Effects Of Treadmill Running And Rutin On Lipolytic Signaling Pathways And Endoplasmic Reticulum Stress In Diet Induced Obese Mice

Obesity and overweight are health issue globally. China is one of the countries in which obesity and overweight are growing fast. For now, there are over 300 million people facing the challenge of overweight or obesity in China. One of the most significant pathological changes under obese condition is the increase in white adipose tissue numbers and adipose tissue dysfunction. Adipose tissue dysfunction will lead to insulin resistance, consequently increases the risks metabolic chronic diseases such as type 2 diabetes. Thus, it is of importance to further explore effective strategies for improving adipose tissue dysfunction as well as the potential underlying mechanisms.Objective: We aimed to explore the effects of rutin and treadmill exercise intervention independently and in combination on high fat diet(HFD) induced disrupted lipolytic signaling, endoplasmic reticulum(ER) stress, adiponectin and TRPV4 related proteins in diet induced obese mice, and whether the depot-specific effects exist.Methods: Sixty C57BL/6J mice were randomized into 5 groups: chow group(mice fed with diet containing 10% fat, 70% carbohydrate and 20% protein), HFD(mice fed with diet containing 60% fat, 20% carbohydrate and 20% protein), HFD plus rutin intervention group(HR, mice fed with HFD diet with 0.1% rutin), HFD combined with treadmill running group(HE, mice fed with HFD diet and underwent treadmill training at a intensity of 15m/min, 60 min per day, 5 times/week), HFD combined with treadmill running and rutin intervention group(HRE). During the intervention, we weekly recorded mice’s weight and food intake. At the end of the 16 weeks’ intervention, we examined:1. Basic index and blood biochemical index : intraperitoneal glucose(GTT) and insulin tolerance tests(ITT) were performed, changes in glucose over time were plotted, and the area under the curve(AUC) was calculated for each. We also determined serum markers, including fasting blood glucose level(FBG), serum insulin level(FIns), serum adiponectin level, serum glycerol and free fatty acid content.2. Adipose tissue lipolysis : we incubated adipose tissue explants ex vivo, and isoproterenol stimulated lipolysis were determined. In vivo lipolytic markers from adipose tissue were measured by western blotting.3. Adipose tissue ER stress and adiponectin secretion: ER stress markers, that is GRP78、CHOP and p-JNK, were measured by western blotting; Adiponectin and PPAR-γm RNA expression were measured by real-time PCR.4. Adipocyte morphology and TRPV4 related proteins: we observed the size and morphology of adipocyte from both epididymal(EPI) and subcutaneous adipose tissue(SC) of each intervention group by hematoxylin-eosin staining method. TRPV4 related proteins were measured by western blotting.Results:1. Basic index and blood biochemical indexDuring the experiment, all groups’ body weight grew steadily, and HFD group’s mice were significantly heavier than CHOW group since the third week, while the body weight of HR, HE and HRE group had no significant change compared with CHOW, but were apparently smaller than HFD group(p<0.05). FBG and Fins of HFD group were significantly higher than CHOW group, other intervention methods decreased FBG and Fins at different degrees. Serum adiponectin levels of HFD, HR, HE, HRE group showed significant decrease compared with CHOW group. In GTT experiment, HRE group have significantly decreased AUC compared with HFD group, and in ITT experiment, HR, HE and HRE groups’ AUC decreased notably compared with HFD group.2. Adipose tissue lipolysisIn epididymal adipose tissue(EPI), HFD resulted in significant reduction in the phosphorylation of hormone sensitive lipase at serine660(p-HSL660) and perilipin A protein expression(p<0.05). Exercise and exercise plus rutin intervention completely restored p-HSL660 and perilipin A protein expression to normal level. Ex vivo rutin or exercise intervention restored blunted catecholamine stimulated lipolysis caused by HFD. In subcutaneous adipose tissue(SC), HFD had no effect on p-HSL660 and perilipin A protein expression(p>0.05).3. Adipose tissue ER stress and adiponectin secretionIn EPI, HFD resulted in induction of ER stress markers including GRP78, CHOP and p-JNK, reduction in adiponectin m RNA expression, PPAR-γ and Dsb A-L protein expression in vivo(p<0.05). HRE group had a significant decrease in GRP78 and p-JNK expression. HE and HRE group had no significant change in PPAR-γ and Dsb A-L protein expression compared with CHOW group(p>0.05).In SC, HFD resulted in induction of ER stress markers, including GRP78 and p-JNK, adiponectin and PPAR-γ m RNA expression, reduction in Dsb A-L protein expression in vivo,(p<0.05). HE and HRE group restored p-JNK and PPAR-γ protein expression to normal level.4. Adipocyte morphology and TRPV4 related proteinsIn EPI, HFD and HR group have reduced expression of PGC-1α+β, exercise and exercise plus rutin completely restored PGC-1α+β expression to normal level.In SC, an significant increase of TRPV4 protein expression was observed in HFD group, HE and HRE intervention decreased TRPV4 protein expression significantly compared with HFD group(p<0.05). HR, HE and HRE group showed a significant increase in PGC-1α+β protein expression compared with CHOW and HFD group(p<0.05).Conclusions:1. Rutin plus exercise intervention could improve high fat diet induced elevated FBG and Fins, as well as glucose intolerance and insulin intolerance.2. Rutin plus exercise intervention significantly improved lipolysis disorder in EPI caused by HFD.3. Rutin plus exercise intervention effectively improved ER stress status in EPI, while in SC, HR, HE and HRE intervention only reduced p-JNK protein expression.4. Exercise and rutin plus exercise intervention improved PPAR-γ and Dsb A-L protein expression caused by HFD in EPI, while in SC, HFD resulted in induction of adiponectin m RNA and PPAR-γ protein expression.5. In the aspect of TRPV4 and PGC-1α+β expression, an increase of TRPV4 was observed in subcutaneous adipose tissue and a decrease of PGC-1α+β in epididymal adipose tissue was observed under HFD. Exercise and rutin plus exercise could restore increased TRPV4 expression to normal level and increase PGC-1α+β.6. Depot-specific effects existed in regards to how interventions affect lipolytic signaling, in vivo ER stress markers, adiponectin m RNA expression, TRPV4 and PGC-1α+β expression.