The in vitro culture of ovarian follicles or cumulus-oocyte complexes (COC) is used to study the factors that regulate follicular development and may have potential use in artificial reproductive technology (ART). Before ovulation, the follicle is formed by oocyte and cell populations known as granulosa cells (GCs). These cells build the internal and external mass of the follicular wall. Oocyte growth and proliferation of the surrounding cells depend on the gap junctions between the oocyte and the GCs. Maintenance of the optimal in vitro culture system allowing for preservation of follicle architecture and granulosa-oocyte interaction may be critical for success in vitro maturation of follicles. Recently many studies have focused on a culture of GCs, which have important functions related to steroidogenesis. Granulosa cells maintained in in vitro conditions exhibit stem cell properties making it important to consider in vitro culture (IVC) methods of the GC population.

The ovary is commonly known as an endocrine gland responsible for sex steroid production. One of the outstanding cells in ovarian microenvironment - granulosa cells (GCs) are responsible for converting the androgens to estrogens during follicular growth and secreting progesterone after ovulation. These secretory processes within the ovary are directly involved in hormonal signaling pathways, and they depend on different stages of cholesterol and lipid biosynthesis during the ovarian cycle. The understating of the regulation and further investigation into the processes taking part in ovary will expose new clinical advantages in detection and treatment of female reproductive system diseases associated with sex hormone abnormalities. The expression of genes belonging to ontology groups associated with steroid biosynthesis and metabolism, such as “cholesterol biosynthetic process” (GO:0006695, “regulation of lipid biosynthetic process” (GO:0046890), “regulation of lipid metabolic process” (GO:0019216), “response to insulin” (GO:0032868) and “response to lipopolysaccharide” (GO:0032496) were analyzed by using the microarray approach. The patterns of gene expression in human GCs at days 1-day, 7-day, 15-day, and 30-day of primary in vitro culture have been analyzed. Based on the microarray results, a group of upregulated genes have been selected: CCL20, CXCL5, STAR, MSMO1, and AADAC. The genes STAT5B, OPA3, PPARG, PROX1, and SEC14L2 were decreased across all the experimental groups during the 30-day cell cultivation period. These results suggest that, the GCs in cell culture under in vitro express steroidogenic markers and it is important to understand associations with lipid and liposaccharide synthesis relative to reproductive medicine.

Several hypotheses have been proposed, relating to the potential genesis of follicular cells in the ovarian niche. Reports using mice as an experimental model have suggested that the ovaries may contain stem cells that are likely involved in the formation of new follicles in adult reproductive life. Over recent years, various types of ovarian cells have been identified and described to confirm or disprove the existence of ovarian adult stem cells. Most research is focused on granulosa cells (GCs), which are essential for follicular development and maturation of female germ cells (oocytes). GCs exhibit the features of stem cells, such as expression of stem cell markers: OCT-4, Sox-2, Nanog as well as certain markers of mesenchymal stem cells, including CD29, CD44, CD90, CD105, CD117, and CD166. Another discovery in favor of the potential stemness of GCs is their ability to transdifferentiate towards other cell lines and high telomerase (TERT) activity in dividing compartments of the follicle during its maturation.