| According to the international diabetes federation?IDF?,there are currently 425 million diabetics in the world,and the number is expected to rise to 629 million by 2045.Type 2diabetes accounts for about 90%of total diabetes.The two main pathogenetic factors of type2 diabetes mellitus are insulin resistance and islet?cell failure.B group vitamins has certain effect in the treatment of type 2 diabetes mellitus and its complications,but its therapeutic mechanism is not clear.The RIN-GIPR cell model was constructed in this paper.To detect the effects of B vitamins on RIN-GIPR cells,and to detect the effect of B vitamins on?-cell autophagy using cellular immunofluorescence and western blot.The results are as follows:1.The plasmid pCMV6-AC-GIPR/GFP was transferred into RIN-m5F cells,and the monoclonal cell line was obtained by G418 screening.The cell line could be inherited stably by multiple passages.Under the fluorescence microscope,we can see that the distribution of green fluorescence is uniform.Fluorescence spots appeared in 20min treated with?D-Ala2?GIP.The results showed that the screening model of GIPR agonists was successfully constructed.The model was applied to evaluation of B vitamins,according to the results in B vitamins(1×10-5mol/L?1×10-6mol/L?1×10-7mol/L)on the cell?10 min,20 min,40 min?,fluorescence spots did not activate the model.The cells treated with GIP?D-Ala2?showed fluorescent spots in both 20min and 40min.The results showed that the action site of vitamins B ware not GIP receptor and had no effect on RIN-GIPR cells.2.Group B vitamins?4 concentration gradients each?were treated for 24 h.The results of cellular immunofluorescence assay showed that the macrophages induced by VitaminB1did not increase.The cell autophagy was significantly increased in VitaminB2,VitaminB3,VitaminB5,VitaminB6,VitaminB7,VitaminB9 and VitaminB12.Using western blot detecting autophagy related proteins and found VitaminB2,VitaminB3,VitaminB5,VitaminB6,VitaminB7,VitaminB9,VitaminB12 RIN-m5F cells after processing,its LC3?/LC3?ratio increases,P62 protein levels drop,AMPK phosphorylation levels rise,mTOR phosphorylation is changeless,that seven kinds of vitamins are AMPK islet beta cells induced by autophagy. |
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