I. Introduction IN THE adult ovary, folliculogenesis starts when follicles leave the pool of resting follicles (RF) to enter the growth phase. From there, the early growing follicle undergoes a developmental process including a dramatic course of cellular proliferation and differentiation. In primates, only one follicle commonly reaches the preovulatory stage every cycle; most follicles fail to complete this maturation scheme, dying in the process termed atresia. In recent years, a picture has emerged depicting the classic endocrine control of ovarian function by LH and FSH, entangled in a maze of regulatory systems hinging on cell-cell interactions between follicular cells, via action of a variety of molecules (1–3). Different types of cell-cell interactions have been described. In paracrine regulations, a molecule synthesized by one cellular type is released into the interstitial milieu to act directly on another cellular type. In autocrine regulations, molecules synthesized by one cellular type are rel...

/pdf/regulation-of-ovarian-follicular-development-in-primates-4srrm0fi7f.pdf

Regulation of ovarian follicular development in primates: facts and hypotheses.

Paracrine factors secreted by oocytes and somatic cells regulate many important aspects of early ovarian follicle development in mammals. From activation of dormant primordial follicles to selection of secondary follicles, locally acting factors have been identified that appear to exert important effects on the growth and differentiation of oocytes and granulosa cells. This article summarizes evidence to support a model for bi-directional paracrine communication that is based on developmental regulation of the delivery and reception of paracrine factors at the oocyte-granulosa cell interface. Transzonal projections that originate from granulosa cells and terminate at the oocyte plasma membrane provide a polarized means to orient the secretory organelles of somatic cells. Characterization of transzonal projections in follicles from normal and genetically modified mice reveals dynamic changes in the density and stability of transzonal projections. On the basis of new data analysing the orientation and cytoskeletal content of transzonal projections in mammalian oocytes, a model is proposed for regulation of paracrine growth factor secretion by follicle-stimulating hormone. These findings have immediate implications for ovarian hyperstimulation protocols and follicle culture models as related to the production of mammalian embryos by assisted reproductive technologies.

/pdf/cellular-basis-for-paracrine-regulation-of-ovarian-follicle-21l4gd8u2f.pdf

Cellular basis for paracrine regulation of ovarian follicle development

The assembly of the primordial follicles early in ovarian development and the subsequent development and transition of the primordial follicle to the primary follicle are critical processes in ovarian biology. These processes directly affect the number of oocytes available to a female throughout her reproductive life. Once the pool of primordial follicles is depleted a series of physiological changes known as menopause occur. The inappropriate coordination of these processes contributes to ovarian pathologies such as premature ovarian failure (POF) and infertility. Primordial follicle assembly and development are coordinated by locally produced paracrine and autocrine growth factors. Endocrine factors such as progesterone have also been identified that influence follicular assembly. Locally produced factors that promote the primordial to primary follicle transition include growth factors such as kit ligand (KL), leukaemia inhibitory factor (LIF), bone morphogenic proteins (BMP's), keratinocyte growth factor (KGF) and basic fibroblast growth factor (bFGF). Factors mediating both precursor theca-granulosa cell interactions and granulosa-oocyte interactions have been identified. A factor produced by preantral and antral follicles, Mullerian inhibitory substance, can act to inhibit the primordial to primary follicle transition. Observations suggest that a complex network of cell-cell interactions is required to control the primordial to primary follicle transition. Elucidation of the molecular and cellular control of primordial follicle assembly and the primordial to primary follicle transition provides therapeutic targets to regulate ovarian function and treat ovarian disease.

/pdf/regulation-of-primordial-follicle-assembly-and-development-40bemy3bu9.pdf

Regulation of primordial follicle assembly and development

Theca cells function in a diverse range of necessary roles during folliculogenesis; to synthesize androgens, provide crosstalk with granulosa cells and oocytes during development, and provide structural support of the growing follicle as it progresses through the developmental stages to produce a mature and fertilizable oocyte. Thecal cells are thought to be recruited from surrounding stromal tissue by factors secreted from an activated primary follicle. The precise origin and identity of these recruiting factors are currently not clear, but it appears that thecal recruitment and/or differentiation involves not just one signal, but a complex and tightly controlled combination of multiple factors. It is clear that thecal cells are fundamental for follicular growth, providing all the androgens required by the developing follicle(s) for conversion into estrogens by the granulosa cells. Their function is enabled through the establishment of a vascular system providing communication with the pituitary axis throughout the reproductive cycle, and delivering essential nutrients to these highly active cells. During development, the majority of follicles undergo atresia, and the theca cells are often the final follicular cell type to die. For those follicles that do ovulate, the theca cells then undergo hormone-dependent differentiation into luteinized thecal cells of the corpus luteum. While the theca is an essential component of follicle development and ovulation, we do not yet fully understand the control of recruitment and function of theca cells, an important consideration since their function appears to be altered in certain causes of infertility.

/pdf/theca-the-forgotten-cell-of-the-ovarian-follicle-557ura2rt1.pdf

Theca: the forgotten cell of the ovarian follicle.

https://www.fertstert.org/article/S0015-0282(06)00098-7/pdf

Obesity and androgens: facts and perspectives

Anchorage-Independent Culture of Bovine Granulosa Cells: The Effects of Basic Fibroblast Growth Factor and Dibutyryl cAMP on Cell Division and Differentiation

Granulosa cell death is an early feature of atresia; however, there are many apparent contradictions in the literature concerning the mode of granulosa cell death. We have therefore examined this process in bovine healthy and atretic antral follicles, using a variety of established techniques. Light and electron microscopic observations indicated the presence of pyknotic or shrunken nuclei in both the membrana granulosa and the antrum. In the membrana granulosa, these nuclei were frequently crescent shaped and uniformly electron dense and were approximately the same size as healthy nuclei, all of which are typical of early apoptosis. However, these nuclei were within the membranes of a healthy granulosa cell, suggesting that phagocytosis by a neighboring granulosa cell is an unusually early event in the apoptotic pathway of granulosa cells. In the membrana granulosa, pyknotic nuclei stained intensely with hematoxylin but weakly with the DNA-intercalating stain propidium iodide. A percentage of these pyknotic nuclei stained by TUNEL (terminal deoxyUTP nick end-labeling). However, in the antrum, the pyknotic nuclei and larger globules of DNA stained intensely with both hematoxylin and propidium iodide, but were not TUNEL positive. The comet assay of cell death produced a streak tail of randomly nicked DNA, rather than the plume of low mol wt apoptotic DNA. Globules collected from fresh follicular fluid stained intensely with propidium iodide and were shown by PAGE to contain DNA, the majority of which was high mol wt. In conclusion, granulosa cells within the membrana granulosa die by apoptosis, with phagocytosis by a neighboring cell preceding any potential budding of the nucleus or cell itself. Granulosa cells near the antrum are sloughed off into the antrum, and their death has features more consistent with that of other cell types that undergo death as a result of terminal differentiation. (Endocrinology 140: 2602‐2612, 1999)

Evidence for alternative pathways of granulosa cell death in healthy and slightly atretic bovine antral follicles.

Dynamics of the membrana granulosa during expansion of the ovarian follicular antrum

Development of the ovarian follicular epithelium

We have previously postulated that granulosa cells of developing follicles arise from a population of stem cells. Stem cells and cancer cells can divide indefinitely partly because they express telomerase. Telomerase is a ribonucleoprotein enzyme that repairs the ends of telomeres that otherwise shorten progressively upon each successive cell division. In this study we carried out cell cycle analyses and examined telomerase expression to examine our hypothesis. Preantral (60-100 microm) and small (1 mm) follicles, as well as granulosa cells from medium-sized (3 mm) and large (6-8 mm) follicles, were isolated. Cell cycle analyses and expression of Ki-67, a cell cycle-related protein, were undertaken on follicles of each size (n = 3) by flow cytometry; 12% to 16% of granulosa cells in all follicles were in the S phase, and less than 2% were in the G(2)/M phase. Telomerase activity (n = 3) was highest in the small preantral follicles, declining at the 1-mm stage and even further at the 3-mm stage. In situ hybridization histochemistry was carried out on bovine ovaries, and telomerase RNA was detected in the granulosa cells of growing follicles but not primordial follicles. Two major patterns of staining were observed in the membrana granulosa of antral follicles: staining in the middle and antral layers, and staining in the middle and basal layers. No staining was detected in oocytes. Our results strongly support our hypothesis that granulosa cells arise from a population of stem cells.

/pdf/evidence-for-ovarian-granulosa-stem-cells-telomerase-sag8n4aqgn.pdf

Evidence for Ovarian Granulosa Stem Cells: Telomerase Activity and Localization of the Telomerase Ribonucleic Acid Component in Bovine Ovarian Follicles

T. C. Lavranos

Papers

Anchorage-Independent Culture of Bovine Granulosa Cells: The Effects of Basic Fibroblast Growth Factor and Dibutyryl cAMP on Cell Division and Differentiation

Evidence for alternative pathways of granulosa cell death in healthy and slightly atretic bovine antral follicles.

Dynamics of the membrana granulosa during expansion of the ovarian follicular antrum

Development of the ovarian follicular epithelium

Evidence for Ovarian Granulosa Stem Cells: Telomerase Activity and Localization of the Telomerase Ribonucleic Acid Component in Bovine Ovarian Follicles