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Myocardial interstitial fibrosis in the noninfarcted region remained to be an inevitable complication of myocardial infarction (MI) resulting in cardiac dysfunction and even heart failure. Cannabinoid receptor type II (CB2) activation has been reported to ameliorate fibrosis in hepatic cirrhotic rat. Whereas, the anti-fibrotic effect of CB2 activation in infarcted hearts was illusive. Our study was designed to investigate whether AM1241, a CB2 selective agonist, is capable of alleviating myocardial fibrosis post MI in vivo and in vitro.
C57BL/6 mice were randomly allocated into four experiment groups: (1) sham group, (2) sham+AM1241 group, (3) MI group, (4)MI+AM1241 group. MI mice model were established by coronary artery ligation surgery. AM1241(20mg/kg/d) or vehicles were injected intraperitoneally for 7 consecutive days after MI. Four weeks later, echocardiograph was conducted to assess cardiac function. Sirius-red staining and Western blot were applied to examine fibrosis and fibrosis related markers such as collagen I, fibronectin and tissue inhibitor of metalloprotease(TIMP)-1. Hypoxia/serum deprivation (H/SD) injury was used to stimulate MI in primary cultured cardiac fibroblasts(CFs). Western blot assay and immunofluorescent staining were performed to observe the fibroblast-myofibroblast transformation and secretion of CFs under different conditions. Nrf-2 siRNA was used to explore the role of Nrf-2 in anti-fibrosis process of CB2 agonist.
Our results showed that AM1241 significantly improved cardiac function as compared with that of the MI group (EF:60.04±5.93% vs 42.87±3.16%, p<0.05; FS:26.17±1.88% vs 21.64±1.03%, p<0.05), alleviated interstitial fibrosis and suppressed the expression of fibrosis related proteins (p<0.05) such as collagen I, fibronectin and TIMP-1 in mice with MI. In cardiac fibroblasts subjected to H/SD injury, AM1241 also suppressed the increased level of α-SMA, collagen I and collagen III (p<0.05) induced by H/SD, which were partially abolished by Nrf-2 siRNA transfection. Additionally, AM1241 accelerated the Nrf-2 into nucleus and inhibited TGFβ-1/Smad3 pathway in a Nrf-2 depended manner.
In conclusion, our results revealed that CB2 selective agonist AM1241 ameliorated cardiac fibrosis largely through Nrf2 -mediated down-regulation of TGFβ-1/Smad3 pathway. Thus, CB2 activation might serve as a promising therapeutic target for hindering cardiac fibrosis induced by MI.