Atrial and C-type natriuretic peptides amplify growth factor activity in primary aortic smooth muscle cells

Objectives: The objective of the current study was to determine whether atrial natriuretic peptides enhance the mitogenic effect of FGF-2, based on a previous study showing that NO enhances the mitogenic effect of FGF-2. Methods: Primary rat aortic smooth muscle cells were used for all experiments. Mitogenic activity was determined by (H-3)thymidine incorporation and cell counting. Cyclic GMP was measured by radioimmunoassay. Messenger RNA was measured by Northern blotting. FGFR-1 receptor protein was measured by Western blotting. Results: ANP and CNP had no consistent mitogenic effect of their own but they both enhanced FGF-2-induced DNA synthesis and/or cell proliferation by 2-3 fold. ANP enhanced the increase of c-fos mRNA induced by FGF-2. ANP, alone or in combination with FGF-2, had no effect on FGF receptor protein levels. HS-142-1, a specific antagonist of guanylyl cyclase-linked A- or B-type ANP receptors, inhibited the co-mitogenic effect of ANP. Exogenous cGMP was also co-mitogenic, whereas two peptides that bind selectively to the ANF C-receptor, cANF and des[Cys(105),Cys(121)]rANF(104-126), had no mitogenic or co-mitogenic effect. The co-mitogenic effect of ANP gradually disappeared as the subculture number of the cells was increased, indicating that it was selective for primary cells. ANP enhanced the mitogenic response of primary aortic smooth muscle cells to EGF whereas that to IGF-1 and PDGF was either not increased, or increased modestly. Conclusions: ANP enhances the mitogenic effect of FGF-2, via a mechanism that may involve elevation of immediate early gene expression but not enhancement of FGF receptor protein levels. We speculate that ANP and CNP released from macrophages and endothelial cells could enhance the mitogenic effect of FGF-2 or EGF in vivo.