Study On The Mechanism Of Tmao Aggravating Heart Failure In Patients With Hypertrophic Cardiomyopathy

Objective: Hypertrophic cardiomyopathy(HCM)is a common genetic myocardial disease,which is the primary cause of Sudden cardiac death(SCD)in adolescents and athletes.Patients with myocardial fibrosis and myocardial interstitial fibrosis resulting in decreased cardiac function and heart failure also increase the risk of sudden death.Sudden death often occurs suddenly,which is the difficulty of forensic cause of death.With the increase of clinical intervention in HCM patients,heart failure is becoming a common cause of death in HCM patients.Intestinal flora imbalance has been found to be closely related to the occurrence and development of cardiovascular diseases in recent years.This study mainly discussed the expression of intestinal flora metabolite Trimethylamine n-oxide(TMAO)in HCM patients as well as the effect on the structure and function of t tubules in cultured adult mouse cardiomyocytes.Methods: 1.TMAO concentration in plasma of HCM patients and healthy control group was detected by ELISA.TMAO concentration in plasma of 19 HCM patients and 20 healthy people was detected by ELISA kit,and the relationship between TMAO concentration and cardiac function was analyzed.2.T tubule confocal imaging and calcium ion imaging of adult mouse cardiomyocytes.Langendorff perfusion method was used to collect cardiac cells from adult mice.The cells were divided into the TMAO treatment group and the control group,respectively,and the t-tubule structure and the frequency of calcium spark were observed by confocal imaging.The integrity of t tubule was quantitatively analyzed by IDL image analysis program.3.The expression and aggregation of microtubulin in mouse cardiomyocytes were detected by western blotting and immunofluorescence.Western blotting was used to detect the expression of affinity protein 2(JPH2)and calpain I.The localization and expression of tubulin and JPH2 were detected by immunofluorescence.The structure of JPH2 was processed by using NIH image quantitative microtubule density and IDL image analysis program.Free and polymerized tubulin was detected using the tubulin/tubulin in vivo assay kit.4.Echocardiography in small animals.Mice were divided into TMAO feeding group and control group.The former was fed with 0.12% TMAO in the feed,while the control group was fed regular chow.Cardiac ultrasound was performed at 6 weeks.5.statistical analysis.The data were expressed as mean±SD.Univariate analysis of variance was used to compare the groups.T test was used to compare the two groups.The ROC curve was drawn in R language.SPSS 15.0 software was used for statistical analysis.P <0.05 was considered statistically significant.Results: 1.Plasma TMAO concentration of HCM patients was significantly higher than that of healthy control group.The expression of n-terminal b-type natriuretic peptide precursor in serum of HCM patients was significantly higher than that in healthy control group.ROC curve showed that the area under the TMAO concentration curve(AUC)of HCM patients group was 0.972.2.Within the detection concentration range,TMAO showed a concentration-dependent damage trend on the t tubule tissue of mouse myocardial cells.The concentration of TMAO at 3 mol/L and 10 mol/L significantly reduced the density of t tubule and the power value of t tubule.3.After 24 h of TMAO(3micron mol/L)culture of mouse cardiomyocytes,the average amplitude of calcium transient was significantly decreased.The time to reach the calcium transient peak and the dyssynchrony index increased significantly.The frequency of spontaneous calcium spark and the amplitude of calcium spark increased obviously.4.After 24 hours of TMAO treatment of mouse cardiomyocytes,the distribution of JPH2 in tissues was changed compared with the control group.In the TMAO treatment group,the power of JPH2 was decreased and obvious accumulation of JPH2 was observed at the edge of cardiomyocytes.5.Compared with the control group,the density of microtubule network was significantly increased after TMAO treatment.Western blot results showed that the aggregation of myocardial microtubules in mice treated with TMAO was significantly enhanced,suggesting that TMAO promoted the aggregation of myocardial microtubules.6.The results of cardiac ultrasound in mice showed that the cardiac systolic function of mice in the TMAO feeding group was significantly lower than that of mice in the normal feeding control group,and the left ventricular ejection fraction was also significantly lower,suggesting that TMAO could damage the cardiac function of mice.Conclusions: 1.Decreased cardiac function in patients with HCM is associated with TMAO concentration in vivo.2.TMAO impinges cardiac function by promoting tubulin polymerization,JPH2 translocation,and t tubule remodeling.