Increased TGFβ1 and SMAD3 Contribute to Age-Related Aortic Valve Calcification

AIMS: Calcific aortic valve disease (CAVD) is a progressive heart disease that is particularly prevalent in elderly patients. The current treatment of CAVD is surgical valve replacement, but this is not a permanent solution, and it is very challenging for elderly patients. Thus, a pharmacological intervention for CAVD may be beneficial. In this study, we intended to rescue aortic valve (AV) calcification through inhibition of TGFβ1 and SMAD3 signaling pathways. METHODS AND RESULTS: The klotho gene, which was discovered as an aging-suppressor gene, has been observed to play a crucial role in AV calcification. The klotho knockout (Kl(–/–)) mice have shorter life span (8–12 weeks) and develop severe AV calcification. Here, we showed that increased TGFβ1 and TGFβ-dependent SMAD3 signaling were associated with AV calcification in Kl(–/–) mice. Next, we generated Tgfb1- and Smad3-haploinsufficient Kl(–/–) mice to determine the contribution of TGFβ1 and SMAD3 to the AV calcification in Kl(–/–) mice. The histological and morphometric evaluation suggested a significant reduction of AV calcification in Kl(–/–); Tgfb1(±) mice compared to Kl(–/–) mice. Smad3 heterozygous deletion was observed to be more potent in reducing AV calcification in Kl(–/–) mice compared to the Kl(–/–); Tgfb1(±) mice. We observed significant inhibition of Tgfb1, Pai1, Bmp2, Alk2, Spp1, and Runx2 mRNA expression in Kl(–/–); Tgfb1(±) and Kl(–/–); Smad3(±) mice compared to Kl(–/–) mice. Western blot analysis confirmed that the inhibition of TGFβ canonical and non-canonical signaling pathways were associated with the rescue of AV calcification of both Kl(–/–); Tgfb1(±) and Kl(–/–); Smad3(±) mice. CONCLUSION: Overall, inhibition of the TGFβ1-dependent SMAD3 signaling pathway significantly blocks the development of AV calcification in Kl(–/–) mice. This information is useful in understanding the signaling mechanisms involved in CAVD.