Activating Transcription Factor 1 Directs Mhem Atheroprotective Macrophages Through Coordinated Iron Handling and Foam Cell Protection

Abstract

Rationale: Intraplaque hemorrhage (IPH) drives atherosclerosis through the dual metabolic stresses of cholesterol-enriched erythrocyte membranes and pro-oxidant heme/iron. When clearing tissue hemorrhage, macrophages are typically seen storing either iron or lipid. We have recently defined hemorrhage-associated macrophages (HA-mac) as a plaque macrophage population that responds adaptively to IPH.Objective: This study aimed to define the key transcription factor(s) involved in HO-1 induction by heme. Methods and Results:To address this question, we used microarray analysis and transfection with siRNA and plasmids. To maintain physiological relevance, we focused on human blood-derived monocytes. We found that heme stimulates monocytes through induction of activating transcription factor 1 (ATF-1). ATF-1 coinduces heme oxygenase-1 (HO-1) and Liver X receptor beta (LXR-␤). Heme-induced HO-1 and LXR-␤ were suppressed by knockdown of ATF-1, and HO-1 and LXR-␤ were induced by ATF-1 transfection. ATF-1 required phosphorylation for full functional activity. Expression of LXR-␤ in turn led to induction of other genes central to cholesterol efflux, such as LXR-␣ and ABCA1. This heme-directed state was distinct from known macrophage states (M1, M2, Mox) and, following the same format, we have designated them Mhem. Key Words: macrophages Ⅲ atherosclerosis Ⅲ heme oxygenase-1 Ⅲ lipids Ⅲ activating transcription factor-1 A therosclerosis is an inflammatory disease of large artery walls, driven mainly by lipid peroxidant stress from oxidized low-density lipoprotein (OxLDL). 1 Intraplaque hemorrhage (IPH) is particularly important in promoting both atherosclerotic lesion progression and destabilization. 2 Thus, erythrocytes provide a potent combination of cholesterolenriched membrane lipids and heme-iron, which together pose a serious metabolic challenge in a pathology largely driven by oxidized cholesterols. 2 Along with intracranial hemorrhage, IPH is one of the most important examples of tissue damage due to extravasated blood. 2 Heme-iron is an effective peroxidant catalyst, through hydrogen peroxide coordination and Fenton chemistry 3 ; cholesterol is modified by peroxidation to potently cytotoxic and inflammatory oxysterols (5Ј, 6Ј-epoxycholesterol, 7Ј-ketocholesterol, and 21Ј-keto-cholesterol), 4 and macrophages are abundant in atherosclerosis and generate hydrogen peroxide when activated. The combination of cholesterol loading, heme/iron loading, and macrophage activation would therefore promote lipid peroxidation. 5 How monocytes entering plaques differentiate adaptively to clear hemorrhage-related iron and lipid may therefore be a key transcriptional decision in atherosclerosis. Conclusions:Heme oxygenase-1 (HO-1) is a vital enzyme for iron homeostasis and protection from oxidant stress. It catalyzes pro-oxidant heme and generates biliverdin, free iron and carbon monoxide as reaction products. 6 HO-1 activity also stimulates upregulation of ferritin genes, leading to the safe chelation of iron. 6 Biliverdin is pro...

The Mononuclear Phagocyte System in Homeostasis and Disease: A Role for Heme Oxygenase-1

“…Recently, HA-mac have been renamed Mhem, and will be referred to as such hereafter (32). Mhem are co-dependent on coordinated regulation of iron homeostasis and lipid handling (Table 1).…”

“…Macrophages treated with Hp:Hb are protected against ROS formation induced by oxLDL. However, this effect is reversed when HO-1 expression is blocked with siRNA (32). Free iron is a redox active byproduct of heme degradation by HO-1.…”

“…Mhem fail to develop when Nrf2 and ATF-1 are depleted by siRNA (32,33). These transcription factors both induce HO-1 expression, suggesting that HO-1 may regulate macrophage differentiation directly and as a consequence of its ability to suppress ROS.…”

Enhanced Expression of Hemoglobin Scavenger Receptor CD163 in Accumulated Macrophages Within Filtered Debris Between Acute Coronary Syndromes and Stable Angina Pectoris

“…Hb-derived iron can act as a catalyst in the formation of oxygen free-radicals. 12,13) In an autopsy study, IPH was seen most frequently in vulnerable thin-cap fibroatheroma (TCFA), leading to ACS, as compared with stable lesions. 1) This finding suggests IPH contributes to rapidly progressive plaque destabilization.…”

Cell-Free Hemoglobin and Its Scavenger Proteins: New Disease Models Leading the Way to Targeted Therapies

“…The available data suggest that these Hb/heme-driven macrophages constitute a unique phenotype (referred to as M-hem or M-Hb), with distinct and nonoverlapping features, within the known spectrum of macrophage phenotypes extending from proinflammatory (M1) to anti-inflammatory (M2) macrophages. The Hb/heme-driven macrophage phenotype polarization appears to be triggered by increased intracellular heme levels via a pathway involving signaling by Nrf-2 and the activating transcription factor 1 (ATF1), with subsequent coinduction of HO-1 and liver-Xreceptor b (LXRb) (Kaempfer et al 2011;Boyle et al 2012). The functional significance of the profoundly suppressed levels of HLA class 2 in these macrophages has not been evaluated, but the spectrum of functional consequences of this intriguing Hb/heme-induced condition may include enhanced control of autoimmunity and/or increased susceptibility to infection.…”