Anti-TLR2 antibody triggers oxidative phosphorylation in microglia and increases phagocytosis of β-amyloid

Microglia are multifunctional cells that are primarily neuroprotective and a deficit in their functional integrity is likely to be a contributory factor in the deteriorating neuronal function that occurs with age and neurodegeneration. One aspect of microglial dysfunction is reduced phagocytosis, and this is believed to contribute to the accumulation of amyloid-β (Aβ) in Alzheimer’s disease (AD). Therefore, improving phagocytosis should be beneficial in limiting the amyloidosis that characterises AD

Macrophage Migration Inhibitory Factor Deficiency Ameliorates High-Fat Diet Induced Insulin Resistance in Mice with Reduced Adipose Inflammation and Hepatic Steatosis

<div><p>Macrophage infiltration is a critical determinant of high-fat diet induced adipose tissue inflammation and insulin resistance. The precise mechanisms underpinning the initiation of macrophage recruitment and activation are unclear. Macrophage migration inhibitory factor (MIF), a pro-inflammatory cytokine, displays chemokine-like properties. Circulating MIF levels are elevated during obesity however its role in high-fat diet induced adipose inflammation and insulin resistance remains elusive. Wildtype and MIF^−/− C57Bl\6J mice were fed chow or high-fat diet. Body weight and food intake was assessed. Glucose homeostasis was monitored by glucose and insulin tolerance tests. Adipose tissue macrophage recruitment and adipose tissue insulin sensitivity was evaluated. Cytokine secretion from stromal vascular fraction, adipose explants and bone marrow macrophages was measured. Inflammatory signature and insulin sensitivity of 3T3-L1-adipocytes co-cultured with wildtype and MIF^−/− macrophage was quantified. Hepatic triacylglyceride levels were assessed. MIF^−/− exhibited reduced weight gain. Age and weight-matched obese MIF^−/− mice exhibited improved glucose homeostasis coincident with reduced adipose tissue M1 macrophage infiltration. Obese MIF^−/− stromal vascular fraction secreted less TNFα and greater IL-10 compared to wildtype. Activation of JNK was impaired in obese MIF^−/−adipose, concomitant with pAKT expression. 3T3-L1-adipocytes cultured with MIF^−/− macrophages had reduced pro-inflammatory cytokine secretion and improved insulin sensitivity, effects which were also attained with MIF inhibitor ISO-1. MIF^−/− liver exhibited reduced hepatic triacyglyceride accumulation, enhanced pAKT expression and reduced NFκB activation. MIF deficiency partially protects from high-fat diet induced insulin resistance by attenuating macrophage infiltration, ameliorating adipose inflammation, which improved adipocyte insulin resistance <i>ex vivo</i>. MIF represents a potential therapeutic target for treatment of high-fat diet induced insulin resistance.</p></div

Lack of MIF improves HFD-induced hepatic steatosis and hepatic insulin sensitivity.

<p>Liver tissue was harvested from lean and obese WT and MIF^−/− mice. (a) Weight of liver tissue expressed as a percentage of total body weight. Lean = 4, obese = 4. (b) Fasting plasma ALT levels in lean and obese mice. (c) Hepatic triacylglyceride levels, lean = 4, obese = 4. (d) Hematoxylin &amp; eosin staining to visualize hepatic lipid accumulation (representative of n = 3 images per group). (e) Gene expression analysis of markers of lipogenesis and lipid storage as determined by RT-PCR. Lean = 4, obese = 4. (f) Immunoblot analysis of phosphorylated AKT and β-actin from livers of WT and MIF^−/− mice ± insulin. Lean = 3, obese = 3. (g) Unstimulated tissue phosphorylated-NFkB were determined by immunoblot analysis. Lean = 3, obese = 3. WT mice represented by white bars, MIF^−/− mice represented by black bars in all graphs. Data are mean ± SEM, *p&lt;0.05, **p&lt;0.01 and ***p&lt;0.001 w.r.t obese WT.</p

MIF deficiency partially protects from HFD-induced obesity and improves glucose homeostasis.

<p>(a) GTT (1.5 g glucose/kilogram (kg) body weight (BW)) in fasted lean and obese WT and MIF^−/− mice (white circles = WT lean; black circles =  MIF^−/− lean; white squares = WT obese; black squares = MIF^−/− obese). Lean n = 9, obese n = 9. (b) ITT (0.75 U insulin/kg BW) in fasted lean and obese WT and MIF^−/− mice (white circles = WT lean; black circles = MIF^−/− lean; white squares = WT obese; black squares = MIF^−/− obese). Lean n = 18, obese n = 18–33. (c&amp;d) AUC for WT (white bars) and MIF^−/− (black bars) mice over the course of GTT and ITT expressed as arbitrary units (AU). Lean n = 9, obese n = 18–33. (e) Plasma insulin levels over time in response to glucose challenge (white circles = WT lean; black circles = MIF−/− lean; white squares = WT obese; black squares = MIF−/− obese). Lean n = 9, obese n = 9. (f) Accumulative weight of WT (white circles) and MIF^−/− (black circles) mice. (g) Accumulative food intake of WT (white circles) and MIF^−/− (black circles). (h) Body mass composition (white bars = lean fat mass; black bars = fat fat mass). Lean n = 3–4, obese n = 8–9. Data are mean ± SEM, *p&lt;0.05, **p&lt;0.01 and ***p&lt;0.001 w.r.t obese WT.</p

MIF mRNA is elevated in adipose tissue of obese WT mice, while adipose tissue lack of MIF reduced ATM recruitment into adipose tissue following HFD.

<p>(a) Immunoblot analysis of MIF and control β-actin in adipose of lean and obese WT mice (n = 4). (b) Densitometry analysis quantifying phosphorylated MIF protein expression relative to β-actin expressed in arbitrary units (AU) in lean (white bars) and obese (black bars) mice. (c) Gene expression analysis determined by RT-PCR of <i>Mif</i> in the epididymal adipose tissue (EAT), visceral adipose tissue (VAT), stromal vascular fraction (SVF), adipocyte fraction and liver of lean (white bars) and obese (black bars) WT mice. Lean n = 3–6, obese n = 4–7. (c) Gene expression analysis of known MIF receptors <i>Cxcr4, Cxcr2, Cd74</i> in EAT of lean (white bars) and obese (black bars) mice, lean = 4–8, obese n = 3–5. (d) Recruitment of ATM into adipose tissue: cells triple positive (F4/80⁺/CD11B⁺/CD11C⁺) were classified as M1 macrophages. Cells double positive (F4/80⁺/CD11B⁺/CD11C⁻) were classified as M2 macrophages. (e) Recruitment of T-cells into adipose tissue: Cells double positive (CD3⁺/CD4⁻/CD8⁺) were classified as cytotoxic T-cells. Cells double positive (CD3⁺/CD4⁺/CD8⁻) were classified as helper T-Cells. Recruitment of cells is presented as percentage of total SVF cells. Lean WT (white bars) = 9, lean MIF^−/− (grey bars) = 9, obese WT (dark grey bars) n = 15–17, obese MIF^−/− (black bars) n = 15–17. (f) TNFα and (g) IL-10 cytokine secretion from SVF and adipocyte fraction from lean and obese WT (white bars) and MIF^−/− (black bars). Lean = 12, obese n = 12. Data are mean ± SEM, *p&lt;0.05, **p&lt;0.01 and ***p&lt;0.001 w.r.t obese WT.</p

Plasma metabolic profile and adipose tissue depot weights.

<p>Plasma was isolated from overnight fasted WT and MIF^−/− mice by cardiac puncture and metabolic markers were analyzed enzymatically. (*p&lt;0.05, **p&lt;0.01 w.r.t. WT obese; #p&lt;0.05, ##p&lt;0.01 and ###p&lt;0.001 w.r.t. w.r.t. respective lean).</p><p>Plasma metabolic profile and adipose tissue depot weights.</p