REPRODUCTIVE ENDOCRINOLOGY: CLINICAL P-272 Tuesday, October 31, 2017 FOLLICLE STIMULATINGHORMONE(FSH)REGULATES CYTOSKELETAL PROTEIN EXPRESSION AND CELL STRUCTURE DURING HUMAN GRANULOSA CELL E. Hobeika,b DIFFERENTIATION. S. Baumgarten,a N. Winston,c M. A. Fierro,b A. M. Zamah,d H. Scoccia,e C. Stocco.f aObstetrics and Gynecology, Mayo Clinic, Rochester, MN; bDepartment of Obstetrics and Gynecology, University of Illinois at Chicago, Chicago, IL; c University of Illinois at Chicago, Chicago, IL; dObstetrics and Gynecology, University of Illinois at Chicago, Chicago, IL; eObstetrics and Gynecology, University of Illinois, Chicago, IL; fUIC, Chicago, IL. OBJECTIVE: To understand the mechanisms by which follicle stimulating hormone (FSH) drives human granulosa cell differentiation during ovarian follicle development. DESIGN: Cumulus granulosa cells were collected from women undergoing IVF for male factor infertility at the University of Illinois at Chicago Fertility Center, and primary cultures of the cells were used for in vitro studies of FSH action during granulosa cell differentiation. MATERIALS AND METHODS: Cumulus cells were cultured in serumfree and phenol red-free media for 24 hours before treatment with FSH or no treatment (control) for 48 hours. Total RNA was isolated, and microarray analysis was conducted to assess global gene expression following the various treatment conditions in 3 different patients. Cells from additional patients were cultured and immunofluorescence was used to evaluate the expression, localization, and arrangement of cytoskeletal proteins in human granulosa cells undergoing differentiation. RESULTS: Microarray analysis revealed that FSH regulates the expression of over 850 genes in human granulosa cells, including genes encoding proteins that are crucial for cholesterol metabolism, steroid hormone synthesis, and proliferation. Interestingly, genes encoding proteins for all three cytoskeletal filament types—microfilaments, intermediate filaments, and microtubules—were highly down-regulated by treatment with FSH. Tubulins, which make up microtubules, keratin 18, a key component of intermediate filaments, and actins, which make up microfilaments, were significantly downregulated by 3-fold, 2.4-fold, and 3-fold, respectively, after FSH treatment. Immunofluorescence staining of granulosa cells revealed that the arrangement and morphology of actin filaments were significantly changed after treatment with FSH for 48 hours. In untreated cells, actin filaments were arranged in parallel uniformly throughout the cell; however, after treatment with FSH, the actin filaments were bundled and unevenly distributed throughout the cell. CONCLUSIONS: In human granulosa cells, FSH stimulates the expression of key genes for cholesterol metabolism and steroid hormone synthesis. At the same time, FSH significantly downregulates the expression of cytoskeletal genes and triggers a rearrangement of the cytoskeleton. The cytoskeletal changes elicited by FSH could allow for the changes in organelle size and number necessary to support the enhanced steroidogenic capacity of differentiated granulosa cells. Supported by: This project was funded by the NIH grants R56HD086054 and R01HD057110 (CS). SCB was funded by NIH training grant T32HL07692. P-273 Tuesday, October 31, 2017 SCREENING FOR PRE-DIABETES IN INFERTILE WOMEN: HOW PREDICTIVE IS HEMOGLOBIN A1C? A. Shapiro,a E. C. Holden,a,b P. G. McGovern,a,b D. Alderson,a S. Morelli.a,b aObstetrics, Gynecology and Women’s Health, Rutgers - New Jersey Medical School, Newark, NJ; bUniversity Reproductive Associates, Hasbrouck Heights, NJ. OBJECTIVE: Identification of pre-diabetes (pre-DM) represents an important intervention point for the prevention of diabetes (DM). The prevalence of pre-DM among reproductive-age women in the US is about 18% and may be even greater in infertile women. Though the 2-hour glucose tolerance test (2hGTT) is considered by many to be the gold standard for the diagnosis of pre-DM, multiple screening methods exist. The objective of this study was to determine whether Hemoglobin A1C (HbA1C) predicts pre-DM in a population of women with infertility and/or recurrent pregnancy loss (RPL), when considering the 2hGTT as the gold standard. DESIGN: Retrospective chart review.
FERTILITY & STERILITYÒ
MATERIALS AND METHODS: Pre-DM was diagnosed by one of the following: Fasting plasma glucose (FPG) 110-125 mg/dL, 2hGTT 140-199 mg/dL or Hemoglobin A1c (HbA1C) 5.7-6.4%. DM was defined as one of the following: FPG R126 mg/dL, 2hGTT R200 mg/dL or HgbA1C R6.5%. All patients presenting with infertility and/or RPL to a universityaffiliated reproductive endocrinology practice between ages 18-46 with a 2hGTT result were included. Patients with known diagnosis of DM or preDM were excluded. RESULTS: 242 patient charts were reviewed. The mean age was 33.7 years and mean BMI was 31.6. PCOS was diagnosed in 61% of our study population. The prevalence of pre-DM and DM was 43% (95% CI 36.7-49.5) and 3.7% (95% CI 1.7-6.9), respectively. 240 patients had R1 screening test performed, of which 188 (78.3%) had both a 2hGTT and HgbA1C. Of those 188 patients, 104 (55.3%) were identified as having pre-DM. 69/104 (66%) patients with pre-DM would have been missed with only a 2hGTT, and 6/104 (5.8%) would have been missed with only a HbA1C. When using the 2hGTT as the gold standard for the diagnosis of pre-DM, the sensitivity, specificity, positive and negative predictive values of HgbA1C compared to 2hGTT was 70% (95% CI 45.7-88.1), 58.9% (95% CI 51.1-66.4), 16.9% (95% CI 9.5-26.7) and 94.3% (95% CI 88-97.9), respectively. Cohen’s kappa coefficient for HgbA1C and 2hGTT was 12.1 (95% CI 2.1-22.1). CONCLUSIONS: Although a normal HbA1C was highly predictive of a normal 2h-GTT, the two tests demonstrate poor agreement in the identification of pre-DM in women with infertility and/or RPL. The clinical relevance of an elevated HbA1C in this population needs to be better defined. P-274 Tuesday, October 31, 2017 EFFECT OF GONADOTROPHIN RELEASING HORMONE AGONIST (GNRHA) - INDUCED PSEUDOMENOPAUSE ON SERUM 25-HYDROXY VITAMIN D3 LEVEL AND HEALTH RELATED QUALITY OF LIFE (HRQOL) IN ENDOMETRIOSIS. S. M. Bhattacharya,a A. Basu,b B. Biswas.c aObstetrics and Gynecology, S. C. Das Memorial Medical & Research Center, KPC Medical College, Kolkata, India; bPathology, West Virginia University School of Medicine, Morgantown, WV; cObstetrics and Gynecology, Apollo Clinic, Kolkata, India. OBJECTIVE: There are evidences that 25 hydroxy vitamin D3 (D3) sufficiency is required for optimal health1 . Our objective was to study the effects of two doses of Gonadotrophin Releasing Hormone Analogue (GnRHa) (Inj.Triptorelin)- induced pseudomenopause in endometriosis on D3 level and Health related quality of life (HRQOL) (using Menopause Rating Scale). DESIGN: Open label prospective interventional study. MATERIALS AND METHODS: Study was done in Kolkata, India between 01.10.13 to 31.1.15 after obtaining informed written consents and ethical approval. Based on a pilot study, a sample size of 60 was estimated to have 90% power and 95% confidence level with 2-sided test of significance to detect a mean difference of 3 in the total HRQOL score (164.2 before v/s 195.8 after treatment). Following laparoscopic diagnosis and fertility sparing surgery of endometriosis, 68 patients (age 17-37 years) were studied and were given inj.GnRHa (two subcutaneous doses of injection Triptorelin, 3.75 mg, one month apart to induce pseudomenopause). Serum D3 (ng/ml) level was measured twice - at baseline and one month after the second dose. HRQOL was assessed using the Menopause Rating Scale (MRS) at the two time-points. For data analysis, paired t test was done. RESULTS: No significant change in D3 level was found after the two doses. However, there were significant increases in the total MRS scores (4.2, 95% CI ¼ 2.21, 4.21), along with increases in the psychological (2.65, 95% CI ¼ 1.67, 5.39) and urogenital (1.27, 95% CI ¼ 0.46, 3.13) components at the study end-point, demonstrating significant deterioration in HRQOL. CONCLUSIONS: GnRHa induced pseudomenopause in endometriosis does not cause any significant change in D3 level but causes significant deterioration in HRQOL. Studies are needed to assess the effects of GnRHa on other systems involved in D3 homeostasis and effects of supplementation of D3 in addition to calcium on HRQOL in those with hypovitaminosis D3. CTRI registration no.- CTRI/2013/08/003927. Reference: 1. Verbrugge H, Gielen E, Kilisen K et al. Who Should Receive Calcium and Vitamin D Supplementation.Age Ageing. 2012; 41; 576-80