Primary Study On The Association Of Expression Between SIRT1 And FOXO1 In Beta Cells Of Islet In Rats

As reported in most animal experiments, a fairly long term calorie restriction(CR) coulddramatically extend the life span of laboratory rodents, and improve their insulin sensitivity andinsulin resistance to lessen insulin secretion, so CR is much better perspective for the Type 2Diabetes Mellitus. The laboratory rodents experiments indicate that 30%～50% less calories ofnormal diet every day from the juvenile will never cause cacotrophy, however it could extendover 30% of the normal lifespan. Many studies in yeast and nematode reveal that the higherexpression of the silent information regulator 2 (SIR2) could be introduced by calorie restriction,and through which extends their life span, while the mammals do it by higher expression ofSIRTl-the mammalian homologous gene of SIR2.SIR2(silent information regulator 2) is a NAD~+-dependent histone deacetylase, which isthought as a longevity gene, and responsible for the modulation of the organism aging process,chromatosome silence, genetic transcription, genetic recovery and so on. FOXO family, themammalian homologous gene of DAF-16 in yeast, as the target molecule of Akt, is a key gene ofthe insulin and/or insulin-like growth factor(IGF)-like signalling (INS/IGF-1) pathway, and is amajor gene in the regulation of energy metabolism and cell senescence. The FOXOs arerelocated in cell cytoplasm from cell nucleus when they are repressed by phosphorylation of Akt,while also regulated by the deacetylase of SIR2. Many researches confirm that the signalINS/IGF-1 negatively regulates the gene DAF-16 to control the energy metabolism, organismgrowth and development in worms, however, in mammals the longevity gene SIRT1 indirectlyinforces the signal INS/IGF-1 by reducing the expression level of IGT-1 binding protein(IGFBP 1), or directly by its deacetylase activity for the FOXOs, to regulates the process of cellsenescence, and both suppress the FOXOs transcription activities to reduce the expression levelsof apoptosis and cell senescence related genes and improve the oxidative stress tolerance.T2DM is a common disease characterized with insulin resistance and defect secretion ofbeta cell of islet, and it has been quiet a serious problem for public health all over the world. Atpresent, we know little about the mechanism in the regulation of cell senescence of pancreaticislet cells in mammals, and the mechanism is not clear now. Therefore, a further study in the betacell senescence mechanism is much significant for the improvement in prevention and cure of the Type 2 Diabetes Mellitus.Previous experiments in our laboratory topic have show that cell senescence of pancreaticislet cells increases and insulin secretion reduces with aging. The clinic and epidemiologicsurvey also indicate the carbohydrate tolerance fall off and the morbidity rate of T2DM andIGT(impaired glucose tolerance) increase with aging. Animal studies also imply the beta cellsenescence is association with the morbidity of T2DM introduced by high fat diet in mousemodel. Therefore, we suppose that the longevity gene SIRT1 regulates the pancreatic islet cellssenescence through the downstream gene FOXO family members. We will introduce the highexpression of SIRT1 to reach the goal of delaying cell senescence by calorie restriction, anddetect the expression level of SIRT1 and FOXO1 by immunohistochemistry. Subsequently theimages collected from immunohistochemistry will be further analyzed by image handlingsoftware to explore the correlation between SIRT1 and FOXO1. So that, we can recognized thepathogenetic mechanism of T2DM.Content and Methods1. 11 male SD rats of 18 months old were randomized into two groups: 6 in calorie restriction(CR) group and 5 in the control group. The CR group is given 60% of calorie in nomaldiet while the control group nomal calorie diet, both were fed for 6 months.2. The cauda pancreatis paraffin section of the two groups SD rats were collected for beta-galstain and immunohistochemical stain of SIRT1, FOXO1 and insulin. And the correlationbetween the cell senescence and above indexes were analyzed.Result1. The protein SIRT1 was located specificly in the pancreatic islet cells. SIRT1 was located in thepancreatic islet cell cytoplasm and nucleus, mainly in the cytoplasm. CR group expresseshigher SIRT1 protein compared with the control group (P＜0.05).2. The transcription factor FOXO1 was also located specificly in the pancreatic islet cellcytoplasm and nucleus. The two groups have no difference on the expression of FOXO1, butthe expression level of FOXO1 located in the cell nucleus of CR group is lower than that ofthe control group (P＜0.01). 3. The insulin protein was expressed in the pancreatic islet cell cytoplasm, and the insulinexpression level of CR group was less than that of the control group (P＜0.05).4. The staining of beta-gal of the pancreatic islet cell presented blue, and located in the cellcytoplasm. Both two groups were positive, but CR group presented less stain compared withthe control group (P＜0.01).Conclusion1. Calorie restriction could delay the pancreatic islet cell senescence, and induce the highexpression of SIRT1 protein in the rat beta cell of islet.2. The transcription factor FOXO1 is a key molecule regulated the pancreatic islet cellsenescence process, and SIRT1 maybe delay senescence process of rat beta cell through therepression of transcription factor FOXO1 activity, so a further study in the beta cellsenescence mechanism is much significant for the improvement in prevention and cure of theType 2 Diabetes Mellitus.3. Calorie restriction may improve the organism insulin sensibility, and reduce the insulinsecretion of beta cell of islet.