropins namely follicle stimulating hormone (FSH) and luteinizing hormone (L H). Several reports indicated that the process of initial recruitment of primordial follicles to the growing follicles is not gonadotropin-dependent but Bone morphogenetic protein (BMP)-dependent. However, this has not been unequivocally confirmed. The aim of this study was to investigate the temporo-spacial protein expression of the BMP receptors 1B (BMPR1b), FSHR and L HR in several stages of follicle development. While the localization of all receptors was found in granulosa cell membrane of the follicles the temporal expression was varied. BMPR1b was expressed in all follicle stages, FSHR was detected in primary follicles onward and L HR was absent in both primordial and primary follicles but appeared in later stages. Quantitative analysis based on the intensity of fluorescent signals showed that the expression of BMPR1b, FSHR and L HR significantly (p< 0.001 p< 0.0001 p< 0.0001 respectively) increased with follicular development. We have concluded that the combination of sensitive immunofluorescence labeling and computerized 3D image analysis proves efficient tools for in situ detection and quantification of the expression of small amount of protein in a complex tissue structure.