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Background: Sheng-Mai Yin (SMY), a modern Chinese formula based on Traditional Chinese Medicine theory, has been used to treat cardiovascular diseases in Eastern Asia. Our study focuses on the cardioprotection of SMY against doxorubicin (DOX)-induced cardiac toxicity in vivo., Methods: Rats were injected with DOX (2.5 mg/kg) in six injections over a 2-week period. SMY was administrated intragastrically at the dose of 8.35, 16.7 and 33.4 g/kg, or 16.7 g/kg only twice a day concurrently with DOX for the 2-weeks. A series of assays were performed to detect the effects of SMY on: (i) heart weight index (HWI) and left ventricular mass index (LVMI); (ii) cardiac function; (iii) heart tissue morphology; (iv) the contents of carboxy terminal propeptide of procollagen typeI (PICP), amino terminal propeptide of procollagen type III (PШNP), transforming growth factor-β1 (TGF-β1), B-type natriuretic peptide (BNP), monocyte chemoattractant protein-1 (MCP-1), interferon gamma (INF-γ) and interleukin 6 (IL-6) by ELISA; (v) the mRNA levels of TGF-β1 and toll-like receptor-2 (TLR2); and (vi) protein level of TGF-β1., Results: Rats treated with SMY displayed the reductions of BNP and CK-MB increased by DOX in a dose-dependent manner. Moderate dose of SMY exhibited the correction for the increased HWI, LVMI, and the injured cardiac function, as well as the collagen accumulation. In addition, cardioprotection of SMY against DOX-induced cardiac toxicity was demonstrated by the reduction of myocardial fibrosis, characterized by the suppression of PICP, PШNP and TGF-β1, as well as the anti-inflammation and the regulation for cardiac immune microenvironment, characterized by the inhibition of TLR2, MCP-1, INF-γ and IL-6., Conclusions: SMY may protect heart function through the restriction of myocardial fibrosis induced by DOX, which suggests the potentially therapeutic effect of SMY on DOX-induced cardiomyopathy.