K Ca 3.1 Drives Pro-Fibrotic Activation and Represents a Novel Therapeutic Target in Aortic Stenosis
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Introduction
Aortic stenosis (AS) is characterised by progressive aortic valve (AV) leaflet fibrosis and calcification, yet no medical therapies exist to slow disease progression. AV interstitial cells (VICs) that differentiate into myofibroblasts are central drivers of fibrosis. The Ca 2+ -activated K + channel K Ca 3.1 promotes pro-fibrotic signalling in several fibrotic diseases, however its role in AS remains unknown.
Methods
K Ca 3.1 protein expression was examined in paraffin embedded tissue by Immunohistochemistry from control and AS valve tissue. VICs were isolated, cultured and phenotypically characterised as myofibroblasts from AV tissue obtained from patients with severe tricuspid AS undergoing surgical AV replacement (n=19). K Ca 3.1 mRNA and protein expression were assessed by qRT-PCR and immunohistochemistry, and functional channel activity confirmed using patch-clamp electrophysiology. The effects of transforming growth factor-β1 (TGFβ1) stimulation and pharmacological inhibition with the selective K Ca 3.1 blocker senicapoc were examined.
Results
Immunoreactive K Ca 3.1 channels and smooth muscle actin were detected in both control and AS aortic valve tissue, localised to elongated, nucleated interstitial cells, with significantly higher expression observed in AS tissue compared to control. Isolated VICs exhibited an activated myofibroblast phenotype, expressing THY-1, vimentin, collagen and α-smooth muscle actin (αSMA) (n=9). Myofibroblasts expressed K Ca 3.1 mRNA and protein and demonstrated functional plasma membrane channels. TGFβ1 stimulation increased K Ca 3.1, αSMA and collagen type I mRNA expression, while K Ca 3.1 blockade with senicapoc (100 nM) significantly attenuated TGFβ1-induced αSMA expression, stress fibre formation and collagen gel contraction. Senicapoc had no effect on myofibroblast proliferation or migration.
Conclusions
We show for the first time that functional K Ca 3.1 channels are expressed in human AS tissue and AV myofibroblasts, where they regulate myofibroblast contraction, α-SMA expression, and differentiation, promoting pro-fibrotic activity. These responses are attenuated by the selective K Ca 3.1 inhibitor senicapoc. Given its established safety in phase 3 clinical trials, K Ca 3.1 inhibition represents a promising and readily translatable anti-fibrotic therapeutic strategy for AS.
