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Background: Whereas severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-specific antibody tests are increasingly being used to estimate the prevalence of SARS-CoV-2 infection, the determinants of these antibody responses remain unclear.Objectives: Our aim was to evaluate systemic and mucosal antibody responses toward SARS-CoV-2 in mild versus severe coronavirus disease 2019 (COVID-19) cases.

Objective-Sepsis is a major cause of death for intensive care patients. High concentrations of inflammatory cytokines are characteristic of severe systemic inflammation and activated monocytes are their predominant cellular source. To identify targets for antiinflammatory intervention, we investigated the response of human macrophages to inflammatory and antiinflammatory mediators. Methods and Results-We profiled gene expression in human macrophages exposed to lipopolysaccharide (LPS) and interferon (IFN)-␥ in the presence or absence of recombinant activated protein C (APC) or IL-10 and identified Wnt5A as one of the transcripts most highly induced by LPS/IFN-␥ and suppressed by APC and IL-10. We confirmed regulation of Wnt5A protein in macrophages and detected it in sera and bone marrow macrophages of patients with severe sepsis. We established that a functional Wnt5A/frizzled-5/CaMKII signaling pathway was essential for macrophage inflammatory activation. To prove the essential contribution of Wnt5A we measured inflammatory cytokines after stimulation with Wnt5A, silenced Wnt5A by siRNA, and blocked receptor binding with soluble Frizzled-related peptide-1 (sFRP1). Key Words: geneexpression Ⅲ macrophages Ⅲ activated protein C S epsis is a suspected or proven infection with a systemic inflammatory response. In severe sepsis, organ dysfunction also occurs and it is associated with a high mortality and morbidity. Severe sepsis still causes about 9.3% of all deaths in the USA. 1,2 Conclusion-Wnt5A is critically involved in inflammatory macrophage See accompanying article on page 400During sepsis, the extent of plasma protein C depletion correlates with the severity of the outcome. 3 In animal studies 4 and clinical trials APC prevented death from severe sepsis or septic shock. 5 Although this beneficial effect of APC is mostly ascribed to its anticoagulant properties, antiinflammatory effects of APC have also been proposed. 6 The direct modulation of inflammation by APC has recently been described in gene expression profiling studies with human endothelial cells. 7,8 Recently, recombinant human APC has been introduced as a therapeutic agent for treatment of patients with severe sepsis because of its unique anticoagulant and antiinflammatory properties; however, the exact mechanism of antiinflammatory action is still unknown. 9 Macrophages play a central role in inflammation by responding to and releasing of numerous inflammatory cytokines and chemokines, leading to severe systemic inflammation and septic shock. However, the knowledge of antiinflammatory interactions on the level of monocytes/macrophages is scant. Therefore, we decided to expand our investigations on antiinflammatory effects of APC on this cellular system. In the present study, we were using a whole genome expression analysis approach, to define novel targets of APC in an in vitro model of inflammatory macrophage activation. Using probes obtained from human macrophages stimulated by INF-␥ (IFN-␥) and endotoxin (LPS), we consistently found Wnt5A to ...

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Hemolysis, which occurs in many disease states, can trigger a diverse pathophysiologic cascade that is related to the specific biochemical activities of free Hb and its porphyrin component heme. Normal erythropoiesis and concomitant removal of senescent red blood cells (RBC) from the circulation occurs at rates of approximately 2 × 106 RBCs/second. Within this physiologic range of RBC turnover, a small fraction of hemoglobin (Hb) is released into plasma as free extracellular Hb. In humans, there is an efficient multicomponent system of Hb sequestration, oxidative neutralization and clearance. Haptoglobin (Hp) is the primary Hb-binding protein in human plasma, which attenuates the adverse biochemical and physiologic effects of extracellular Hb. The cellular receptor target of Hp is the monocyte/macrophage scavenger receptor, CD163. Following Hb-Hp binding to CD163, cellular internalization of the complex leads to globin and heme metabolism, which is followed by adaptive changes in antioxidant and iron metabolism pathways and macrophage phenotype polarization. When Hb is released from RBCs within the physiologic range of Hp, the potential deleterious effects of Hb are prevented. However, during hyper-hemolytic conditions or with chronic hemolysis, Hp is depleted and Hb readily distributes to tissues where it might be exposed to oxidative conditions. In such conditions, heme can be released from ferric Hb. The free heme can then accelerate tissue damage by promoting peroxidative reactions and activation of inflammatory cascades. Hemopexin (Hx) is another plasma glycoprotein able to bind heme with high affinity. Hx sequesters heme in an inert, non-toxic form and transports it to the liver for catabolism and excretion. In the present review we discuss the components of physiologic Hb/heme detoxification and their potential therapeutic application in a wide range of hemolytic conditions.

Abstract-Heme toxicity contributes to the pathogenesis of chronic inflammatory diseases, atherosclerosis, and hemolysis associated vasculopathy. Macrophage clearance of cell free hemoglobin (Hb) is thus an essential homeostatic function of these cells. We examined the transcriptional response of human PBMC derived macrophages to Hb by gene array analysis. The observed noninflammatory macrophage response was characterized by induction of an antioxidative and antiinflammatory gene expression pattern with most prominent induction of the inducible heme oxygenase (HO-1). The metabolically active Hb-CD163-HO-1 pathway resulted in synthesis of ferritin-1 of the antioxidative and antiinflammatory end products linked to heme breakdown by HO-1. This response was mediated by the Hb scavenger receptor CD163 and heme and was not related to Hb mediated depletion of reduced glutathione. In contrast to other cellular responses induced by CD163, there was no role of protein phosphorylation dependent CD163 signaling in the protective macrophage response to Hb. Instead, CD163 acted as an Hb transporter, which undergoes constitutive and ligand independent internalization and recycling between the cell surface and early endosomes.

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 adhesion-induced, transient expression and secretion of procalcitonin and CGRP in vitro may play an important role during monocyte adhesion and migration in vivo. PBMC-derived macrophages may contribute to the marked increase in circulating procalcitonin by recruiting parenchymal cells within the infected tissue, as exemplified with adipocytes.