| Myocardial infarction, as a serious clinical type of coronary heart disease, has a rather poor prognosis.About 50% of cardiovascular deaths are attributed to myocardial infarction. With the global process of acceleration, myocardial infarction, has soon become the world's biggest cause of death. Arrhythmias, heart failure and other complications are the cause of death post myocardial infarction. Among them, malignant arrhythmias, especially ventricular arrhythmia is the main reason for sudden death in patients with myocardial infarction. Researches show more than 50% myocardial infarction survivors died of malignant ventricular arrhythmia events. Therefore, the mechanism and intervetion strategy has drawed wild attention by clinical physicians and scientists.Previous studies indicated that ventricular, electrical and neural remodeling are potentially involved in the occurrence and development of ventricular arrhythmias and sudden cardiac death after myocardial infarction. In recent years, the effect of gap junctions on cardiac electrophysiology has sparked the attention of many researchers. Gap junctions are an important communication pathway among myocardial cells, as they provide substrates for electrical and chemical communication between cells. Many studies show that alterations in the quantity, distribution and function of gap junctions under pathological conditions, which is collectively known as gap junction remodeling. It correlates with ventricular arrhythmias and are the main cause of death in myocardial infarction. Connexin 43 (Cx43) is, by far, the most predominant type in ventricular myocytes. Reduced expression of Cx43 in the epicardial border zone contributed to the increased anisotropy and reentrant arrhythmias after myocardial infarction?Studies supported that sympathetic regeneration is closely related to inflammation process after myocardial infarction. Macrophages play an important role in the inflammatory response of the pathological process of neuroplasticity. Under different environmental stimuli, macrophages phenotype can shift, M1 macrophages and M2 macrophages, which exert different functions. M1 macrophages participate in the trigger and maintaining of inflammatory response; and M2 macrophages generate anti-inflammatory cytokines, terminate pro-inflammatory process and repair damaged tissuee; which has a protective effect on the body. Studies found that in early stage myocardial infarction, M1 macrophages play a leading role. Therefore, inhibition of the inflammatory responses that triggered by M1 macrophages as soon as possible after the occurrence of myocardial infarction,which may regulate sympathetic regeneration, may be an effective means of prevention and treatment of malignant ventricular arrhythmias.The clinical use of angiotensin receptor blocker (ARB) is mainly based on the attenuation of pathological left ventricular remodeling and the improvement of left ventricular systolic function. LIFE, TRACE and AFFIRM trial tests observed that the new incidence of atrial fibrillation was significantly reduced in the patients of cardiac dysfunction, hypertension and left ventricular hypertrophy with the administration of angiotensin-converting enzyme inhibitors (ACEI) and/or ARB, suggesting that both ACEI and ARB may have some anti-arrhythmic effects; and studies have shown that these effects may be associated with ACEI and / or ARB can improve atrial gap junction remodeling, but the research of its effect of post-MI myocardium cardiac electrophysiology is less. Therefore, we speculate that ARB early intervention after MI can effectively regulate gap junction remodeling after MI, and providing a new theoretical and experimental basis to support prevention and treatment of malignant ventricular arrhythmias.Hydroxymethyl-coenzyme A (HMG-CoA) reductase inhibitors, also known as statins, are widely used in clinical practice among patients with high cholesterol. Meanwhile,statins also have other effects outside the regulation of blood lipids,such as anti-platelet aggregation, inhibition of monocyte-macrophage adhesion and secretion, and inhibit lipid outside the proliferation and migration of vascular smooth muscle cells and other anti-arrhythmia. Emerging evidences also indicate that statins exert anti-arrhythmogenic effects by improving sympathetic nerve sprouting after myocardial infarction, but the exact mechanism of this effect remains unclear.More importantly, treatment with statins also changes macrophage phenotypes, shifting MltoM2 ratio and reducing levels of markers that are linked to macrophage differentiation.Meanwhile, inflammation after myocardial infarction causes cardiac nerve sprouting and consequent ventricular arrhythmias; macrophages are involved in the entire inflammation response process and serve as a link between inflammation and sympathetic hyper-innervation by regulating nerve growth factor expression;and the aim of the present study was also to explore the mechanism by which atorvastatin ameliorates cardiac sympathetic nerve sprouting via regulating macrophage polarization.Part 1. Effect and mechanism of valsartan on ventricular arrhythmias after myocardial infarction in ratsObjectives:To investigate the effect of prolonged valsartan treatment on gap junction remodeling and ventricular arrhythmia susceptibility in rats with myocardial infarction.Methods:SD rats were randomly divided into 3 groups:Sham group, MI group and ARB group which treated with valsartan after myocardial infarction. The animal models were created by ligating the left coronary artery. The rats were anesthetized with normal saline and valsartan (30mg/kg/day) the next day. All the surviving rats were subjected to hemodynamic test and received electrical stimulation test to induce ventricular arrhythmias. Ten rats of each group were selected,and the hearts were dissected into the infarcted border zone(BZ)and the non-infarcted left ventricle free wall(LVFW).Cx43 expression was studied using immunostaining, Western blot and transmission electron microscopy. The isolated ventricular myocytes were isolated from the remaining rats of each group;patch clamp technique was used to record the transient outward potassium current (Ito) current density which was used to evaluate the function of the gap junction channels.Results:1. Eight weeks after the MI, the incidence of inducible ventricular arrhythmias was significantly greater in the MI group than in the Sham group(P<0.01).Ejection fraction (EF) and cardiac output (CO) were significantly lower than those in Sham group, left ventricular systolic pressure (LVSP) and the maximal rate of the increase/decrease of left ventricular pressure(±dp/dtmax) were also significantly decreased (P<0.05). Cx43 expression was decreased in BZ of the MI group compared with the Sham group(0.215±0.017,0.330±0.024, P<0.05);Ventricular transient outward potassium current density significantly decreased compared with Sham group(P<0.05);2. ARB treatment decreased the arrhythmia scores of post-MI ventricular arrhythmias compared with the rats without treatment(P<0.05). FE,CO and ±dp/dtmax increased significantly(P<0.05);while the left ventricular end diastolic pressure (LVEDP) and LVSP decreased (P<0.05).The Cx43 expression levels were higher in the ARB group(P<0.05). Meanwhile, the MI-induced abnormal distribution of Cx43-positive gap junctions appeared to be reversed by ARB treatment;ventricular transient outward potassium current density was partially restored.3. The LVFW of each group had a normal pattern of Cx43-positive gap junction distribution, but there was a significant reduction of the AOD in the MI group compared with the Sham and ARB group (P<0.05). No significant difference was found between the Sham group and the ARB group.4. Western blotting data also supported the above results. Meanwhile, the electron microscope examination revealed that, ARB intervention after myocardial infarction may restore abnormal Cx43 distribution to some extent.Conclusions:1. Gap junction remodeling may be one of the mechanisms responsible for ventricular arrhythmias induced by myocardial infarction.2. Valsartan treatment could decrease the high incidence of ventricular arrhythmias after myocardial infarction partly by reversing gap junction remodeling.Part2. Effect and mechanism of atorvastatin on ventricular arrhythmias after myocardial infarction in ratsObjectives:To investigate the relationship between macrophage phenotype transformation and sympathetic nerve remodeling after MI, to clarify the effect of statin therapy on the polarity of macrophages after MI and the relationship with sympathetic nerve remodeling after MI. And provide a new therapeutic idea for the prevention and treatment of malignant arrhythmia caused by sympathetic nerve remodeling, and provide theoretical basis and basic experimental support for clinical application of statins.the mechanism by which atorvastatin ameliorates sympathetic nerve sprouting via regulating macrophage polarization.Methods:Wistar rats were randomly divided into 3 groups:Sham group, MI group and Statin group which treated with atorvastatin after myocardial infarction. Rat models of MI were created by ligating the left coronary artery. MI-operated rats received either atorvastatin or phosphate-buffered saline (PBS) for 7 days. The association between VAs and MI was evaluated using programmed electrophysiological stimulation. Peritoneal macrophages were isolated, and then were stimulated with lipopolysaccharide+interferon-y (LPS+IFN-y)and interleukin-4(IFN-y);one well of the LPS+IFN-y group was stimulated with atorvastatin,the purity of the detached cells was as ssessed via flow cytometric analysis; and the polarityof the macrophages was also examined;the mRNA levels of inducible nitric oxide synthase (iNOS), arginase-1(Arg1), tumor necrosis factor-a (TNF-a)and interleukin-1?(IL-1?) were examined by RT-PCR.Immunohistochemical analyses and immunofluorescence staining were used to analyze macrophage infiltration and to detect the distribution and density of growth-associated protein-43 (GAP-43) and tyrosine hydroxylase(TH) innerve fibers in peri-infarct zones after myocardial infarction. The protein levels of NGF were detected via western blot analysis, and the concentrations of NGF in the supernatants were determined via enzyme-linked immunosorbent assay.The mRNA levels of NGF, was examined by quantitative real-time polymerase chain reaction (PCR).Results:1. When macrophages were stimulated with LPS+IFN-y, IL-4 and atorvastatin, the positive rate of CD86/CD68 was significantly different, indicating that the cells shifted toward the M1 phenotype in LPS+IFN-y-stimulated cultures and toward the M2 phenotype in IL-4-stimulated cultures. In addition, the addition of atorvastatin to LPS-stimulated cultures decreased the levels of CD86/CD68-positive cells, demonstrating that atorvastatin indeed directed macrophages toward M2 phenotype (CD86, a hall mark of classical Ml macrophages, and CD68, a common marker of macrophages).2. INOS and Arglwere significantly upregulated in the LPS+IFN-y-stimulated group and the IL-4-stimulated grouprespectively than in the control group (P<0.01). Compared to low levels in the control group, the mRNA levels of IL-1? and TNF-a were higher in the LPS+IFN-y stimulated group(P<0.01)and in the IL-4-stimulated group(.P<0.01), and both were expressed at lower levels in the IL-4-stimulated group than in the LPS+IFN-y stimulated group. In addition, as shown by the higher level of Arg 1 relative to iNOS in the atorvastatin group, the addition of atorvastatin to the LPS+IFN-y-stimulated cultures decreased the levels of IL-1? and TNF-a(P<0.01).3. The mass of infiltrated cells shown by positive staining for the macrophage marker CD68 was significantly greater than in the Sham group (P<0.01). Moreover, the measurements of CD163/CD68-positive cells counted in each region at the third day post MI, which indicates the percentage of M2 macrophages, suggest that this time period was when macrophages differentiated within the myocardium. As a result, compared to the MI group, a relative increase in the level of CD 163-positive macrophages levels in the infarct area were observed (P<0.01), demonstrating that atorvastatin can induce a phenotypic switch in macrophages to M2 in the early phase.4. GAP-43-positive nerve fibers had almost no expression in Sham group. In the infarct borders of the MI group, we observed substantially more GAP-43-positive nerve fibers (P<0.01). Moreover, in MI group, the density of GAP-43-positive nerve fibers noticeably decreased (P<0.01).Compared to low density in the normal regions in the MI group, the density of TH-positive nerve fibers at the infarct borders was significantly upregualted (P<0.01), and the distribution was disordered. In addition, in Statin group, a significantly lower density of TH-positive nerve fibers was observed (P<0.01).5. As compared with sham hearts, MI hearts showed increased mRNA and protein level of NGF; and the level was decreased after atorvastatin treatment (P<0.05).Conclusions:1. The conversion of macrophages polar after myocardial infarction may affect the expression of NGF, which could regulate sympathetic remodeling.2. Atorvastatin can promote the transformation of M1 macrophages to M2 macrophages, and reduced expression of NGF, and ultimately suppress sympathetic regeneration.3. Early treatment of atorvastatin after myocardial infarction can regulate macrophage polarization,improve sympathetic remodeling and exert its anti-arrhythmic effects. |
PDF Full Text Request