Effects of methotrexate on plasma cytokines and cardiac remodeling and function in postmyocarditis rats

Zhengang Zhang, Pei Zhao, Aihua Li, Xiaolei Lv, Yang Gao, Hongguang Sun, Yongling Ding, Jian Liu, Zhengang Zhang, Pei Zhao, Aihua Li, Xiaolei Lv, Yang Gao, Hongguang Sun, Yongling Ding, Jian Liu

Abstract

Excessive immune activation and inflammatory mediators may play a critical role in the pathogenesis of chronic heart failure. Methotrexate is a commonly used anti-inflammatory and immunosuppressive drug. In this study, we used a rat model of cardiac myosin-induced experimental autoimmune myocarditis to investigate the effects of low-dose methotrexate (0.1 mg/kg/d for 30 d) on the plasma level of cytokines and cardiac remodeling and function. Our study showed that levels of tumor necrosis factor-(TNF-)alpha and interleukin-6 (IL-6) are significantly increased in postmyocarditis rats, compared with the control rats. Methotrexate treatment reduced the plasma levels of TNF-alpha and IL-6 and increased IL-10 level, compared to saline treatment. In addition, postmyocarditis rats showed significant cardiac fibrosis characterized by increased myocardial collagen volume fraction, perivascular collagen area, and the ratio of collagen type I to type III, compared with the control rats. However, MTX treatment not only markedly attenuated cardiac fibrosis, diminished the left ventricular end-diastolic dimension, but also increased the left ventricular ejection fraction and fractional shortening. Collectively, these results suggest that low-dose methotrexate has ability to regulate inflammatory responses and improves cardiac function and hence contributes to prevent the development of postmyocarditis dilated cardiomyopathy.

Figures

Figure 1
Figure 1
Changes of plasma cytokines levels. The levels of TNF-alpha, IL-6, and IL-10 were determined by ELISA. All results are expressed as mean ± SD (n = 10). *P < .01 versus control group; #P < .01 versus EAM60 group; △P < .01 versus EAM30 group.
Figure 2
Figure 2
Macroscopic and microscopic assessment of the hearts. (a) Representative gross appearance of the heart from each group of rats. (b) Representative photomicrographs of the ventricular sections (HE staining; ×200). In hearts from the control group, myocytes is well organized. In EAM60 group, the cardiac myocytes exhibited extensive necrosis, degeneration, and disorder. MTX treatment led to a significant decrease in the size of these lesions.
Figure 3
Figure 3
Masson's trichrome staining. Representative photomicrographs of the ventricular sections from the three groups of rats are shown (×200). Green staining represents the collagen fibers (a)–(c). Myocardial collagen volume fraction (CVF; (d)) and perivascular collagen area (PVCA; (e)) were calculated in the control, EAM60 and EAM-MTX groups, respectively. All results are represented as mean ± SD (n = 10). *P < .05, #P < .01 versus control group; ▲P < .01 versus EAM60 group.
Figure 4
Figure 4
Collagen types I and III staining. Representative photomicrographs of ventricular sections from the three groups of rats are shown ((a); ×200). The areas of collagen types I and III were, measured (b) and the ratios of collagen type I/III were calculated (c). All results are represented as mean ± SD (n = 10). *P < .05, # P < .01 versus control group; △P < .05, ▲P < .01 versus EAM60 group.

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