The Expression Of Ovn In Yak Oviduct During Estrous Cycle And The Effect Of EGF On Its Expression

Oviductin(Ovn) plays crucial role during fertilization and early embryonic development. However, there are little studies on the molecular characteristics of Ovn and its expression during estrous cycle. In this study, we cloned Ovn c DNA by RT-PCR. And detected the expression of oviductin in oviductal ampulla and isthmus of yak in early follicular, late follicular, early luteal and late luteal by q RT- PCR and immunohistochemistry, analyzed its different expression during the estrous cycle from m RNA and protein level. Based on the above studies, we supplied different concentration of EGF to oviduct epithelial cells of yak cultured in vitro, the expression levels of Ovn, HIF-1α, Bcl-2 and Bax in different groups were detected by q RT-PCR, which was to investigate the regulation function of EGF on the secretion of oviduct epithelial cells of yak. Results showed that:(1) Oviductin c DNA sequences of yak shared very high homology with cattle and buffalo. The phylogenetic tree based on amino acid sequences of oviductin indicated that yak oviductin was closely related to its cattle counterpart, reached 99.1% with Bos tauru. The oviductin c DNA of yak was successfully cloned, and the Gen Bank accession number was KF303541.(2) The expression of oviductin m RNA was highest in the stage of late follicle, decreased stepwise in the stages of early luteal and late luteal, and was lowest in the stages of early follicle. The prominent immunolabeling were observed in the late follicle and early luteal stages, which was reduced in other stages. There was no significant differences of expression of oviductin m RNA between ampulla and isthmus.(3) The adherent time and growth of yak primary oviduct epithelial cells were slow, oviductin was mainly distributed in cytoplasm, and 90% FBS + 10% DMSO cryopreservation was better than traditional method.(4) With the addition of different concentration of EGF to oviduct epithelial cells cultured in vitro, there would be found an increased m RNA relative expression of Ovn, HIF-1α and Bcl-2 and a declined m RNA relative expression of Bax, with a dose-dependent effect, the optimal concentration was 50 ng?m L-1.In this study, we obtained c DNA sequence of yak oviductin containing an open reading frame and defined the differences of Ovn m RNA expression in different stages of estrous cycle. Found that EGF may regulating HIF-1α to inhibit the apoptosis of oviduct epithelial cells and promote the secretion of oviductin. The results in this study provide theoretical foundation to explore the functions of EGF during yak oocytes maturation and preimplantation embryo development, which will be helpful to improve the technique of yak embryo cultured in vitro.