Mechanism Of Cuscuta Chinensis Lam. Acting As Estrogen Based On PI3K/AKT Signaling Pathway

Cuscuta chinensis is the mature dry seed of the Convolvulaceae plant Cuscuta chinensis Lam.It is sweet and pungent.It is belongs to tonic drug.Cuscuta chinensis can regulate the level of estrogen in the body and has a good effect on the lack of kidney qi caused by the perimenopausal.The previous research showed that the 95% ethanol extract of cuscuta chinensis from Heilongjiang has significant estrogen-like activity.Among them,four components:hyperoside,kaempferol,quercetin,and apigenin are the prototype components in the blood.These are cuscuta chinensis’s "Main medicinal components" that play the role of estrogen-like activity.But the mechanism of estrogen-like activity is still unclear.In order to further investigate four components of hyperoside,kaempferol,quercetin,and apigenin,as well as the 95% ethanol extract of cuscuta chinensis to play an estrogen-like activity mechanism.The main research contents and results of this article are as follows:(1) Based on network pharmacology predicting the targets and molecular mechanisms of estrogen-like effects of four main components in cuscuta chinensis.There are total of 198 targets related to estrogen deficiency.Four compounds can act on AKT1、IL6、EGFR、ESR1 and other targets to play an estrogen-like activity;The results of KEGG enrichment analysis show that the mechanism of cuscuta chinensis’s estrogen-like activity may be related to the PI3K-AKT signal pathway,estrogen signal pathway,EGFR signal pathway and other estrogen-related signal pathway.(2) Taking the human breast cancer cell MCF-7 (ER+) as the research object.Cell proliferation was detected by MTT experiment.Cell cycle detected by flow cytometry.Western blot experiment to detect the expression of estrogen-related protein.Clarify the mechanism of four main components to estrogen-like activity.Experimental results show that the four main components can promote the proliferation of MCF-7 cells when administered alone (P<0.05,P<0.01),and the combined administration of four main components can also promote the proliferation of MCF-7 cells (P<0.05,P<0.01).Compared with single component,the combined of four main components reach the highest proliferation rate can reduce dosage.The results of flow cytometry showed that the ratio of cells in G0/G1 phase decreased,while the ratio of cells in S phase increased (P<0.05) after the combined administration of the four main components on the cells for 48 hours.The results of Western Blot experiments showed that the combined administration of the four main components on MCF-7 cells can up-regulate the expression level of ERa and PI3K (P<0.05,P<0.01).(3) Establish a model of estrogen deficiency in ovariectomized female rats,and randomly divided them into positive drug group,cuscuta chinensis administration group,model group and sham operation group.Western blot and immnohistochemistry experiments were used to investigate the mechanism of the 95% ethanol extract of cuscuta chinensis’s estrogen-like activity.Western Blot experiment results showed that the ethanol extract of cuscuta chinensis can up-regulate the expression levels of ERα,PI3K,and p-AKT protein in rat uterine tissue.Compared with the model group,the difference was statistically significant (P<0.05,P<0.01).The results of immunohistochemical experiments showed that the ethanol extract of cuscuta chinensis can up-regulate the expression levels of EGFR and p-AKT in rat uterine tissues.Compared with the model group,the difference was statistically significant (P<0.05,P<0.01).The ethanol extract of cuscuta chinensis can play an estrogen-like effect on ovariectomized female rats by activating the ER-mediated PI3K/AKT signaling pathway.(4) Establish a model of premature ovarian failure in female rats,and randomly divide them into positive drug group,cuscuta chinensis administration group,model group and control group.Western Blot and immunohistochemistry experiments were used to investigate the mechanism of the 95% ethanol extract of cuscuta chinensis’s estrogen-like activity.Western Blot experiment results showed that the ethanol extract of cuscuta chinensis can up-regulate the expression levels of ERα,PI3K,and p-AKT protein in rat uterine tissue.Compared with the model group,the difference was statistically significant (P<0.05,P<0.01).The results of immunohistochemical experiments showed that the ethanol extract of cuscuta chinensis can upregulate the expression levels of EGFR and p-AKT in rat uterine tissue.Compared with the model group,the difference was statistically significant (P<0.05,P<0.01).The ethanol extract of cuscuta chinensis can play an estrogen-like effect on premature ovarian failure rats by activating the ER-mediated PI3K/AKT signaling pathway.