Clin Res Cardiol (2022). https://doi.org/10.1007/s00392-022-02002-5

Inhibition of neutral sphingomyelinase 2 by GW4869 reduces the development of aortic valve stenosis in mice.
L. Reese1, S. T. Niepmann1, M. Bulic1, P. Düsing1, M. R. Hosen1, G. Nickenig1, S. Zimmer1, F. Jansen1, A. Zietzer1
1Medizinische Klinik und Poliklinik II, Universitätsklinikum Bonn, Bonn;
Introduction: Aortic valve stenosis (AVS) is the most prevalent valvular disease in humans. To date, there is no medical treatment available for AVS and consequently replacement of the aortic valve is the only treatment option. Immune cell infiltration and calcification are two important pathophysiological hallmarks of the development of AVS. Ceramides are known to contribute to inflammation and calcification. In the context of acute inflammation ceramides are produced to a relevant part by neutral sphingomyelinase 2 (nSMase2). It has been shown that the nSMase inhibitor GW4869 reduces the inflammatory response of monocytes and inflammation in the context of atherosclerotic plaque development.

Methods and Results: AVS was induced in mice through a wire-injury. For this model a coronary angiography wire was introduced into the left ventricle over the right carotid artery. The wire was then rotated on the aortic valve level to generate a defined injury. For the sham procedure the wire was only inserted into the right carotid artery. The mice were treated with GW4869 at 2 mg/kg Body weight in DMSO or DMSO intraperitoneally three times a week starting directly prior to the injury. In two different experiments the mice were sacrificed 21 days (A-C) or 42 days (D-E) after the wire-injury. 21 days after the wire-injury GW4869 significantly reduced the aortic valve area (A). Immune histological staining with CD68 revealed less macrophage infiltration into the aortic valve when treated with GW4869 (B). Von Kossa staining showed only low calcification and no difference between the groups 21 days after wire-injury (C). Peak velocity was measured echocardiographically 14 days, 28 days and 42 days after wire-injury. We detected a significantly reduced peak velocity with GW4869 treatment compared to DMSO at 28 and 42 days (D). In addition to the aforementioned histological analysis, we found that 42 days after the wire-injury GW4869 treatment reduced calcification significantly (E). 42 days after the wire-injury monocyte infiltration and aortic valve area were already reduced towards baseline and not significantly different anymore. In vitro experiments with human valvular endothelial cells (VECs) show that GW4869 reduces VCAM-1 expression and the adhesion of THP-1 monocytes to VECs after stimulation with 100ng/ml lipopolysaccharide.

Conclusion: Our experiments indicate that nSMase is involved in immune cell infiltration and valvular calcification. GW4869 treatment is able to reduce aortic valve thickening, immune cell infiltration and calcification in vivo. Our results show that this reduces the development of AVS as the peak velocity is significantly reduced with GW4869. In conclusion, pharmacological inhibition of nSMase2 is a promising concept to counteract the development of AVS.


 

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