Establishment and validation of a model for non-luteinized human mural granulosa cell culture

•Cell culture techniques of human mural granulosa cells (MGCs) serve as a major in vitro tool.•We establish a standardized protocol for the culture of MGCs as a model for different stages of folliculogenesis.•Four days incubation of luteinized MGCs achieve genes expression levels similar to those of early non-luteinized follicles.•FSH stimulation for 48 h showed ovulatory genes expression similar to the pre-ovulatory non-luteinized follicles.•Stimulated cells responded to hCG stimulation with pattern similar to the response of pre-ovulatory follicles.

Cell culture techniques of human mural granulosa cells (MGCs) serve as a major in vitro tool. However, the use of luteinized MGCs has major limitations due to their luteinized state. Our aim was to establish a standardized protocol for the culture of MGCs as a model for different stages of folliculogenesis. We showed that early-non-luteinized, preovulatory-non-luteinized and luteal-MGCs have distinct gene expression pattern. After 4 days of incubation of luteinized-MGCs, ovulatory genes mRNA’s achieve expression levels similar to the early non-luteinized follicles. FSH stimulation for 48 h of these 4 days cultured MGCs showed ovulatory genes mRNA’s expression similar to the pre-ovulatory non-luteinized follicles. These FSH-stimulated cells responded to hCG stimulation in a pattern similar to the response of pre-ovulatory follicles.This novel model may provide a standardized research tool for delineation of the molecular processes occurring during the latter stages of follicular development in the human ovary.