Natriuretic peptides have been implicated in the regulation of cardiac remodeling in vivo. Disruption of the type-B natriuretic peptide (BNP) gene or its receptor, the type-A natriuretic peptide receptor (NPR-A), gene in mice results in cardiac fibrosis and enhanced fibrotic response to stress. Here, we demonstrate that BNP inhibits TGFβ-induced fibrosis in cultured human cardiac fibroblasts. Human BNP dose-dependently induced intracellular cyclic GMP production in primary human cardiac fibroblasts, consistent with expression of NPR-A by these cells and activation by the peptide. TGFβ induced a modest increase (∼50%) in fibroblast proliferation that was dramatically inhibited by BNP. Using cDNA microarray, we show that BNP globally opposed TGFβ-regulated genes related to fibrosis (collagen 1, fibronectin, CTGF, PAI-1, TIMP3), myofibroblast conversion (α-smooth muscle actin), proliferation (PDGFα, IGF1, IGFBP10) and inflammation (COX2, IL6) without any significant effects on cells not stimulated with TGFβ. These results were confirmed by real-time PCR. Furthermore, Western blot analyses show that TGFβ-induced collagen 1 and fibronectin expression was inhibited (75–80%) by BNP. These findings provide the first demonstration that BNP has a direct effect on cardiac fibroblasts to inhibit fibrotic responses, suggesting that BNP functions as an anti-fibrotic factor in the heart to prevent cardiac remodeling in pathological conditions.