Cardioprotective effect of prostaglandin-E2 receptor-4 (EP4) stimulation on the ischemic heart has been demonstrated. Its effect on the heart affected by myocarditis, however, remains uncertain. In this study, we investigated therapeutic effect of EP4 stimulant using a mouse model of autoimmune myocarditis (EAM) that progresses to dilated cardiomyopathy (DCM). EP4 was present in the hearts of EAM mice. Treatment with EP4 agonist (ONO-0260164: 20 mg/kg/day) improved an impaired left ventricular (LV) contractility and reduction of blood pressure on day 21, a peak myocardial inflammation. Alternatively, DCM phenotype, characterized by LV dilation, LV systolic dysfunction, and collagen deposition, was observed on day 56, along with activation of matrix metalloproteinase (MMP)-2 critical for myocardial extracellular matrix disruption, indicating an important molecular mechanism underlying adverse ventricular remodeling after myocarditis. Continued treatment with ONO-0260164 alleviated the DCM phenotype, but this effect was counteracted by its combination with a EP4 antagonist. Moreover, ONO-0260164 inhibited in vivo proteolytic activity of MMP-2 in association with up-regulation of tissue inhibitor of metalloproteinase (TIMP)-3. EP4 stimulant may be a promising and novel therapeutic agent that rescues cardiac malfunction during myocarditis and prevents adverse ventricular remodeling after myocarditis by promoting the TIMP-3/MMP-2 axis.