Objective. Panax ginseng is used widely for treatment of cardiovascular disorders in China. Ginsenoside Re is the main chemical component of P. ginseng. We aimed to investigate the protective effect of ginsenoside Re on isoproterenol-induced myocardial fibrosis and heart failure in rats. Methods. A model of myocardial fibrosis and heart failure was established by once-daily subcutaneous injection of isoproterenol (5 mg/kg/day) to rats for 7 days. Simultaneously, rats were orally administrated ginsenoside Re (5 or 20 mg/kg) or vehicle daily for 4 weeks. Results. Isoproterenol enhanced the heart weight, myocardial fibrosis, and hydroxyproline content in rat hearts. Ginsenoside Re inhibited (at least in part) the isoproterenol-induced increase in heart weight, myocardial fibrosis, and hydroxyproline content. Compared with the isoproterenol group, treatment with ginsenoside Re ameliorated changes in left ventricular systolic pressure, left ventricular end diastolic pressure, and the positive and negative maximal values of the first derivative of left ventricular pressure. Ginsenoside Re administration also resulted in decreased expression of transforming growth factor (TGF)-β1 in serum and decreased expression of Smad3 and collagen I in heart tissue. Conclusion. Ginsenoside Re can improve isoproterenol-induced myocardial fibrosis and heart failure by regulation of the TGF-β1/Smad3 pathway.