The Expression And Hormonal Regulation Of Vascular Endothelial Growth Factor-A In The Of Goat Embryo Implantation

Embryo mortality is the main factor affecting the fertility,production and economic efficiency of ruminants.Most pregnancy failures occur in early pregnancy and are related to embryo implantation failure.Vascular endothelial growth factor A（Vascular endothelial growth factor-A,VEGF-A）is the most effective and critical angiogenic factor known so far.It plays an important role in the reproduction process,but the role of VEGF-A in goat embryo implantation is incomplete clear.In this experiment,goats were used as experimental animals to establish embryo implantation-related models in order to clarify the expression and regulation of VEGF-A in goat uterine early pregnancy,evaluate the role of VEGF-A in embryo implantation,and provide basis for studying the mechanism of ruminant embryo implantation,it is of far-reaching significance to improve animal reproduction and thus increase the economic value of production.Objective:to evaluate the role of VEGF-A in goat embryo implantation by detecting the expression and regularity of VEGF-A in goat uterus during early pregnancy and estrous cycle;to detect the regulation of VEGF-A by IFNτthrough uterine perfusion of IFNτmodel,to evaluate whether VEGF-A is regulated by IFNτ;and to detect whether VEGF-A is regulated by estrogen and progesterone by ovariectomized subcutaneous injection of hormone.In vitro culture goat uterine epithelial cells to detect the effect of hormones on VEGF-A expression levels in uterine epithelial cells.This study is used to clarify VEGF-A’s expression and related hormone regulation in goat embryo attachment.Materials and methods:Materials and methods:the uterine tissues of Heilongjiang native goats were taken on the 0th,3rd,6th,8th and 11th days of estrous cycle,respectively,and the expression of VEGF-A was detected by immunohistochemical method.In the model of early pregnancy,the uterine tissues of D0,D3,D6,D9,D16,D19,D25,D30 caruncular and D30 inter caruncular,D50th and D60 were selected to detect the expression of VEGF-A in goat uterus by in situ hybridization,immunohistochemistry and Western blotting,and the expression of VEGF-A in trophoblast of early embryo on D16,D30,D50 and D60 was detected by immunohistochemistry.Interferon-τwas infused into the uterus of goats,and serum protein was infused into the control group.The expression of VEGF-A in the control group and the experimental group was detected by immunohistochemical method.After bilateral ovariectomy,E₂,P₄ and oil（control）were subcutaneously injected into the body after hormone metabolism was exhausted,and the uterine tissue was taken to detect the effect of hormone on the expression of VEGF-A in goat uterus by immunohistochemistry,q PCR and Western blotting.Goat uterine epithelial cells were isolated and cultured.The epithelial cells were treated with E₂,P₄ and E₂+P₄ respectively.The expression of VEGF-A and estrogen receptor I and progesterone receptor in goat uterine epithelial cells were detected by q PCR,and the effects of estrogen and progesterone on goat uterine epithelial cells were evaluated.Results:during the estrous cycle,the expression of VEGF-A protein was the strongest on ED6and ED8 during the estrous cycle,and there was no significant difference in the expression of VEGF-A protein between the deep and shallow glands.In the early pregnant uterus of goats,VEGF-A was mainly expressed in the luminal epithelium and glandular epithelium,and was highly expressed in the adhesion stage（D16）,but not in the uterine glands of D50 and D60,but highly expressed in the caruncle matrix.VEGF-A is highly expressed in D16,while D30,D50 and D60are relatively weak.In the goat uterine perfusion IFNτmodel,compared with the control group,VEGF-A was highly expressed in the uterus perfused with IFNτ,especially in the deep glands,indicating that IFNτstimulated the expression of VEGF-A.In vivo hormone treatment model,the results showed that both estrogen and progesterone down-regulated the expression of VEGF-A in goat uterus.Goat uterine epithelial cells isolated in vitro were cultured with hormone.The results showed that there was no significant change in VEGF-A in E₂ group and P₄ group,but E₂+P₄ treatment significantly upregulated the expression of VEGF-A m RNA,and E₂ group,P₄ group and E₂+P₄group all up-regulated ESR1（p<0.05）.PR was significantly up-regulated by E₂.P₄ alone upregulated the expression of PR（p<0.05）,but not significantly.E₂+P₄ combined treatment could also up-regulate the expression of PR,but the effect was not as obvious as E₂ alone.The response of goat uterine epithelial cells cultured in vitro to estrogen and progesterone was diff erent from that of uterine tissue in vivo to estrogen and progesterone.Conclusion:（1）VEGF-A is mainly expressed in luminal epithelium and glandular epithelium in goat endometrium,and is highly expressed in the adhesion stage（D16）.（2）VEGF-A is highly expressed in D50 and D60 fleshy areas.（3）VEGF-A is highly expressed in trophoblast cells of gestational D16 embryo.（4）IFNτcan up-regulate the expression of VEGF-A.（5）In vivo,VEGF-A in goat uterine tissue was down-regulated by estrogen and progesterone alone,and the expression of VEGF-A in goat uterine epithelial cells treated with estrogen and progesterone in vitro was not significantly different from that in the control group,but the expression of VEGF-A m RNA was up-regulated by estrogen and progesterone combined treatment.Our study p rovides a reference for elucidating the mechanism of physiological changes in the uterus during implantation of ruminant embryos.