Heather M. Wilson

Suppressor of cytokine signaling (SOCS)1 is a key determinant of differential macrophage activation and function

Macrophages become activated by their environment and develop polarized functions: classically activated (M1) macrophages eliminate pathogens but can cause tissue injury, whereas alternatively activated (M2) macrophages promote healing and repair. Mechanisms directing polarized activation, especially in vivo, are not understood completely, and here, we examined the role of SOCS proteins. M2 macrophages activated in vitro or elicited by implanting mice i.p. with the parasitic nematode Brugia malayi display a selective and IL‐4‐dependent up‐regulation of SOCS1 but not SOCS3. Using siRNA‐targeted knockdown in BMDM, we reveal that the enhanced SOCS1 is crucial for IL‐4‐induced M2 characteristics, including a high arginase I:iNOS activity ratio, suppression of T cell proliferation, attenuated responses to IFN‐γ/LPS, and curtailed SOCS3 expression. Importantly, SOCS1 was essential in sustaining the enhanced PI3K activity that drives M2 activation, defining a new regulatory mechanism by which SOCS1 controls M2 polarization. By contrast, for M1 macrophages, SOCS1 was not only an important regulator of proinflammatory mediators (IL‐6, IL‐12, MHC class II, NO), but critically, for M1, we show that SOCS1 also restricted IL‐10 secretion and arginase I activity, which otherwise would limit the efficiency of M1 macrophage proinflammatory responses. Together, our results uncover SOCS1, not only as a feedback inhibitor of inflammation but also as a critical molecular switch that tunes key signaling pathways to effectively program different sides of the macrophage balance.

Unique Expression of Suppressor of Cytokine Signaling 3 Is Essential for Classical Macrophage Activation in Rodents In Vitro and In Vivo

On infiltrating inflamed tissue, macrophages respond to the local microenvironment and develop one of two broad phenotypes: classically activated (M1) macrophages that cause tissue injury and alternatively activated macrophages that promote repair. Understanding how this polarization occurs in vivo is far from complete, and in this study, using a Th1-mediated macrophage-dependent model of acute glomerulonephritis, nephrotoxic nephritis, we examine the role of suppressor of cytokine signaling (SOCS)1 and SOCS3. Macrophages in normal kidneys did not express detectable SOCS proteins but those infiltrating inflamed glomeruli were rapidly polarized to express either SOCS1 (27 ± 6%) or SOCS3 (54 ± 12%) but rarely both (10 ± 3%). Rat bone marrow-derived macrophages incubated with IFN-γ or LPS expressed SOCS1 and SOCS3, whereas IL-4 stimulated macrophages expressed SOCS1 exclusively. By contrast, incubation with IFN-γ and LPS together suppressed SOCS1 while uniquely polarizing macrophages to SOCS3 expressing cells. Macrophages in which SOCS3 was knocked down by short interfering RNA responded to IFN-γ and LPS very differently: they had enhanced STAT3 activity; induction of macrophage mannose receptor, arginase and SOCS1; restoration of IL-4 responsiveness that is inhibited in M1 macrophages; and decreased synthesis of inflammatory mediators (NO and IL-6) and costimulatory molecule CD86, demonstrating that SOCS3 is essential for M1 activation. Without it, macrophages develop characteristic alternatively activated markers when exposed to classical activating stimuli. Lastly, increased glomerular IL-4 in nephrotoxic nephritis inhibits infiltrating macrophages from expressing SOCS3 and was associated with attenuated glomerular injury. Consequently, we propose that SOCS3 is essential for development of M1 macrophages in vitro and in vivo.

Alterations in the concentration of an apolipoprotein E-containing subfraction of plasma high density lipoprotein in coronary heart disease

The concentrations of high density lipoprotein (HDL) subfractions in 100 healthy male subjects were compared with 100 newly presenting patients with myocardial infarction (MI) within 12 h of the onset of chest pain. A subfraction of HDL enriched in apolipoprotein E (apo E), separated by heparin-Sepharose affinity chromatography, was present in lower concentrations (P < 0.001) in the plasma of the coronary patients than in the control subjects. This finding was confirmed by a lower content (P < 0.02) of apo E, measured by ELISA, in the total HDL fraction isolated from the coronary patients. Gradient gel electrophoresis of the total HDL demonstrated that the coronary patients had a significantly decreased concentration of the large HDL particles, HDL2b, of mean diameter 10.57 nm and a higher concentration of the smaller-sized HDL3, especially HDL3c, of mean diameter 7.62 nm. The coronary patients had a lower concentration of HDL cholesterol than the control subjects, attributable to the HDL2 fraction, with no difference in HDL2a between the two groups. There was no difference in the concentration of plasma cholesterol or triglyceride. The distribution of apo E phenotypes was similar in the two groups. HDL2b produced the highest discriminant power between the two groups, followed by apo E-rich HDL, HDL2 and HDL3c Plasma cholesterol correlated strongly with apo E-rich HDL for control subjects but not for MI survivors. This study demonstrates that the inverse relationship between HDL cholesterol and coronary risk shown in epidemiological studies is attributable to the large, apo E-containing HDL subspecies which under some circumstances are implicated in cholesterol removal by reverse cholesterol transport. This study also suggests that the concentration of the large, apo E-containing HDL may provide a sensitive predictor for subjects at risk of developing coronary heart disease.

Macrophages and the kidney.

Purpose of reviewMacrophage infiltration is a hallmark of all forms of inflammatory and non-inflammatory renal injury. However, the classical view of macrophages as cells that cause injury has been superseded with evidence of their heterogeneous role, i.e. with involvement in all stages of the inflammatory process including tissue repair and healing. This review summarizes the major advances in macrophage biology achieved in the last year, highlighting the different activation states, how these are regulated, and their relevance in renal disease. Recent findingsNew concepts have emerged concerning the factors controlling monocyte recruitment into inflamed tissue and their subsequent differentiation into activated macrophages. There is now compelling evidence for the heterogeneity of macrophages in clinical disease, i.e. they appear to be able to both promote and downregulate inflammation. An increased understanding of the factors regulating the expression of pro-inflammatory or reparative characteristics by macrophages is establishing how their function can be manipulated to attenuate renal inflammation in experimental models. SummaryAn understanding of the role of macrophages at different time-points in renal inflammation, and the development of techniques for modulating macrophage activation in vivo, will provide a powerful method for exploiting the reparative attributes of these cells in clinical settings, restoring regulation to the inflammatory process and promoting healing.

Inhibition of Macrophage Nuclear Factor-κB Leads to a Dominant Anti-Inflammatory Phenotype that Attenuates Glomerular Inflammation in Vivo

Infiltrating macrophages (mphi) can cause injury or facilitate repair, depending on how they are activated by the microenvironment. Studies in vitro have defined the roles of individual cytokines and signaling pathways in activation, but little is known about how macrophages integrate the multiple signals they receive in vivo. We inhibited nuclear factor-kappaB in bone marrow-derived macrophages (BMDMs) by using a recombinant adenovirus expressing dominant-negative IkappaB (Ad-IkappaB). This re-orientated macrophage activation so they became profoundly anti-inflammatory in settings where they would normally be classically activated. In vitro, the lipopolysaccharide-induced nitric oxide, interleukin-12, and tumor necrosis factor-alpha synthesis was abrogated while interleukin-10 synthesis increased. In vivo, fluorescently labeled BMDMs transduced with Ad-IkappaB and injected into the renal artery significantly reduced inducible nitric oxide synthase and MHC class II expression when activated naturally in glomeruli of rats with nephrotoxic nephritis. Furthermore, although they only comprised 15% of glomerular macrophages, their presence significantly reduced glomerular infiltration and activation of host macrophages. Injury in nephrotoxic nephritis was also decreased when assessed morphologically and by severity of albuminuria. The results demonstrate the power of Ad-IkappaB-transduced BMDMs to inhibit injury when activated by acute immune-mediated inflammation within the glomerulus.

Macrophages heterogeneity in atherosclerosis – implications for therapy

•  Introduction •  Current concepts in monocyte heterogeneity •  Pro‐atherogenic role of plaque macrophages •  Anti‐atherogenic role of plaque macrophages •  Macrophage heterogeneity and in vitro classification •  Macrophage heterogeneity within atherosclerotic plaques •  Macrophage heterogeneity in obesity and obesity‐associated disorders •  Macrophage modulation by pathogens and tumours •  Pharmacological modulation of macrophage function •  Macrophages as a target to non‐invasively image vulnerable plaques •  Summary

Macrophages: Promising Targets for the Treatment of Atherosclerosis

Atherosclerosis is now recognised as a chronic inflammatory disease occurring within the artery wall and ultimately responsible for myocardial infarction, stroke and peripheral vascular disease. A crucial step in atherogenesis is the infiltration of monocytes into the subendothelial space of large arteries where they differentiate into macrophages and become functionally active. Macrophage accumulation within plaques is a hallmark of all stages of atherosclerosis, indeed recent studies have shown their presence has the potential to act as a non-invasive marker of disease activity and plaque stability. Activated macrophages are major players in all stages of lesion development. They not only accumulate lipids but also express effector molecules that are pro-inflammatory, cytotoxic and chemotactic. Furthermore, they secrete enzymes that degrade extracellular matrix leading to plaque destabilisation and increased risk of rupture. However, macrophages are heterogeneous and when appropriately activated they have the potential to drive tissue remodelling and ultimately vascular repair. Pharmacological modulation of macrophage activities therefore represents an important strategy for the prevention and treatment of atherosclerosis and other inflammatory diseases. The aim of this review is to give a brief overview of our current understanding of macrophage activation, distribution and function within inflamed tissue. This will provide the basis for highlighting already available and future methods to exploit specifically activated macrophages as diagnostic and therapeutic targets for atherosclerosis.

Plasminogen activator inhibitor-1 and haemostasis in obesity.

The connection between obesity and disordered haemostasis is well established, but incompletely understood. There is a strong link between inhibition of fibrinolysis and obesity, and elevation of the plasma inhibitor, plasminogen activator inhibitor-1 (PAI-1), is regarded as a central factor. Here we explore the increased risk of atherothrombotic disorders in obese subjects, and the evidence for metabolic and genetic causes. There is a clear relationship between plasma PAI-1 and obesity, and adipose tissue synthesises PAI-1, as has been shown in mouse and rat models, and more recently in human material. This tissue also produces several effector molecules that can up regulate PAI-1. These molecules include transforming growth factor beta, tumour necrosis factor alpha, angiotensin II and interleukin 6, all of which up regulate PAI-1 in various cell types. The issue of whether adipose tissue directly contributes to plasma PAI-1, or whether it primarily contributes indirectly, its products stimulating other cells to produce PAI-1 that feeds into the plasma pool, is not yet resolved. Finally, we briefly examine other proteins of haemostasis that are products of adipose tissue. Further studies are needed to define the regulation of these proteins, in adipose tissue itself and in other cells influenced by its products, in order to extend recent insights into the links between obesity and haemostasis.

A critical role for suppressor of cytokine signalling 3 in promoting M1 macrophage activation and function in vitro and in vivo.

Macrophages respond to their microenvironment and develop polarized functions critical for orchestrating appropriate inflammatory responses. Classical (M1) activation eliminates pathogens while alternative (M2) activation promotes regulation and repair. M1 macrophage activation is strongly associated with suppressor of cytokine signalling 3 (SOCS3) expression in vitro, but the functional consequences of this are unclear and the role of SOCS3 in M1‐macrophage polarization in vivo remains controversial. To address these questions, we defined the characteristics and function of SOCS3‐expressing macrophages in vivo and identified potential mechanisms of SOCS3 action. Macrophages infiltrating inflamed glomeruli in a model of acute nephritis show significant up‐regulation of SOCS3 that co‐localizes with the M1‐activation marker, inducible nitric oxide synthase. Numbers of SOCS3hi‐expressing, but not SOCS1hi‐expressing, macrophages correlate strongly with the severity of renal injury, supporting their inflammatory role in vivo. Adoptive transfer of SOCS3‐short interfering RNA‐silenced macrophages into a peritonitis model demonstrated the importance of SOCS3 in driving production of pro‐inflammatory IL‐6 and nitric oxide, while curtailing expression of anti‐inflammatory IL‐10 and SOCS1. SOCS3‐induced pro‐inflammatory effects were due, at least in part, to its role in controlling activation and nuclear accumulation of nuclear factor‐κB and activity of phosphatidylinositol 3‐kinase. We show for the first time that SOCS3 also directs the functions of human monocyte‐derived macrophages, including efficient M1‐induced cytokine production (IL‐1β, IL‐6, IL‐23, IL‐12), attenuated signal transducer and activator of transcription 3 activity and ability of antigen‐loaded macrophages to drive T‐cell responses. Hence, M1‐associated SOCS3 was a positive regulator of pro‐inflammatory responses in our rodent models and up‐regulated SOCS3 is essential for effective M1‐macrophage activation and function in human macrophages.

Alterations in plasma lipids, lipoproteins and high density lipoprotein subfractions in peripheral arterial disease

The concentrations of the major lipoprotein classes and of high density lipoprotein (HDL) subfractions in 63 male patients with arteriosclerosis of the lower limbs (claudication) were determined and compared with values from 63 healthy controls. The patients with peripheral arterial disease (PAD) had reduced levels of total HDL-cholesterol and HDL2b of large particle size, increased levels of small HDL3c particles and a high ratio of total plasma-cholesterol to HDL-cholesterol (coronary risk factor). The PAD patients, however, had lower levels of low density lipoprotein (LDL)-cholesterol but higher concentrations of very low density lipoprotein (VLDL)-cholesterol and plasma triglyceride than healthy subjects. This study therefore suggests that in PAD, the protective effect of HDL may be more important than the atherogenic effect of LDL. It further suggests that while HDL-cholesterol HDL2b and the ratio of total plasma-cholesterol to HDL-cholesterol may provide valid indices for identifying individuals at risk of PAD, other factors, such as LDL and total cholesterol, may not provide such an appropriate risk indicator.