Abstract 159: Neurotrophin 3 Plays a Critical Role in TLR4-mediated Fibrogenic Response in Human Aortic Valve Interstitial Cells

Abstract

Aortic stenosis is a chronic inflammatory condition, and valvular fibrosis and calcification are significant pathological changes in the progression of aortic stenosis. Aortic valve interstitial cell (AVIC) proliferation and over-expression of extracellular matrix (ECM) proteins promote valvular fibrosis. Although pro-inflammatory stimuli are the culprits of aortic valve fibrosis, the mechanism by which they induce aortic valve fibrogenic change remains incompletely understood. Our recent findings suggest that AVICs from diseased aortic valves express higher levels of neurotrophin 3 (NT3) protein and that NT3 is capable of up-regulating AVIC proliferation and ECM protein expression. In this study, we tested the hypothesis that pro-inflammatory stimulation up-regulates NT3 production to enhance the pro-fibrogenic activity in human AVICs. Methods and Results: Human AVICs isolated from normal valves were treated with Toll-like receptor 4 (TLR4) agonist lipopolysaccharide (LPS, 0.20 μg/ml) for 3 to 28 days in the presence or absence of a neutralizing antibody against NT3. LPS elevated BrdU incorporation, collagen III expression and collagen deposition. Neutralization of NT3 markedly reduced these pro-fibrogenic changes. Interestingly, LPS up-regulated NT3 levels in human AVICs in a TLR4-dependent fashion, and inhibition of either Akt or ERK1/2 attenuated the effect of LPS on NT3 expression. Recombinant NT3 induced human AVIC proliferation, collagen III expression and collagen deposition through the Trk receptors. Inhibition of the Trk receptors abrogated the pro-fibrogenic effect of LPS on AVICs. Conclusions: Stimulation of TLR4 in human AVICs elevates the pro-fibrogenic activity through up-regulating the production of neural growth factor NT3. The Akt and ERK1/2 pathways are involved in TLR4-mediated NT3 up-regulation, and the Trk receptors mediate the pro-fibrogenic effect of TLR4 stimulation in human AVICs. These findings reveal a molecular mechanism underlying pro-inflammatory elevation of AVIC pro-fibrogenic activity and indicate the therapeutic potential of antagonizing NT3 for suppression of aortic valve fibrosis in an inflammatory setting.