| Objective:Obesity gradually becomes a global epidemic with the improvement of living standards.Obesity is a major risk factor for type 2 diabetes mellitus(T2DM).Therefore,weight control can help improve the complications of diabetes and delay the progression of the disease.Previous studies have confirmed that HIIT has a significant fat-reducing effect.Exercise reduces obesity by inducing browning of white fat and increasing energy consumption.The mitochondrial content of adipocytes is a key factor in increasing its oxygen consumption.Mitophagy plays an important role in regulating the number of mitochondria.However,the relationship between the effect of HIIT on adipocyte autophagy and browning remains unclear.Therefore,this paper explores the effect of HIIT on adipose tissue mitophagy and browning by establishing a T2 DM mouse model,and provides a reliable theoretical basis and new therapeutic targets for supporting the clinical application of HIIT in the treatment of T2 DM.Methods:4-week-old C57BL/6J male mice(N=40)were selected and fed adaptively with normal standard diet for 1 week,and then randomly divided into normal diet control group(CON group,N=10)and high-fat diet group(HFD group),N=30).The HED group continued to maintain high-fat feeding and received intraperitoneal injection of streptozotocin(STZ)after 12 weeks to induce T2 DM,and then were randomly divided into the diabetic quiet group(SED,N=10)and the diabetic high-intensity exercise group(HIIT,N=10).During the 8-week intervention period,the HIIT group received high-intensity interval training,and neither the SED group nor CON involved exercise intervention.After the intervention,(1)the epididymal adipose tissue of the mice was taken and weighed,and the tissue was embedded in paraffin for HE staining to observe the effect of HIIT on the morphology of epididymal adipose tissue cells;(2)UCP1 antibody was used for immunohistochemical staining to observe the effect of HIIT on adipose tissue browning;(3)Fluorescence quantitative PCR was performed on the total RNA of epididymal adipose tissue to detect insulin signaling pathway-related factors(PI3K,AKT,IRS-1,GLUT4),browning-related genes(PRDM16,UCP1,DIO-2,PPAR,CEBP),autophagy-related genes(ATG7,Parkin,P62,Beclin),mitochondrial synthesis-related genes(Sirt1,PGC-1α,Nrf-1),mitochondrial thermogenesis genes(Cidea,COX8 b,Elovl3),inflammatory cytokines(IL-4,IL-6,IL-10,TNF-α)and oxidative stress-related factors(Prdx1,Sod1,NOX2,NOX4).Results:(1)Establishment of T2 DM mouse model induced by high fat diet combined with STZ injection After feeding C57 BL / 6J mice with high-fat diet for 12 weeks,the weight of HFD group increased significantly compared with con group(P<0.01).After intraperitoneal injection of STZ,the fasting blood glucose of HFD group was significantly higher than that of con group(P<0.01).After intraperitoneal injection of glucose and insulin,the fasting blood glucose of HFD group was always higher than 16.7mmol/l,The area under the curve(AUG)was significantly higher than that in con group(P<0.05).Therefore,the T2 DM mouse model was successfully established.(2)The effect of HIIT on insulin signaling pathway in T2 DM mice Compared with the CON group,the m RNA expressions of GLUT4,IRS-1 and AKT,the main genes of insulin signaling pathway in the SED group were significantly decreased(P<0.05),while the PI3 K gene expression in the SED group did not reach a significant difference(P<0.05),but there was still a downward trend;compared with the SED group,the m RNA expressions of GLUT4,IRS-1,AKT,and PI3 K in HIIT were significantly increased(P<0.05).(3)The effect of HIIT on body composition in T2 DM mice After 8 weeks of high-intensity interval exercise,compared with the mice in the SED group,the body weight and body fat percentage of the mice in the HIIT group were significantly decreased(P<0.05).After 8 weeks of high-intensity intermittent exercise,there was no significant difference in fasting blood glucose between the HIIT group and the SED group(P<0.05),and the area under the curve of the glucose tolerance test and insulin test were significantly lower than those in the SED group(P<0.05).(4)The effect of HIIT on adipocyte morphology in T2 DM mice The results of HE staining showed that,compared with the CON group,the SED group had fat compartments with large vacuoles,and the shape was irregular,the nucleus,organelles and cytoplasm were squeezed to one side to form a thin ring,and the characteristics of white fat were obvious;compared with the SED group,HIIT group mouse e WAT cells appeared multi-chambered,the cells became smaller and rounder,the tissue became more dense,the number of cells increased significantly,and there was a tendency of browning.In BAT,mice in the SED group exhibited larger,single vacuolar fat compartments with reduced cell numbers.Compared with the CON group,the average area of fat in the SED group was significantly increased(P<0.01),and compared with the SED group,the average area of fat in the HIIT group was significantly decreased(P<0.01).In the frequency map of adipocyte size distribution,it was shown that the HIIT group had the highest frequency of adipocyte size in the range of 0-1000 μm2.(5)The effect of HIIT on the browning of white adipose tissue in T2 DM mice Immunohistochemical results showed that the expression of UCP1 protein in epididymal adipose tissue in the HIIT group was significantly increased,and the positive staining was brownish-yellow granular;while the CON group and the SED group hardly expressed UCP1.Consistent with the results of immunohistochemistry,RT-PCR results showed that compared with CON group,CEBP(P<0.01),DIO-2(P<0.05),PRDM-16(P<0.01),UCP1(P<0.05)in SED group were significantly decreased.Although there was no significant difference between PPAR-α and CON group,compared with SED group,the expression of PPAR-α in HIIT group was significantly increased(P<0.05).In addition,the expressions of DIO-2,PRDM-16 and UCP1 in the HIIT group were significantly increased(P<0.05).(6)The effect of HIIT on mitochondria in T2 DM mice RT-PCR results showed that compared with the CON group,in addition to the significant decrease in the m RNA expression of P62(P<0.05),the expressions of other mitophagy-related genes ATG7,Beclin and Parkin in the SED group were significantly increased(P<0.05).Compared with the SED group,the expressions of ATG7,Beclin and Parkin in the HIIT group were significantly decreased(P<0.05),and the expression of P62 was significantly increased(P<0.05).Compared with the CON group,the mitochondrial thermogenesis-related genes Cidea,COX8 b and Elovl3 in the SED group were significantly decreased(P<0.05);compared with the SED group,the Cidea,COX8 b and Elovl3 in the HIIT group were significantly increased(P<0.05).In addition,the mitochondrial biosynthesis-related indicators PGC-1α,Sirt1,and Nrf-1 in the SED group were significantly lower than those in the CON group(P<0.05);Compared with the SED group,the m RNA expressions of PGC-1α,Sirt1,and Nrf-1 were increased significantly in the HIIT group(P<0.05).(7)Correlation between visceral adipose tissue p62 and UCP1 The correlation between visceral fat browning marker gene UCP1 and mitochondrial autophagy genes parkin and p62 in T2 DM mice was analyzed.The results showed that parkin was moderately negatively correlated with the expression of UCP1 m RNA(r =-0.58,P = 0.012),and p62 was moderately positively correlated with the expression of UCP1 m RNA(r = 0.7553,P = 0.0003)(8)The effect of HIIT on adipose tissue inflammation in T2 DM mice RT-PCR results showed that compared with the CON group,except for IL-4 and IL-6,there was no significant difference(P>0.05);the inflammatory factors IL-10(P<0.01)and TNF-α/(P<0.05)was significantly increased;compared with the SED group,the levels of IL-4(P<0.05),IL-6(P<0.05),IL-10(P<0.05)and TNF-α(P<0.01)m RNA expression were significantly reduced.(9)The effect of HIIT on oxidative stress in adipose tissue of T2 DM mice Compared with the CON group,the NOX4 in the SED group increased significantly(P<0.05),although there was no significant difference in NOX2,but there was an obvious upward trend;Prdx decreased significantly(P<0.05),although there was no significant difference in Sod1,there was A clear downward trend.Compared with the SED group,NOX2 and NOX4 in the HIIT group were significantly decreased(P<0.01);Sod1 and Prdx were significantly increased(P<0.05).Conclusions:(1)High intensity intermittent exercise inhibited the mitochondrial autophagy of white fat in T2 DM,increased the number of mitochondria,fatty acid oxidation,heat production and energy consumption,decreased the level of oxidative stress,inflammation and insulin sensitivity.(2)High intensity intermittent exercise promoted the browning of white fat and the increase of UCP1 expression in T2 DM,which may be related to the decrease of mitochondrial autophagy and the increase of mitochondrial content in white adipose tissue... |
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