This paper demonstrates the role of CD36 in the TSP-1 mediated effects in ovarian folliculogenesis. Based on the results of this study, CD36 appears to be an important regulator of ovarian angiogenesis, and follicular and luteal development. Disruption of expression of CD36 significantly alters ovarian morphology and expression of factors related to ovarian cell proliferation, survival, and angiogenesis. CD36 is expressed in granulosa and cumulus cells of rodents, bovines  and humans [27, 28], although the specific function of the receptor is not completely understood.
In vitro, CD36 TSP-1 receptors are known to co-localize with VEGF receptors on cell membranes, and evidence suggests that these receptors may directly associate with each other and regulate signaling activity. We showed that knockdown of CD36 in ovarian granulosa cells resulted in an increase in expression of phosphorylated VEGFR2. The three type I repeat region of TSP-1 has been shown to reduce VEGFR2 phosphorylation and inhibit VEGF signal transduction  and the association between CD36, β1 integrins, and TSP-1 is thought to be important in mediating this inhibition . In the CD36 null mice, there was a significant increase in ovarian blood vessel density compared to WT controls. In these mice, there was an increase in ovarian expression of VEGF and VEGFR2 and the in vitro data in this paper suggests that removal of the inhibitory influence of CD36 would allow for enhanced phosphorylation of VEGFR2, resulting in an increase in peri-follicular and luteal angiogenesis.
Knockdown of CD36 has been shown to increase proliferation and expression of survival and angiogenic in endothelial and tumor cells . In this study, CD36 knockdown in vitro resulted in increased granulosa proliferation and survival, which was associated with increased granulosa cell expression of phosphorylated Akt. Akt phosphorylation is an important promoter of granulosa cell proliferation and viability, which are the driving forces behind follicle growth and maturation [31, 32]. In vitro, the increased expression of phosphorylated Akt was associated with increased granulosa cell proliferation and decreased apoptosis. In vivo, CD36 null mice had increased granulosa cell proliferation and a greater number of growing follicles, compared to WT controls. Akt phosphorylation promotes follicle activation and survival  and inhibits follicle atresia [34, 35]. Increased activity of the PI3K/Akt pathway in the absence of CD36 could also have contributed to the increased number of follicles present in the ovaries of the CD36 null mice. Thrombospondin signaling has been linked to follicle atresia previously [15, 23, 24, 36] although the mechanisms have been unclear. This paper suggests that the PI3/Akt signaling pathway may be an important mediator of TSP-1’s effects in the ovary.
Normal function of the ovary is dependent on the tightly regulated angiogenic mechanisms that facilitate folliculogenesis, luteogenesis, and dissemination of the steroid hormones generated within the ovary. Angiogenesis within the ovary is a balance between and expression of pro- and anti-angiogenic factors. We have shown that members of the VEGF family and members of the anti-angiogenic TSP-1 are coordinately expressed during the ovarian cycle and have profound impacts on the angiogenic processes that occur throughout the cycle [15, 16]. Within the ovary, cytokine action causes a downregulation of CD36, which is necessary to facilitate the explosive angiogenesis that occurs during luteal formation . We showed in this study that CD36 null mice had significantly higher ovarian microvessel density compared to wildtype controls. CD36 appears to be critical in the regulation of physiological and pathophysiological angiogenesis. CD36 mediated TSP-1 signaling maintains corneal vascularity and CD36 deficiency leads to age-related corneal neovascularization while activation of CD36 can induce regression of inflammatory corneal angiogenesis . CD36 has been implicated in the regulation of tumor angiogenesis. TSP-1, and other proteins containing the thrombospondin type-1 repeats (TSR), are known to endogenously inhibit the angiogenesis that occurs to facilitate tumor growth. CD36 has been shown to be required for the anti-angiogenic activity of these proteins and is absence impairs their angiogenesis inhibition and tumor vascularity increases and tumor growth is accelerated [17, 39].
The morphological changes in the ovaries in the CD36 null mice somewhat replicate those seen in the condition of polycystic ovarian syndrome (PCOS) in which there is an increased number of primary follicles which remain preovulatory and do not progress to ovulation and formation of the corpus luteum . In our study, CD36 null mice had an increased number of preovualtory follicles, concomitant with fewer corpora lutei, compared to WT controls. Elevated blood vessel density, and increased VEGF signaling were seen in the ovaries of the CD36 null mice and these are hallmarks of the pathogenesis of PCOS [41, 42]. It has been shown that by increasing TSP-1 signaling, ovarian hypervascularization can be reversed as a method to treat PCOS . Interestingly, CD36 null mice had elevated serum FSH, with suppressed LH, compared to WT controls. In classic PCOS, the hyperandrogenism and elevated GnRH generally results in an increase in the LH/FSH ratio . In our mice, the elevated FSH may have been responsible for the increased number of recruited, but unovulated follicles as FSH is known to stimulate follicle development [45, 46] and protect against follicle atresia [47, 48]. FSH is a potent activator of the PI3K/akt signaling pathway  and the elevated circulating FSH seen in the CD36 null mice may have contributed to the increased akt phosphorylation observed in the ovaries from these mice. Reduced circulating LH in the CD36 null mice may have reduced the ovulatory stimulus, resulting in the increased number of pre-ovulatory follicles and fewer corpora lutei seen in these mice. CD36 has also been implicated in the pathogenesis of PCOS due to its role in regulating metabolism. CD36 is expressed in tissues regulated to fatty acid metabolism, including adipocytes . Women with PCOS often exhibit higher levels of visceral obesity  and elevated CD36 expression in adipose tissue is seen in women with PCOS , further implicating the receptor in the pathogenesis of this disease. Levels of soluble CD36 are elevated in PCOS patients, and they are associated with the altered insulin sensitivity seen in this disease . The metabolic profile of the CD36 null mice was not evaluated in this study, but will be the subject of investigation in the future. Transcriptome analysis has been performed on follicular cells from PCOS patients and healthy women. Haouzi et al.  showed that cumulus cells surrounding the oocyte had differential expression of a number of growth factors and genes involved in steroid metabolism. Granulosa cell expression of genes involved in inflammation, metabolism, and coagulation has also been implicated in the pathogenesis of PCOS [54, 55]. After review of the gene lists published in the various transcriptome papers, CD36 was not specifically mentioned, although the cellular process that it is involved in have been implicated. A closer evaluation and comparison of CD36 expression in samples from PCOS patients and healthy women may be warranted.