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Acute viral myocarditis can progress to chronic myocarditis leading to dilated cardiomyopathy (DCM). Persistent CD4 + T-cell-mediated autoimmunity triggered by infection plays a critical role in this progression. Increasing evidence demonstrates that effector memory CD4 + T (CD4 + T EM ) cells, a subset of memory CD4 + T cells, are crucial pathogenic mediators of many autoimmune diseases. However, the role of CD4 + T EM cells during the progression from acute viral myocarditis to DCM remains unknown. In this study, we observed an increase in CD4 + T EM cells both in the periphery and the heart, and memory CD4 + T cells were the predominant sources of IL-17A and IFN-γ among inflamed heart-infiltrating CD4 + T cells during the progression from acute myocarditis to chronic myocarditis and DCM in CVB3-induced BALB/c mice. Moreover, splenic CD4 + T EM cells sorted from DCM mice induced by CVB3 were found to respond to cardiac self-antigens ex vivo. Additionally, adoptive transfer experiments substantiated their pathogenic impact, inducing sustained myocardial inflammation, tissue fibrosis, cardiac injury, and impairment of cardiac systolic function in vivo. Our findings illustrate that long-lived CD4 + T EM cells are important contributors to the progression from acute viral myocarditis into DCM., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Although most patients with acute viral myocarditis recover spontaneously, some patients progress to heart failure. Perturbations in innate immunity may partially explain the heterogeneity of clinical outcomes. As the most abundant immune cells in the heart, cardiac macrophages have heterogeneous origins, including embryonic-derived resident macrophages (ResMϕs) and monocyte-derived macrophages (MoMFs). However, the time course change and role of cardiac macrophage subsets has not been fully explored. In the present study, we found that BALB/c mice had prolonged MoMF accumulation and low proportions of ResMϕs that could not be restored to normal levels. MoMFs of BALB/c mice generally exhibit an M1-dominant functional phenotype. Moreover, the preferential depletion of MoMF by a C-C chemokine receptor type 2 (CCR2) inhibitor resulted in improved acute myocarditis and chronic fibrosis, as well as the recovery of ResMϕs number and reduced CD4 + T cell expansion. Hence, immunomodulatory therapy that targets the balance among cardiac macrophages and modulates their function is expected to prevent the progression of cardiac injury to overt heart failure and improve adverse outcomes., (© 2023 Wiley Periodicals LLC.)

Background: Acute viral myocarditis (AVMC) is an inflammatory disease of the myocardium. Evidence indicates that dysbiosis of gut microbiome and related metabolites intimately associated with cardiovascular diseases through the gut-heart axis., Methods: We built mouse models of AVMC, then applied 16 S rDNA gene sequencing and UPLC-MS/MS metabolomics to explore variations of gut microbiome and disturbances of cardiac metabolic profiles., Results: Compared with Control group, analysis of gut microbiota showed lower diversity in AVMC, decreased relative abundance of genera mainly belonging to the phyla Bacteroidetes, and increased of phyla Proteobacteria. Metabolomics analysis showed disturbances of cardiac metabolomics, including 62 increased and 84 decreased metabolites, and mainly assigned to lipid, amino acid, carbohydrate and nucleotide metabolism. The steroid hormone biosynthesis, cortisol synthesis and secretion pathway were particularly enriched in AVMC. Among them, such as estrone 3-sulfate, desoxycortone positively correlated with disturbed gut microbiome., Conclusion: In summary, both the structure of the gut microbiome community and the cardiac metabolome were significantly changed in AVMC. Our findings suggest that gut microbiome may participate in the development of AVMC, the mechanism may be related to its role in dysregulated metabolites such as steroid hormone biosynthesis., (© 2023. The Author(s).)

Viral myocarditis (VMC), which is defined as inflammation of the myocardium with consequent myocardial injury, may develop chronic disease eventually leading to dilated cardiomyopathy (DCM). Molecular mechanisms underlying the progression from acute VMC (aVMC), to chronic VMC (cVMC) and finally to DCM, are still unclear. Here, we established mouse models of VMC and DCM with Coxsackievirus B3 infection and conducted NMR-based metabolomic analysis of aqueous metabolites extracted from cardiac tissues of three histologically classified groups including aVMC, cVMC and DCM. We showed that these three pathological groups were metabolically distinct from their normal counterparts and identified three impaired metabolic pathways shared by these pathological groups relative to normal controls, including nicotinate and nicotinamide metabolism; alanine, aspartate and glutamate metabolism; and D-glutamine and D-glutamate metabolism. We also identified two extra impaired metabolic pathways in the aVMC group, including glycine, serine and threonine metabolism; and taurine and hypotaurine metabolism Furthermore, we identified potential cardiac biomarkers for metabolically distinguishing these three pathological stages from normal controls. Our results indicate that the metabolomic analysis of cardiac tissues can provide valuable insights into the molecular mechanisms underlying the progression from acute VMC to DCM.

Although B cells are essential for humoral immunity and show noteworthy immunomodulatory activity through antibody-independent functions, the role of B cells in regulating Treg cell responses remains controversial. Tregs (CD4+CD25+Foxp3+) are considered to play an immunoprotective role in viral myocarditis (VMC) by controlling autoimmune effector T cells. Here, we proved that B-cell knockout can not only lead to significant reductions in Tregs in the spleen, blood, and heart of VMC mice but also decrease the activation and immune function of splenic Tregs, which was reversed by adoptive transfer of B cells; the transcription levels of TGF-β and Foxp3 in the myocardium were also significantly reduced. B-cell depletion by anti-CD20 impaired the anti-inflammatory function of splenic Tregs and the homeostasis of myocardial Tregs population. Moreover, B cells can convert CD4+CD25- T cells into Foxp3+ and Foxp3-, two functionally suppressive Treg subgroups. Although the reduction in myocardial inflammation in BKO mice indicates that B cells may play a proinflammatory role, the beneficial side of B cells cannot be ignored, that is, to control autoimmunity by maintaining Treg numbers. The results observed in the animal model of VMC highlight the potential harm of rituximab in the nonselective depletion of B cells in clinical applications., (© The Author(s) 2022. Published by Oxford University Press on behalf of the British Society for Immunology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Viral myocarditis (VMC) is the main cause of sudden acute heart failure and cardiac death in adolescents; however, treatment for VMC is limited. Trehalose is a natural non-reductive disaccharide that protects against cardiovascular diseases by inducing autophagy. The protective effect of trehalose on VMC and the specific mechanism remains unclear. In this study, we established a VMC mouse model, treated with trehalose in vivo, and cultured B cells from VMC mice with trehalose in vitro to elucidate the effect of trehalose on B cells in acute VMC. Trehalose alleviated myocardial injury in VMC mice and increased the number of autophagosomes, LC3II/LC3I ratio, and expression level of LAMP2, whereas it decreased the expression of p62 in VMC-B cells. Bafilomycin A1 suppressed VMC-B cell autophagy induced by trehalose. At the mechanistic level, trehalose treatment significantly upregulated the phosphorylation of AMPK and ULK1 in VMC-B cells. Dorsomorphin and SBI-0206965 abolished the increased phosphorylation level and altered the expression levels of autophagy-related proteins. In conclusion, trehalose alleviates myocardial inflammatory damage of VMC by inducing B cell autophagy, mediated by the AMPK/ULK1 signalling pathway. Thus, trehalose may be a potentially useful molecule for alleviating myocardial injury in VMC., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Viral myocarditis (VMC) is an inflammatory heart condition which can induce dilated cardiomyopathy (DCM). However, molecular mechanisms underlying the progression of VMC into DCM remain exclusive. Here, we established mouse models of VMC and DCM by infecting male BALB/c mice with Coxsackievirus B3 (CVB3), and performed NMR-based metabonomic analyses of mouse sera. The mouse models covered three pathological stages including: acute VMC (aVMC), chronic VMC (cVMC) and DCM. We recorded 1 D 1 H-NMR spectra on serum samples and conducted multivariate statistical analysis on the NMR data. We found that metabolic profiles of these three pathological stages were distinct from their normal controls (CON), and identified significant metabolites primarily responsible for the metabolic distinctions. We identified significantly disturbed metabolic pathways in the aVMC, cVMC and DCM stages relative to CON, including: taurine and hypotaurine metabolism; pyruvate metabolism; glycine, serine and threonine metabolism; glycerolipid metabolism. Additionally, we identified potential biomarkers for discriminating a VMC, cVMC and DCM from CON including: taurine, valine and acetate for aVMC; glycerol, valine and leucine for cVMC; citrate, glycine and isoleucine for DCM. This work lays the basis for mechanistically understanding the progression from acute VMC to DCM, and is beneficial to exploitation of potential biomarkers for prognosis and diagnosis of heart diseases.

Viral myocarditis (VMC) is the major cause of sudden death in adolescents. To date, no effective treatment has been identified for VMC. Studies have shown that T helper (Th) cells such as Th1, Th2, Th17, and Th22 cells are involved in the pathogenesis of VMC. However, the role of B cells and their impact on Th cells in VMC is unclear. In this study, we investigated the role of B cells in Th cell differentiation in myocardial damage in an animal model of VMC. C57BL/6 mice were infected with Coxsackievirus B3 (CVB3) intraperitoneally or injected with phosphate-buffered saline as a control condition. At day 7, samples from these mice were analyzed by histology, ELISA, flow cytometry, and gene expression assays. We found that TNF-α-, IL-6-, and IL-17-producing B cell numbers were significantly increased, while IL-4-producing B cell population was significantly reduced in acute VMC. Furthermore, we performed B cell knockout (BKO), SCID, and SCID+B cells reconstitution experiments. We found that BKO alleviated the cardiac damage following CVB3 infection, may hamper the differentiation of Th1 and Th17 cells, may promote the differentiation of Th2 cells, and proved ineffective for the differentiation of Th22 cells. In contrast, SCID+B cells reconstitution experiment exacerbated the cardiac damage. Ex vivo studies further revealed that B cells promote the differentiation of Th1 and Th17 cells and inhibit the differentiation of Th2 cells. Our study shows that B cells are activated and have strong abilities of antigen presentation and producing cytokines in VMC; B cells not only play a pathogenic role in VMC independent of T cells but also promote Th1 and Th17 cell differentiation, and hamper Th2 cell differentiation in VMC., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

The immune system and the nervous system depend on each other for their fine tuning and working, thus cooperating to maintain physiological homeostasis and prevent infections. The cholinergic system regulates the mobilization, differentiation, secretion, and antigen presentation of adaptive and innate immune cells mainly through α7 nicotinic acetylcholine receptors (α7nAChRs). The neuro-immune interactions are established and maintained by the following mechanisms: colocalization of immune and neuronal cells at defined anatomical sites, expression of the non-neuronal cholinergic system by immune cells, and the acetylcholine receptor-mediated activation of intracellular signaling pathways. Based on these immunological mechanisms, the protective effects of cholinergic system in animal models of diseases were summarized in this paper, such as myocardial infarction/ischemia-reperfusion, viral myocarditis, and endotoxin-induced myocardial damage. In addition to maintaining hemodynamic stability and improving the energy metabolism of the heart, both non-neuronal acetylcholine and neuronal acetylcholine in the heart can alleviate myocardial inflammation and remodeling to exert a significant cardioprotective effect. The new findings on the role of cholinergic agonists and vagus nerve stimulation in immune regulation are updated, so as to develop improved approaches to treat inflammatory heart disease., (Copyright © 2021 The Authors. Published by Elsevier B.V. All rights reserved.)

Aims: This work aimed to evaluate the regulatory function of IL-10-producing B cells in viral myocarditis (VMC)., Main Methods: We adoptively transferred purified IL-10-producing B cells to VMC mice via the tail vein. We observed the inflammatory responses and cardiac lesions by histological analysis, examined the proportions of spleen Th1 and T17 cells by flow cytometry and expression levels of related transcription factors (T-bet and RORγt) by reverse transcription polymerase chain reaction (RT-PCR), and calculated the cardiac pathological scores and the mean survival times., Key Findings: IL-10-producing B cells were found to be T cell-dependent in the pathogenesis of VMC. They mainly downregulated T-bet and RORγt mRNA levels to decrease the proportions of Th1 and Th17 cells, thereby restraining the inflammation and damage in the myocardium in B cell-deficient VMC mice. Adoptive transfer of IL-10-producing B cells before VMC induction also normalized the inflammatory responses and prolonged the survival time in wild-type (WT) VMC mice. While the transfer of IL-10-producing B cells on day 3 of VMC alleviated the severity of disease, it did not extend the mean survival time of VMC mice. By contrast, IL-10-producing B cells showed no effect on day 7 of VMC. In conclusion, IL-10-producing B cells downregulate the proportion of Th1 and Th17 cells to alleviate inflammatory damage in the myocardium during VMC before the induction or the early phase of disease., Significance: These findings suggest that IL-10-producing B cells may be a new therapeutic target for modulating the immune response in VMC., (Copyright © 2019 Elsevier Inc. All rights reserved.)

The role of B cells in viral myocarditis (VMC) remains controversial. In order to establish a role and mechanism of action for B cells in acute VMC, we established an acute VMC mouse model by intraperitoneal injection of Coxsackie virus group B type 3 (CVB3). At day 7, mice were analyzed using myocardial histopathology, and the presence of M2 macrophages in spleen and heart. Mice were divided into four groups, all having a C57BL/6 background: control group; wild-type (WT) VMC; mMt/mMt (-/-) VMC (BKO), and BKO + B cell VMC. A role for B cells was demonstrated by a significant reduction in myocardial pathological score and an increase in the frequency of M2 macrophages in the BKO group, when compared to the WT group. Once BKO mice underwent B cell reconstitution with isolated WT B cells, the myocardial pathological score was increased significantly, while the frequency of M2 macrophages decrease. Our findings demonstrate that B cells increase myocardial inflammation by suppressing M2 polarization in acute VMC in vivo.

The cluster of differentiation protein complex, CD80/CD86, regulates Th1/Th2 differentiation in autoimmune disease. In order to establish the effects of CD80/CD86 on Th17 cell differentiation in acute viral myocarditis (VMC), we infected C57BL/6 mice with Coxsackie virus B3 (CVB3) and examined the effects of the treatment with anti-CD80/CD86 monoclonal antibodies (mAbs) on Th17 cell differentiation in vivo. The effects of anti-CD80/CD86 mAbs on Th17 cell differentiation were further evaluated in vitro. The treatment with anti-CD80 mAb induced marked suppression of Th17 cell differentiation and ROR-γt mRNA expression, whereas anti-CD86 mAb alone had no effect, both in vivo and in vitro. Our finding that CD80 regulates Th17 differentiation supports the potential utility of anti-CD80 mAb as an effective new immunotherapeutic target in acute VMC.

Objective To analyze the migration and expression of CD34 + VLA-4 + cells under the induction of vascular cell adhesion molecule-1 (VCAM-1) in a murine model of acute viral myocarditis (VMC). Methods Frequency of CD34 + VLA-4 + cells in the myocardial tissues and peripheral blood were examined by flow cytometry. The mRNA and protein of VCAM-1 in the myocardial tissues were analyzed by real-time quantitative PCR and Western blotting. Results In the acute VMC mice, CD34 + VLA-4 + cell population in the myocardial tissues significantly increased at day 3, peaked at day 7, and then decreased, but it was still higher than that in the control group at day 14 and 28. It decreased in the peripheral blood at day 3, and then increased to the peak at day 7, thereafter it decreased, but was still higher than that in the control group at day 14 and 28. We found a high expression of VCAM-1 in the myocardial tissues of the acute VMC mice, paralleling the mobilization of CD34 + VLA-4 + cells in the myocardial tissues. Conclusion VCAM-1 promotes CD34 + VLA-4 + cell mobilization into the damaged myocardial tissues.

Objectives: Bone marrow-derived cells (BMCs) have recently been identified to play a vital role in repairing damaged myocardium; however, it is not known whether or not mobilization of BMCs is involved in the pathogenesis of acute viral myocarditis (VMC). Thus, we analyzed the expression of CD45+CD34+VLA-4+ cells and vascular cell adhesion protein-1 (VCAM-1) in a murine model of acute VMC., Methods: Male BALB/c mice were intraperitoneally infected with coxsackievirus B3 to establish acute VMC. The frequency of CD45+CD34+VLA-4+ cells in the heart, peripheral blood, and bone marrow was examined by flow cytometry 3, 7, 14, and 28 days after injection. Cardiac VCAM-1 and pathology scores were determined by immunohistochemistry, and myocardial VCAM-1, IL-1β, and TNF-α were analyzed by RT-PCR and Western blot., Results: In mice with acute VMC, the CD45+CD34+VLA-4+ cell population in the heart was significantly increased by day 7 and then decreased; in contrast, the CD45+CD34+VLA-4+ cell population decreased in the bone marrow and peripheral blood by day 3 and then increased. High expression of VCAM-1 was detected in the heart in parallel with CD45+CD34+VLA-4+ cell expression., Conclusions: In mice with acute VMC, VCAM-1-induced CD45+CD34+VLA-4+ cell mobilization into the injured heart is involved in the repair of injured myocardium., (© 2017 S. Karger AG, Basel.)

Background: Interleukin-10 (IL-10)-producing B cells, a subset of regulatory B cells, play critical roles in autoimmune and infectious diseases. However, the role of IL-10-producing B cells in acute viral myocarditis (AVMC) remains unknown., Methods: BALB/c mice were intraperitoneally (i. p.) infected with coxsackievirus B3 (CVB3) to establish AVMC models (AVMC group), while control mice (control group) were treated with phosphate-buffered saline (PBS) i. p. According to the time after injection, the AVMC group mice or control group mice were randomly separated into 1 week and 2 week subgroup. Myocardial histopathological changes were observed by hematoxylin and eosin staining and the frequency of splenic IL-10-producing B cells was measured by flow cytometry., Results: Histopathologic examination of heart tissues showed that mice infected with CVB3 developed AVMC. Compared with control group, the frequency of splenic IL-10-producing B cells was increased significantly in the AVMC group, with the 1 week AVMC subgroup (3.58 ± 0.47%) higher than the 2 week AVMC subgroup (2.50 ± 0.42%) (all P < 0.05)., Conclusions: IL-10-producing B cells are increased in CVB3-induced AVMC, indicating that IL-10-producing B cells may play an important role in the pathogenesis of CVB3-induced AVMC.

Background: A new subset of T helper (Th) cells, named IL-22-producing Th22 cells, was identified recently. Th22 cells have been implicated in immunity and inflammation. However, the role of these cells in the progression from acute viral myocarditis (AVMC) to dilated cardiomyopathy (DCM) and myocardial fibrosis remains unknown., Methods: BALB/c mice were repeatedly i.p. infected with Coxsackie virus B3 (CVB3) to establish models of AVMC, chronic myocarditis and DCM. On week 2, 12 and 24 post initial injection, the percentage of splenic Th22 cells, the levels of plasma IL-22, cardiac IL-22 receptor (IL-22R) expression, and indicators of myocardial fibrosis were measured. Further, mice with AVMC and chronic myocarditis were treated with an anti-IL-22 neutralizing antibody (Ab). The collagen volume fraction (CVF), the percentage of splenic Th22 cells, plasma IL-22 levels, cardiac IL-22R expression and indicators of myocardial fibrosis were then monitored., Results: Compared to control mice at the same time points, AVMC, chronic myocarditis and DCM mice have higher percentage of splenic Th22 cells, higher plasma IL-22 levels, increased cardiac IL-22R, as well as increased collagen typeI-A1 (COL1-A1), collagen type III-A1 (COL3-A1) and matrix metalloproteinase-9 (MMP9) expression. However, the expression of tissue inhibitor of metalloproteinase-1(TIMP-1) was decreased. Treatment of AVMC and chronic myocarditis mice with an anti-IL-22 Ab decreased the survival rate and exacerbated myocardial fibrosis. The percentage of splenic Th22 cells, plasma IL-22 levels and cardiac IL-22R expression also decreased in anti-IL-22 Ab treatment group as compared to IgG and PBS treated groups of AVMC and chronic myocarditis mice. Moreover, increased expression of COL1-A1, COL3-A1, MMP9 but decreased expression of TIMP-1 were observed in anti-IL-22 Ab mouse group., Conclusions: Th22 cells play an important role in the pathogenesis of CVB3-induced mouse chronic myocarditis and DCM. IL-22 is a myocardium-protective cytokine by inhibiting myocardial fibrosis. Therefore, Th 22 cells may be considered as potential therapeutic targets for DCM.

Objective: To explore the effect of interleukin-17A (IL-17A) on the serum level of antiheart autoantibodies in mice with viral myocarditis., Methods: Male wild-type (WT) and IL-17A-deficient (IL-17A(-/-)) BALB/c mice were intraperitoneally injected with Coxsackie virus B3 (CVB3) for establishing VMC models (VMC-WT group and VMC-IL-17A(-/-) group). Meanwhile, a control group (WT group) of WT mice were established by i.p. administration of phosphate buffered saline (PBS). Paraffin sections of cardiac tissues were made 14 days after CVB3 injection. Myocardial histopathologic changes were evaluated by HE staining. The levels of anti-adenine nucleotide translocator (ANT) autoantibody, anti-β-myosin heavy chain (β-MHC) autoantibody and anti-cardiac L-type calcium channel (CACH2) autoantibody in sera were measured by ELISA., Results: Compared with WT group, the levels of anti-ANT-autoantibody and anti-β-MHC-autoantibody significantly increased in VMC-WT group (P<0.01, P<0.05), while the concentration of anti-CACH2-autoantibody showed no significant difference between WT and VMC-WT groups (P>0.05). Compared with VMC-WT group, the level of anti-ANT-autoantibody was reduced in VMC-IL-17A(-/-) group (P<0.05), while the levels of anti-β-MHC-autoantibody and anti-CACH2-autoantibody showed no significant difference between them (P>0.05)., Conclusion: IL-17A contributed to the secretion of anti-ANT-autoantibody of VMC mice, but had no effect on the secretion of anti-β-MHC-autoantibody and anti-CACH2-autoantibody in VMC mice.

Objective: Interleukin-27 (IL-27) has been reported to reduce the levels of interleukin-17 (IL-17) and alleviate the severity of experimental autoimmune myocarditis. IL-17, an important tissue-protective cytokine in viral myocarditis (VMC), has been reported to increase synovial expression of IL-27 in rheumatoid arthritis. However, the influence of IL-17 on IL-27 expression in murine model of VMC remains unknown., Methods: Wild-type (WT) and IL-17A-deficient (IL-17A(-/-)) mice on the BALB/c background were intraperitoneally (i.p) injected with coxsackievirus B3 (CVB3) for establishing VMC models. Cardiac tissue was obtained on day 7 after CVB3 injection. Myocardial histopathologic changes were observed by hematoxylin-eosin (HE) stained myocardial sections.Expression of IL-27 in heart and serum was measured by real-time reverse transcription-polymerase chain reaction (RT-PCR) and enzyme linked immunosorbent assay (ELISA), respectively. Furthermore, splenic lymphocytes and peritoneal macrophages were purified 1 week after injection from WT mice.Isolated lymphocytes were cultured in the presence of different concentrations (0 and 25 ng/ml) of recombinant IL-17 (rIL-17) for 24 h. Macrophages were cultured with different concentrations of rIL-17 (0 and 10 ng/ml) for 48 h.IL-27 mRNA expression of cultured cells was assayed by RT-PCR, and their protein level in the culture supernatant was measured by ELISA., Results: Compared with WT mice, significantly less cardiac inflammation was evidenced in the heart of IL-17A-/- mice (0.9 ± 0.3 vs.1.9 ± 0.5) , relative cardiac IL-27 p28 mRNA expressions (1.11 ± 0.24 vs.3.1 ± 0.8) and serum IL-27 protein[(72 ± 18) pg/ml vs.(95 ± 25) pg/ml] were also significantly lower in IL-17A-/- mice (all P < 0.05).In the culture lymphocytes, the relative mRNA (1.02 ± 0.13 vs.1.32 ± 0.21) and protein [(49 ± 9) pg/ml vs.(52 ± 11) pg/ml]expressions of IL-27 p28 and were similar post treatment with 0 and 25 ng/ml rIL-17 (all P > 0.05). Compared with 0 ng/ml rIL-17 culture with macrophages, higher relative mRNA (8.5 ± 3.1 vs.2.2 ± 0.7) and protein [(368 ± 95) pg/ml vs.(150 ± 38) pg/ml] expressions of IL-27 p28 were detected in 10 ng/ml rIL-17 group (all P < 0.05)., Conclusion: Our data indicates that cytokine IL-17 may contribute to the secretion of IL-27 in VMC mice.Furthermore, macrophages but not lymphocytes may be the important IL-27-producing immune cells and major target cells for IL-17. Thus,IL-27 and IL-17 might be actively involved in the pathogenesis of VMC.

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.

Interleukin (IL)-22 has either proinflammatory or tissue‑protective properties, depending on the nature of the affected tissue and the local cytokine milieu, including the presence or absence of IL-17A co-expression. We have previously demonstrated that IL-22 has critical anti-inflammatory and antiviral roles in mice with coxsackievirus B3 (CVB3)‑induced acute viral myocarditis (AVMC) in the presence of IL-17A. However, whether IL-17A determines the function of IL-22 in AVMC remains unknown. Therefore, the present study, in continuation of our previous investigations, aimed to determine whether IL-22 plays a distinctly different role in the absence of IL-17A in AVMC by using IL-17A-deficient mice. Results demonstrated that the neutralization of IL-22 in IL-17A‑deficient mice alleviated the severity of myocarditis. This was demonstrated by the lower pathological scores of heart sections and ratios of heart weight/body weight (HW/BW), reduced production of activator of transcription 3 (STAT3) and proinflammatory cytokines TNF-α and IL-6, followed by increased viral replication and decreased levels of the antiviral cytokine IFN-γ. Furthermore, the correlation between cardiac CVB3 RNA and IL-22 mRNA or IFN-γ mRNA was negative. In conclusion, IL-22 exacerbated the severity of AVMC and restrained viral replication in the absence of IL-17A. Spleen lymphocytes cultured with recombinant IL-17 (rIL-17) increased the production of IL-22. Combined with our previous data, these results indicate that IL-17A is not involved in regulating the antiviral role, however, may mediate the tissue-protective versus pathogenic properties of IL-22 in CVB3-induced AVMC in mice.

Background: Recently, a new subset of T helper (Th) cell that predominantly secret cytokine interleukin-22 (IL-22) is identified, termed Th22 cells. The Th22 subset has been demonstrated to be involved in immunity and tissue inflammation. However, the existence of Th22 cells and role of IL-22 in acute viral myocarditis (AVMC) remain unknown., Methods: BALB/c mice were intraperitoneally (i.p) infected with CVB3 for establishing AVMC models. Control mice were treated with phosphate-buffered saline (PBS) i.p. On day 14 post injection, frequencies of splenic Th22 cells were determined, productions of IL-22 and expressions of IL-22R (IL-22 receptor) were measured. To further investigate the effects of IL-22, AVMC mice treated with Anti-IL-22 neutralizing antibody were explored. The severity of AVMC were monitored; the frequencies of Th22 cells, the expressions of IL-22 and IL-22R were investigated; in addition to IFN-γ, inflammatory cytokines IL-17, TNF-α, IL-6 as well as IL-1β, were evaluated. Cardiac viral replication were detected., Results: Compared with control group, significant elevations of circulating Th22 cells and IL-22, cardiac protein and mRNA of IL-22, and IL-22R1 were demonstrated in AVMC group. Treatment of AVMC mice with Anti-IL-22 Ab exacerbated the severity of viral myocarditis, verified by lower survival rate, higher HW/BW ratios and cardiac pathological scores. Anti-IL-22 Ab decreased the frequencies of Th22 cells and the levels of IL-22, and increased the expressions of cardiac IL-22R1. Up-regulations of IL-17, IL-6 and TNF-α, down-regulations of IFN-γ proteins and gene expressions in the plasma and myocardium, were observed in Anti-IL-22 Ab group. Furthermore, neutralization of IL-22 significantly promoted cardiac viral replication., Conclusions: Our data indicate that the increased frequencies of IL-22-producing Th22 cells may play an important role in the pathogenesis of CVB3-induced mice AVMC, IL-22 may act as an myocardium-protective cytokine via the IL-22-IL-22R pathway, and suggest that targeting the Th22 cell and IL-22-IL-22R pathway could provide new therapeutic modalities for the treatment of CVB3-induced AVMC.

Background: Recently, a new subset of CD4+T helper(Th) cell that predominantly secret cytokine interleukin-9(IL-9) is identified, termed Th9 cell. It has been reported to participate in tissue inflammation and autoimmune responses, and induce disease which differed from Th17 cells. Th17 cells have been shown to play a critical role in viral myocarditis (VMC), but whether Th9 cells are involved in the pathogenesis of VMC remains unclear. Results: BALB/c mice were intraperitoneally (i.p) injected with coxsackie virus B3(CVB3) for establishing VMC models. Control mice were treated with phosphate-buffered saline i.p. On day 0,7,14,21,28,35,42 after injection, myocardial histopathological changes were evaluated by hematoxylin-eosin staining. Splenic Th17 and Th9 cells subsets were analyzed by flow cytometry. And cardiac IL-17, IL-9 mRNA were measured by semi-quantitative reverse transcription-PCR and nested PCR, respectively. Results showed the levels of Th17 cells and IL-17 mRNA obviously increased in VMC mice on 7 day after infection, peaked on day 28, and highly persisted to at least day 42 (p < 0.05). While the frequencies of Th9 cells and IL-9 mRNA showed no significant difference between VMC and control group throughout the course of the experiment(p > 0.05). Conclusions: It was differentiated Th17 but not Th9 cells significantly elevated in the development of CVB3-induced VMC. The microenvironment of VMC seemed to contribute to the differentiation and proliferation of Th17 rather than Th9 cells. Our preliminary data implied Th9 cells could not protect against VMC nor promote the disease. [ABSTRACT FROM AUTHOR]