Role of transient receptor potential channels in mammalian oviduct and uterine epithelia

Calcium is an important secondary messenger and plays a major role in cell function, including proliferation, cell growth, secretion and death . It also plays a critical role in uterine smooth muscle contraction and embryo implantation. This thesis is concerned with calcium homeostasis in epithelial tissue lining the oviduct and uterus which are key players in early reproductive events, being involved in gamete transport, sperm capacitation and providing the micro-environment for the gametes and early embryo. Calcium transport across epithelial cells is either via tight junctions or calcium channels, specifically, members of the transient receptor potential (TRP) channel superfamily and the Na+/Ca2+ exchanger. TRP channels are an important class of calcium channels with more than 28 identified members and their potential involvement in calcium transport in uterus and oviduct epithelia has yet to be determined. The aim of this study was to discover which TRPC isoforms are expressed in epithelial cells lining the female reproductive tract in the bovine and human. Gene expression of TRPC channels changes was measured throughout the estrous cycle in bovine oviduct and uterine epithelial cells using Real-Time PCR, while immunohistochemistry, immunocytochemistry and western blotting were used to discover the localization of TRPC channels in oviduct/uterine epithelium and changes in protein expression of TRPC isoforms induced by sex hormones. to The physiological role of TRPC isofoms in regulating intracellular calcium concentration in bovine oviduct epithelial cells was determined using a calcium assay approach and finally. the potential clinical relevance of a possible role of TRP channels in female reproduction was investigated. OF 7 members of TRPC family, TRPC1, 2, 3, 4 and 6 were expressed in bovine oviduct and uterine epithelia. In human endometrium, TRPC1, 6 and 7 genes were detected. Expression levels of all TRPC isoforms present in both bovine oviduct and uterine epithelia changed throughout the estrous cycle. 17β-estradiol, FSH and LH individually and in combination up-regulated gene expression of TRPC isoforms in bovine oviduct epithelial cells. However, progesterone inhibited the upregulatory effect of 17β-estradiol, FSH and LH on TRPCs gene expression. TRPC1 and TRPC6 which are the common TRPC isoforms in bovine oviduct/uterine epithelium and human endometrium were localized on the apical, basal and lateral membranes of the epithelial tissue in bovine oviduct/uterus and human endometrium. TRPC isoforms were physiologically active in bovine oviduct epithelial cells (BOEC). SKF96365 which is a general TRP channel blocker inhibited the calcium influx into BOEC. Furthermore, Hyperforin which is a TRPC6 channel activator increased the intracellular calcium concentration in BOEC. TRPC1, 6 and 7 expression in endometrium of patients being treated for infertility by IVF illustrated that gene expression of TRTPC1 and 6 were up regulated in the endometrium of the IVF patients compared to controls. However, gene expression of TRPC7 in IVF patients was downregulated compared to that of the endometrium of the control group. Gene expression of TRPC6 and 7 in endometrium of women with Poly Cystic Ovarian Syndrome (PCOS) who have higher level of LH and normal FSH level, alongside the absence of the post-ovulatory increase in progesterone secretion, were up -regulated compared to that of the control group. However, the expression level of TRPC1 in endometrium of PCOS patients was not significantly different compared to the control group. Gene expression of TRPC isoforms in the epithelia lining the female reproductive tract is possibly regulated by sex hormones via nuclear factor-kappa B (NF-КB) signalling pathway. However, further investigation is required to determine the mechanisms underlying the endocrine regulation of TRPC channels.