A High-fat Diet Impairs Mitochondrial Dynamics And The Respiratory Chain Complex In Rat Myocardial Tissues

BackgroundWith the improvement of people's living standards,great changes have taken place in the diet structure,gradually changing to the western diet.The Western Diet is rich in saturated fatty acids and cholesterol,and lacks of vitamins and dietary fiber.According to the latest "nutrition and health status of Chinese residents",high dietary energy,high fat intake and lack of exercise are closely related to the occurrence of overweight,obesity,diabetes and dyslipidemia.The intake of FAT(FAT),especially saturated trans fatty acids,is associated with increased risk of with obesity,diabetes and Cardiovascular Disease(CVD,Cardiovascular diseases)[1].In human trials,found that even a 25%-35%of total calories than fat energy,it could cause adipose tissue deposition,while in men from carbohydrate fat newborn ratio is low.This means that most of the lipid accumulation in the body comes from the diet[2l.Studies have shown that increased FA intake in the diet will affect the size of the cardiac esterified fatty acid pool and the composition of fatty acids.TG(triglyceride)content in cardiomyocytes increased with FA uptake.TG content in muscle cells is considered as an indicator of lipid toxicity.Nevertheless,the underlying molecular mechanisms between increased TG in the heart and changes in cardiac function are poorly understood.Unlike other tissue cells,myocardial cells mainly depends on FFA(free fatty acid oxidation of power,to generate ATP maintain myocardial cell shrinkage.The mitochondria plays a main role in the process.However,chronic exposure to excessive amounts of fatty acids and may lead to mitochondrial dysfunction and heart failure.Previous studies have focused on more,cause the release of ROS(ROS)lipid accumulation,make easily damage caused by ROS mitochondria want to participate in the apoptosis process.Mitochondrial quality control in maintaining normal cardiac function plays a key role.Mitochondrial dynamics,including mitochondrial fission and fusion mitochondria,mitochondrial quality control is the heart of the main regulatory mechanism.Mitochondrial dynamics not only needed to make cardiovascular mature,but also closely associated with a variety of CVD,In CVD,the structure and function of mitochondria undergo a series of changes[3].So whether a high-fat diet can lead to mitochondrial structure and function obstacle?Whether a high-fat diet induced cardiac function obstacle is through the dynamic unbalance of mitochondria way?Such studies have reported less.This subject combines previous clinical trials and epidemiological investigations to raise questions.A high-fat diet can lead to a build-up of myocardial lipids,which can lead to cardiac dysfunction.The function and structure of mitochondria were evaluated to elucidate the role of mitochondrial dynamics in cardiac dysfunction induced by high-fat diet in rats.To further explore the molecular mechanism of mitochondrial kinetics disorders in promoting the occurrence of cardiovascular diseases,and to provide a new idea for clinical targeted treatment of cardiac dysfunction diseases from the perspective of improving energy metabolism and mitochondrial kinetics.Objective:1?Whether a high-fat diet causes heart toxicity of fat(including cardiac lipid accumulation and cardiac function evaluation).2?Whether the number?shape and function of mitochondria is changed in high-fat diet induced cardiac function dysfunction.3?Whether mitochondrial dynamics associated with cardiac function obstacle.Research methods:In order to study on a high-fat diet on myocardial mitochondrial function and the effect of 40 SD rats were randomly divided into two groups,(1)normal diet group(n=20)feed containing 10%fat,70%carbohydrate,20%protein;(2)a high-fat diet group(n = 20)feed quantity of heat is 5.24 kcal/g,containing 60%fat,20%carbohydrate,20%protein.Building time of 28 weeks.After the test of myocardial lipid content were determined.Using the cardiac color doppler ultrasound to two groups to evaluate cardiac function in rats.Superfine projection electron microscope is used to assess mitochondrial morphology.Using high performance liquid chromatography(HPLC)method for rat myocardial adenosine metabolism in system evaluation,mitochondrial dynamics related gene quantitative polymerase chain reaction and western blot method used for determination.Mitochondrial proteomic analysis is used to study the effects of a high-fat diet the mitochondrial protein group.Results:1?A high-fat diet rats caused by myocardial lipid accumulation Pericardial fat tissue(PFT)was less in CON group,while that of HFD group was significantly increased,which was wrapped around the apex of the heart and the surface of coronary arteries.Compared with CON group(0.33 ± 0.047 g),and PFT in HFD group rats increased three times as heavy(0.95 ± 0.102 g),the difference was statistically significant.(as shown in figure 2.F).TG content in the HFD group left ventricular muscle(45.8 ± 9.3 umol/ug protein)is twice as much as CON group(21.3± 6.9 umol/ug protein)of more than two times,the difference was statistically significant(P<0.05).And Compared with CON group(0.18 ±0.04 nmol/ug protein),FFA content in HFD group rats myocardial(0.26 ±0.07 nmol/ug protein)increases(P<0.05).2?Cardiac structure and function change caused by high fat diet ratsGeneral view and organization weighing shows:compared with CON group,HM(heart mass,1130 ± 36.84 mg vs 1327 ± 25.62 mg,p<0.05)increased,the LVM(left ventricle mass,943.67 ± 33.29 mg vs 1127.58 ± 30,78 mg,p<0.05)increased in HFD group.The length of the femur was used as corrected,compared with CON group,HMI(heart mass index,24.77 ± 1.84 mg/mm vs.30.92 ± 1.57 mg/mm),LVMI(left ventricular mass index,20.49 ± 1.85 mg/mm vs 26.23 ± 2.08 mg/mm)also increased obviously(p<0.05)in HFD group.Echocardiography showed that the left ventricle in CON group was elliptical,the wall thickness was uniform and the motion was good.In HFD group,the cardiac cavity was enlarged,the myocardium was hypertrophic and the motion was weak.(as shown in figure 1.C-D).Statistical analysis shows that,compared with CON group,ventricular septal thickness(IVSd,2.16 ± 0.15 vs 2.48 ± 0.08 mm,P<0.05),end systolic left interior diameter(LVIDs,3.18 ± 0.32 vs 3.84 ± 0.52 mm,P<0.05),left ventricular posterior wall thickness(LVPWd,2.10 ± 0.10 mm vs 2.52 ± 0.25 mm,P<0.05),end systolic left ventricular posterior wall thickness(LVPWs,2.82 ±0.36 vs 3.30 ±0.25 mm,P<0.05)significantly increased,ejection fraction(EF,82.7 ±5.3 vs 73.3 ±2.0%),short axial shortening rate(FS,46.5 ±5.1 vs 37.3 ± 1.7%,P<0.05)significantly decreased in HFD group.3.A high-fat diet caused abnormal mitochondria structure,the number and function in ratsUltrastructure in myocardium were observed under transmission electron microscopy in the myocardial ultrastructure showed that myocardial CON group rats coarse wire and thin muscle wire arrangement rule,sarcomere is normal,Z line is clear,next to the muscle fiber bundle containing a large number of mitochondria,circular or elliptic,rendering the mitochondria double membrane structure integrity,mitochondrial crest structure are clear and complete within crest density is normal;HFD group of rats,the myocardial muscle thickness wire arrangement is not clear,sarcomere display is not clear,Z line disappear;Mitochondria arrangement is irregular,visible mitochondrial membrane structure is not complete.Some mitochondria length increases,even across multiple sarcomere,part of the mitochondrial swelling significantly,mitochondria crest sparse,matrix particles significantly reduced,some mitochondria crest fracture,dissolved,internal appear more bright area or vacuoles.Some mitochondria presents fragmentation,the structure is not complete.MtDNA copy number of rats in the HFD group was 39.1±6.1%of the mtDNA copy number in the CON group(P<0.05),indicating that the number of mitochondria in the myocardium of rats in the HFD group decreased significantly.To evaluate the efficiency of mitochondrial oxidative phosphorylation in,we tested the rats left ventricular myocardial cells mitochondrial Complex ?,Complex ?,Complex?(mitochondrial enzyme ?,?,?)and CS(citric acid)composite enzyme activity,the results show that compared with CON group of rats,HFD group Complex in the mitochondria of rats ?(19.5 ± 5.1 nmol/min·mgpro vs 6.2 ± 1.3 nmol/min·mgpro,P<0.05),the Complex ?(21.9 ± 4.1 nmol/min·mgpro 13 ± 1.4 v nmol/min·mgpro,P<0.05),ComplexIII(125.3 ± 5.7 nmol/min·mgpro vs 80.0 ± 8.4 nmol/min·mgpro,CS(P<0.05),and 371.2 ± 38.5 nmol/min·mgpro vs 244.0 ± 35.0 nmol/min·mgpro,P<0.05)activity decline,difference is statistically significant.Especially in Complex I activity decline significantly.Composite enzyme activity decreased,mitochondrial oxidative phosphorylation.To detect a high-fat diet influence on rat myocardial energy metabolism,we use the high performance liquid chromatography(HPLC)analysis in the rat myocardial ATP(adenosine triphosphate),ADP(adenosine diphosphate),AMP(AMP)content,and calculate the AMP/ATP,TAN(total adenosine phosphate),EC(Dutch).Analysis results show that,compared with CON group,TAN(5.97 ± 0.34 ug/ml vs.5.18 ±0.71ug/ml,P>0.05)remains unchanged,but the ATP(2.83 ± 0.4 ug/ml vs.1.78 ±0.45 ug/ml,ADP(P<0.05)and 2.50 ± 0.15 ug/ml vs.2.03 ± 0.23 ug/ml,P<0.05)content decreased in the HFD group,EC(0.67 ± 0.03 ug/ml vs.0.55 ± 0.05 ug/ml,P<0.05)decreased in the HFD group;Compared with CON group,concentration of AMPin HFD group was obviously higher(0.78 ± 0.14 ug/ml vs.1.3 ± 0.36 ug/ml,P<0.05),the ratio of AMP/ATP(0.26 ± 0.03 vs.0.75 ± 0.04,P<0.05)significantly higher in HFD group.A high-fat diet causes myocardial cell energy metabolism disorder as a whole.The detection of mitochondrial dynamics related gene mRNA showed:compared with CON group,the expression of mitochondrial fusion gene MFN2 mRNA,OPA1 mRNA,the expression of mitochondrial fission gene FIS 1 mRNA genes related to mitochondrial decreased in HFD group.The difference is statistically significant.We extract the left ventricular myocardial total protein and left ventricular myocardial mitochondria,dynamics of mitochondria related protein in Western Blot showed that compared with CON group,mitochondria fusion genes MFN1?MFN2?OPA1 protein expression decreased,mitochondrial fission related protein P-DLP?FIS expression increased,DLP expression unchanged in HFD group.And,obviously,In myocardial mitochondria,concentration of mitochondrial fusion proteins and mitochondrial fission proteins related to higher and more obvious changes?4?High fat diet make mitochondria of rats proteomics involved a series of metabolic substrates,oxidative phosphorylation related compound enzymes,transporters,mitochondrial ATP dynamic changes:(1)energy substrate related:fatty acid oxidation related protein expression decreased obviously;Aerobic oxidation of glucose and glycolysis related protein expression has a tendency to rise;The Krebs cycle related protein expression.That metabolic substrate by free fatty acids into glucose.The change of the substrate is also an important metabolic sign of heart?(2)protein involved in oxidative phosphorylation of the mitochondrial respiratory chain complex enzymes I,V related expression fall.Oxidative phosphorylation efficiency decline.(3)transport related protein subunits expression decreased ATP,the difference was statistically significant(P<0.05),such as ADP/ATP transferase,electron transfer flavoprotein-coenzyme Q oxidoreductase,electron transfer protein subunits alpha huang and so on.(4)mitochondrial dynamics related proteins:mitochondria fusion protein MFN2(difference was statistically significant,P<0.05)expression decrease MFN1,OPA1 expression.(there was no statistically significant difference,P>0.05);Mitochondrial division related FIS1 expression of albumen(difference was statistically significant,P<0.05).That reduce mitochondrial fusion,split,dysfunction of the mitochondrial dynamicsConclusion1?A high-fat diet of 28 w increases the rats in the pericardium and the myocardial content of TG,causes the rat myocardial FFA,increased levels of fatty group a drop in the cardiac remodeling and cardiac function in rats.2?Myocardial mitochondria ultrastructure observation found here were morphological abnormalities and structural damage in myocardial mitochondria of rats in HFD group;HFD cause myocardial mitochondrial copy number decrease;Composite enzyme in mitochondria ?,? compound enzyme,compound enzyme ?,citrate synthase activity decline;Under the condition of constant total adenosine,HFD group in the rat myocardial ATP content reduce,can be converted into ATP content of ADP,AMP content increasing,Energy Charge fell,the myocardial cell Energy metabolism disorder as a whole.HFD group of rats appeared myocardial cell mitochondrial structure and function disorder.3?HFD result in rats myocardial dysfunction of the mitochondrial dynamics,the mitochondria fusion and division of dynamic balance is upset,quality and quantity is difficult to maintain,mitochondria ATP produced shortage,energy depletion,cause cardiac function obstacle.4?Mitochondrial proteomics research,there were changes in energy metabolic substrates and decrease in electron transport chain-related proteins in high-fat diet group;Also found that the mitochondrial dynamics related proteins MFN1,MFN2,OPA1 expression decreased,FIS1 rise.Dysfunction of the mitochondrial dynamics,mitochondrial structure and the number of normal difficult to maintain.Mitochondrial dysfunction and impaired mitochondrial dynamics in the cardiac dysfunction induced by high fat plays an important role,this is a potential target for the treatment of heart disease.