Effects Of Follic Acid On Endothelial Function Of Thoracic Aortea And Myocardioal Aroptosis In Rats With Type 2 Diabetes Mellitus

Objective:(1)To develop an ideal rat model of type 2 diabetes mellitus.(2)To investigate the effect of long-term folic acid supplementation on endothelial function of thoracic aortea in rats with type 2 diabetes mellitus and explore its mechanisms.(3)To study the effect of long-term folic acid supplementation on myocardial apoptosis in rats with type 2 diabetes mellitus and explore its mechanisms.Methods:(1)The rat model of type 2 diabetes mellitus was established by a single injection of low dose streptozotocin (STZ) via tail vein after 6 weeks of high calorie diet in 51 rats. The other 11 healthy rats were employed as normal control group (NC group). The oral glucose tolerance, fasting blood glucose, fasting serum insulin and other corresponding indexes were measured to evaluate the quality of the model.(2)The rats with type 2 diabetes mellitus were randomly divided into model control group (MC group), low-dose folic acid group (LDF group)and high-dose folic acid group (HDF group).11 healthy rats in NC group were chosen to compare with. After 11 weeks of folic acid supplementation, blood of the rats was collected for determination of the levels of NO, SOD and MDA in the serum. The thoracic aortea were dissected from the rats and cut into 3 mm rings for measurement of vascular reactivity.Acetylcholine-induced endothelium-dependent vascular relaxation (EDVR) of aorta rings was tested. The hearts were harvested as well. Terminal deoxynuleotidyl transferase-mediated dUTP nick end labeling (TUNEL) and StreptAvidin-Biotin Complex (SABC) immunohistochemical staining were used respectively to detect myocardial apoptosis and the expression of Bcl-2, Bax and Fas genes in the myocardium.Result:(1)After 6 weeks of high calorie diet,the body weight and fasting serum insulin level of the rats were obviously higher and the insulin sensitivity index(ISI) was markedly lower than those of the rats fed with normal diet.There was no significant difference in the level of fasting blood glucose and oral glucose tolerance between the two group.1 week after injection of STZ,the rate of model making was 76.47%,the mortality was 13.73% and the rate of spontaneous amelioration was 5.13%.After 16 weeks,there was no significant difference in the body weight and fasting serum insulin level between the rats in NC group and the rats with diabetes mellitus,but the level of fasting blood glucose in the rats with diabetes mellitus was obviously higher and ISI was markedly lower than those of the rats in NC group. (2) After 12 weeks of type 2 diabetes mellitus,the levels of NO and SOD in serum were significantly lower and the level of MDA in serum was notably higher in MC group than those in NC group;After 11 weeks of folic acid supplementation, compared with MC group,the levels of NO and SOD in serum significantly increased in LDF group and HDF group,while the level of MDA in serum decreased markedly.(3)The results of isolated aorta rings'reactivity showed:Compared with NC group, EDVR responses to Ach of each concentration were obviously weakened in MC group.Compared with MC group,EDVR responses to Ach of each concentration were significantly improved in LDF group and HDF group,and further increment of folic acid leaded to no further improvement of EDVR response.(4)The results of TUNEL assay showed:Compared with NC group,the rate of myocardial apoptosis increased notably in MC group,LDF group and HDF group. Compared with MC group,the rate of myocardial apoptosis decreased markedly in LDF group and HDF group,and further increment of folic acid leaded to further decrease of myocardial apoptosis rate.(5)The results of immunohistochemistry test showed:Compared with NC group,ODs of Bcl-2,Bax and Fas protein significantly increased in MC group, LDF group and HDF group.Compared with MC group,ODs of Bcl-2 protein increased markedly in LDF group and HDF group,and further increment of folic acid leaded to further increase.Compared with MC group,ODs of Bax and Fas protein significantly decreased in LDF group and HDF group,and further increment of folic acid leaded to further decrease.Conclusion:(1)This rat model of type 2 diabetes mellitus owns the character of insulin resistance which resembles the clinical phenotype of patients with type 2 diabetes . mellitus.It is an ideal animal model of type 2 diabetes mellitus.(2)EDVR response is impaired in the rat with type 2 diabetes mellitus,and the decrease of NO activity and oxidative stress may be related to the impairment. Long-term folic acid supplementation can effectively prevent the impairment.The effect may be associated with the increase of NO formation and its antioxidant effect.(3)The rate of myocardial apoptosis increases in the rat with type 2 diabetes mellitus,and it may be regulated by the Bcl-2, Bax and Fas genes. Oxidative stress may be an imporant cell death signal in the diabetic heart.The decrease of NO activity may play a key role in the development of diabetic myocardial apoptosis. Long-term folic acid supplementation can effectively inhibit myocardial apoptosis in the rat with type 2 diabetes mellitus.It can up-regulate the expression of Bcl-2 gene and down-regulate the expression of Bax and Fas genes in the diabetic myocardium.The mechanism that folic acid inhibits myocardial apoptosis in the rat with type 2 diabetes mellitus may be associated with the increase of NO formation and its antioxidant effect.