Study On Regulatory Effect Of Enalapril On TNF-α、Th1/Th2、Treg/Th17of Post Myocardial Infraction In Mice Which Was Induced By A Novel Model Method

Objective: The purpose of this study was to introduce and comprehensively describe a novel and rapid surgical method to induce MI and directly compare it to the classic method.we use this new method to establish MI model in mice for investigation of the regulatory effect of enalapril on TNF-α、Th1/Th2、Treg/Th17post myocardial infraction.Methods:176Male C57/B6mice were grouped into4groups at random: new method MI (N-M,n=86) or sham (N-Sham,n=20) and classic method MI (C MI,n=50) or sham (C-Sham,n=20). In the new method, hearts of total86were manually exposed through a small incision of the left fourth intercostal space MI was induced. In the classic method, MI was induced through a thoracotomy. In group of MI, the left anterior descending coronary artery was ligated. Hearts from group of Sham were without ligation. To investigate the inflammatory damage/response of these2methods, the plasma level of TNF-awas measured at3days、7days、14days and28days after surgery. Echocardiography was used as noninvasive method to measure cardiac function4weeks after permanent coronary artery occlusion.The survivors of new method MI were devided into two groups at random: Myocardial infarction group(N-MI)、Enalapril intervention group(N-Ena) which administration enalapril by gavage once a day with a dose of7.5mg/kg.The mices of three groups((N-Ena、N-MI、N-Sham) were killed at3days、7days、14days and28days after surgery respectively and collecting blood by the heart punctures. the plasma level of TNF-α and Th1/Th2、Treg/Th17were evaluated by ELISA and flow cytometry. Echocardiography was used to measure cardiac function4weeks after surgery.Statistical analysis:SPSS13.0for windows statistic software is used. All statistical analysis are expressed as mean±tandard deviation(SD) of the indicated number of experiments. Groups are compared using one-way analysis of variance or T test. Single factor data are compared using one-way analysis of variance as indicated, and are followed by a pos-hoc least significant difference (LSD) test. We use Welch test and Dunnett’s T3when variances are not homogeneous. Data are evaluated at the significance level P<0.05.Results:1. This novel MI procedure is rapid, with an average ligation of left coronary artery procedure time of3.0±0.5minutes and an average ventilated time of5.0±0.8minutes, whereas the classic method requires16±1.5minutes per ligation of left coronary artery procedure and31±2.5minutes ventilated time. Surgical mortality was16.7%in N-M and15.9%in C-MI.2. To compared the inflammatory damage/response of these2methods, the plasma level of TNF-α was measured at3days and7days after surgery. The plasma concentration of TNF-α are33.99±2.75pg/mL>66.25±3.30pg/mL、14.98±1.13pg/mL、24.03±1.40pg/mL in N-MI、C-MI、N-Sham、C-Sham respectively at3days. The variance of each groups is homogeneous (P:0.211), the results of single variance analysis are:The plasma concentration of TNF-α have significantly different in four groups (F:521.23, P<0.001). In MI groups of both2methods The plasma concentration of TNF-α were increased compared with Sham groups (P<0.001). The plasma level of TNF-α in N-MI deceased significantly when compared with C-MI (P<0.001) and N-Sham is lower than C-Sham. At7days,The plasma concentration of TNF-α are41.35±2.66pg/mL、84.56±2.49pg/mL、8.27±0.88pg/mL、8.32±0.79pg/mL in N-MI、C-MI、N-Sham、C-Sham respectively. The variance of each groups is homogeneous (P:0.424), the results of single variance analysis are:The plasma concentration of TNF-α have significantly different in four groups (F:1917.25, P<0.001). In MI groups of both2methods The plasma concentration of TNF-α are increased compared with Sham groups (P<0.001), The plasma level of TNF-ain N-Sham have no significant when compared with C-Sham (P:0.971)The plasma level of TNF-α at3days、7days、14days and28days were compared in each four groups. The variance of each groups was homogeneous (P:0.426, P:0.641, P:0.537, P:0.084), the results of single variance analysis are: The plasma concentration of TNF-α have significantly different in four groups (P<0.001). Expressions of TNF-α were found to be significantly decrease at7days in N-Sham、C-Sham (P<0.001),and then stay this level till last.Echocardiography was used to measure LVEDd、LVESd、LVFS、LVEF4weeks after surgery. The variance of each groups was homogeneous(P:0.297、P:0.254、P:0.682、P:0.382). Both MI groups had significantly increased LV diastolic (LVEDd) and systolic (LVESd) inner chamber dimensions and accordingly decreased LV ejection fraction (LVEF) and fractional shortening (LVFS) when compared with Sham (P<0.001), There were no differences between the N-Sham and C-Sham(P:0.523、P:0.723、P:0.751、P:0.605) nor between the MI-N and MI-C groups (P:0.613、P:0.698、P:0.665、P:0.232)2. To investigate the difference of three groups(N-MI、N-Ena、N-Sham) about plasma concentration of TNF-α,separate analyses were carried out at3days.7days.14days.28days. The variance of plasma concentration of TNF-α was unequal at3days (P:0.034).the results of Welch test was P<0.001. The plasma concentration of TNF-α in N-Ena didn’t had significant difference compared with N-MI (P:0.175) The variance of each groups was homogeneous(P:0.392, P:0.068, P:0.111) at7 days%14days、28days. the results of single variance analysis are: The plasma concentration of TNF-a have significantly different in four groups (F:409.71, F:159.18, F:82.61, P<0.001). The plasma concentration of TNF-a in N-Ena had significantly decreased when compared with N-MI (P<0.001). The plasma level of TNF-α in N-Ena and N-MI had significantly increased when compared with N-Sham (P<0.001). The plasma concentration of TNF-α in N-Ena had significantly decreased when compared with N-MI (P<0.001) at14days and28days. The plasma level of TNF-a in N-Ena and N-MI had significantly increased when compared with N-Sham (P<0.001) at14days and28days.We analysis of Treg/Th17in three differents groups(N-Sham、N-Ena、N-MI) at28days. The variance of rate of Treg and Th17in each groups’ was homogeneous(P:0.085, P:0.406). the results of single variance analysis are: the rate of Treg and Th17had significantly different in three groups (F:583.47, F:478.80, P <0.001). The rate of Treg in N-Ena and N-MI had significantly reduced while rate of Th17increased when compared with N-Sham (P<0.001). The rate of Treg in N-Ena had significantly increased and rate of Th17decreased when compared with N-MI (P<0.001). The variance of Treg/Th17ratio was unequal in three groups(P:0.002). the results of Welch test was P<0.001. Treg/Th17ratio in N-Ena had significantly increased when compared with N-MI (P<0.001)We analysis of Th1/Th2in three differents groups(N-Sham、N-Ena、N-MI) at28days. The variance of rate of Thl and Th2in each groups’ was homogeneous (P:0.956, P:0.076). the results of single variance analysis are: the rate of Thl and Th2had significantly different in three groups (F:531.03, F:925.6, P<0.001).The rate of Thl in N-Ena and N-MI had significantly increased while rate of Th2decreased when compared with N-Sham (P<0.001). The variance of Th1/Th2ratio was unequal in three groups(P:0.002). the results of Welch test was P<0.001. Th1/Th2ratio in N-Ena had significantly reduced when compared with N-MI (P<0.001) To measure LVEDd、LVESd、LVEF、LVFS4weeks after surgery and medical intervention by means of Echocardiography. The variance of each groups was homogeneous(P:0.103、P:0.069、P:0.646、P:0.724).the results of single variance analysis are: LVEDd、LVESd、LVEF and LVFS had significantly different in three groups (P<0.001). Both N-MI and N-Ena group had significantly increased LVEDd、 LVESd and decreased LVEF and LVFS (P<0.001). LVEDd and LVESd in N-Ena reduced when compared with N-MI (P<0.001), on the contry, LVEF and LVFS increased in N-Ena (P<0.001)Conclusions:1. This new rapid method of MI in mice represents a more efficient and less tissue damaging model of myocardial infarction compared with the classic method. Pectoral muscles remain intact after surgery.2. This new method of MI induces the same degree of cardiac dysfunction as the classic method and can be demonstrate by echocardiography.3. Enalapril could have the effect of decrease the The plasma concentration of TNF-a in mice with MI.4. Th1/Th2、Treg/Th17imbalance exist in peripheral blood of mice with MI, T h1/Th2ratios were significantly increased and Treg/Thl7ratios were significantly reduced.5. Enalapril could have the effect of regulation Th1/Th2、Treg/Th17imbalance in peripheral blood of mice with MI at28days.