The Preliminary Research Of The Impact Of Estrogen On Glycolipid Metabolism And Islet Function In CETP Transgenic Mice

Objective:Estrogen deficiency is closely associated with the onset of type 2 diabetes. So study about effect of estrogen on glucose metabolism and islet function is vital to prevention of type 2 diabetes and its complications. CETP, cholesteryl ester transfer protein, which acts as a protein transfering cholesteryl ester, plays a crucial role in the process of reverse cholesterol transport. CETP has been demonstrated to have great significance on glycolipid metabolism and islet function. C57BL/6J mice or SD rats, which were used as the animal models to study the effect of estrogen, express no or little CETP gene. Therefore, mice expressing the similar CETP gene as humuns, combined with hyperinsulinemic euglycemic clamp test, were adopted to investigate the impact of estrogen on glycolipid metabolism and islet function in CETP transgenic mice, thus taking a further look at the role that estrogen plays on development of type 2 diabetes.Method:1. Two groups of mice with different genetic backgrounds, wild-type C57BL/6J mice and NFR-CETPTg mice were selected for the first part of this study. Plasma CETP activity and genotypes identification measurements were conducted among these animals before experiments. Characteristic of tissue expression were measured in NFR-CETPTg mice.2. In the second part of this study, sham group, ovariectomized (OVX) group and estradiol group were included. Plasma CETP activity was measured to evaluate the influence of estrogen on CETP activity. Mice were fed on a normal diet since eight 12 weeks old of age until eight weeks after operation. Daily food intake and body weight were kept a record of to observe the effect of estrogen on body weight and metabolism in CETP transgenic mice. Levels of fasting glucose were measured four and eight weeks after surgery, and serum lipid levels were detected to find out the effect of estrogen on the lipid profile and glucose metabolism in CETP transgenic mice. Intraperitoneal injection of glucose tolerance test (IPGTT) was done eight weeks after surgery. Mice were sacrificed and islets of the three groups of CETP transgenic mice were collected to conduct glucose-stimulated insulin secretion experiments (GSIS).3. In the third part of this study, sham group, ovariectomized (OVX) group and estradiol group were included. Mice accepted intraperitoneal injection of insulin tolerance test (IPITT) and hyperinsulinemic euglycemic clamp eight weeks after surgery, in order to investigated the influence of estrogen on insulin sensitivity in CETP transgenic mice. Furthermore, pancreas biopsies were done and islets morphological changes were observed in these mice in order to figure out the effect of estrogen on the islet function and morphology in CETP transgenic mice.Results:1. Mice genotype identificationNFR-CETPTg mice showed specific 500bp bands. NFR-CETPTg mice and human plasma CETP activity were both significantly higher, the results were statistically significant. Characteristic of tissue expression in NFR-CETP transgenic mice illustrated that the tissue distribution of CETP mRNA expression in NFR-CETPTg mice was the same as that in human.2. Effects of estrogen on CETP activity.Determination of serum CETP activity suggested that there was no significant difference in serum CETP activity among sham group, ovariectomized group and estrogen group.3. Comparison of food consumption body weight during 8 weeksOvariectomized mice consumed more food than the sham group (P<0.05) during eight weeks of feeding, and food consumption of estrogen group decreased. Ovariectomized mice gained more weight compared with the sham group, and there was significantly difference between the two groups of mice seven weeks significant (P<0.05) after surgery. Besides, weight gain of estrogen group increased comparatively slower.4. Effects of estrogen on lipid profile and blood gucose levels of CETP transgenic mice.Ovariectomized mice had a trendency of elevated TC compared with the sham mice, but with no statistically significance. Levels of serum TG, HDL-C, LDL-C were not significantly different among these three groups of mice. There was no significant difference of fasting blood glucose among the sham group, ovariectomized group and estrogen group of mice four weeks after surgery; while ovariectomized mice displayed elevated fasting blood glucose compared with sham mice eight weeks after surgery, and estrogen replacement could partially compensate the difference.5. Comparison of glucose tolerance.Results of IPGTT illustrated that ovariectomized mice had significantly higher blood glucose levels at each time point compared with sham mice (P<0.05) eight weeks after surgery, while blood glucose levels of estrogen group improved significantly at 0 min,15 min and 30 min. Furthermore, there was no significantly difference between the mice of estrogen group and sham group at 60min and 120min.6. Comparison of islet function.Results of GSIS showed that eight weeks after surgery, ovariectomized mice tended to have impaired insulin secretion function than sham mice; while estrogen replacement could partially relieve the impaired islet function.7. Effects of estrogen on insulin sesitivity.Results of IPITT illustrated there was no statistical significance among the three groups in blood glucose levels eight weeks after ovariectomy, probably because IPITT could just assess insulin sensitivity relatively roughly. Mice were operated on hyperinsulinemic euglycemic clamp and we found that glucose infusion rate (GIR) of ovariectomized mice reduced eight weeks after surgery, estrogen may improve insulin resistance.8. Effects of estrogen on pancreatic morphologyThe number of isolated islets of ovariectomized mice smaller compared to the sham group, and the results were statistically significant (P<0.05). Estrogen replacement could increase the number of islets to some extent. Results of hematoxylin-eosin (HE) of pancreatic biopsy suggested that, islet size and the number of cells of ovariectomized mice decreased compared to sham mice, and estrogen replacement could somewhat enlarged the islet size and increased the number of cells in islets.Conclusions:1. NFR-CETPTg mouse is an ideal animal model for studies of estrogen effects on glucose metabolism.2. Hyperinsulinemic euglycemic clamp is the gold standard assessing insulin sensitivity.3. Estrogen deficiency causes disorders of glucose metabolism, and estrogen replacement could alleviate the harmful effects on islet P cells to some extent.