Objective: To investigate the effects of calcium supplementation on the inflammatory response of vascular endothelial cells and the possible role and mechanism of G protein-coupled estrogen receptor(GPER)under the physiological conditions of estrogen deficiency(ovary removal).
Methods: Forty healthy 8-week-old female C57 BL/6 mice were divided into 4 groups by random number table according to body weight, which were respectively the control group, ovarian removal group, ovarian removal high calcium group, and G1 ovarian removal high calcium group, each group ten. One week after the basal diet, the control group underwent sham surgery, and the ovaries-removed group, ovarian-removed high-calcium group, and G1 ovarian-removed high-calcium group were treated for ovarian removal. The control group and the ovary-removed group were fed basic feed(AIN-93 G, calcium content 0. 5%), the ovarian-removed high-calcium group and the G1 implanted ovarian-removed high-calcium group were fed high-calcium feed(adjusted AIN-93 G, calcium content 1. 5%). Three months later, mice in the control, ovarian-removed, and ovarian-reduced high-calcium groups were implanted with a placebo subcutaneously, and G1 ovarian-removed high-calcium groups were implanted with a GPER specific agonist G1, intervention for 28 days, under the anesthesia state, collect blood from 40 mice, isolate serum to measure vascular endothelial adhesion factor-1(VCAM-1), tumor necrosis factor-α(TNF-α), estradiol and blood calcium concentration; take aortic arch for immunization histochemical detection of transient receptor potential channel 1(TRPC1), phosphorylated-extracellular regulated kinase(p-ERK)1/2, VCAM-1 and CD68 expression levels. SPSS software performs statistical analysis and draws conclusion.
Results: The differences in weight gain between the three groups and the control group were statistically significant(P<0. 05), while the differences in weight between the three groups were not statistically significant(P>0. 05). There were statistically significant differences in serum estradiol levels between the three groups, the ovariectomized high calcium group, and the G1 ovariectomized group implanted with the control group(P<0. 05). The difference was not statistically significant(P>0. 05). The differences in serum VCAM-1 concentrations between the control group and the other three groups were statistically significant(P<0. 05). The difference in serum VCAM-1 concentration was statistically significant between the ovariectomized group and the implanted G1 ovariectomized high calcium group and the ovariectomized high calcium group(P<0. 05). There was no significant difference in serum VCAM-1 concentration in the group(P>0. 05). There was no significant difference in TNF-α concentration between the four groups(P>0. 05). By immunohistochemistry, the percentages of TRPC1, p-ERK1/2, VCAM-1, and CD68-positive cell areas in the ovarian high-calcium group were significantly higher than those in the ovarian-removed group and the G1-ovarian-removed high-calcium group. The difference was statistically significant(P<0. 05), and the difference in the percentage of positive cell area between the ovarian-removed group and the G1-removed high-calcium group was not statistically significant(P>0. 05).
Conclusion: In the state of estrogen deficiency, calcium supplementation causes an increase in inflammatory response, which may be related to the change in GPER activity, and then affect the TRPC1/ERK1/2 pathway.
Keywords: G protein-coupled estrogen receptor; calcium supplementation; ovariectomy; vascular endothelial adhesion factor-1.