Myosin Injection Research Articles

Human umbilical cord mesenchymal stem cells alleviate interstitial fibrosis and cardiac dysfunction in a dilated cardiomyopathy rat model by inhibiting TNF‑α and TGF‑β1/ERK1/2 signaling pathways.

Dilated cardiomyopathy (DCM) is a disease of the heart characterized by pathological remodeling, including patchy interstitial fibrosis and degeneration of cardiomyocytes. In the present study, the beneficial role of human umbilical cord-derived mesenchymal stem cells (HuMSCs) derived from Wharton's jelly was evaluated in the myosin-induced rat model of DCM. Male Lewis rats (aged 8-weeks) were injected with porcine myosin to induce DCM. Cultured HuMSCs (1×106 cells/rat) were intravenously injected 28 days after myosin injection and the effects on myocardial fibrosis and the underlying signaling pathways were investigated and compared with vehicle-injected and negative control rats. Myosin injections in rats (vehicle group and experimental group) for 28 days led to severe fibrosis and significant deterioration of cardiac function indicative of DCM. HuMSC treatment reduced fibrosis as determined by Masson's staining of collagen deposits, as well as quantification of molecular markers of myocardial fibrosis such as collagen I/III, profibrotic factors transforming growth factor-β1 (TGF-β1), tumor necrosis factor-α (TNF-α), and connective tissue growth factor (CTGF). HuMSC treatment restored cardiac function as observed using echocardiography. In addition, western blot analysis indicated that HuMSC injections in DCM rats inhibited the expression of TNF-α, extracellular-signal regulated kinase 1/2 (ERK1/2) and TGF-β1, which is a master switch for inducing myocardial fibrosis. These findings suggested that HuMSC injections attenuated myocardial fibrosis and dysfunction in a rat model of DCM, likely by inhibiting TNF-α and the TGF-β1/ERK1/2 fibrosis pathways. Therefore, HuMSC treatment may represent a potential therapeutic method for treatment of DCM.

Human umbilical cord mesenchymal stem cells alleviate acute myocarditis by modulating endoplasmic reticulum stress and extracellular signal regulated 1/2-mediated apoptosis.

Acute myocarditis is a non-ischemic inflammatory disease of the myocardium, and there is currently no standard treatment. Mesenchymal stem cells (MSCs) can alleviate myosin-induced myocarditis; however, the mechanism has not been clearly elucidated. In the present study, the authors investigated the ability of human umbilical cordMSCs (HuMSCs) to attenuate myocardial injury and dysfunction during the acute phase of experimental myocarditis. Male Lewis rats (aged 8 weeks) were injected with porcine myosin to induce myocarditis. Cultured HuMSCs (1×106 cells/rat) were intravenously injected 10 days following myosin injection. A total of 3 weeks following injection, this resulted in severe inflammation and significant deterioration of cardiac function. HuMSC transplantation attenuated infiltration of inflammatory cells and adverse cardiac remodeling, as well as reduced cardiomyocyte apoptosis. Furthermore, it was identified that HuMSC transplantation suppressed endoplasmic reticulum stress and extracellular signal-regulated kinase (ERK)1/2 signaling in experimental autoimmune myocarditis (EAM). The reduced number of TUNEL-positive apoptotic cells in myocardial sections from HuMSC-treated EAM rats compared with control demonstrates HuMSCs' anti-apoptotic function. Based on these data, the author suggested that treatment with HuMSCs inhibits myocardial apoptosis in EAM rats, ultimately protecting them from myocardial damage. The conclusion demonstrated that HuMSC transplantation attenuates myocardial injury and dysfunction in a rat model of acute myocarditis, potentially via regulation of ER stress, ERK1/2 signaling and induction of cardiomyocyte apoptosis.

Curcumin induces M2 macrophage polarization by secretion IL-4 and/or IL-13

AimsTo address the underlying mechanisms by which curcumin facilitates M2 phenotype polarization of macrophages and its roles in the protective effects during experimental autoimmune myocarditis (EAM). Methods and resultsThe expression of classic M2 markers, including macrophage mannose receptor (MMR), arginase-1 (Arg-1) and peroxisome proliferator-activated receptor-γ (PPAR-γ) was upregulated in curcumin-treated Raw264.7 macrophages. Curcumin increased interleukin-4 (IL-4) and interleukin-13 (IL-13) mRNA expression and protein secretion. Curcumin notably increased STAT6 phosphorylation. Leflunomide, a STAT6 inhibitor, and IL-4 and/or IL-13 neutralizing antibodies antagonized the induction of MMR, Arg-1 and PPAR-γ by curcumin in Raw264.7 cells. In vivo, 6-week old male Lewis rats were used to induce EAM and orally administrated with curcumin or corn oil for 3weeks after myosin injection. Cardiac functional parameters, including left ventricular fractional shortening (LVFS), ejection fraction (EF), left ventricular end-systolic diameter (LVEDs) and heart rate (HR) were significantly improved by curcumin treatment. Curcumin also reduced the inflammatory cell infiltration and myocardial mRNA levels of interleukin-1β (IL-1β) and inducible nitric oxide synthase (iNOS). Meanwhile, the myocardial mRNA levels of MMR and Arg-1 were markedly up-regulated by curcumin. Immunofluorescence assay showed that the number of CD68+ MMR+ and CD68+ Arg-1+ double positive macrophages in curcumin-treated myocardial tissue was significantly higher than untreated control. The number of CD68+ iNOS+ double positive macrophages was increased obviously in EAM group, but decreased markedly by curcumin treatment. ConclusionsTaken together, these results show that curcumin induces macrophage M2 polarization by secretion of IL-4 and/or IL-13. Curcumin ameliorates EAM by reducing infiltration inflammatory macrophages and by polarizing M0 and M1 macrophages to M2 phenotype.

Abstract 17425: Curcumin Ameliorates Experimental Autoimmune Myocarditis by Inducing M2 Macrophage Polarization

AIMS: Our previous study showed that curcumin, the main active ingredient in turmeric, induces the polarization of macrophages to anti-inflammatory M2 phenotype. In this study, we addressed the underlying mechanisms and observed whether curcumin exerts its protective effects on experimental autoimmune myocarditis (EAM) by facilitating the M2 phenotype polarization of macrophages. METHODS AND RESULTS: The mRNA and protein expression of M2 markers, including MMR, Arg-1, KLF-4 and PPAR-γ, were obviously up-regulated in curcumin-treated RAW264.7 macrophages. Curcumin increased not only IL-4 and IL-13 mRNA expression, but also protein content of these two cytokines in supernatant. The STAT6 inhibitor leflunomide antagonized the induction of MMR, Arg-1, KLF-4 and PPAR-γ by curcumin in RAW264.7 cells, which is suggested that curcumin induces macrophages M2 polarization through secretion of IL-4 and IL-13 in an autocrine manner. In vivo, 6-week-old male Lewis rats were immunized with myosin to induce EAM and orally administrated with curcumin (50mg/kg/day) or corn oil for 3 weeks after myosin injection. Cardiac functional parameters, including LVFS, EF, LVESD and HR, were significantly improved by curcumin treatment. Curcumin also reduced the heart weight-to-body weight ratio, inflammatory cell infiltration and the myocardial mRNA level of IL-1β and iNOS. Meanwhile, the myocardial mRNA level of KLF4, MMR and Arg-1 were markedly up-regulated by curcumin. Immunofluorescence assay showed that the number of CD68+MMR+ and CD68+Arg-1+ double positive macrophages in curcumin-treated myocardial tissue was obviously more than vehicle-treated ones. CONCLUSIONS: Taken together, these results show that curcumin ameliorates EAM by attenuating inflammation and by inducing macrophage M2 polarization.

Beneficial effect of a synthetic prostacyclin agonist, ONO-1301, in rat autoimmune myocarditis model

Injury to the heart can result in cardiomyocyte hypertrophy, fibrosis, and cell death. Myocarditis sometimes progresses to dilated cardiomyopathy. We previously reported that ONO-1301, a synthetic prostacyclin agonist with thromboxane-synthase inhibitory activity, promotes production of hepatocyte growth factor (HGF) from various cell types and ameliorates ischemia-induced left ventricle dysfunction in the mouse, rat and pig. Here, we investigated the therapeutic efficacy of ONO-1301 in a rat model of myosin-induced experimental autoimmune myocarditis, in which the heart transits from an acute inflammatory phase to a chronic dilated cardiomyopathy phase. Four weeks after myosin injection to Lewis rats, ONO-1301 (6mg/kg/day) was orally administered for 4 weeks (ONO-1301 group). Hemodynamic parameters and plasma brain natriuretic peptide (BNP) level were significantly improved by ONO-1301. Histological analysis revealed that capillary density in the myocardium was significantly increased by ONO-1301. ONO-1301 increased circulating endothelial progenitor cells (EPC) as determined by FACS analysis. These beneficial effects of ONO-1301 were partially abrogated by a neutralizing anti-HGF antibody (8mg/kg/dose). These findings indicate beneficial effects of ONO-1301 in a rat experimental autoimmune myocarditis model.

Inhibition of cardiac oxidative and endoplasmic reticulum stress-mediated apoptosis by curcumin treatment contributes to protection against acute myocarditis

Curcumin is used anecdotally as an herb in traditional Indian and Chinese medicine. In the present study, the effects and possible mechanism of curcumin in experimental autoimmune myocarditis (EAM) rats were further investigated. They were divided randomly into a treatment and vehicle group, and orally administrated curcumin (50 mg/kg/day) and 1% gum arabic, respectively, for 3 weeks after myosin injection. The results showed that curcumin significantly suppressed the myocardial protein expression of inducible nitric oxide synthase (iNOS) and the catalytic subunit of nicotinamide adenine dinucleotide phosphate reduced (NADPH) oxidase. In addition, curcumin significantly decreased myocardial endoplasmic reticulum (ER) stress signaling proteins and improved cardiac function. Furthermore, curcumin significantly decreased the key regulators or inducers of apoptosis. In summary, our results indicate that curcumin has the potential to protect EAM by modulating cardiac oxidative and ER stress-mediated apoptosis, and provides a novel therapeutic strategy for autoimmune myocarditis.

Curcumin Ameliorates Cardiac Inflammation in Rats with Autoimmune Myocarditis

Curcumin is a natural polyphenolic compound abundant in the rhizome of the perennial herb turmeric, Curcuma longa. It is commonly used as a dietary spice and coloring agent in cooking, and is used anecdotally as an herb in traditional Indian and Chinese medicine. It has been reported that curcumin has the potential to protect against cardiac inflammation through suppression of GATA-4 and nuclear factor-κB (NF-κB); however, no study to date has addressed the effect of curcumin on experimental autoimmune myocarditis (EAM) in rats. In this study, 8-week-old male Lewis rats were immunized with cardiac myosin to induce EAM. They were then divided randomly into a treatment or vehicle group and orally administrated curcumin (50 mg/kg/d) or 1% gum arabic, respectively, for 3 weeks after myosin injection. We performed hemodynamic, echocardiographic, hematoxylin and eosin staining, mast cell staining and Western blotting studies to evaluate the protective effect of curcumin in the acute phase of EAM. Cardiac functional parameters measured by hemodynamic and echocardiographic studies were significantly improved by curcumin treatment. Furthermore, curcumin reduced the heart weight-to-body weight ratio, area of inflammatory lesions and the myocardial protein level of NF-κB, interleukin (IL)-1β, tumor necrosis factor (TNF)-α and GATA-4. Our results indicate that curcumin has the potential to protect against cardiac inflammation through suppression of IL-1β, TNF-α, GATA-4 and NF-κB expresses, and may provide a novel therapeutic strategy for autoimmune myocarditis.

Sulfated Polysaccharide Fucoidan Ameliorates Experimental Autoimmune Myocarditis in Rats

Homing of cardiac myosin-specific CD4-positive T cells into the myocardium is the initial pathologic event of experimental autoimmune myocarditis (EAM). Subsequently, various bystander inflammatory cells are recruited into the myocardium crossing vascular endothelial cell walls. Sulfated polysaccharide fucoidan binds selectin nonselectively and blocks its function. Therefore, this study was designed to evaluate whether in vivo fucoidan treatment can improve EAM. A 21-day infusion of physiological saline or fucoidan was administrated intraperitoneally to the rats with sham operation (sham-saline, n = 5; sham-fucoidan, n = 6) or those with cardiac myosin injection (EAM-saline, n = 10; EAM-fucoidan, n = 10). After 3 weeks, fucoidan treatment improved left ventricular ejection fraction (79.04 ± 2.81 vs 65.94% ± 3.22%; P < .01 vs EAM-saline) with a reduced ratio of heart weight to body weight (4.016 ± 0.239 vs 4.975 ± 0.252 mg/g; P < .05 vs EAM-saline) in EAM. Furthermore, fucoidan treatment decreased serum levels of BNP (292.0 ± 53.4 vs 507.4 ± 89.2 ng/mL; P < .05 vs EAM-saline) and the myocarditis area (31.66 ± 1.53 vs 42.51% ± 3.24%; P < .01 vs EAM-saline) in EAM. These beneficial effects of fucoidan were accompanied by inhibition of both macrophage and CD4-positive T-cell infiltration into the myocardium. Fucoidan, a nonselective selectin blocker, attenuates the progression of EAM. This observation may be explained, at least in part, by blocking the extravasation of inflammatory cells into the myocardium.

A novel phenylpyridazinone, T-3999, reduces the progression of autoimmune myocarditis to dilated cardiomyopathy

Regardless of the origin, injury to the heart can result in cardiomyocyte hypertrophy, fibrosis, and cell death. Myocarditis often progresses to dilated cardiomyopathy (DCM), a major cause of heart failure. In our study, we used a rat model of myosin-induced experimental autoimmune myocarditis (EAM), in which the heart transits from an acute phase (inflammatory myocarditis) to a chronic phase (remodeling and DCM). Our objective was to investigate whether T-3999, a novel phenylpyridazinone, can reduce this progression. Four weeks after myosin injection, T-3999 was administered daily to male Lewis rats in two doses (3 and 10 mg/kg, orally). Four weeks later, treatment was terminated; hemodynamic and echocardiographic measurements were performed; hearts were excised for histopathology and estimation of histamine, mRNA, and protein levels. Mortality rate was reduced by drug treatment. T-3999 reduced % fibrosis and tissue collagen III. Profibrotic markers-transforming growth factor-β(1), tumor necrosis factor-α, and galectin-3--were attenuated by treatment. Mast cell density and degranulation, and tissue histamine concentration were also reduced. This indicates an anti-inflammatory effect of the drug in reducing fibrosis. Hypertrophy was reduced as reflected by reduced myocyte diameter and natriuretic peptide expression. T-3999 treatment increased the sarcoendoplasmic reticulum Ca(2+) ATPase 2 protein level and improved several cardiac function parameters. The reduction of the remodeling process and improvement in myocardial function suggest an effect of T-3999 in attenuating ventricular remodeling in post-myocarditis DCM.

Infusion of adrenomedullin improves acute myocarditis via attenuation of myocardial inflammation and edema

Our aim was to assess whether adrenomedullin (AM), a potent vasodilator peptide with a variety of cardioprotective effects, has a therapeutic potential for the treatment of acute myocarditis in a rat model. One week after myosin injection, rats received a continuous infusion of AM or vehicle for 2 weeks, and pathological and physiological investigations were performed. AM treatment significantly reduced the infiltration of inflammatory cells in myocarditic hearts, and decreased the expressions of macrophage chemoattractant protein-1, matrix metalloproteinase-2 and transforming growth factor-beta. Myocardial edema indicated by increased heart weight to body weight ratio and wall thickness was attenuated by AM infusion (5.7+/-0.5 vs. 6.5+/-0.4 g/kg, and 1.9+/-0.3 vs. 2.8+/-0.5 mm, respectively). Infusion of AM significantly improved left ventricular maximum dP/dt and fractional shortening of myocarditic hearts (4203+/-640 vs. 3450+/-607 mm Hg/s, and 21.3+/-4.1 vs. 14.7+/-5.1%, respectively). Infusion of AM improved cardiac function and pathological findings in a rat model of acute myocarditis. Thus, infusion of AM may be a potent therapeutic strategy for acute myocarditis.

Transplantation of mesenchymal stem cells attenuates myocardial injury and dysfunction in a rat model of acute myocarditis

Acute myocarditis is a non-ischemic inflammatory disease of the myocardium for which there is currently no specific treatment. We have previously shown that mesenchymal stem cells (MSC) can ameliorate heart injury during acute ischemia and in dilated cardiomyopathy; however, the therapeutic potential in acute myocarditis is unclear. In this study, we investigated the ability of MSC to attenuate myocardial injury and dysfunction during the acute phase of experimental myocarditis. Ten-week-old male Lewis rats were injected with porcine myosin to induce myocarditis. Cultured MSC (3×106 cells/rat) were injected intravenously 7 days after myosin injection. At 3 weeks, myosin injection resulted in severe inflammation and significant deterioration of cardiac function. MSC transplantation attenuated increases in CD68-positive inflammatory cells and monocyte chemoattractant protein-1 (MCP-1) expression in myocardium, and improved cardiac function in this model. Furthermore, myocardial capillary density was higher in myocarditis tissue, and was further increased by MSC transplantation. In vitro, cultured adult rat cardiomyocytes were injured in response to MCP-1, whereas this effect was attenuated by MSC-derived conditioned medium, suggesting cardioprotective effects of MSC acting in a paracrine manner. MSC transplantation attenuated myocardial injury and dysfunction in a rat model of acute myocarditis, at least in part through paracrine effects of MSC.

FK506 suppressed the inflammatory change of EAM in SJL/J mice

Experimental autoimmune myositis (EAM) is a good model of human inflammatory myopathy. We induced EAM in SJL/J mice by injection with myosin and treated inflammatory changes with FK506. The mice developed inflammatory changes after the fifth myosin injection. After treatment with FK506, inflammation was suppressed and central nuclei of the muscle fibers increased. These findings indicate that FK506 is effective in the treatment of EAM. The data suggests that FK506 inhibits interaction with calcineurin. Intercellular adhesion molecule-1 (ICAM-1) positive cells were present in the inflammatory and non-inflammatory areas of EAM. The FK506-treated group stained more weakly for ICAM-1 than the untreated EAM group.

Programmed Cell Death (Apoptosis) in the Myocardium with Acute Myocarditis.

Acute myocarditis is a potentially lethal disease. However, the precise mechanism of myocardial damage in myocarditis is still unknown, although its pathogenesis seems to be involved in the result of natural killer cells, cytotoxic T cells and autoantibodies. Recently, it has been reported that programmed death of cardiac myocytes can be induced by several pathological conditions including infarction and end-stage cardiac failure. However, little is known about the myocardial cell death in myocarditis. In this study, we investigated whether myocardial cell death via apoptosis occurs in the heart tissue with myocarditis.Since it has been reported that the pathogenesis of the tissue damage in viral myocarditis resembles that in experimental autoimmune myocarditis induced by myosin, we used an experimental model for autoimmune myocarditis induced in Lewis rat with the use of myosin as the antigen. Rats were sacrificed on days 14, 17 and 21 after myosin injection. The tissue specimens were taken from ventricle of the heart.