Changes in Plasma TGF-β1 Levels in a Murine Model of Aortic Stenosis (Surgical Constriction of the Ascending Aorta) in C57Bl/6 (wild-type) Mice and Mice with a Targeted Deletion of Platelet TGF-β1

Abstract 1065

Transforming growth factor-β1 (TGF-β1) is a multifunctional cytokine that controls cell proliferation, differentiation, apoptosis, immune responses, and tissue fibrosis. Circulating platelets contain a high concentration of TGF-β1 in their α -granules and release it as an inactive (latent) complex when activated. We previously reported that platelet TGF-β1 is responsible for ∼45% of basal plasma TGF-β1; that shear force can activate TGF-β1 in vitro; and that mice with megakaryocyte-specific deletion of TGF-β1 [PF4-Cre/Tgfb1flox (Tgfb1^flox)], which have ∼15% of normal platelet TGF-β1, are partially protected from developing cardiac hypertrophy, fibrosis, and systolic dysfunction induced by transverse aortic constriction (TAC) surgery. Since the TAC model involves placing a constriction distal to the innominate artery, which produces high shear both at the site of stenosis and in innominate artery, in this study we tried to simulate better the hemodynamics of human valvular aortic stenosis by constricting the ascending aorta (AAC) of both C57Bl/6 wild-type (WT) mice and Tgfb1^floxmice.

The AAC surgery employed a 27 gauge needle guide and reduced the diameter of the ascending aorta by >80% in both WT and TGFb1^flox mice (p<0.001). The constriction increased the shear force at the site of stenosis by ∼20-fold in both genotypes (Table). In WT mice, total TGF-β1 levels increased from 1.02 ng/ml (n=23) before surgery to 1.26 ng/ml (24% increase) and 1.35 (32% increase) at 1 and 4 weeks after AAC surgery, respectively (p=0.02 and p=0.08). Baseline TGF-β1 levels in Tgfb1^flox mice were 0.72 ng/ml, ∼30% lower than in WT mice. Although there was a significant increase in total TGF-β1 levels in Tgfb1^flox mice after surgery, their levels were 35% and 53% lower than those in WT mice at 1 week and 4 weeks after surgery (0.82 ng/ml at both 1 and 4 weeks; n=20; p<0.01 at both time points). Shear force at the site of the stenosis correlated positively with plasma total TGF-β1 levels in WT mice (r=0.45, p=0.048 at 1 week and r=0.61, p=0.007 at 4 weeks), but not in Tgfb1^flox mice (r=0.30, p=0.26 at 1 week and r=0.36, p=0.17 at 4 weeks). Both WT and Tgfb1^flox mice had 11% decreases in platelet counts 4 weeks after surgery. Ejection fraction (EF) measured by echocardiography decreased in both groups by ∼27% at both 1 and 4 weeks after AAC surgery. The decrease in EF correlated with the increase in total TGF-β1 levels in WT mice 4 weeks after surgery (r=0.61, p=0.01), but not in Tgfb1^flox mice (r=0.18, p=0.73). The decrease in EF also correlated with the reduction in platelet count after 4 weeks in both groups (r=0.77, p=0.02 in WT and r=0.50, p=0.03 in Tgfb1^flox).

We conclude that: 1. the AAC model results in increased plasma TGF-β1 levels, 2. both platelet and non-platelet sources of TGF-β1 appear to contribute to the increase, 3. in WT, but not Tgfb1^floxmice, shear force and EF are correlated with plasma TGF-β1 levels, and 4. reducing platelet TGF- β1 by ∼85% does not protect against the reduction in systolic function (as judged by EF) induced by ACC.