Moesin is activated in cardiomyocytes in experimental autoimmune myocarditis and mediates cytoskeletal reorganization with protrusion formation

Abstract

Acute myocarditis is a self-limiting disease. Most patients with myocarditis recover without cardiac dysfunction in spite of limited capacity of myocardial regeneration. Therefore, to address intrinsic reparative machinery of inflamed hearts, we investigated the cellular dynamics of cardiomyocytes in response to inflammation using experimental autoimmune myocarditis (EAM) model. EAM was induced by immunization of BALB/c mice with α-myosin heavy chain peptides twice. The inflammatory reaction was evoked with myocardial damage with the peak at 3 wk after the first immunization (EAM3w). Morphological and functional restoration started from EAM3w, when active protrusion formation, a critical process of myocardial healing, was observed in cardiomyocytes. Shotgun proteomics revealed that cytoskeletal proteins were preferentially increased in cardiomyocytes at EAM3w, compared with preimmunized (EAM0w) hearts, and that moesin was the most remarkably upregulated among them. Immunoblot analyses demonstrated that the expression of both total and phosphorylated moesin was upregulated in isolated cardiomyocytes from EAM3w hearts. Immunofluorescence staining showed that moesin was localized at cardiomyocyte protrusions at EAM3w. Adenoviral vectors expressing wild-type, constitutively active and inactive form of moesin (wtMoesin, caMoesin, and iaMoesin, respectively) were transfected in neonatal rat cardiomyocytes. The overexpression of wtMoesin and caMoesin resulted in protrusion formation, while not iaMoesin. Finally, we found that cardiomyocyte protrusions were accompanied by cell-cell contact formation. The expression of moesin was upregulated in cardiomyocytes under inflammation, inducing protrusion formation in a phosphorylation-dependent fashion. Moesin signal could be a novel therapeutic target that stimulates myocardial repair by promoting contact formation of cardiomyocytes.

Autoimmune myocarditis is not associated with left ventricular systolic dysfunction

“…20 Interestingly, various previous studies reported little or no decrease in LV systolic function in EAM, 21,22 whereas others found a significant systolic dysfunction. 20 Interestingly, various previous studies reported little or no decrease in LV systolic function in EAM, 21,22 whereas others found a significant systolic dysfunction.…”

“…Our results from day 21 are well in line with the results of a recent study by Pistulli et al, who found that EAM is associated with LV-hypertrophy and diastolic dysfunction, but not with a deterioration in LV systolic function, when immunized animals are compared to healthy age-matched controls. 20 Interestingly, various previous studies reported little or no decrease in LV systolic function in EAM, 21,22 whereas others found a significant systolic dysfunction. 23,24 In humans, myocarditis can result in a variety of echocardiographic findings including decreased LV systolic function, LV-hypertrophy, diastolic dysfunction or regional wall motion abnormalities.…”

“…3,25,26 Interestingly, Felker et al found significant differences between the clinical syndromes of fulminant and acute myocarditis that portray different courses of the same disease. Interestingly, the authors of the studies reported that little or no decrease in LV systolic function in EAM (Pistulli et al and Miyawaki et al 20,22 ) used the same amino acid sequence (MyHC-α 614-629: Ac-SLKLMATLFSTYASAD) for the immunization of the animals that was used in our study. 27 Considering the diverging echocardiographic findings from previous studies, we speculate that there might be similar "syndromes" in mice with EAM that portray different courses of the autoimmune disease.…”

Adult murine cardiomyocytes exhibit regenerative activity with cell cycle reentry through STAT3 in the healing process of myocarditis

“…Recently, we have reported that cytoskeletal proteins are upregulated in cardiomyocytes and that cardiomyocytes actively form protrusions, a morphological feature of myocardial regeneration 62 , in the resolution phase of EAM 63 . The most remarkably upregulated cytoskeletal protein, moesin, potently promoted the protrusion formation of cardiomyocytes, and the protrusions mediated cardiomyocyte cell-cell contact.…”

“…Flow-through suspensions were then filtered through a 40 μm cell strainer and the fraction which remained on the 40 μm cell strainer was considered a cardiomyocyte-rich fraction. By this isolation strategy, cardiomyocytes were substantially enriched (pre-isolation: 39.7%, post-isolation: 78.0%) 63 .…”