缝隙连接蛋白43在老年大鼠缺血性室性心律失常中的作用

Objective To investigate the expression of connexin43 ( Cx43 ) and effect of vagal nerve stimulation (VNS) on ventricular tachyarrhythmias during acute myocardial ischemia( MI )in aged rats. Methods Male Sprague-Dawley rats[Adult group ( ≤ 4 months) and Aged group ( ≥24 months)]:MI (n = 15 ):ligated left anterior descending coronary for 30 minutes; MI-vagal nerve stimulation(VNS) ( n = 15 ); MI-VNS-atropine (0. 5 mg/kg, n = 13 ); MI-VNS-carbenoxolone ( 10 mg/kg, n = 11 ); sham operation (SO, n = 10):without coronary ligation. Ventricular arrhythmias were monitored by an electrocardiogram. Cx43 protein expression was analyzed by Western blot. Results During the 30 minutes ligation,incidences of ventricular tachycardia (VT) and ventricular fibrillation(VF) in aged rats increased significantly compared to those of adult rats ( P ＜ 0. 05 ). VNS did not affect the occurrence of VT and VF ( both P ＞ 0.05 ); however, VNS suppressed the occurrence of irreversible VF ( P ＜ 0. 05 ); both atropine and carbenoxolone ( a gap junction inhibitor) could abolish the effect of VNS on ischemia-induced irreversible VF ( both P ＜0. 05). Ischemia did not result in changes of total Cx43 amount in adult and aged rats compared to that of SO group,respectively. The amount of nonphosphorylated Cx43 was increased markedly in adult and aged rats compared to that of SO group,respectively.Cx43 dephosphorylation induced by ischemia was significantly suppressed by VNS in adult and aged rats( P ＜0. 05 ). However,the amount of total Cx43 of SO group in aged rats was significantly decreased by 50% compared to that of SO group in adult rats ( P ＜ 0. 05 ). Conclusion The present study suggested that the incidence of ischemia-induced ventricular tachyarrhythmias increased markedly and the anti-arrhythmic effect of VNS was decreased significantly in aged rats, which may be associated with reduction of Cx43 protein of ventricle in aged rats.     

Effect and mechanism of Irbesartan on occurrence of ventricular arrhythmias in rats with myocardial ischemia through connexin43(cx43)

Objective: To explore the effect and mechanism of angiotensin Ⅱ receptor blockers-Irbesartan on occurrence of ventricular arrhythmias in rats with myocardial ischemia. Methods: Rats with embryonic cardiomyocytes-H9c2 were randomly divided into control group, ischemia group, Irbesartan group and Irbesartan+ischemia group. The cell viability of rats in each group was tested using MTT. Real-time PCR was employed to detect the expression of connexin43(Cx43) mR NA and western blot to detect the expression of Cx43 and phosphorylated Cx43. SD rats were randomly divided into the sham-operation group(SO), myocardial infarction group(MI), Irbesartan group and MI+ Irbesartan group, with 10 rats in each group. HE staining was employed to observe the change in the pathomorphology of left ventricular tissue and TUNEL method to analyze the cell apoptosis in the tissue. The immunofluorescence was adopted to observe the expression and distribution of Cx43 in the left ventricular myocardium and study the change in the expression of Cx43 in the cardiac muscular tissue at mR NA and protein level. Results: The intervention of Irbesartan in the condition of ischemia indicated the significant decrease in the number of necrotic cells. The expression of Cx43 was significantly decreased under the culture of ischemia(P0.05), but in the presence of Irbesartan, the expression of Cx43 was increased compared with the ischemia group(P0.01). The results of WB assay showed the similar trend of change at mRNA level. There was the significant difference in the score of ventricular arerythmia between MI group and SO group(P0.01). The incidence of ventricular tachycardia or ventricular fibrillation was significantly increased compared with the one in SO group(P0.05). There was the significant difference in the overall score between MI+Irbesartan group and MI group(P0.05). The expression of Cx43 in the cardiac muscular tissue in MI group was significantly decreased(P0.01 vs SO group). But the expression of Cx43 was increased after the treatment with Irbesartan. Conclusions: Irbesartan can inhibit the injury of H9c2 cardiomyocytes and the decreased expression of Cx43 that are induced by the ischemic myocardial infarction. Irbesartan can also improve the reconstruction of Cx43 in rats with ischemic myocardium to inhibit the myocardial infarction-induced arrhythmias.     

Effects of Ang Ⅱ perfusion on transmural heterogeneous of Cx43 in acute myocardial ischemia reperfusion

Objective:To observe the effects of angiotensin Ⅱ(Ang Ⅱ) pefusion on transmural heterogeneity of Cx43 expression in the rabbit model with acute myocardial ischemia reperfusion(MIR),and investigate the role of rennin-angiotensin system in malignant ventricular arrhythmia induced by MIR.Methods:Twenty rabbits were randomly divided into MIR group(n=10) and Ang Ⅱ group(n=10).MIR model was produced with traditional ligation and opening of the anterior descending coronary artery in all animal.The hearts in vitro in the MIR group and the Ang Ⅱ group were perfused with simply improved Tyrode's solution and containing Ang Ⅱ Tyrode's solution respectively.90%monophasic action potential repolarization duration,transmural dispersion of repolarization.Cx43 protein(Cx43-pro) and mRNA(Cx43-Cq) expression in subepicardial,midmyocardial and subendocardial myocardium were measured in both groups.The greatest differences of Cx43-pro and Cx43-Cq among three myocardial layers were calculated and shown with △Cx43-pro and △Cx43-Cq respectively.Results:After Ang Ⅱ perfusion,90%monophasic action potential repolarization duration among three myocardial layer were significantly prolonged(P 0.05 and P 0.01),and transmural dispersion of repolarization also significantly increased compared with the MIR group(P 0.05).Compare with the MIR group,three myocardial Cx43-pro and Cx43-Cq expression in the Ang Ⅱ group were significantly decreased(P 0.05 and P 0.01).but△Cx43-pro and △Cx43-Cq were significant increased.Conclusions:Renin-angiotensin system increases transmural heterogeneity of Cx43 expression in the rabbit model with MIR by Ang Ⅱ,and enlarge transmural dispersion of repolarization among three myocardial layers of left ventricular which induces malignant ventricular arrhythmia.     

抗心律失常肽与溶血磷脂酸对兔室性心律失常影响的研究

Objective To investigate the effect and potential mechanism of lysophosphatidic acid (LPA) and antiarrhythmic peptide (AAP10) on rabbit ventricular arrhythmia. Methods Twenty-four rabbits were randomly divided into three groups (n =8 each): control group, LPA group and AAP10 + LPA group. Using arterially perfused rabbit ventricular wedge preparations, transmural ECG and action potentials from both endocardium and epicardium were simultaneously recorded in the whole process of all experiments with two separate floating microeletrodes. The incidence of ventricular arrhythmia post S1S2 stimulation was recorded. Protein levels of nonphosphorylated Cx43 and total Cx43 were evaluated by Western blot. The distribution of nonphosphorylated Cx43 was observed by confocal immunofluorescence microscopy. Results Compared with the control group, the QT interval, endocardial action potential duration, transmural repolarization dispersion (TDR) and incidence of ventricular arrhythmia were significantly increased and nonphosphorylated Cx43 expression was significantly upregulated in the LPA group. Compared with the LPA group, cotreatment with AAP10 can reduce the QT interval, endocardial action potential duration, TDR and incidence of ventricular arrhythmia (25.0% vs 62. 5%, P ＜ 0. 01 ) and downregulate nonphosphorylated Cx43. Conclusions LPA could promote the arrhythmia possibly by upregulating nonphosphorylated Cx43 and subsequent gap junction transmission inhibition. Gap junction enhancer AAP10 could attenuate the proarrhythmic effect of LPA probably by downregulating myocardial nonphosphorylated Cx43 expression.     

抗心律失常肽与溶血磷脂酸对兔室性心律失常影响的研究

Objective To investigate the effect and potential mechanism of lysophosphatidic acid (LPA) and antiarrhythmic peptide (AAP10) on rabbit ventricular arrhythmia. Methods Twenty-four rabbits were randomly divided into three groups (n =8 each): control group, LPA group and AAP10 + LPA group. Using arterially perfused rabbit ventricular wedge preparations, transmural ECG and action potentials from both endocardium and epicardium were simultaneously recorded in the whole process of all experiments with two separate floating microeletrodes. The incidence of ventricular arrhythmia post S1S2 stimulation was recorded. Protein levels of nonphosphorylated Cx43 and total Cx43 were evaluated by Western blot. The distribution of nonphosphorylated Cx43 was observed by confocal immunofluorescence microscopy. Results Compared with the control group, the QT interval, endocardial action potential duration, transmural repolarization dispersion (TDR) and incidence of ventricular arrhythmia were significantly increased and nonphosphorylated Cx43 expression was significantly upregulated in the LPA group. Compared with the LPA group, cotreatment with AAP10 can reduce the QT interval, endocardial action potential duration, TDR and incidence of ventricular arrhythmia (25.0% vs 62. 5%, P ＜ 0. 01 ) and downregulate nonphosphorylated Cx43. Conclusions LPA could promote the arrhythmia possibly by upregulating nonphosphorylated Cx43 and subsequent gap junction transmission inhibition. Gap junction enhancer AAP10 could attenuate the proarrhythmic effect of LPA probably by downregulating myocardial nonphosphorylated Cx43 expression.     

Effects of Perindopril on Left Ventricular Remodeling and Osteopontin Expression in Rats With Myocardial Infarction

Objectives To observe the effects of perindopril on left ventricular remodeling and myocardial osteopontin expression in rats with myocardial infarction. Methods In this study male adult SD rats were randomly divided into 3 groups: sham-operation group, MI-saline group and MI-perindopril group. Left anterior descending artery was ligated to generate myocardial infarction. Perindopril (2 mg/kg body weight/day) was administered from the next day of MI. Four weeks later, left ventricular diameter (LVEDD and LVESD) and left ventricular ejection fraction was estimated with echocardiography, LVSP, LVEDP and±dp/dtmax was detected with hemodynamic measurement, cardiomyocyte diameter and interstitial fibrosis infiltration were evaluated with histological methods, and myocardium osteopontin protein expression level was detected with western blot. Results ①Compared with the sham-operation group, all rats with MI developed significant systolic and diastolic dysfunction, as was indicated by decreased LVEF, LVSP and±dp/dtmax, as well as increased LVEDP. ②Rats with MI showed significantly dilated left ventricles and higher ventricular weight / body weight ratio, significantly increased cardiomyocyte diameter and marked interstitial fibrosis in the non-infarction area. ③Perindopril treatment partly prevented cardiac dysfunction and left ventricular remodeling as indicated by the parameters mentioned above. ④No osteopontin protein was detected in myocardium of sham-operation rats. In rats with MI, high level osteopontin protein expression was significantly inhibited by perindopril treatment. Conclusions In rats with MI, perindopril treatment significantly prevented left ventricular remodeling and myocardium osteopontin protein expression.     

Effect of ischemic preconditioning on P-selectin expression in hepatocytes of rats with cirrhotic ischemia-reperfusion injury

AIM: To investigate the effects and mechanisms of ischemic preconditioning (IPC) on the ischemia/reperfusion (I/R) injury of liver cirrhosis in rats and the effect of IPC on P-selectin expression in hepatocytes.METHODS: Forty male SD rats with liver cirrhosis were randomly divided into sham operation group (SO group),ischemia/reperfusion group (I/R group), ischemic preconditioning group (IPC group), L-Arginine preconditioning group (APC group), L-NAME preconditioning group (NPC group), eight rats in each group. Hepatocellular viability was assessed by hepatic adenine nucleotide level and energy charge (EC) determined by HPLC, ALT, AST and LDH in serum measured by auto- biochemical analyzer and bile output.The expression of P-selectin in the liver tissue was analyzed by immunohistochemical technique. Leukocyte count in ischemic hepatic lobe was calculated.RESULTS: At 120 min after reperfusion, the level of ATP and EC in IPC and APC groups was higher than that in I/R group significantly. The increases in AST, ALT and LDH were prevented in IPC and APC groups. The livers produced more bile in IPC group than in I/R group during 120 min after reperfusion (0.101±0.027 versus 0.066±0.027 ml/g liver,P=0.002). There was a significant difference between APC and I/R groups, (P=0.001). The leukocyte count in liver tissues significantly increased in I/R group as compared with SO group (P＜0.05). The increase in the leukocyte count was prevented in IPC group. Administration of L-arginine resulted in the same effects as in IPC group. However,inhibition of NO synthesis (NPC group) held back the beneficial effects of preconditioning. Significant promotion of P-selectin expression in hepatocytes in the I/R group was observed compared with the SO group (P＜0.01). IPC or L-arginine attenuated P-selectin expression remarkably (P＜0.01). However, inhibition of NO synthesis enhanced Pselectin expression (P＜0.01). The degree of P-selectin expression was positively correlated with the leukocyte counts infiltrating in liver (r=0.602, P=0.000).CONCLUSION: IPC can attenuate the damage induced by I/R in cirrhotic liver and increase the ischemic tolerance of the rats with liver cirrhosis. IPC can abolish I/R induced leukocyte adhesion and infiltration by preventing postischemic P-selectin expression in the rats with liver cirrhosis via a NO-initiated pathway.     

Expression of vimentin and glial fibrillary acidic protein in central nervous system development of rats

Objective: To investigate the distribution and contents of vimentin(Vim) and glial fibrillary acidic protein(GFAP) immunoreactivities in the central nervous system(CNS)of normal newborn, adult and aged rats.Methods: In this study, thirty healthy and normal Sprague–Dawley rats were simply classified into three groups: Newborn(7 days aged), adult(5 months aged) and aged(24 months aged) rats. Brains and spinal cord were dissected and cut into frozen sections. The expression of Vim and GFAP in CNS were detected by confocal immunofluorescence.Results: In each group, Vim was expressed in all the regions of CNS including the hippocampal, cerebral cortex, the third ventricle and spinal cord, and the expression was highest in neuron-like cell of newborn rats, while Vim was mainly expressed in cell bodies in adult and aged rats. GFAP was expressed in all the regions of CNS including the hippocampal, cerebral cortex, the third ventricle and spinal cord, and the expression was in astrocytes of aged rats. In the third ventricle, Vim was detected in all groups, and only observed in neuron-like cells of newborn. Meanwhile, the GFAP expression showed no significant differences between adult and aged rats in this region. The co-localization of Vim and GFAP were mainly observed in hippocampus and cerebral cortex of newborn,but this co-localization was found in the third ventricle of the rats in all groups.Conclusion: Our data demonstrate for the first time that the expression of Vim and GFAP in the rat's CNS during development. This data may provide a foundation for the further mechanistic studies of these two main intermediate filaments during development of CNS.     

Effect of β blockers on β_3-adrenoceptor mRNA Expression in the Rats with Chronic Heart Failure

Objectives To investigate the changes of β3-adrenoceptor (β3-AR) mRNA expression in the rats with chronic heart failure (CHF), and to explore the effect of β blockers (βBs) on β3 mRNA expression. Methods Thirty-four rats were randomly divided into Sham group (n = 10) and heart failure group (n = 24). Rat model was established by aortic constriction. The survival rats in heart failure group were divided into heart failure control group (HF group, n = 6), metoprolol group (MET group, n = 8) and carvedilol group (CAR group, n = 8) three months after operation. Metoprolol tartarte was started orally with 12 mg·kg-1·d-1, carvedilol with 6 mg·kg-1·d-1, isometric saline was started in HF group. After three months of drug therapy, measurement of hemodynamics, index of ventricular mass, the level of β3-AR mRNA expression were performed. Results Compared with Sham group, left ventricular end systolic pressure (LVESP), and the absolute values of maximal rate of rise and fall ( ± dp/dtmax) of left ventricular pressure were all significantly decreased (P < 0.01), left ventricular end diastolic pressure (LVEDP) was significantly increased in HF group (P < 0.01). The hemodynamic parameters were improved by βBs, and carvedilol was more effective than metoprolol (P < 0.01). The index of ventricular mass was higher in HF group than MET group, CAR group and Sham group (P < 0.01). βBs significantly decreased the index of left ventricular mass (LVMI), and Carvedilol was more effective than metoprolol (P < 0.01). The index of right ventricular mass (RVMI) did not change in MET group (P > 0.05), but significant decrease could be seen in CAR group (P < 0.01). The level of β3-AR expression in left ventricle was greater than that in right ventricle whether in the failing heart or in the non-failing heart. Compared with Sham group, the level of β3-AR mRNA expression was significantly increased in HF group (P < 0.01). The levels of β3-AR mRNA expression showed a remarkable decrease in CAR group(P < 0.01), but was not seen in MET group. Conclusions The β3-AR expression level remarkably increases in the rat's left and right failing ventricles. Carvedilol is more effective on improving hemodynamics and attenuating ventricular remodeling than metoprolol in the rats with CHF. Carvedilol rather than metoprolol downregulates β3-AR expression in the rat's failing ventricles. The beneficial effect of carvedilol in CHF maybe partly due to the downregulation of β3-AR expression in the failing heart.     

老龄小鼠心肌梗死后wnt信号通路表达的变化及与心脏破裂的关系

Objective To observe the association between wnt signal pathway and post infarction left ventricular remedeling/rupture in mice with various ages. Methods Three months-old (young group, n=116) and 18 months-old (aged group, n = 116) male C57/BL mice were studied. Seventy mice underwent ligation of left coronary artery, 10 sham-operation and echocardiography and hemodynamics were performed 7 d post-infarction, 36 infarcted mice were used for detecting expression of dvl-1, β-catenin and connexin 43 in left ventricular (LV) myocardium and infarction region at 3 d, 7 d, 14 d post infarction (n = 12 each). Results Incidence of cardiac rupture was significantly higher in aged mice than in young mice (36.7% vs. 16.7% ,P <0.05) and degree of LV dilation and contractile dysfunction was significantly severer in aged mice than those in young mice post infarction. Expression of dvl-1, β-catenin in left ventricle was upregulated in MI group compared with sham group(P <0.05), expression of dvl-1 and β-catenin in infarction region in MI 3d group in aged mice was significantly downregulated than in young mice (P < 0.05). Expression of connexin 43 is 2.15 fold higher in young sham mice than in aged sham mice (P < 0.05) and decreased significantly post infarction (P < 0.05). Expression of connexin 43 in infarction region in mice 3 d and 14 d post infarction was significantly lower in aged mice than in respective young mice(all P < 0.05). Conclusion Reductant activation of wnt signal pathway post infarction in aged mice might be responsible for increased incidence of cardiac mpture and aggravated remodeling.