Murine Viral Myocarditis Research Articles

Modulation of myocardial inflammation, fibrosis development and ventricular arrhythmogenicity by endurance exercise in murine coxsackievirus B3 myocarditis

Abstract Aims Nonischaemic myocardial fibrosis is associated with progressive deterioration of myocardial function and forms the substrate for arrhythmias in atria and ventricles. In the absence of a specific aetiology, its origin is commonly attributed to preceding viral myocarditis. Athletes presenting with ventricular arrhythmias often demonstrate nonischaemic fibrosis on magnetic resonance imaging. Previous translational studies have shown an adverse effect of exercise on the course of acute viral myocarditis. Nevertheless, the effect on myocardial fibrosis development, a frequent and important consequence of viral myocarditis, has not yet been investigated. In the present study, we evaluate for the first time the impact of endurance exercise on longer-term myocardial inflammation, myocardial fibrosis and arrhythmogenicity using a murine viral myocarditis model. Methods and results Male C57BL/6J mice (11 weeks of age, n=72) were randomly assigned to 8 weeks of treadmill running (EEX) or without exercise (SED). Two weeks into the study (training or control), animals were injected intraperitoneally with either coxsackievirus B3 to induce acute viral myocarditis (CVB) or vehicle (PBS). Exercising myocarditis mice showed lower mortality (11% vs. 27%), albeit without statistical significance (P=0.23). At sacrifice (i.e. 6 weeks after inoculation), prominent myocardial inflammatory infiltration and cardiomyocyte loss was observed in both CVB groups, without difference on semiquantitative histological scoring. Nevertheless, the infiltrating cells in the CVB-EEX group were more often proinflammatory in nature (predominantly iNOS-reactive macrophages and positive CD8+/CD4+ ratio) compared to these in CVB-SED (predominantly arginase-1-reactive macrophages and higher CD4+/CD8+ ratio). In addition, the EEX group showed more haemosiderin-laden macrophages and spindled fibroblasts as signs of organisation. At that time, the majority of virus was cleared from the heart. Treadmill running during myocarditis enhanced development of interstitial fibrosis (with limited or extensive distribution)(82.4% in CVB-EEX vs. 56.3% in CVB-SED; P=0.049) and perivascular and/or interstitial fibrosis with extensive distribution (64.7% & 64.7% in CVB-EEX vs. 50% & 31.3% in CVB-SED; P=0.048). The highest scar counts were observed in CVB-EEX, but the mean count (1.9 vs. 1.2; P=0.19) did not significantly differ between the myocarditis groups. In CVB-EEX, the lesions contained denser and more organised collagen bundles. In vivo electrophysiology studies showed similar ventricular arrhythmia inducibility (P>0.20) and arrhythmia burden (P=0.49) in the myocarditis groups, although the longest arrhythmias and highest cumulative burden occurred in CVB-EEX. Conclusion Endurance exercise training during viral myocarditis modulates the longer-term inflammatory process and promotes perivascular and interstitial fibrosis development, potentially enhancing ventricular arrhythmogenicity.

Abstract P1049: The Impact Of Endurance Exercise On Longer-term Myocardial Fibrosis Development And Ventricular Arrhythmogenicity In Murine Coxsackievirus B3 Myocarditis

Aims: Diagnostic work-up of athletes often reveals nonischaemic myocardial fibrosis, which is associated with cardiac dysfunction and malignant arrhythmias. Its isolated finding on magnetic resonance imaging is commonly attributed to preceding viral myocarditis. Previous murine studies have shown a negative effect of exercise early in the course of viral myocarditis. In this study, we investigate, for the first time, the impact of endurance exercise on myocardial fibrosis development and arrhythmogenicity in a murine viral myocarditis model. Methods & Results: Eleven-week-old male C57BL/6J mice (n=72) were randomised to 8 weeks of treadmill running (EEX) or without exercise (SED). After 2 weeks, animals received a single injection with either coxsackievirus B3 to induce acute viral myocarditis (CVB) or vehicle as control (PBS). Exercise resulted in a milder clinical disease course, with less weight loss and better preserved running capacity. In addition, mortality was lower in exercising myocarditis mice (11 vs. 27%), however without statistical significance (P=0.23). Exercise during myocarditis increased the occurrence of interstitial fibrosis (limited or extensive distribution) at sacrifice ( i.e. 6 weeks after inoculation)(82.4% in CVB-EEX vs. 56.3% in CVB-SED; P=0.049). Additionally, exercise enhanced development of perivascular and/or interstitial fibrosis with extensive distribution (64.7% & 64.7% in CVB-EEX vs. 50% & 31.3% in CVB-SED; P=0.048). CVB-EEX animals demonstrated more myocardial scars on average, albeit not significantly (1.9 vs. 1.2; P=0.19), with similar scar distribution. Despite manifest fibrotic remodelling, collagen and fibrogenetic genes were not upregulated in the myocarditis groups at the time of sacrifice. In vivo right ventricular programmed electrical stimulation showed comparable ventricular arrhythmia inducibility in the myocarditis groups (P>0.20). The longest arrhythmias and highest cumulative arrhythmia burden occurred in CVB-EEX, however, without reaching statistical significance when compared to CVB-SED (P=0.49). Conclusion: Endurance exercise during viral myocarditis enhances perivascular and interstitial fibrosis development, which may promote ventricular arrhythmogenicity.

Sex-Specific Effects of Plastic Caging in Murine Viral Myocarditis.

Background: Myocarditis is an inflammatory heart disease caused by viral infections that can lead to heart failure, and occurs more often in men than women. Since animal studies have shown that myocarditis is influenced by sex hormones, we hypothesized that endocrine disruptors, which interfere with natural hormones, may play a role in the progression of the disease. The human population is exposed to the endocrine disruptor bisphenol A (BPA) from plastics, such as water bottles and plastic food containers. Methods: Male and female adult BALB/c mice were housed in plastic versus glass caging, or exposed to BPA in drinking water versus control water. Myocarditis was induced with coxsackievirus B3 on day 0, and the endpoints were assessed on day 10 post infection. Results: We found that male BALB/c mice that were exposed to plastic caging had increased myocarditis due to complement activation and elevated numbers of macrophages and neutrophils, whereas females had elevated mast cell activation and fibrosis. Conclusions: These findings show that housing mice in traditional plastic caging increases viral myocarditis in males and females, but using sex-specific immune mechanisms.

Interleukin-37 Ameliorates Coxsackievirus B3-induced Viral Myocarditis by Modulating the Th17/Regulatory T cell Immune Response.

Myocarditis is a heterogeneous group of disorders defined by inflammation of the heart muscle with an excessively activated immune response. Numerous interventions have been investigated for the treatment of myocarditis while success is limited. Interleukin-37 (IL-37), a novel member of the IL-1 cytokine family, is a natural inhibitor of innate immunity associated with autoimmune diseases. However, the modulatory effect of IL-37 in myocarditis is unknown. In this study, we investigated the immunological regulation of IL-37 in the coxsackievirus B3-induced model of murine viral myocarditis. The results show that IL-37 significantly ameliorates the signs of myocarditis with increased survival rate and bodyweight, improved histological changes, reduced activities of MB isoenzyme of creatine kinase and cardiac troponin I, and a suppressed response of Th17 cells and enhanced response of regulatory T cells (Tregs) in the spleen. Moreover, IL-37 down-regulates the expression of Th17-related cytokines IL-6 and IL-17A, while promoting Treg-related cytokine IL-10 levels in the heart. Therefore, IL-37 may exhibit anti-inflammatory activity in the murine model of myocarditis by regulating the balance between Th17 and Treg cells, thereby providing a possible novel therapeutic target in myocarditis.

Right Cervical Vagotomy Aggravates Viral Myocarditis in Mice Via the Cholinergic Anti-inflammatory Pathway.

The autonomic nervous system dysfunction with increased sympathetic activity and withdrawal of vagal activity may play an important role in the pathogenesis of viral myocarditis. The vagus nerve can modulate the immune response and control inflammation through a ‘cholinergic anti-inflammatory pathway’ dependent on the α7-nicotinic acetylcholine receptor (α7nAChR). Although the role of β-adrenergic stimulation on viral myocarditis has been investigated in our pervious studies, the direct effect of vagal tone in this setting has not been yet studied. Therefore, in the present study, we investigated the effects of cervical vagotomy in a murine model of viral myocarditis. In a coxsackievirus B3 murine myocarditis model (Balb/c), effects of right cervical vagotomy and nAChR agonist nicotine on echocardiography, myocardial histopathology, viral RNA, and proinflammatory cytokine levels were studied. We found that right cervical vagotomy inhibited the cholinergic anti-inflammatory pathway, aggravated myocardial lesions, up-regulated the expression of TNF-α, IL-1β, and IL-6, and worsened the impaired left ventricular function in murine viral myocarditis, and these changes were reversed by co-treatment with nicotine by activating the cholinergic anti-inflammatory pathway. These results indicate that vagal nerve plays an important role in mediating the anti-inflammatory effect in viral myocarditis, and that cholinergic stimulation with nicotine also plays its peripheral anti-inflammatory role relying on α7nAChR, without requirement for the integrity of vagal nerve in the model. The findings suggest that vagus nerve stimulation mediated inhibition of the inflammatory processes likely provide important benefits in myocarditis treatment.

Effect of carvedilol on expression of tumor necrosis factor-α, monocyte chemoattractant protein-1 in murine viral myocarditis

Objective To investigate the role of tumor necrosis factor-α(TNF-α) and monocyte chemoattractant protein-1(MCP-1) in the pathogenesis of viral myocarditis(VMC) and the effect of carvedilol. Methods Eighty male Balb/c mice were randomly divided into 3 groups: control group(n=20), myocarditis group(n=30) and carvedilol group(n=30). The latter 2 groups were intraperitoneally inoculated with Coxsackie virus B3(CVB3) to induce VMC, while control group with virus-free Eagle's medium instead.Carvedilol was poured 5 mg/kg from the next day after injecting virus in carvedilol group, 9 g/L saline was used in control group and myocarditis group, twice a day.On the 7th and 14th day after inoculation, 8 mice were sacrificed after taking blood sample which were randomly selected from each group.Histological cross sections of hearts were stained with hematoxylin-eosin(HE), and the content of serum TNF-α was assayed with enzyme-linked immunoadsordent assay, while the expression of MCP-1 mRNA in myocardial tissue was detected with reverse transcriptase-polymerase chain reaction(RT-PCR) method. Results The pathological score of myocarditis group was significantly higher than that in control group(P<0.01), while carvedilol group was significantly lower than that in myocardial group(P<0.05). The serum level of TNF-α of VMC mice in myocarditis was significantly higher than that in control group(P<0.01) and positively correlated with the pathological score of the myocardium(r=0.42, P<0.05), and serum TNF-α in carvedilol group was remarkably decreased compared with myocarditis group(P<0.05). The expression level of MCP-1 mRNA in myocarditis group was increased significantly compared with control group(P<0.01), and remarkably decreased in carvedilol group compared with myocarditis group(P<0.05). Conclusions TNF-α and MCP-1 may be involved in the pathogenesis of VMC, and carvedilol may protect myocardium against CVB3-induced VMC by down-regulating the expression of TNF-α and MCP-1. Key words: Carvedilol; Viral myocarditis; Tumor necrosis factor-α; Monocyte chemoattractant protein-1

MicroRNA-21 and -146b are involved in the pathogenesis of murine viral myocarditis by regulating TH-17 differentiation

Viral myocarditis (VMC) is a common cardiovascular disease, and microRNAs (miRNAs) have been postulated to be involved in its pathology. Using microarrays, we observed that miRNA-21 and -146b were upregulated in a murine model of VMC. We also found that miRNA-451 was downregulated. In vivo silencing of miRNA-21 and -146b resulted in less-severe VMC. Overexpression of miRNA-451 did not ameliorate the severity of VMC. Further work revealed that inhibition of miRNA-21 and -146b decreased the expression levels of Th17 and RORγt . Overexpression of miRNA-451 had no effect on IL-17 and RORγt expression. Inhibition of miRNA-21 and -146b might ameliorate myocardium inflammation by mediating downregulation of RORγt expression, indicating that these miRNAs are involved in the pathogenesis of murine VMC.

Ivabradine inhibits the production of proinflammatory cytokines and inducible nitric oxide synthase in acute coxsackievirus B3-induced myocarditis

The role of β-adrenergic stimulation on viral myocarditis has been investigated in animal models. The beneficial action of the β-blocker carvedilol in murine viral myocarditis can be explained partly by the resulting heart rate reduction and the inhibition of proinflammatory cytokine production. The modulation of myocardial necrosis and contractile dysfunction by proinflammatory cytokines may be partially mediated by the production of nitric oxide (NO). The selective If current inhibitor ivabradine reduces the heart rate without affecting cardiac contractility and has been shown to be cardioprotective in failing hearts. However, little is known about the effects of ivabradine in viral myocarditis, and in particular, its effects on inducible NO synthase (iNOS) have not been investigated. This study was therefore designed to examine the effects of ivabradine in murine viral myocarditis. In a coxsackievirus B3 murine myocarditis model, the effects of ivabradine and carvedilol on the myocardial histopathological changes and fibrosis, NO production, iNOS protein and cytokine levels were studied. Both ivabradine and carvedilol similarlyattenuated myocardial lesions and fibrosis, inhibited NO synthesis by iNOS, and decreased the production of TNF-α and IL-6. These results show that ivabradine has a therapeutic benefit in murine CVB3-induced myocarditis. The beneficial effects of ivabradine in viral myocarditis are partially mediated by the inhibition of both the production of proinflammatory cytokines and the synthesis of NO by iNOS.

Abstract 68: Role of Substance P Signaling in the Pathogenesis of Murine Viral Myocarditis

Viral myocarditis is an important cause of heart failure in adolescents and young adults for which no specific treatment is currently available. Substance P (SP) is a neuropeptide and pain transmitter that activates RhoA, a small G-protein, shown to induce hypertrophy of neonatal rat ventricular myocytes. The current studies were performed to determine if SP signaling contributes to the pathogenesis of encephalomyocarditis virus (EMCV) infection, a well-established murine model of viral myocarditis. Aprepitant (1.2 mg/kg), a SP receptor antagonist, or fasudil (10mg/kg), a Rho A inhibitor, or saline control, was administered daily to mice orally by gavage 3 times for 3 days prior to intraperitoneal infection with 50 PFU of EMCV, and the mice monitored for 14 days. Pretreatment with aprepitant reduced mortality by 40% (25±7% vs. 65±7%), reduced heart-to-body weight ratio by 26% (0.0088±0.0001 vs. 0.0119±0.0001), reduced end systolic diameter (ESD) by 21% (3.0±0.2mm vs. 3.8±0.4mm), improved ejection fraction by 72% (57.2±4.7% vs. 33.2 ±9.3%), reduced chamber volume by 46% (34±5.0 μl vs. 62.7±30.8 μl), increased peak aortic flow velocity by 41% (87.0±24.7 vs. 61.7±20.1), and reduced cardiomyocyte diameter size by13% (11.0±0.1 μM vs. 12.7± 0.2 μM); p&lt;.05 for all, Student’s t-test). Pretreatment with fasudil reduced heart-to-body weight ratio by 18% (0.0097±0.0001 vs. 0.0119±0.0001) and reduced cardiomyocyte diameter size by 12% (11.14±0.51 μM vs. 12.7± 0.2 μM; p&lt;.05 for both), but did not significantly impact heart functional changes or mortality. These findings indicate that SP contributes to cardiac remodeling and dysfunction following ECMV infection and raises the possibility that inhibition of SP signaling may be of benefit in the treatment of patients with viral myocarditis.

The heart-protective mechanism of nitronyl nitroxide radicals on murine viral myocarditis induced by CVB3

Our previous researches showed that nitronyl nitroxyl derivatives, NNP and NNVP were good anti-oxidants and provided radioprotective effects in C6 cells. The objective of the present study is to investigate the possible antiviral effects and underlying pharmacological of the two nitronyl nitroxide radicals against CVB3 invitro and invivo. The results showed that NNP and NNVP were some of the most potent compounds in terms of their antiviral effects by protecting myocardial cells against oxidative damage of free radicals. Treatment with NNP or NNVP could decrease the intracellular ROS level invitro. They could lead to a significant decrease in activities of biochemical markers AST, CK and LDH in infected murine serum and could increase SOD and CAT activities and decreased MDA activities compared with infected control in vivo. NNP and NNVP could reduce NO production in infected mice by reacting with NO to produce the imino nitroxides which was confirmed by ESR spectrometry. In addition, NNP and NNVP could both decrease the mRNA expression of proinflammatory cytokines, TNF-α, IL-2 and IL-6. In conclusion, nitronyl nitroxide radicals NNP and NNVP were shown to have antiviral activities against CVB3 and they may represent potential therapeutic agents for viral myocarditis.

Expression of IL-23/Th17 Pathway in a Murine Model of Coxsackie Virus B3-Induced Viral Myocarditis

BackgroundThe IL-23/Th17 pathway is implicated in the pathogenesis of a number of chronic inflammatory and autoimmune diseases. Whether it regulates the viral myocarditis (VMC) is unknown.ResultsTo examine the pathogenesis role...

Expression of il-23/th17 pathway in a murine model of coxsackie virus b3-induced viral myocarditis

BackgroundThe IL-23/Th17 pathway is implicated in the pathogenesis of a number of chronic inflammatory and autoimmune diseases. Whether it regulates the viral myocarditis (VMC) is unknown.ResultsTo examine the pathogenesis role of IL-23/Th17 axis in VMC, we used male BALB/c mice to induced VMC by Coxsackie virus B3 (CVB3) peritoneal injection. IL-23, IL-17, and signal transducer and activator of transcription 3 (STAT3) mRNA in the myocardium of VMC mice were assessed by semi-quantitative RT-PCR. IL-23 and IL-17 protein from blood serum were evaluated by ELISA. Phosphorylated-STAT3 (p-STAT3) protein expression in the myocardium was evaluated by immunohistochemical staining. Flow cytometric analysis was used to evaluate the frequencies of Th17 subsets. Isolated CD4+ T cells from VMC mice were cultured with recombinant IL-23(rIL-23) in vitro. In addition, a STAT3-specific inhibitor (S3I-201) was used to test whether regulation of STAT3 could be partly responsible for Th17 diminution. Results showed that expression of IL-23, IL-17, STAT3 mRNA and protein increased in VMC mice. When purified CD4+ T cells derived from VMC mice were cultured in vitro with rIL-23, the frequency of Th17 cells was dramatically increased, accompanied by significantly enhanced production of IL-17 in the supernatants of cultured CD4+ T cells. S3I-201 significantly restrained Th17 cell proliferation.ConclusionsThe IL-23/Th17 pathway axis is strongly expressed in murine VMC, identifying a novel pathway of potential significance in viral myocarditis.

The heart-protective mechanism of Qishaowuwei formula on murine viral myocarditis induced by CVB3

Aim of study The heart-protective effect and mechanism of Qishaowuwei formula (QSW), a Traditional Chinese Medicine formula composed of Radix Astragali, Radix Paeoniae Rubra and Fructus Schisandrae was investigated on murine model of viral myocarditis (VMC) induced by Coxsackievirus B3 (CVB3). Materials and methods Mice were randomly divided into infected control group, QSW high dose group, QSW medium dose group, QSW low dose group and Vitamin C plus Ribavirin treatment group. 50 mice were included in each group. The day of virus inoculation was defined as day 0 and the drug treatment continued once a day for 14 days. Mice were sacrificed on days 3, 7, 14, 21 postinoculation (p.i.). The histopathological changes of myocardium, CVB3 RNA copies in the myocardium, cardiomycytic apoptosis, the serum level of superoxide dismutase (SOD) and maleic dialdehyde (MDA) and the phenotype of T lymphocytes subsets in peripheral blood was analyzed. Results QSW treatment significantly increase the survival rate ( p < 0.05) in VMC model. Histopathology and flow cytometry inspection revealed low ratio of cardiomyocytes necrosis and apoptosis in QSW treated mice with dose dependent manner. The cardiomyocytic ultra-structure observed by transmission electron microscope also supported the above results. The ameliorated tissue damage was consistent with reduced CVB3 copy numbers detected by real-time PCR in the myocardium of QSW treated mice. The antioxidant effect of QSW was proved by elevated activity of SOD and reduced level of MDA in the serum. Furthermore, the disturbed balance of CD4+ and CD8+ subsets in peripheral blood was restored. Conclusion These results demonstrated QSW had potent protective effect against CVB3-induced heart injury and this effect might be mediated by its inhibition on viral replication, antioxidant activity and immunoregulation mechanism.

L‐arginine ameliorates experimental autoimmune myocarditis by maintaining extracellular matrix and reducing cytotoxic activity of lymphocytes

It was previously shown that administration of the nitric oxide synthase inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME) aggravated murine viral myocarditis by increasing myocardial virus titres. Experimental autoimmune myocarditis in mice and rats mimics human fulminant myocarditis. The effects of L-arginine, a precursor of nitric oxide, upon heart failure in experimental autoimmune myocarditis were evaluated. Dietary L-arginine (L-arginine group) and L-arginine plus N(G)-nitro-L-arginine methyl ester (L-arginine + l-NAME group) were administered to C57BL/6 mice immunized with porcine cardiac myosin over 3 weeks. An untreated myocarditis group was prepared. Cardiac damage was less in the L-arginine group compared with the other two groups, as was incidence of heart failure. In addition, extracellular matrix change was less prominent in the L-arginine group. Plasma concentrations of nitric oxide were elevated in the L-arginine group. Cytotoxic activities of lymphocytes were lower in L-arginine group than in other two groups. L-arginine treatment may be effective in preventing the development of heart failure in experimental myocarditis by maintaining extracellular matrix and reducing the cytotoxic activity of lymphocytes.

Immunohistochemical characterization of infiltrating mononuclear cells in the rat heart with experimental autoimmune giant cell myocarditis.

The pathogenesis of giant cell myocarditis remains unclear. Subsets of inflammatory infiltrating cells may reflect the pathogenesis and etiology of the disease. Therefore, we examined subsets of infiltrating mononuclear cells in the heart of the rat with experimental giant cell myocarditis. Lewis rats were immunized with cardiac myosin in Freund's complete adjuvant (FCA). Severe myocarditis characterized by congestive heart failure and multinucleated giant cells were elicited. The lesions were composed of predominant mononuclear cells, polymorphonuclear neutrophils and fragments of degenerated myocardial fibres. The subsets of infiltrating mononuclear cells were investigated using MoAbs against rat CD4+ T cell (W3/25), CD8+ T cell (CX8), B cell (OX33) and macrophage (OX42). By serial examination, bound immunoglobulin could only be found on degenerated myocardial fibres. In this model, most infiltrating mononuclear cells were composed of macrophages and CD4+ T cells. The frequencies of macrophages and CD4+ T cells were 73.7% and 13.8%, respectively. CD8+ T cells were scarce and B cells were rare in the lesions. The frequencies of CD8+ T cells and B cells were 4.5% and 0.4%, respectively. The dominance of macrophages and CD4+ T cells was the constant finding among the sites of the lesions and throughout the course of the disease. These characteristic subsets of infiltrating cells were in contrast to those of murine viral myocarditis which were mainly composed of natural killer (NK) cells and CD8+ T cells. Clarifying the subsets of infiltrating cells in myocarditis may contribute to differential diagnosis of myocarditis between viral and autoimmune types. From this study, the pathogenesis of experimental autoimmune giant cell myocarditis seemed to be closely related to CD4+ T cells and macrophages.

Antiviral effects of sophoridine against coxsackievirus B3 and its pharmacokinetics in rats

Coxsackievirus B3 (CVB3) is a major pathogen for acute and chronic viral myocarditis. The aim of this study was to investigate the antiviral effects of sophoridine, an alkaloid extracted from Chinese medicinal herb, Sophora flavescens, against CVB3, and the underlying pharmacokinetics. First, we determined the antiviral effects of sophoridine against CVB3 in in vitro (primarily cultured myocardial cells), in vivo (BALB/c mice) and serum pharmacological experiments. Then, we determined the pharmacokinetic behavior in serum samples of SD rats after oral administration by HPLC. Finally, we determined the effects of sophoridine on the production of cytokines in a murine viral myocarditis model by measuring mRNA expression of some important cytokines in hearts of infected BALB/c mice by RT-PCR. We found that sophoridine exhibited obvious antiviral effects both in vitro and in vivo, and serum samples obtained from rats with oral administration of sophoridine reduced the virus titers in infected myocardial cells. The serum concentration profile correlated closely with antiviral activity profile. Moreover, sophoridine significantly enhanced mRNA expression of IL-10 and IFN-γ, but decreased TNF-α mRNA expression. In conclusion, sophoridine possesses antiviral activities against CVB3, by regulating cytokine expression, and it is likely that sophoridine itself, not its metabolites, is mainly responsible for the antiviral activities. Therefore, sophoridine may represent a potential therapeutic agent for viral myocarditis.

Collagen degradation in a murine myocarditis model: relevance of matrix metalloproteinase in association with inflammatory induction.

Myocardial collagen degradation is regulated by matrix metalloproteinases (MMPs) and tissue inhibitors of matrix metalloproteinase (TIMPs). The possible relevance of MMPs in association with the inflammatory induction was investigated in a murine coxsackievirus B3 myocarditis model. Hearts from viral infected and sham-infected BALB/c mice were analyzed by semi-quantitative RT-PCR, picrosirius red staining, Western blot analysis, and immunohistochemistry. In viral infected mice, both mRNA and protein abundance for collagen type I remained unaltered. In addition, picrosirius red staining showed the unchanged total collagen content. However, degraded soluble fraction of collagen type I protein was increased. Moreover, the mRNA abundance for MMP-3 and MMP-9 was upregulated, whereas the mRNAs for TIMP-1 and TIMP-4 were downregulated, respectively. The upregulation of MMP-3/MMP-9 and downregulation of TIMP-4 were confirmed at the protein level, and were associated with significantly increased mRNA levels of interleukin 1beta, tumor necrosis factor-alpha, transforming growth factor-beta1 and interleukin-4. The increment of MMPs in the absence of counterbalance by TIMPs may lead to a functional defect of the myocardial collagen network by posttranslational mechanisms. This may contribute significantly to the development of left ventricular dysfunction in murine viral myocarditis. The inflammatory response with induction of cytokines may mediate the dysregulation of the myocardial MMP/TIMP systems.

Oral administration of L-arginine prevents congestive heart failure in murine viral myocarditis.

It was previously demonstrated that administration of the nitric oxide synthase inhibitor NG-nitro-l-arginine methyl ester (l-NAME) aggravated viral myocarditis in mice. In the current study, the effects of l-arginine, a precursor of nitric oxide, on congestive heart failure (CHF) in myocarditis were evaluated. Dietary l-arginine and l-arginine plus l-NAME (l-arginine + l-NAME group) were administered to encephalomyocarditis virus-infected BALB/c mice over 4 weeks (experiment I) and to encephalomyocarditis virus-infected DBA/2 mice from the 4th through 12th weeks after the virus inoculation (experiment II). An infected control was prepared in each experiment. In experiment I, survival was higher in the l-arginine group compared with the other two groups, and cardiac damage was less, as was incidence of CHF. In addition, extravasated fibrin was less prominent in the l-arginine group. Plasma concentrations of l-arginine and nitric oxide were elevated in the l-arginine group. In experiment II, plasma cardiomyopathic lesions in the l-arginine group were less prominent and were associated with lower plasma catecholamine and lower myocardial collagen concentrations compared with the other two groups. l-arginine treatment may be effective in preventing the development of CHF in viral myocarditis by modifying postmyocarditic architectural remodeling.

Expression of tumor necrosis factor ligand superfamily costimulatory molecules CD27L, CD30L, OX40L and 4-1BBL in the heart of patients with acute myocarditis and dilated cardiomyopathy

Background: T-cell-mediated myocardial damage is known to be involved in acute myocarditis and dilated cardiomyopathy. Recently, we found that tumor necrosis factor (TNF) ligand superfamily costimulatory molecules, especially 4-1BBL, played an important role in the myocardial damage of murine acute viral myocarditis. Methods and results: To investigate the roles for CD27L, CD30L, OX40L and 4-1BBL, which belong to TNF ligand superfamily, in the development of acute myocarditis and dilated cardiomyopathy, we analyzed the expression of these antigens in the myocardial tissues of patients with acute myocarditis and dilated cardiomyopathy. We also examined expression of the receptors for these molecules, CD27, CD30, OX40 and 4-1BB, which belong to TNF receptor superfamily, on the infiltrating cells. Strong expression of CD27L, CD30L and 4-1BBL and weak to moderate expression of OX40L was found in the cardiac myocytes of patients with acute myocarditis. Moderate expression of CD27L, CD30L and 4-1BBL and weak expression of OX40L was found on the cardiac myocytes of patients with dilated cardiomyopathy. Most of the infiltrating cells expressed CD27, CD30 and 4-1BB and a part of the infiltrating cells expressed OX40. Conclusions: Our findings suggest that expression of TNF ligand superfamily costimulatory molecules on cardiac myocytes may play a role in the cell-mediated myocardial damage in patients with acute myocarditis and dilated cardiomyopathy as in murine viral myocarditis.

Effect of prevention and treatment of murine acute viral myocarditis and protection of lymphoid organ atrophy with xinkang oral liquid"Equation missing"

The effect of prevention and treatment of Xinkang oral liquid, ( , XKOL) on experimental coxsackievirus B3 (CVB3) myocarditis mice model were investigated. The mice were inoculated intraperitoneally with 0.3 ml of 105 TCID50 of CVB3 to induce acute viral myocarditis model. These mice were divided into model control group (Group A), prevention high dosage group (Group B) and prevention low dosage group (Group C), treatment high dosage group (Group D) and treatment low dosage group (Group E), respectively. In addition, XKOL control group (Group F) and normal control group (Group G) were not infected with CVB3 intraperitoneally. The administration of XKOL in Group B and C began 2 days before virus infection. All animals were sacrificed on day 20 for evaluation. Histological examination showed extensive myocardial necrosis and cell infiltration in most of Group A mice, but necrosis and cell infiltration were less severe in Group B,C,D and E mice. Thymus weight in Group B,C,D and E mice were heavier and less cell depletion occurred than those in Group A. The XKOL could effectively inhibit myocardial CVB3 replication, reduce the myocardial inflammatory response, lower incidence rate of myocarditis and prevent the disease associated lymphoid organ atrophy in this animal models.