Regular Exercise Regulates The Effect Of High-fat Diet-mediated Apolipoprotein B On The Heart Function And Exercise Capacity Of Drosophila

1 Research purposeApolipoprotein B is one of the lipid components of blood.It is a lipid-carrying protein.It mainly ingests low-density lipoprotein.It has a close relationship with cardiovascular diseases.Obesity is one of the important causes of cardiovascular disease,especially in the environment of systemic lipid metabolism disorders.At present,the heart has become an important organ for systemic lipid regulation.Although regular exercise can regulate lipid metabolism,the molecular mechanisms between them are still poorly understood.In this study,Drosophila model organisms were used,and by detecting Drosophila heart function,systemic triglyceride levels,oil red O staining,apoLpp mRNA expression level,and Drosophila climbing ability,to explore:（1）The effect of apoLpp inhibition on the heart function and exercise ability of Drosophila under different dietary conditions;（2）Regulation of regular exercise on the expression of apoLpp mRNA under different dietary conditions.Provide a theoretical basis for apoLpp to resist the cardiac function damage induced by high-fat diet.2 Research methodsThe female fruit flies of the UAS-apoLpp^RNAistrain were crossed with the male fruit flies of W¹¹¹⁸.The female fruit flies that emerged within 12 hours of the F1 generation were collected,and the flies were randomly divided into the following 4 groups:NFD+W¹¹¹⁸>apoLpp^RNAi（normal diet control group）,HFD+W¹¹¹⁸>apoLpp^RNAi（high-fat diet control group）,NFD+E+W¹¹¹⁸>apoLpp^RNAi（normal diet exercise group）,HFD+E+W¹¹¹⁸>apoLpp^RNAi（high-fat diet exercise group）.Each group contains 200 fruit flies.The female fruit flies of the UAS-apoLpp^RNAistrain were crossed with the male fruit flies of Hand-Gal4,the female fruit flies that emerged within 12 hours of the F1 generation fruit were collected,and the fruit flies were randomly divided into the following four groups:NFD+Hand>apoLpp^RNAi（normal diet Heart apoLpp knockdown group）,HFD+Hand>apoLpp^RNAi（high-fat diet heart apoLpp knockdown group）,NFD+E+Hand>apoLpp^RNAi（normal diet heart apoLpp knockdown exercise group）,HFD+E+Hand>apoLpp^RNAi（high Fat diet cardiac apoLpp knock-down exercise group）.Each group contains 200 fruit flies.The female fruit flies of the UAS-apoLpp^RNAistrain were crossed with the male fruit flies of Arm-Gal4.The female fruit flies that emerged within 12 hours of the F1 generation were collected,and the fruit flies were randomly divided into the following four groups:NFD+Arm>apoLpp^RNAi（normal diet Whole body apoLpp knockdown group）,HFD+Arm>apoLpp^RNAi（high-fat diet whole body apoLpp knockdown group）,NFD+E+Arm>apoLpp^RNAi（normal diet whole body apoLpp knockdown exercise group）,HFD+E+Arm>apoLpp^RNAi（high Fat diet whole body apoLpp knock-down exercise group）.Each group contains 200 fruit flies.When raising fruit flies,all fruit flies are placed in transparent glass tubes,20-30 per tube.NFD-fed fruit flies are placed in a HWS intelligent constant temperature and humidity incubator（25°C,50%humidity,12-hour day-night cycle）,HFD-fed fruit flies need to be reared at 21-22°C.Drosophila in the exercise group used Drosophila exercise device for exercise intervention every morning from 10:00 to11:30.Note:When the fruit flies of the high-fat exercise group undergo regular exercise intervention,the fruit flies need to be transferred to a clean glass tube and labeled,and then transferred to the original culture medium after the exercise is completed.Detection index:The collected female flies were first raised in NFD medium for 5 days,and then transferred to HFD medium for 5 days.At the same time,they carried out regular exercise for 5 days.At the 11th day of age,the fruit flies were taken out.Use M-mode to detect heart rate,cardiac cycle,diastolic interval,systolic interval,arrhythmia index,diastolic diameter,systolic diameter,fractional shortening,and fibrillary tremor to assess cardiac function.Real-time fluorescence quantification（q RT-PCR）was used to detect the relative expression of apoLpp mRNA in cardiomyocytes.The climbing index is used to evaluate the climbing ability of fruit flies.ELISA detects systemic triglyceride levels.Oil red O staining locates the expression status of lipids.3 Research results3.1 Drosophila M-mode test resultsUsing independent samples T test and two-way analysis of variance and LSD multiple comparison.Compared with the NFD+W¹¹¹⁸>apoLpp^RNAigroup,the NFD+E+W¹¹¹⁸>apoLpp^RNAigroup had no significant difference in HR（P>0.05）,no significant difference in HP（P>0.05）,no significant difference in DI（P>0.05）,and no difference in SI Significant difference（P>0.05）,AI was significantly reduced（P<0.05）,DD was not significantly different（P>0.05）,SD was not significantly different（P>0.05）,FS was significantly increased（P<0.01）,and FL was not significant Difference（P>0.05）.HFD+W¹¹¹⁸>apoLpp^RNAigroup HR significantly increased（P<0.001）,HP significantly decreased（P<0.001）,DI significantly decreased（P<0.05）,SI significantly decreased（P<0.05）,AI significantly increased（P<0.05）,DD was significantly reduced（P<0.01）,SD was significantly reduced（P<0.05）,FS was not significantly different（P>0.05）,and FL was significantly increased（P<0.001）.HFD+E+W¹¹¹⁸>apoLpp^RNAigroup has no significant difference in HR（P>0.05）,HP has no significant difference（P>0.05）,DI has no significant difference（P>0.05）,SI has no significant difference（P>0.05）,AI is significant Decrease（P<0.05）,DD significantly increased（P<0.001）,SD was not significantly different（P>0.05）,FS was significantly increased（P<0.001）,and FL was not significantly different（P>0.05）.Compared with the HFD+W¹¹¹⁸>apoLpp^RNAigroup,the HFD+E+W¹¹¹⁸>apoLpp^RNAigroup had a significant decrease in HR（P<0.001）,a significant increase in HP（P<0.001）,a significant increase in DI（P<0.001）,and a significant increase in SI High（P<0.01）,AI significantly decreased（P<0.01）,DD significantly increased（P<0.001）,SD significantly increased（P<0.001）,FS significantly increased（P<0.001）,FL significantly decreased（P<0.01）.Compared with the NFD+W¹¹¹⁸>apoLpp^RNAigroup,the NFD+Arm>apoLpp^RNAigroup had no significant difference in HR（P>0.05）,no significant difference in HP（P>0.05）,no significant difference in DI（P>0.05）,and a significant increase in SI（P<0.01）,AI was not significantly different（P>0.05）,DD was significantly increased（P<0.05）,SD was not significantly different（P>0.05）,FS was not significantly different（P>0.05）,FL was significantly increased（P<0.001）.Compared with the NFD+Arm>apoLpp^RNAigroup,the NFD+E+Arm>apoLpp^RNAigroup had a significant decrease in HR（P<0.01）,a significant increase in HP（P<0.05）,a significant increase in DI（P<0.01）,and a significant decrease in SI（P<0.01）,AI was significantly reduced（P<0.05）,DD was not significantly different（P>0.05）,SD was not significantly different（P>0.05）,FS was not significantly different（P>0.05）,FL was significantly reduced（P>0.05）<0.001).In the HFD+Arm>apoLpp^RNAigroup,HR was significantly increased（P<0.05）,HP was not significantly different（P>0.05）,DI was not significantly different（P>0.05）,SI was significantly decreased（P<0.001）,and AI was not significantly different（P>0.05）,DD was not significantly different（P>0.05）,SD was significantly reduced（P<0.05）,FS was significantly increased（P<0.05）,and FL was significantly reduced（P<0.05）.HFD+E+Arm>apoLpp^RNAigroup HR significantly decreased（P<0.001）,HP significantly increased（P<0.001）,DI significantly increased（P<0.001）,SI significantly decreased（P<0.001）,and no significant difference in AI（P>0.05）,DD was not significantly different（P>0.05）,SD was significantly increased（P<0.05）,FS was significantly decreased（P<0.05）,and FL was significantly decreased（P<0.001）.Compared with the HFD+Arm>apoLpp^RNAigroup,the HFD+E+Arm>apoLpp^RNAigroup had a significant decrease in HR（P<0.001）,a significant increase in HP（P<0.001）,a significant increase in DI（P<0.001）,and no significant SI Difference（P>0.05）,AI significantly decreased（P<0.05）,DD significantly increased（P<0.05）,SD significantly increased（P<0.001）,FS significantly decreased（P<0.001）,FL significantly decreased（P<0.001）.Compared with the NFD+W¹¹¹⁸>apoLpp^RNAigroup,there was no significant difference in HR in the NFD+Hand>apoLpp^RNAigroup（P>0.05）,no significant difference in HP（P>0.05）,no significant difference in DI（P>0.05）,and a significant increase in SI（P<0.05）,AI was not significantly different（P>0.05）,DD was significantly increased（P<0.01）,SD was significantly increased（P<0.01）,FS was not significantly different（P>0.05）,FL was significantly increased（P<0.05）.Compared with the NFD+Hand>apoLpp^RNAigroup,the NFD+E+Hand>apoLpp^RNAigroup significantly reduced HR（P<0.05）,HP significantly increased（P<0.05）,DI significantly increased（P<0.05）,and SI significantly decreased（P<0.05）,AI was significantly reduced（P<0.05）,DD was not significantly different（P>0.05）,SD was not significantly different（P>0.05）,FS was significantly increased（P<0.05）,and FL was significantly reduced（P<0.01）.HFD+Hand>apoLpp^RNAigroup HR significantly increased（P<0.01）,HP significantly decreased（P<0.05）,DI showed no significant difference（P>0.05）,SI significantly decreased（P<0.001）,AI significantly increased（P<0.05）,DD was not significantly different（P>0.05）,SD was not significantly different（P>0.05）,FS was not significantly different（P>0.05）,and FL was not significantly different（P>0.05）.HFD+E+Hand>apoLpp^RNAigroup has no significant difference in HR（P>0.05）,no significant difference in HP（P>0.05）,no significant difference in DI（P>0.05）,SI significantly decreased（P<0.001）,and no significant difference in AI There was no significant difference in DD（P>0.05）,no significant difference in SD（P>0.05）,no significant difference in FS（P>0.05）,and no significant difference in FL（P>0.05）.Compared with the HFD+Hand>apoLpp^RNAigroup,the HFD+E+Hand>apoLpp^RNAigroup has no significant difference in HR（P>0.05）,no significant difference in HP（P>0.05）,no significant difference in DI（P>0.05）,and significant difference in SI Decrease（P<0.001）,AI was significantly reduced（P<0.01）,DD was significantly reduced（P<0.01）,SD was not significantly different（P>0.05）,FS was not significantly different（P>0.05）,and FL was not significantly different（P>0.05）.3.2 The relative expression level of apoLpp mRNA in DrosophilaUsing independent sample T test and two-way analysis of variance and LSD multiple comparison.Compared with the NFD+W¹¹¹⁸>apoLpp^RNAigroup,the whole body apoLpp mRNA expression level in the NFD+E+W¹¹¹⁸>apoLpp^RNAigroup was significantly increased（P<0.001）;the heart apopp mRNA expression level in the HFD+W¹¹¹⁸>apoLpp^RNAigroup was significantly increased（P<0.001）;HFD+E+W¹¹¹⁸>apoLpp^RNAigroup has no significant difference in the expression level of apoLpp mRNA in the whole body（P>0.05）.Compared with the NFD+W¹¹¹⁸>apoLpp^RNAigroup,the whole body apoLpp mRNA expression level in the NFD+Arm>apoLpp^RNAigroup was significantlyreduced（P<0.001）.Comparedwiththe NFD+Arm>apoLpp^RNAigroup,the whole body apoLpp mRNA expression level in the NFD+E+Arm>apoLpp^RNAigroup was significantly increased（P<0.001）;there was no significant difference in the whole body apoLpp mRNA expression level in the HFD+Arm>apoLpp^RNAigroup（P>0.05）;there was no significant difference in the expression level of apoLpp mRNA in the whole body of the HFD+E+Arm>apoLpp^RNAigroup（P>0.05）.Compared with the HFD+Arm>apoLpp^RNAigroup,the relative expression level of apoLpp mRNA in the whole body of the HFD+E+Arm>apoLpp^RNAigroup had no significant difference（P>0.05）.Compared with the NFD+W¹¹¹⁸>apoLpp^RNAigroup,the whole body apoLpp mRNA expression level in the NFD+Hand>apoLpp^RNAigroup was significantly reduced（P<0.001）.Compared with the NFD+Hand>apoLpp^RNAigroup,the whole body apoLpp mRNA expression level in the NFD+E+Hand>apoLpp^RNAigroup was significantly increased（P<0.001）;there was no significant difference in the whole body apoLpp mRNA expression level in the HFD+Hand>apoLpp^RNAigroup（P>0.05）;there was no significant difference in the expression level of apoLpp mRNA in the whole body in the HFD+E+Hand>apoLpp^RNAigroup（P>0.05）.Compared with the HFD+Hand>apoLpp^RNAigroup,the HFD+E+Hand>apoLpp^RNAigroup had no significant difference in the expression level of apoLpp mRNA in the whole body（P>0.05）.3.3 Climbing ability test resultsUsing independent samples T test and two-way analysis of variance and LSD multiple comparison.Compared with the NFD+W¹¹¹⁸>apoLpp^RNAigroup,the climbing index of the NFD+E+W¹¹¹⁸>apoLpp^RNAigroup was not significantly different（P>0.05）;the climbing index of the HFD+W¹¹¹⁸>apoLpp^RNAigroup was significantly lower（P<0.01）;HFD+E The climbing index of+W¹¹¹⁸>apoLpp^RNAigroup had no significant difference（P>0.05）.Compared with the HFD+W¹¹¹⁸>apoLpp^RNAigroup,the climbing index of the HFD+E+W¹¹¹⁸>apoLpp^RNAigroup increased significantly.Compared with the NFD+W¹¹¹⁸>apoLpp^RNAigroup,the climbing index of the NFD+Arm>apoLpp^RNAigroup was significantly increased（P<0.01）.Compared with the NFD+Arm>apoLpp^RNAigroup,the climbing index of the NFD+E+Arm>apoLpp^RNAigroup was significantly increased（P<0.05）;the climbing index of the HFD+Arm>apoLpp^RNAigroup was significantly decreased（P<0.001）;HFD+E The climbing index of+Arm>apoLpp^RNAigroup had no significant difference（P>0.05）.Compared with the HFD+Arm>apoLpp^RNAigroup,the climbing index of the HFD+E+Arm>apoLpp^RNAigroup was significantly increased（P<0.001）.Compared with the NFD+W¹¹¹⁸>apoLpp^RNAigroup,the climbing index of the NFD+Hand>apoLpp^RNAigroup was significantly higher（P<0.05）.Compared with the NFD+Hand>apoLpp^RNAigroup,the climbing index of the NFD+E+Hand>apoLpp^RNAigroup was not significantly different（P>0.05）;the climbing index of the HFD+Hand>apoLpp^RNAigroup was significantly lower（P<0.001）;HFD+E The climbing index of+Hand>apoLpp^RNAigroup had no significantdifference（P>0.05）.Comparedwiththe HFD+Hand>apoLpp^RNAigroup,the climbing index of the HFD+E+Hand>apoLpp^RNAigroup was significantly increased（P<0.001）.3.4 Triglyceride test resultsUsing independent samples T test and two-way analysis of variance and LSD multiple comparison.Compared with the NFD+W¹¹¹⁸>apoLpp^RNAigroup,the systemic triglycerides of the NFD+E+W¹¹¹⁸>apoLpp^RNAigroup were significantly reduced（P<0.05）;the systemic triglycerides of the HFD+W¹¹¹⁸>apoLpp^RNAigroup were significantly increased（P<0.001）;HFD+E+W¹¹¹⁸>apoLpp^RNAigroup has no significant difference in systemic triglycerides（P>0.05）.Compared with the HFD+W¹¹¹⁸>apoLpp^RNAigroup,the systemic triglycerides of the HFD+E+W¹¹¹⁸>apoLpp^RNAigroup were significantly lower（P<0.001）.Compared with the NFD+W¹¹¹⁸>apoLpp^RNAigroup,the NFD+Arm>apoLpp^RNAigroup had significantly lower systemic triglycerides（P<0.001）.Compared with the NFD+Arm>apoLpp^RNAigroup,there was no significant difference in systemic triglycerides in the NFD+E+Arm>apoLpp^RNAigroup（P>0.05）;the systemic triglycerides in the HFD+Arm>apoLpp^RNAigroup were significantly increased（P<0.01）.There was no significant difference in systemic triglycerides in the HFD+E+Arm>apoLpp^RNAigroup（P>0.05）.Compared with the HFD+Arm>apoLpp^RNAigroup,the systemic triglycerides of the HFD+E+Arm>apoLpp^RNAigroup were significantly lower（P<0.05）.Compared with the NFD+W¹¹¹⁸>apoLpp^RNAigroup,the NFD+Hand>apoLpp^RNAigroup had significantly lower systemic triglycerides（P<0.05）.Compared with the NFD+Hand>apoLpp^RNAigroup,there was no significant difference in systemic triglycerides in the NFD+E+Hand>apoLpp^RNAigroup（P>0.05）.The systemic triglycerides in the HFD+Hand>apoLpp^RNAigroup were significantly increased（P<0.001）.There was no significant difference in systemic triglycerides in the HFD+E+Hand>apoLpp^RNAigroup（P>0.05）.Compared with the HFD+Hand>apoLpp^RNAigroup,the systemic triglycerides of the HFD+E+Hand>apoLpp^RNAigroup were significantly reduced（P<0.001）.3.5 Drosophila belly oil red O staining and morphological observationOil red O staining is an effective method to detect fat distribution and content.We performed Oil Red O staining on NFD-fed and HFD-fed flies.The results showed that the abdominal fat accumulation of NFD-fed flies was not obvious,while the HFD-fed flies accumulated a lot of abdominal fat.At the same time,from a morphological point of view,HFD-fed flies are fatter and have a more pronounced abdominal bulge than NFD-fed flies.4 Conclusion（1）Knockdown of apoLpp mRNA in Drosophila can resist heart function damage and low climbing ability caused by HFD.（2）Regular exercise reduces the expression of apoLpp mRNA,thereby resisting HFD-induced cardiac function damage and low exercise capacity.