The Effect Of Combined Intervention Of Time-restricted Feeding(TRF) And Exercise On Glucose Homeostasis And Hepatic Glycolipid Metabolism In Mice Fed With High-fat Diet

Background:In recent decades,the prevalence of obesity in the global population has increased significantly,and the average body mass index(BMI)of adults has increased,which is a worrying trend.In addition,with the development of society,the emergence of night shifts and nightlife has greatly changed people’s eating patterns,and eating at night has increased the incidence of obesity.Obesity is a major risk factor for a range of chronic diseases such as diabetes,cardiovascular disease and cancer,suggesting that obesity has developed into an extremely serious public health problem.The root cause of overweight and obesity is that calorie intake is greater than calorie consumption.Therefore,increasing calorie expenditure or decreasing caloric intake has been the traditional way to lose weight and improve the metabolic health of patients.Exercise achieves weight loss and improves metabolism by increasing calorie consumption;Caloric restriction(CR)has also been proven to delay aging and increase human life.However,it is difficult for people to adhere to a strict CR strategy for a long time.The time restricted feeding(TRF)came into being.TRF is a novel dietary intervention that refers to the control of the eating window below 12 hours,beyond which the body fasts.It alleviates obesity and improves blood sugar levels by limiting daily eating times.However,TRF intervention may bring some negative effects,such as: studies have pointed out that TRF is insufficient in weight loss and elimination of systemic inflammation;Whether the weight loss effect is long-term remains to be explored;And TRF alone may cause loss of appetite,anxiety,etc.Moderate exercise can not only reduce weight,but also prevent or treat anxiety disorders,which is beneficial to physical and mental health.Therefore,this study combines exercise and TRF strategies,explores the comprehensive effects of the two on the glucose and lipid metabolism of metabolic diseases,and explores whether the combination of TRF and exercise has a synergistic effect on metabolic improvement.Objective: The purpose of this study is to explore the effect and effect of TRF strategy combined with moderate intensity exercise on glycolipid metabolism in mice on a high-fat diet(HFD),and to explore the possible physiological mechanisms.Based on this study,a simple and implementable weight loss strategy was proposed,in order to provide a theoretical basis for non-drug treatment methods for obese people.Methods: 60 male C57BL/6J mice aged 5-6 weeks old were purchased and placed in the SPF-level animal feeding room,with an indoor temperature of 22±2°C,air humidity of 50±10%,12 h day and night cycle,feeding and drinking according to groups.After 3 days of adaptive feeding with conventional feed,the mice were randomly divided into six groups(n=10): regular diet + 24 h feeding group(C),regular diet + 8h feeding group(TC),45% high-fat diet + 24 h feeding group(H),45% high-fat diet + 8h feeding group(TH),45% high-fat diet + 24 h feeding + exercise group(HE),45% high-fat diet + 8h feeding + exercise group(THE).Among them,the C and TC groups were fed with ordinary feed;The remaining four groups were fed a 45% highfat diet.Mice in the C,H and HE groups were fed freely and watered.In addition,TRF intervention was performed on the TC,TH,and THE groups during the dark cycle.The specific TRF scheme is as follows: under the 24-hour system,from 23:00 to 07:00 every day,mice are free to eat and take water,and the rest of the time is fasted and free to take water.During animal feeding,mice are weighed weekly and their feed intake is measured daily.The HE and THE groups underwent 12 weeks of moderate-intensity treadmill exercise;The remaining four groups maintained a quiet lifestyle for 12 weeks.The specific scheme is as follows: the use of small animal treadmills.The mice in the exercise group used 1 week to adapt to the treadmill,exercise for 4 days a week,the speed gradually increased from 6 m/min to 13 m/min,and the duration gradually increased from 15 min to 45 min.From the second week,the mouse training parameters were: 13m/min,60min/day,exercise 5 days a week,rest on Wednesday and Sunday,and the training time lasted 12 weeks.Training time is 08:00-10:00 in the morning.Daily food intake and water intake records of mice;body composition tests before and after exercise interventions;Mice are tested for intraperitoneal insulin tolerance one week before sacrifice.After the experiment,the mice were sacrificed by necking,serum,liver,adipose tissue,skeletal muscle,etc.,and polymerase chain reaction(PCR)and western blot(WB)experiments were carried out according to the experimental requirements.After obtaining the data,Graph Pad Prism 9.0 was used to perform data statistics using two-way ANOVA.Results:(1)Effects on body weight and food intake.Compared with group C,lean body mass in group H was significantly decreased(P<0.0001),fat content was significantly increased(P<0.0001)and body weight was significantly increased(P<0.0001).Compared with TC group,lean body mass in TH group was significantly decreased(P<0.0001),fat content was significantly increased(P<0.0001),and body weight was significantly increased(P<0.05).Compared with group H,lean body mass was significantly increased(P<0.001),fat content was significantly decreased(P<0.001),and body weight was significantly decreased(P<0.0001)in group TH.Compared with group H,lean body mass was significantly increased(P<0.01),fat content was significantly decreased(P<0.01)and body weight was significantly decreased(P<0.0001)in group TH,and lean body mass was significantly increased(P<0.0001),fat content was significantly decreased(P<0.01)and body weight was significantly decreased(P<0.001)in group HE.Compared with TH group,lean body mass in THE group was significantly increased(P<0.01)and fat content was significantly decreased(P<0.01).Compared with HE group,lean body mass in THE group was significantly increased(P<0.05),fat content was significantly decreased(P<0.05)and body weight was significantly decreased(P<0.05).At the early stage of the experiment(1-4 weeks),the weekly caloric intake of mice in each group had no significant change.The weekly caloric intake of each group at the middle stage of the experiment(5-8 weeks)was as follows.Compared with group H,weekly caloric intake in group TH was significantly decreased(P<0.001).Compared with group H,weekly caloric intake in TH group was significantly decreased(P<0.01).Compared with HE group,weekly caloric intake in THE group was significantly decreased(P<0.05).The weekly caloric intake of each group in the later period of the experiment(9-12 weeks)was as follows.Compared with group C,weekly caloric intake in TC group was significantly decreased(P<0.05).Compared with group H,the weekly caloric intake in group TH was significantly decreased(P<0.01).Compared with group H,the weekly caloric intake in group TH was significantly decreased(P<0.01).(2)Effects on glucose metabolism.FBG level: In the initial stage,compared with group C,fasting glucose level was significantly increased in group TC(P<0.01)and group H(P<0.0001).Compared with group H,blood glucose level in HE group was significantly decreased(P<0.0001).Compared with TH group,fasting blood glucose level in HE group was significantly decreased(P<0.001).Compared with HE group,blood glucose level in THE group was significantly increased(P<0.05).In the final stage,compared with group H,blood glucose in HE group was significantly decreased(P<0.01);Compared with TH group,blood glucose level in HE group was significantly decreased(P<0.01).Area under ITT curve: Compared with group C,the area under H suite was significantly increased(P<0.001);Compared with group H,the area under TH suite was significantly reduced(P<0.05).Compared with group H,the area under ITT curve of mice in HE group was significantly decreased(P<0.001).Compared with HE group,THE area under ITT curve in the group was significantly increased(P<0.05).Gluconeogenetic gene: Compared with H group,the expression level of PCK1 gene in TH group was significantly decreased(P<0.05).Compared with H group,the expression of liver PCK1 gene in TH group and HE group was significantly decreased(P<0.05).Compared with TH group,G6 PC gene expression in THE group was significantly decreased(P<0.05).Compared with HE group,G6 PC expression in THE group showed a significant trend of decline(P=0.0552).(3)Effects on lipid metabolism.Compared with group C,In group H,serum total cholesterol level was significantly increased(P<0.0001),serum triglyceride was significantly increased(P<0.01),serum low density lipoprotein was significantly increased(P<0.0001),serum alanine aminotransferase level was significantly increased(P<0.01),liver CD36 expression was significantly increased(P<0.05),liver CPT1 b m RNA and protein expression were significantly decreased(P<0.01),Pgc1 a gene expression level was significantly decreased(P<0.05),PER2 expression was significantly decreased(P<0.05),Pgc1 a protein expression level was significantly increased(P<0.05).The lipid drop area and CPT1 A protein expression in TC group were significantly decreased(P<0.01);Compared with TC group,the serum total cholesterol level in TH group was significantly increased(P<0.01),the serum low density lipoprotein level was significantly increased(P<0.05),the mouse fat drop area was significantly increased(P<0.01)and the Pgc1α protein expression was significantly increased(P<0.05).Compared with group H,In TH group,serum total cholesterol level was significantly decreased(P<0.0001),serum triglyceride was significantly decreased(P<0.05),serum low density lipoprotein was significantly decreased(P<0.0001),serum glutamic oxalacetic transaminase and glutamic-pyrulol transaminase levels were significantly decreased(P<0.05),liver CD36 expression was significantly decreased(P<0.01),PE R2 expression was significantly increased(P<0.01).Compared with group H,In TH group,serum total cholesterol level was significantly decreased(P<0.001),serum triglyceride was significantly decreased(P<0.05),serum low density lipoprotein was significantly decreased(P<0.001),serum glutamic oxaloacetic transaminase and glutamic propylaminotransferase levels were significantly decreased(P<0.05),liver CD36 expression was significantly decreased(P<0.01),liver Pg The expression of c1 a and PER2 were significantly increased(P<0.05).In HE group,serum total cholesterol level was significantly decreased(P<0.05),serum low density lipoprotein was significantly decreased(P<0.001),mouse lipid drop area was significantly decreased(P<0.05),liver CD36 expression was significantly decreased(P<0.01),and Pgc1α protein expression was significantly increased(P<0.05).Compared with TH group,PER2 expression and CPT1 A protein expression in liver of HE group were significantly decreased(P<0.01),CPT1 b m RNA and protein expression in liver of THE group were significantly decreased(P<0.05)and PER2 expression was significantly decreased(P<0.05).Compared with HE group,THE expression level of Elovl3 gene in the group was significantly decreased(P<0.05),and the expression of CPT1 b gene was significantly decreased(P<0.01).Conclusion:(1)In the high-fat diet environment,TRF intervention can reduce body weight,regulate lipid levels and reduce the degree of liver injury;Moderate intensity exercise intervention can reduce body weight,improve dyslipidemia and reduce liver fat deposition.Compared with TRF alone,exercise intervention has better effect on blood glucose control in HFD mice than TRF alone.TRF combined exercise intervention can reduce weight.On the one hand,the effect of weight loss is more prominent than that of exercise intervention alone;on the other hand,the effect of fat reduction and muscle increase is better than that of the two independent actions.(2)TRF or exercise intervention can reduce the expressions of gluconeogenic genes G6 PC and PCK1 and thus improve glucose metabolism in the high-fat diet environment.The improvement of lipid metabolism in high-fat diet mice by TRF or exercise intervention was not due to the activation of PPARα-Pgc1α signaling pathway,but might be achieved through other pathways.The changes of PER2 gene expression in liver suggest that the next step should be to explore its internal mechanism through the pathways of rhythm and nutrition perception.