The role of uremic toxin indoxyl sulfate in the pathophysiology of aortic valve stenosis

Aims Chronic kidney disease (CKD) is closely associated with cardiovascular disease (CVD). This includes aortic valve stenosis (AS), one of the most common valve diseases among adults. CKD leads to the retention of uremic toxins such as indoxyl sulfate (IS), which is known to induce inflammatory and pro-calcific processes. We hypothesise that IS specifically induces AS formation. Methods and Results Stimulation of human valvular interstitial cells (VICs) with IS in addition to phosphate led to increased calcification. RNA sequencing identified naked cuticle homolog 2 (NKD2) as an upregulated gene in VICs under uremic conditions. Knockdown of NKD2 reduced calcification of VICs and upregulation of IL-6. The organic anion transporting polypeptide 3A1 (OAT3A1) was identified to mediate IS uptake as well as upregulation of NKD2 and IL-6. We identified NF-κB signalling to be involved in IS-induced IL-6 upregulation. In vivo, we investigated combined models of adenine-induced kidney injury or oral IS supplementation with wire injury-induced AS in C57BL/6J mice. Echocardiography showed aggravated AS in uremic mice compared to control mice after wire injury. Explanted valves from uremic mice with AS exhibited a significant increase in macrophage infiltration, fibrotic areas and valvular NKD2 expression compared to controls. IS-treated mice showed aggravated AS compared to control mice. This was accompanied by more prominent valve fibrosis, macrophage infiltration, and NKD2 expression in explanted valves of IS-treated mice. In the blood and bone marrow, IS treatment led to the differentiation of monocytes into intermediate and non-classical monocytes. This was paralleled by IS-induced monocyte adhesion to valvular endothelial cells in vitro. Conclusion Uremic conditions aggravate AS development in mice by inducing valvular fibrosis and macrophage infiltration. Indoxyl sulfate is involved in this process and stimulates monocyte differentiation and adhesion to the valvular endothelium. On a cellular level, we hypothesize that IS-mediated NKD2 induction leads to a calcifying and inflammatory response in VICs.