| Objective: Diabetes mellitus is a complex metabolic disease.Recently,the incidence of diabetes has been increasing significantly,only behind cancer and cardiovascular disease,it has become the third largest health killer,so diabetes prevention and treament research have been a hot topic.Estrogen is closely related to diabetes,animal experiments indicat that male animals have a higher probability of getting diabetes,if removal of female ovaries,the risk of developing diabetes increase significantly.Epidemiological surveys show that men are more likely to develop diabetes than women,the incidence of diabetes is significantly higher in postmenopausal women,decreased estrogen levels can cause insulin resistance,once are thought to be one of the causes of diabetes.Some scholars have tried to use estrogen for the prevention and treatment of diabetes,and they have achieved a certain effect,but later studies show that estrogen can increase the risk of breast and ovarium cancer,so some researchers have focused on looking for the natural estrogen alternatives with low side effects but could delay diabetes.Resveratrol is a estrogenic hormone analogues,our laboratory had ever investigated the effects of different doses of resveratrol and 17 β-estradiol on blood glucose,serum insulin malondialdehyde(MAD),and plasma total antioxidant capacity(T-AOC)in type 1 diabetes(T1D),and indicate that resveratrol and 17 β-estradiol can delay and prevent the onset of diabetes induced by streptozotocin(STZ)in ovariectomized mice.On the basis of the previous experiment,we had using molecular biology techniques to study the effect of glucose transport of resveratrol and estradiol on peripheral tissue skeletal muscle and myocardium of STZ-induced diabetic mice and the underlying molecular mechanism.Methods: Female mice after ovariectomy were randomly divided into eight groups as follows: Ovariectomy group(OVX),Ovariectomy group with intraperitoneal injection of 50 mg/kg body weight of STZ;Ovariectomy group with intraperitoneal injection of STZ and subcutaneous injection of 0.1 mg/kg body weight of resveratrol;Ovariectomy group with intraperitoneal injection of STZ and subcutaneous injection of 1 mg/kg body weight of resveratrol;Ovariectomy group with intraperitoneal injection of STZ and subcutaneous injection of 10 mg/kg body weight of resveratrol;Ovariectomy group with intraperitoneal injection of STZ and subcutaneous injection of 0.01 mg/kg body weight of 17β-estradiol;Ovariectomy group with intraperitoneal injection of STZ and subcutaneous injection of 0.1 mg/kg body weight of 17β-estradiol;Ovariectomy group with intraperitoneal injection of STZ and subcutaneous injection of 1 mg/kg body weight of 17β-estradiol.Some groups were intraperitoneally injected STZ at every day lasting for five days.At the same time resveratrol and 17β-estradiol were administrated subcutaneously at every other day for 4 weeks.Mice were killed by dislocation of the cervical spine,skeletal muscle and cardiac muscle were cut and kept at-80°C for Western Blot and RT-PCR experiments;another part of tissues were fixed in the 4% paraformaldehyde for Immunohistochemical tests.The indexes were detected at the present study as follows: the expressions of glucose transporter 4(GLUT4),insulin receptor-1(IRS-1),extracellular signal-regulated kinase(ERK),the phosphorylation of extracellular signal-regulated kinase(p-ERK)and GLUT4 mRNA.Results: Western blot and RT-PCR experiments showed that the expression of GLUT4 protein and mRNA in the ovariectomy with intraperitoneal injection of STZ group was dramatically reduced in both skeletal muscle and myocardial tissue compared with the ovariectomy group(P < 0.001).After treatment with resveratrol and 17β-estradiol the expression of GLUT4 was increased significantly: In skeletal muscle,except the group of resveratrol at the dose of 0.01 mg/kg body weight,the expression of GLUT4 in other groups were significantly increased(P <0.001);in the myocardial tissue,the expression of GLUT4 in the 1 mg/kg body weight of resveratrol group,the 10 mg/kg body weight of resveratrol group,the 0.1 mg/kg body weight of 17β-estradiol group and the 1 mg/kg body weight of 17β-estradiol group were significantly increased(P <0.001).Immunohistochemistry experiment results were as the same with Western blot and RT-PCR experiments.Compared with the ovariectomy group,the expression of IRS-1 protein in the ovariectomy with intraperitoneal injection of STZ group was dramatically reduced.After treatment with resveratrol and 17β-estradiol,the expression of IRS-1 increased significantly: In skeletal muscle,the expression of IRS-1 in the 10 mg/kg body weight of resveratrol group and the 0.1mg/kg body weight of 17β-estradiol group were significantly increased(P <0.001);in the myocardial tissue,the expression of IRS-1 in the 1 mg/kg body weight of17β-estradiol group were increased significantly(P <0.001).Phosphorylation levels of ERK protein in diabetic model mice skeletal muscle and cardiac tissue also had great differences.Compared with the ovariectomy group,the phosphorylation levels of ERK protein in the ovariectomy with intraperitoneal injection of STZ group was dramatically improved(P <0.001).After treatment with resveratrol and 17β-estradiol the phosphorylation levels of ERK protein decreased significantly: In skeletal muscle,the phosphorylation levels of ERK protein in the 10 mg/kg body weight of resveratrol group,the 0.01 mg/kg body weight of 17β-estradiol group,the 0.1 mg/kg body weight of 17β-estradiol group and the 1 mg/kg body weight of 17β-estradiol group were significantly decreased(P <0.001);in the myocardial tissue,the phosphorylation levels of ERK protein in the 10 mg/kg body weight of resveratrol group,the 0.01 mg/kg body weight of 17β-estradiol group,and the 0.1 mg/kg body weight of 17β-estradiol group were significantly decreased(P <0.001).Conclusion:(1)The transport 2of glucose in diabetic mice skeletal muscle and myocardium was abnomal,the expression of GLUT4 and IRS-1 was decreased,the glucose transport declined and blood glucose increased.(2)Resveratrol and 17 β-estradiol can significantly increase the expression of GLUT4 and IRS-1 in skeletal muscle and myocardial tissues,improve the transshipment of glucose,and have delay and preventive effects on the onset of mice diabetes mellitus.(3)The increase of the expression of GLUT4 and IRS-1 induced by resveratrol and 17 β-estradiol in diabetic mice skeletal muscle and myocardium may be associated with the inhibition of ERK protein phosphorylation and the decrease of p-ERK levels.Therefore resveratrol is expected to become a natural substitute for the prevention and treatment of diabetes mellitus. |
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