Marinobufagenin (MBG), an endogenous Na/K-ATPase inhibitor and a vasoconstrictor, induces vascular fibrosis via PKC-dependent inhibition of Fli-1, a nuclear transcription factor and a negative regulator of collagen synthesis. In vascular smooth muscle cells (VSMC), atrial natriuretic peptide (ANP), via cGMP/PKG1 dependent mechanism, reduces sensitivity of Na/K-ATPase to MBG. We hypothesized that VSMC from aged rats have a heightened sensitivity to the pro-fibrotic effect of MBG due to an age-associated down-regulation of cGMP/PKG-dependent signaling.

In response to acute NaCl loading (0.4g/kg), aged (24-month old) Sprague-Dawley rats exhibited exaggerated responses of urinary MBG (5.4±0.4 vs. 1.9±0.2 pmol/hr; P<0.01) and of systolic blood pressure (29 vs. 15 mmHg; P<0.01), and greater inhibition of Na/K-ATPase in aorta vs. 3-month old rats.

In VSMC from young rats on a normal salt diet, 1 nmol/L MBG induced down-regulation of Fli-1 and a 50% increase in the levels of collagen-1, and these effects were blocked by 1 nmol/L ANP. In VSMC from aged rats levels of PKG1 and Fli-1 were markedly reduced. MBG (1 nmol/L) decreased Fli-1 by 60%, increased level of collagen-1 two-fold (P<0.01). In contrast to young rats, ANP failed to oppose these effects. Silencing of the PKG1 gene in VSMC from young rats sensitises these cells to the pro-fibrotic effect of MBG: 1 nmol/L MBG increased levels of collagen-1 2.5-fold (P<0.01).

Thus, the age-associated reduction in vascular PKG1 levels and resultant decline in cGMP signaling sensitize VSMC to the pro-fibrotic effect of MBG. Silencing of PKG1 in young VSMC mimics these effects of aging.