Identification of the Role of Vitamin D and VDR in Human Endometrial Stromal Cells

In women undergoing in vitro fertilization, higher vitamin D status has been associated with improved live birth rates. Among women with polycystic ovarian syndrome (PCOS), vitamin D has been associated with improvements in spontaneous ovulation. Vitamin D receptor (VDR) is a nuclear receptor activated by 1,25 dihydroxyvitamin D which is the most active metabolite of vitamin D. Mice that lack the vitamin D receptor exhibit arrested follicular development, uterine hypoplasia, prolonged estrous cycles, and subfertility. Also, one study of mice reported that the vitamin D receptor is expressed in gonadotropin-releasing hormone (GnRH) neurons, which might suggest a role for vitamin D in hypothalamic signaling. Since the vitamin D receptor knockout mouse is sub fertile with uterine hyperplasia, the most likely cause of vitamin D deficiency may be due to estrogen production by the ovary or estrogen responsiveness of the uterus. So, in this study, we want to identify the effect of vitamin D on human endometrial stromal cells associated with female reproduction. Overall design: Experimental Design: There are 4 groups, and 12 samples in total. All RNAs will be extracted from the human endometrium stromal cell line (T-HESC). 1) Non-targeting siRNA transfection (control) - treated with vehicle (n=3), treated with 1,25-dihydroxyvitamin D3 (n=3) 2) VDR siRNA transfection (VDR knockdown) - treated with vehicle (n=3), treated with 1,25-dihydroxyvitamin D3 (n=3) Data Analysis Strategy: Dr. Tianyuan Wang from the informatics group will help us with the analysis. 1) Remove adaptor 2) Require unique mapping and allow 1 mismatch 3) Remove duplicated reads 4) Down-sampling to the appropriate sequencing depth for each 5) Compare between different groups (siNT vs siVDR, Vehicle vs ligand (1,25-dihydroxyvitamin D3))