Abstract 18328: Neurotrophin 3 Induces the Expression of Osteogenic Proteins in Human Aortic Valve Interstitial Cells via Trk Receptors

Abstract

Calcific aortic valve disease affects a large number of elderly people, and progressive aortic valve leaflet calcification ultimately results in heart failure. However, the molecular mechanism that mediates valvular osteogenic responses is incompletely understood. Aortic valve interstitial cells (AVICs) express osteogenic proteins in response to stimulation of Toll-like receptor (TLR) 2 or 4. We recently found that soluble biglycan induces the expression of transforming growth factor-β1 (TGF-β1) and bone morphogenetic protein-2 (BMP-2) in human AVICs via TLR2. The objective of this study is to identify novel pro-osteogenic mediators and to elucidate the mechanisms underlying their action in human AVICs. Methods and results: We examined gene expression profile in AVICs isolated from 5 normal human aortic valves and 5 diseased human aortic valves using microarray analysis. Among a large group of genes analyzed, mRNA levels of neurotrophin 3 (NT3), a neural growth factor, were markedly up-regulated in AVIC isolates of diseased valves. Immunoblotting analysis confirmed higher levels of NT3 protein in AVICs of diseased valves. Surprisingly, normal human AVICs expressed greater levels of TGF-β1 and BMP-2 when exposed to recombinant human NT3 (0.05 or 0.10 μg/ml) for 3 days. Further, up-regulation of TGF-β1 and BMP-2 expression by NT3 was preceded by phosphorylation of Akt and glycogen synthase kinase 3 beta (GSK3β). The pro-osteogenic effect of NT3 was not affected by blocking antibodies against TLR2 and TLR4, indicating that NT3 is incapable of acting as a danger-associated molecular pattern in human AVICs. However, neural growth factor receptors (TrkA, TrkB, TrkC and p75 NTR ) were detectable in human AVICs. The pan Trk inhibitor K252a (0.2 μmol/L) suppressed Akt and GSK3β phosphorylation, and markedly reduced TGF-β1 and BMP-2 expression following NT3 stimulation. Conclusions: AVICs of diseased human aortic valves express higher levels of NT3. NT3 induces the expression of TGF-β1 and BMP-2 by human AVICs via Trk receptors. Thus, NT3 is a novel pro-osteogenic mediator in human aortic valve, and it may play a role in aortic valve leaflet calcification through up-regulation of the expression of osteogenic proteins by AVICs.