Cardiac fibrosis is a hallmark of heart disease and involves recruitment, proliferation, and differentiation of extracellular matrix-producing fibroblasts, leading to overproduction of collagen within the myocardium. In this study, the effects of relaxin in inhibiting these processes were investigated. We used neonatal rat atrial and ventricular fibroblasts, which respond to pro-fibrotic stimuli (i.e., transforming growth factor-beta and angiotensin II) and naturally express the relaxin receptor LGR7. Relaxin significantly inhibited TGF-beta- and angiotensin II-mediated fibroblast function and collagen production over a 72-h period, while increasing MMP-2 expression and activity in the presence of both profibrotic factors (all P <.05). These studies demonstrate that relaxin may have therapeutic potential in diseased states characterized by cardiac fibrosis.