The Effect Of Capsaicin And Atorvastatin On Myocardium Remodeling

Background and objective:Despite the prominent progress in treatment of chronic heart failure, heart failure isstill a major cause of morbidity and mortality worldwide. Look for better drug andtreatment method remains a challenge in cardiology.Heart failure is a very common clinicalcardiovascular syndrome. Hypertension, coronary heart disease, myocardial infarction,arrhythmia, valvular abnormalities can all lead to heart failure.The study of cardiacstructure and function, has experienced quite a long period of time in heart failure. Until the1990s, people realized that the basis for the development of heart failure is ventricularremodeling. Ventricular remodeling is the determinants of morbidity and mortality inpatients with heart failure. Ventricular remodeling refers to gene expression,changes inquality between the molecules, cells and stormal cells when the heart damaged. Clinicalrealization of the changes in heart size, shape and function.The regulation of ventricularremodeling include RAAS, immunity,inflammation, mechanical stress, oxidative stress,high salt, high sugar, and so on. They can influence ventricular remodeling to a certainextent.Heart energy metabolism can be transformed into mechanical energy stored in the fattyacid and glucose in the chemical to provide energy for contraction and relaxation of theheart. Metabolic substrate from fatty acid to glucose utilization of the adaptive conversionplays an important role in the pathological process of myocardial remodeling. Abnormalglucose and lipid metabolism by affecting myocardial energy metabolism, and promotemyocardial remodeling. Salt is important environmental factors of hypertension, cardiachypertrophy. Salt metabolic abnormalities may contribute to myocardial remodeling byincreased oxidative stress.This study to explore the two aspects of the lipid and high salt onmyocardial remodeling.The peroxisomal proliferator-activator receptor co-activator1α (PGC-1α) is highly expressed in tissues with high metabolic rates, such as heart, muscle and brown adiposetissue. PGC-1α is indispensable for the heart to match the increased demand for ATP andwork output in response to various physiological stimuli, including stress and pressureoverload. Except for peroxisomal proliferator-activator receptorγ, PGC-1α also could reactwith other members of the nuclear receptor transcription factor superfamily and nonnucleartranscription factors to control diverse cellular energy metabolic pathways. For example,PGC-1α interacts with ERRα and activates the expression of pyruvate dehydrogenasekinase4(PDK4), which is a negative regulator of glucose oxidation. It has been reportedthat under the development of cardiac hypertrophy, which is characteristic by increasedutilization of glucose, the expression of PGC-1α and PDK4are repressed. UCP2is highlyexpressed in the myocardium and reduce the production of ROS and reduce oxidative stress.iNOS expression raised to increase myocardial fibrosis, myocardial nitration, and startmyocardial remodeling. PPARdelta can activation of UCP2to reduce the expression ofiNOS, reduce oxidative stress, and improve myocardial remodeling.The statins are widely used lipid-lowering drugs in the clinical, it was discovered thebenefit of statins is much larger than lipid-lowering effect itself. In the present,it has beendiscovered and confirmed the role include improved endothelial function,anti-inflammatory, antioxidant, the stable plaque, improve coagulation, to reduceneurohormonal activation, stimulate endothelial progenitor cell differentiation and reversalof myocardial remodeling, reduce bone resorption,anti-tumorpleiotropic and so on.Salt isan important environmental factors of hypertension, cardiac hypertrophy. Salt intake andthe prevalence of hypertension was a significant dose-effect relationship. The incidence ofhypertension in China was from north to south, may be around the eating habits are closelyrelated. Northern residents eating high-salt, Yunnan, Guizhou, Sichuan and other residentsin addition to eating salty, and hi hot meals. Capsaicin has been reported to be able to adjustthe high-salt-induced high blood pressure.Based on the above background, we propose the following scientific hypotheses:Atorvastatin through the regulation of PGC-1a, to improve the inflammatory status ofmyocardial glucose and lipid metabolism disorders, and thus reverse myocardialremodeling? Capsaicin activation of TRPV1to adjust the high-salt-induced blood pressure,while improve cardiac remodeling induced by high salt? Materials and Methods:Part one: The embryonic rat-heart-derived H9c2cells (Cell Bank, Chinese Academy ofSciences, Shanghai, China) were maintained in growth medium composed of DMEMsupplemented with10%fetal bovine serum. H9c2cells were plated at a density of5,000cells/cm2and allowed to proliferate in growth medium. Medium was changed every2days.After incubation at37°C in humid air (5%CO2and95%O2) for near confluence, the H9c2cells were then deprived of serum and incubated for another24h before TNF-α (100ng/mL)treatment. The present study is involved in vitro experiments. In vivo models include H9c2cells, H9c2+TNF-α, H9c2+TNF-α+Atorvastatin, siPGC-1α were also observed. Part two:The present study includes in vivo and in vitro experiments. In vivo models were C57BL/6Jwild-type (WT) mice and TRPV1-null mutant (TRPV1–/–) mice fed with interventionaldiet. In vitro models include cultured H9c2cells exposured to capsaicin. WT mice andTRPV1–/–were randomly grouped and fed with a normal salt (0.5%sodium, NS) diet, ahigh salt (8%sodium, HS) diet and a high salt plus capsaicin (8%sodium+0.01%capsaicin,HS+Cap) diet for1year beginning at8weeks of age.1. H9c2cardiomyocytes were incubated with tumor necrosis factor-α (TNF-α,100ng/mL) in the presence or absence of atorvastatin. Then, we calculated the glucoseoxidation of each group.2.Immunoblottings of PGC-1α were performed in H9c2cardiomyocytes stimulatedwith TNF-α (100ng/mL) in the presence or absence of different concentration atorvastatin(ATV,1-25μmol/L) for24h. H9c2cardiomyocytes were co-incubated with atorvastain andmevalonate, farnesol, geranylgeraniol in the presence of TNF-α, then investigate the proteinexpression of PGC-1α.3. Protein expression of PDK4and CPT1were conducted in TNF-α-stimulatedcardiomyocytes in the presence or absence of atorvastatin.4. Selective silencing of PGC-1α by RNA interference in H9c2cardiomyocytes wasperformed using a lentiviral system. Furthermore, Protein expression of PDK4and CPT1were conducted in siPGC-1α cardiomyocytes.5. Glucose oxidation was conducted in various groups, including control, GFP+TNF-α,siPGC-1α+TNF-α. 6. Expressions of TRPV1protein in H9c2cells and myocardium were detected byimmunoblotting. Distribution of TRPV1in H9c2cells was shown by immunofluorescence.7. Capsaicin induced [Ca2+]i change was examined by PTI systems. Capsaicininduced a concentration-dependent [Ca2+]i increase in H9c2cells, which was inhibited byiRTX pretreatment.8. Cardiac function was evaluated by echocardiogram.9HW/BW%and Collagen deposition were detected, and expression of nitrotyrosinewere determined by Masson trichrome, Superoxide anion in heart was detected bydihydroethidium (DHE).Cardiac hypertrophy was evaluated by hematoxylin/eosin stainsof different group histological sections.10Protein expressions of PPAR-δ, inos, ucp2in heart were examined by western blot.11. Protein expressions of PPAR-δ in H9c2cells was examined by western blot.Results:1. TNF-α induced a significant increase in glucose oxidation rate with respect tountreated H9c2cells.Whereas, co-incubation of TNF-α-stimulated H9c2cells withatorvastatin abolished this increase.2. Atorvastatin counteracts TNF-α-induced PGC-1αsuppression in a dose-dependentmanner in cardiomyocytes. The effect of atorvastatin on PGC-1α was almost abolished bymevalonate and partially by farnesol, but not by geranylgeraniol.3. The protein levels showed that TNF-α stimulation led to a significant reduction ofPGC-1α target genes, PDK4and CPT1, whereas these changes of metabolic genes weremarkedly inhibited in the presence of atorvastatin.4. Silence of PGC-1α suppressed PDK4and CPT1protein expressions incardiomyocytes.5. Silence of PGC-1α increases the glucose oxidation in TNF-α-stimulatedcardiomyocytes.6. TRPV1existed in both H9c2cells and mice myocardium.7. Capsaicin induced a concentration-dependent [Ca2+]i increase in H9c2cells, whichwas inhibited by iRTX pretreatment.8. Dietary capsaicin improves cardiac function in WT mice on long-term high-salt dietthrough TRPV1activation. However, capsaicin had no effects on TRPV1-/-mice. 9. Activation of TRPV1by dietary capsaicin attenuates cardiac hypertrophy andfibrosis on long-term high salt diet. But, capsaicin had no effects on TRPV1-/-mice.10. Activation of TRPV1by dietary capsaicin upregulates PPARδ, UCP2proteinexpression and decreases iNOS production in mice on long-term high-salt diet.11.TRPV1activation by capsaicin increases PPARδ expression in cardiomyocytes.Conclusions:1. TNF-α induced a significant increase in glucose oxidation rate with respect tountreated H9c2cells.Whereas, co-incubation of TNF-α-stimulated H9c2cells withatorvastatin abolished this increase. Furthermore, the protein levels showed that TNF-αstimulation led to a significant reduction of PGC-1α target genes, PDK4and CPT1,whereas these changes of metabolic genes were markedly inhibited in the presence ofatorvastatin.2. We employ the siRNA to knockdown of PGC-1α in H9c2cells. And investigate theprotein expression of PDK4, CPT1and the glucose oxidation rate in inflammatory status. Itshowed that PGC-1α silence significantly suppressed the protein expressions of PDK4andCPT1in cardiomyocytes. The glucose oxidation rate was markedly increased by PGC-1αsilence in TNF-α-stimulated H9c2cells.3. Atorvastatin inhibits TNF-α-induced glucose oxidation through PGC-1αup-regulation in cardiomyocytes, which might be associated with the regulation ofisoprenoid metabolites.4. Activation of TRPV1by capsaicin attenuates high-salt intake-induced oxidativestress, reduces production superoxide anion, and improves high-salt intake-inducedimpairment of cardiac function, alleviate high-salt intake-induced cardiac hypertrophy.5. Activation of TRPV1by dietary capsaicin upregulates PPARδ, UCP2proteinexpression and decreases iNOS production in mice on long-term high-salt diet. but not inthose of TRPV1-null mice. Vitro studies further support that capsaicin upregulates PPARδthrough TRPV1activation.6. Activation of TRPV1by long-term dietary capsaicin might play a potential role inthe prevention of the development of high-salt intake-induced cardiac hypertrophy. Chronicdietary capsaicin may represent a simple and effective lifestyle intervention in populationswith high-salt diet-induced hypertension and cardiac hypertrophy.