| Background:Perimenopausal women’s saliva secretion disorder is manifested by decreased saliva secretion and changes in composition.Studies have shown that its occurrence is closely related to the decrease of estrogen content.The chloride channel is the original driving force of the saliva secretion process,and α-saliva amylase is one of the most abundant organic substances in saliva.Does the change in estrogen content affect the chloride channel and α-saliva amylase to participate in the salivation disorder of perimenopausal women? The occurrence of is unclear.Objective:Observe the changes of estrogen receptor,chloride channel and α-saliva amylase activity in the submandibular gland under low estrogen conditions through animal experiments,and observe the activity of estrogen receptor GPR30 on chloride ion channels and α-saliva amylase through cell experiments.Explore the possible mechanism of estrogen regulating chloride secretion and α-salivary amylase activity through GPR30.Methods:Animal experiments using 60 adult female SD rats,were randomly divided into SHAM operation group(SHAM),ovariectomized group(OVX),ovariectomized estrogen treatment group(OVX+E),each group of 20.Bilateral oophorectomy was performed in OVX group and OVX+E group.After 2 weeks of recovery,the OVX+E group was injected with estrogen for 4 weeks,vaginal exfoliated cell smears,uterine morphology and radioimmunoassay determined that the ovariectomized model was established successfully.Immunohistochemistry and western blotting were used to detect the expression of estrogen receptor(ERα,ERβ,GPR30),chloride channel protein ANO1,inositol triphosphate receptor IP3 R,vasoactive intestinal peptide VIP and receptor VPAC1 in submandibular gland;Immunofluorescence technology was used to observe whether there is co-expression between ANO1 and IP3R;the kit detects the activity of α-saliva amylase in tissues.In the cell experiment,SMG-C6 cell line was used to specifically agonize and antagonize GPR30.There were four groups: control group(Control),agonist group(G1),antagonist group(G15)and first antagonist and then agonized group(G15+G1).The expression of related proteins was observed by immunohistochemistry.Results:After the rats were ovariectomized,uterine has obviously atrophy,the estrous cycle was disappeared,the estrogen level in the body was significantly reduced,the estrogen receptor ERβ did not change,the ERα expression decreased,and the expression did not increase significantly after estrogen treatment;the expression of GPR30 decreased,and the expression after treatment returned to the level of the SHAM group.The expression of chloride channel protein ANO1 and inositol triphosphate receptor IP3 R both decreased significantly after ovariectomy.After estrogen treatment,the expression increased close to the level of the control group,and the two co-localized on the cell membrane of acinar cells.After ovariectomy,the activity of salivary amylase secreted by the submandibular gland also decreased significantly,and the expression of vasoactive intestinal peptide VIP and its receptor VPAC1 also decreased.After intervention with estrogen,they returned to the level of the control group.After specifically activating the GPR30 receptor in SMG-C6 cells,the expression of IP3 R,ANO1,VIP,and VPAC1 all increased,and the specifically antagonizing GPR30,the expression of the four proteins all decreased.Conclusion:In the low estrogen state,the expression of GPR30 in rat submandibular gland decreases.On the one hand,it may cause the decrease of IP3 R and ANO1 expression,which affect chloride secretion,leading to the decrease of saliva secretion;on the other hand,it may cause the decrease of VIP and VPAC1 expression,which leads to a decrease in the activity of α-salivary amylase and saliva composition is changed finally. |
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