Follicle Stimulating Hormone Receptor In Regulation Of Adipocyte Differentiation And Ovarian Response

Chapter IFollicle Stimulating Hormone Receptor and Adipocyte Differentiation and MetabolismIntroductionThe glycoprotein hormones are usually regarded as pivotal pituitary-endocrine signals. High expression of selective receptors in the pituitary-endocrine axis is key to mammalian endocrine regulation. The occurrence of glycoprotein hormone receptors in non-endocrine organs is an emerging but controversial area of investigation.The peripheral expression of glycoprotein hormone receptors is relatively limited and might be restricted to specific developmental stages. Glycoprotein hormone receptors might regulate differentiation and survival under circumstances with abnormally low or high levels of circulating hormones.FSH is the central glycoprotein hormone receptor of mammalian reproduction, necessary for gonadal development and maturation at puberty and for gamete production during the fertile phase of life. FSH binds to its specific membrane rceptor FSHR, which is located exclusively in granulosa cells. FSH-FSHR interaction elicits intracellular signaling pathways to determint proliferation and differentiation of granulosa cells. In zebrafish, FSHR is expressed in the kidney and liver, in addition to the gonads. In human, FSHR has been found localized to the surface of human osteoblasts. FSH stimulation regulates bone mass by directly increasing osteoclastogensis and resorptin.Obesity is a disorder that results from excess adipose tissue and is a major for risk diabetes and cardiovascular diseases. Sex steroid hormones are involved in the metabolism, accumulation and distribution of adipose tissues. It is now known that estrogen receptor, progesterone receptor, androgen receptor and hCG/LH receptor exist in adipose tissues, so their actions could be direct. The mechanism for the regulation of the amount and distribution of adipose tissues by sex steroid hormones is not clear. One possible mechanism would be the regulation of key proteins in adipose tissues at the genomic level by transcriptional means.FSH is considered a stimulus for estrogen production from ovarian follicles. As FSH secretion is negatively regulated by estrogen feedback, a high FSH level accompanies hypogonadism. FSH has thus been regarded as a marker for the onset of menopause. However, the effect of FSH on adiposity has never been explored.Our study provides the first evidence that FSHR is characterized in human adipocytes. We show that FSH stimulated the differentiation and function of preadipocyte in vitro and in vivo. In 3T3-L1 cells, FSH accelerated adipogenesis in a dose-dependent manner under the control of PPAR-γ. In addition, FSH-treated mice exhibited increased visceral fat mass associated with triglyceride and cholesterol. FSHR might regulate differentiation and metabolism under circumstances with abnormally high FSH in menopause. We anticipate that therapeutic inhibition of adipose FSHR activity in post-menopausal women will reduce lipogenesis and counteract the accumulation of visceral fat and its related metabolic abnormalities. Part IFollicle Stimulating Hormone Receptor in Adipose TissueObjectives: To examine the possible presence of follicle stimulating hormone receptor (FSHR) in human adipose tissue.Materials and Methods: Adipose tissue was obtained from various anatomical regions of man and women at different ages. It was assessed for the presence of FSHR by western blot, RT-PCR and immunohistochemistry.Results: It was clearly demonstrated that the presence of FSHR in human subcutaneous and omental adipose tissue. FSHR was primarily located on the membrane of adipocytes. The expression of FSHR was significantly higher in adipocytes versus preadipocytes. There was no difference of FSHR expression in subcutaneous and omental adipose tissue from premenopausal, perimenopausal and postmenopausal men and women.Conclusions: For the first time, FSHR was characterized in human mature adipocytes. FSHR was found to be localized on the membrane of adipocyte. FSHR might be invariance of expression in adipose tissue. Part IIThe role of follicle stimulating hormone on proliferarion and differentiation of 3T3-L1 preadipocyteObjectives: To investigate the relationship between follicle stimulating hormone (FSH) and preadipocyte proliferarion and differentiation.Materials and Methods: The mouse 3T3-L1 preadipocytes were cultured in the presence of FSH (3-300ng/mL). And FSH (3-300ng/mL) was added during the differentiation of 3T3-L1 in response to dexamethasone, isobutyl methyl xanthine and insulin. The rate of cell proliferation was measured over time (24-48h) by MTT metablism. Adipogenesis was judged by Oil Red 0 staining. The gene expression of peroxisome proliferator-activated receptor gamma (PPARγ) was analyzed by real-time RT-PCR.Results: The amount of mature adipocytes and PPARγmRNA were gradually cumulative during 3T3-L1 differentiation. Treatment of differentiating 3T3-L1 preadipocytes with FSH resulted in a dose-dependent increase in number of mature adipocytes and PPARγexpression. FSH did not alter the proliferation of 3T3-L1.Conclusions:1. In 3T3-L1 cells, FSH accelerates adipogenesis in a dose-dependent manner.2. FSH acts as a potent positive regulator of PPARγ, a master regulator of adipocyte differentiation, and tightly controls adipogenesis.3. FSH has no effect on preadipocyte proliferarion. Part IIIFollicle stimulating hormone regulation of adiposity in ovariectomized miceObjectives: To elucidate metabolic and molecular mechanisms by which follicle stimulating hormone (FSH) regulates fat storage and fat mobilization.Materials and Methods: C57 BL/6 mice were ovariectomized, randomized to FSH (OVX+FSH) and control saline administration (OVX), and pairfed for 14 days. To compare adipose mass, serum cholesterol and triglyceride content, adipose gene expression and serum leptin and adiponectin levels between OVX, OVX+FSH and sham groups.Results: FSH injection increased adipose weight, adipocyte size and serum cholesterol and triglyceride. FSH upregulated the expression of lipogenic genes in adipocyte as ACC, FAS; downregulated of HSL. FSH treatment was also associated with increasing expression of sterol-regulatory element-binding protein 1 and peroxisome proliferator-activated receptor gamma. FSH+OVX mice were with increased leptin and decreased adiponectin level.Conclusions:1. FSH directly stimulates adipocyte differentiation and metabolism.2. FSH promotes adiposity by upregulating targets adipose genes under the control of PPAR-γand SREBBP1.3. FSH regulates secretion of adipokines. Chapter IIFollicle Stimulating Hormone Receptor and Ovarian ResponseIntroductionApproximately 9 to 24 percent of women undergoing IVF respond poorly to standard ovarian stimulation protocol. Such patients, referred to as poor responders, may result in low levels of serum E₂, fewer mature oocytes and reduced pregnancy rates. Although a variety of strategies have been used to improve ovarian response, the optical approach for poor responders still remains controversial. Up to date, poor ovarian response is one of the most difficult and challenging problems in the field of IVF.Advanced maternal age and diminished ovarian reserve may be associated with poor ovarian response. However, some young women with normal ovarian reserve as indexed by basal serum FSH level and antral follicle count do present poor response to ovarian stimulation. Several hypotheses have been proposed for poor ovarian response alternatively. Poor follicular blood flow, dysfunctions of cytokines and growth factor network, and the presence of ovarian autoantibody have been proposed, but none has been proven.Recently, the description of cases of mutation of the FSH receptor (FSHR) has highlighted the importance of FSHR in determining the ovarian response. Women with inactivating mutations of FSHR were reported to constitute a defect in ovulation failure and amenorrhoea. In-vitro studies had demonstrated that these novel specific mutations of FSHR dramatically reduced receptor expression and impaired proper signal transduction. In a study of transfection of adenoviral vector to immature granulosa cells, it was found that differential effects of FSH in granulosa cells were highly dependent on FSHR density and FSHR-induced signaling which depended on the dosage of transfected vector.In the present study, it is revealed that the availability of FSHR in granulosa cells is associated with the individual response to ovarian stimulation. To the best of our knowledge, this is the first report describing the different levels of FSHR in granulosa cells of young women with different response to ovarian stimulation whose basal serum FSH levels and antral follicle counts were normal. We found lower FSHR expression in poor responders, while higher FSHR expression in high responders, as compared with moderate responders. Increasing dose of rFSH could not improve ovarian response. These results suggest that the variability of FSHR necessarily may determine ovarian response to gonadotropin stimulation. Objectives: To explore the importance of follicle stimulating hormone receptor (FSHR) in granulosa cells in the ovarian response to gonadotropin stimulation.Materials and Methods: 60 women undergoing ovarian stimulation were divided into three groups: poor, moderate, and high responders, according to the number of follicles with diameter >= 14 mm. FSHR expression at protein levels were determined by Western blot in granulosa cells. Estradiol (E2) concentrations in serum/follicle fluid were measured by electrochemiluminescene immunoassay.Results: The expression of FSHR was significantly different among three groups with the lowest expression in the poor responders (0.22±0.02), moderate (0.32±0.07), and high responders (0.46±0.03). The level of FSHR protein was positively correlated with peak level of serum E₂ and the number of mature oocytes. FSH levels in follicular fluid and the dosage of rFSH used were significantly different among three groups with highest values in the poor responders.Conclusions:1. Different levels of FSHR expression in granulosa cells result in different ovarian response with the lowest expression in the poor responders and highest in high responders.2. Lower expression of FSHR may account for poor ovarian response to gonadotropin stimulation, suggesting the critical role of FSHR in the ovarian response to gonadotropin stimulation.3. Increasing dose of rFSH could not improve ovarian poor response.