Hyperlipidaemia and IFNgamma/TNFalpha Synergism are associated with cholesterol crystal formation in Endothelial cells partly through modulation of Lysosomal pH and Cholesterol homeostasis.

Yvonne Baumer,Nehal N Mehta,Joel M Gelfand,William A Boisvert,Jie Zhuang,Martin P Playford,Qimin Ng,Yusuke Sekine,Daniella M Schwartz,Andrew Keel,David A Bluemke,Howard S Kruth,Lam C Tsoi,Pradeep K Dagur,Gregory E Sanda,Ankit Saxena,Erin Stempinski,Cristhian A Gutierrez-Huerta ,Heather Teague ,Amit Dey ,Jóhann E Guðjónsson ,Justin Rodante ,Tiffany M Powell‐Wiley ,Marcus Y Chen ,Youssef Elnabawi ,Noor Khalil ,Christopher K E Bleck

doi:10.1016/j.ebiom.2020.102876

Abstract

BackgroundInflammation plays an important role in the development of cardiovascular disease (CVD). Patients with chronic inflammation diseases have high levels of inflammation and early fatal myocardial infarction due to early, unstable coronary plaques. Cholesterol crystals (CC) play a key role in atherogenesis. However, the underlying mechanisms of endothelial cell (EC)-derived CC formation are not well understood in chronic inflammation.MethodsWe utilized a combination of a mouse psoriasis model (K14-Rac1V12 mouse model) and human psoriasis patients to study the effect of inflammatory cytokines on CC formation in ECs. Lysosomal pH, alterations in lipid load and inflammatory proteins were evaluated as potential mechanisms linking inflammatory cytokines to CC formation. Coronary CT angiography was performed (n = 224) to characterize potential IFNγ and TNFα synergism on vascular diseases in vivo.FindingsWe detected CC presence in the aorta of K14-Rac1V12 mice on chow diet. IFNγ and TNFα were found to synergistically increase LDL-induced CC formation by almost 2-fold. There was an increase in lysosomal pH accompanied by a 28% loss in pH-dependent lysosomal signal and altered vATPaseV1E1 expression patterns. In parallel, we found that LDL+IFNγ/TNFα treatments increased free cholesterol content within EC and led to a decrease in SOAT-1 expression, an enzyme critically involved cholesterol homeostasis. Finally, the product of IFNγ and TNFα positively associated with early non-calcified coronary burden in patients with psoriasis (n = 224; β = 0.28, p < 0.001).InterpretationOur results provide evidence that IFNγ and TNFα accelerate CC formation in endothelial cells in part by altering lysosomal pH and free cholesterol load. These changes promote early atherogenesis and contribute to understanding the burden of CVD in psoriasis.FundingFunding was provided by the Intramural Research Program at NIH (NNM) and the National Psoriasis Foundation (NNM and YB).

Highlights

Cardiovascular disease (CVD) is the main cause of death worldwide accounting for more than 25% of all death in the US.[1]
We compared the detected phenotype to aortas of LdlrÀ/À mouse aortas after and ultrastructural analysis using TEM and SEM. (a) Sections of the aorta were visualized by Polarize light microscopy (PLM) and the area directly under the endothelium emitting a birefringent signal, representative of Cholesterol crystals (CC), quantified. (b) The data from (a) in K14-Rac1V12À/+ mouse aortas were subjected to Pearson correlation after normality check and revealed a strong positive association of CC presence in the aorta and severity in animals not being fed a HFD. (c) The aortic arch of a subset of these mice was subjected to SEM
Our work demonstrates a role of interferon gamma (IFNg)/tumor necrosis factor alpha (TNFa) in driving CC formation due to modulation of lysosomal pH and an increase of intracellular free cholesterol load

Summary

Introduction

Cardiovascular disease (CVD) is the main cause of death worldwide accounting for more than 25% of all death in the US.[1] Inflammation is an important player in CVD development[2] given that CVD is the leading cause of death in chronic inflammatory diseases, including rheumatoid arthritis, lupus erythematosus, and psoriasis.[3À5] Psoriasis, a chronic inflammatory skin disease, affects 2À3% of the world’s population and is associated with premature lipid-rich non-calcified coronary plaque and myocardial infarction.[5À8]. Chronic inflammatory diseases including psoriasis have an increased risk for cardiovascular disease (CVD) driven mostly by inflammatory-associated vascular diseases. The product of IFNg and TNFa positively associated with early non-calcified coronary burden in patients with psoriasis (n = 224; b = 0.28, p < 0.001)

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