Cholesterol regulation of Macrophage Inflammation and Vascular Diseases

Va Veterans Administration Hospital, Richmond VA

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Abstract

? DESCRIPTION (provided by applicant): Low grade chronic inflammation associated with obesity is the underlying cause of the development of metabolic diseases including Type 2 diabetes (T2DM) and atherosclerosis. The most important characteristics of chronic inflammation are its persistence and inability to resolve resulting in lack of restoration of homeostasis and development of metabolic diseases. In contrast to classical inflammation which is triggered by an injury or infection, this low grade chronic inflammation is sterile in nature without a direct involvement of a pathogen and is often triggered by nutrients and metabolic surplus and hence also referred to as metaflammation meaning metabolically triggered inflammation. Metaflammation is characterized by abnormal cytokine production especially Interleukin-1? (IL-1?) and persistent activation of a network of inflammatory pathways. Chronic activation of innate immune sentinels or macrophages underlies this abnormal cytokine production however the cellular mechanisms involved are not completely defined. The production and secretion of IL-1? from macrophages is under very tight regulation which requires coordination of two Signals, namely Signal 1: cellular priming required for inducing transcription (via NF-?B), and Signal 2: activation of NLRP3 inflammasome for proteolytic cleavage of pro-IL-1? by active Caspase-1 and secretion of mature IL-1?. Increase in cellular lipid accumulation is one of the prominent features of obesity and intracellular lipid metabolism, especially cholesterol metabolism, is tightly linked to the inflammatory status of macrophages. For example, inability to efficiently efflux excess cholesterol due to deficiency of ABCA1 transporter increases cytokine production from macrophages and increase inflammation. Consistently, we have demonstrated that reduction in macrophage cholesterol content by macrophage-specific transgenic over-expression of cholesterylester (CE) hydrolase (CEH, an enzyme that hydrolyzes intracellular stored CE and rate-limits cholesterol efflux) leads to a dramatic (>20 fold) reduction in plasma IL-1? levels. However, the mechanisms underlying the crosstalk between cellular cholesterol homeostasis and inflammatory pathways have not been established as yet. Both signals required for activation of inflammatory pathways depend on cell surface proteins: signaling via cell surface receptors (e.g., TLR4) associated with membrane lipid rafts or cholesterol- enriched membrane micro-domains is required for Signal 1 and Signal 2 or NLRP3 activation is triggered by K+ efflux regulated by cell surface-associated ion channels. Since the presence/function of these cell surface receptors and channels depends on the cellular/membrane cholesterol or lipid-raft levels, We hypothesize that CEH-mediated reduction in cellular cholesterol content attenuates 1) the priming of macrophages by modulating TLR4 signaling and 2) the activation of the inflammasome via changes in cellular K+ efflux and thereby beneficially modulates macrophage function. We propose the following Aims to test the hypothesis: Aim 1: To delineate the mechanisms by which CEH-mediated changes in cellular cholesterol content can modulate macrophage priming or Signal 1. Aim 2: To delineate the mechanisms underlying CEH-mediated modulation of inflammasome activation or Signal 2 via cellular cholesterol depletion. Aim 3: To evaluate the role of CEH-mediated attenuation of Signal 1 and Signal 2-dependent macrophage activation in modulating inflammation in vivo. These studies will establish that targeted reduction in macrophage cholesterol content would simultaneously attenuate multiple metabolic diseases including Type 2 Diabetes and Coronary Artery Disease (CAD). In addition, the comprehensive approach used will define the pathways by which hydrophobic cholesterol communicates with the inflammatory machinery and identify molecular mechanisms that can be explored as potential therapeutic targets in future (e.g., regulating K+ efflux) to modulate other sterile inflammation based diseases.