Acute kidney injury (AKI) represents the most frequent complication after cardiac surgery. Macrophage migration inhibitory factor (MIF) is a stress-regulating cytokine that was shown to protect the heart from myocardial ischemia-reperfusion injury, but its role in the pathogenesis of AKI remains unknown. In an observational study, serum and urinary MIF was quantified in 60 patients scheduled for elective conventional cardiac surgery with the use of cardiopulmonary bypass. Cardiac surgery triggered an increase in MIF serum concentrations, and patients with high circulating MIF (>median) 12 hours after surgery had a significantly reduced risk of developing AKI (relative risk reduction, 72.7%; 95% confidence interval, 12 to 91.5%; P = 0.03). Experimental AKI was induced in wild-type and Mif^−/− mice by 30 min of ischemia followed by 6 or 24 hours of reperfusion, or by rhabdomyolysis. Mif-deficient mice exhibited increased tubular cell injury, increased regulated cell death (necroptosis and ferroptosis), and enhanced oxidative stress. Therapeutic administration of recombinant MIF after ischemia-reperfusion in mice ameliorated AKI. In vitro treatment of tubular epithelial cells with recombinant MIF reduced cell death and oxidative stress as measured by glutathione and thiobarbituric acid reactive substances in the setting of hypoxia. Our data provide evidence of a renoprotective role of MIF in experimental ischemia-reperfusion injury by protecting renal tubular epithelial cells, consistent with our observation that high MIF in cardiac surgery patients is associated with a reduced incidence of AKI.

Atherosclerosis is the underlying cause of myocardial infarction and ischemic stroke. It is a lipid-triggered and cytokine/chemokine-driven arterial inflammatory condition. We identify D-dopachrome tautomerase/macrophage migration-inhibitory factor-2 (MIF-2), a paralog of the cytokine MIF, as an atypical chemokine promoting both atherosclerosis and hepatic lipid accumulation. In hyperlipidemic Apoe ^–/– mice, Mif-2-deficiency and pharmacological MIF-2-blockade protect against lesion formation and vascular inflammation in early and advanced atherogenesis. MIF-2 promotes leukocyte migration, endothelial arrest, and foam-cell formation, and we identify CXCR4 as a receptor for MIF-2. Mif-2-deficiency in Apoe ^–/– mice leads to decreased plasma lipid levels and suppressed hepatic lipid accumulation, characterized by reductions in lipogenesis-related pathways, tri-/diacylglycerides, and cholesterol-esters, as revealed by hepatic transcriptomics/lipidomics. Hepatocyte cultures and FLIM-FRET-microscopy suggest that MIF-2 activates SREBP-driven lipogenic genes, mechanistically involving MIF-2-inducible CD74/CXCR4 complexes and PI3K/AKT but not AMPK signaling. MIF-2 is upregulated in unstable carotid plaques from atherosclerotic patients and its plasma concentration correlates with disease severity in patients with coronary artery disease. These findings establish MIF-2 as an atypical chemokine linking vascular inflammation to metabolic dysfunction in atherosclerosis.

Atherosclerosis is a chronic inflammatory condition of our arteries and the main underlying pathology of myocardial infarction and stroke. The pathogenesis is age‐dependent, but the links between disease progression, age, and atherogenic cytokines and chemokines are incompletely understood. Here, we studied the chemokine‐like inflammatory cytokine macrophage migration inhibitory factor (MIF) in atherogenic Apoe ^{− / −} mice across different stages of aging and cholesterol‐rich high‐fat diet (HFD). MIF promotes atherosclerosis by mediating leukocyte recruitment, lesional inflammation, and suppressing atheroprotective B cells. However, links between MIF and advanced atherosclerosis across aging have not been systematically explored. We compared effects of global Mif ‐gene deficiency in 30‐, 42‐, and 48‐week‐old Apoe ^{− / −} mice on HFD for 24, 36, or 42 weeks, respectively, and in 52‐week‐old mice on a 6‐week HFD. Mif ‐deficient mice exhibited reduced atherosclerotic lesions in the 30/24‐ and 42/36‐week‐old groups, but atheroprotection, which in the applied Apoe ^{− / −} model was limited to lesions in the brachiocephalic artery and abdominal aorta, was not detected in the 48/42‐ and 52/6‐week‐old groups. This suggested that atheroprotection afforded by global Mif ‐gene deletion differs across aging stages and atherogenic diet duration. To characterize this phenotype and study the underlying mechanisms, we determined immune cells in the periphery and vascular lesions, obtained a multiplex cytokine/chemokine profile, and compared the transcriptome between the age‐related phenotypes. We found that Mif deficiency promotes lesional macrophage and T‐cell counts in younger but not aged mice, with subgroup analysis pointing toward a role for Trem2 ⁺ macrophages. The transcriptomic analysis identified pronounced MIF‐ and aging‐dependent changes in pathways predominantly related to lipid synthesis and metabolism, lipid storage, and brown fat cell differentiation, as well as immunity, and atherosclerosis‐relevant enriched genes such as Plin1 , Ldlr, Cpne7, or Il34, hinting toward effects on lesional lipids, foamy macrophages, and immune cells. Moreover, Mif ‐deficient aged mice exhibited a distinct plasma cytokine/chemokine signature consistent with the notion that mediators known to drive inflamm'aging are either not downregulated or even upregulated in Mif ‐deficient aged mice compared with the corresponding younger ones. Lastly, Mif deficiency favored formation of lymphocyte‐rich peri‐adventitial leukocyte clusters. While the causative contributions of these mechanistic pillars and their interplay will be subject to future scrutiny, our study suggests that atheroprotection due to global Mif ‐gene deficiency in atherogenic Apoe ^{− / −} mice is reduced upon advanced aging and identifies previously unrecognized cellular and molecular targets that could explain this phenotype shift. These observations enhance our understanding of inflamm'aging and MIF pathways in atherosclerosis and may have implications for translational MIF‐directed strategies.

The inflammatory cytokine macrophage migration‐inhibitory factor (MIF) promotes atherosclerosis via lesional monocyte and T‐cell recruitment. B cells have emerged as important components in atherogenesis, but the interaction between MIF and B cells in atherogenesis is unknown. Here, we investigated the atherosclerotic phenotype of Mif‐gene deletion in Apoe ^−/− mice. Apoe ^−/− Mif ^−/− mice fed a Western diet exhibited strongly reduced atherosclerotic lesions in brachiocephalic artery (BC) and abdominal aorta compared with controls. This phenotype was accompanied by reduced circulating B cells. Flow cytometry revealed a B‐cell developmental defect with increased premature and immature B‐cell counts in bone marrow (BM) of Apoe ^−/− Mif ^−/− mice and diminished B‐cell numbers in spleen. This finding was linked with a decreased expression of Baff‐R and differentiation‐driving transcription factors at the immature B‐cell stage, whereas peritoneal B cells exhibited unchanged CD80 and CD86 expression but vastly decreased CD9 and elevated CD23 levels, indicating that the developmental block favors the generation of immature, egressing, and reactive B cells. Mif deficiency did not affect absolute B‐cell numbers in the vessel wall but favored a relative increase of B cells in the atheroprone BC region and the appearance of periadventitial B‐cell‐rich clusters. Of note, Mif ^−/− mice exhibited a significant increase in oxidized low‐density lipoprotein (oxLDL)‐specific antibodies after the injection of oxLDL, indicating that Mif deficiency is associated with higher sensitivity of B cells against natural‐occurring antigens such as oxLDL. Importantly, Apoe ^_l_ mice adoptively transplanted with Apoe ^−/− Mif ^−/− BM showed reduced peripheral B cells compared with Apoe ^−/− BM transplantation but no atheroprotection in the BC; also, whereas there was a selective increase in atheroprotective IgM‐anti‐oxLDL‐antibodies in global Mif deficiency, BM‐specific Mif deficiency also led to elevated proatherogenic anti‐oxLDL‐IgG. Together, these findings reveal a novel link between MIF and B cells in atherogenesis. Protection from atherosclerosis by Mif deficiency is associated with enhanced B‐cell hypersensitivity, which in global but not BM‐restricted Mif deficiency favors an atheroprotective autoantibody profile in atherosclerotic mice. Targeting MIF may induce protective B‐cell responses in atherosclerosis.—Schmitz, C., Noels, H., El Bounkari, O., Straussfeld, E., Megens, R. T. A., Sternkopf, M., Alampour‐Rajabi, S., Krammer, C., Tilstam, P. V., Gerdes, N., Bürger, C., Kapurniotu, A., Bucala, R., Jankowski, J., Weber, C., Bernhagen, J. Mif ‐deficiency favors an atheroprotective autoantibody phenotype in atherosclerosis. FASEB J . 32, 4428‐4443 (2018). www.fasebj.org

To fulfil its orchestration of immune cell trafficking, a network of chemokines and receptors developed that capitalizes on specificity, redundancy, and functional selectivity. The discovery of heteromeric interactions in the chemokine interactome has expanded the complexity within this network. Moreover, some inflammatory mediators, not structurally linked to classical chemokines, bind to chemokine receptors and behave as atypical chemokines (ACKs). We identified macrophage migration inhibitory factor (MIF) as an ACK that binds to chemokine receptors CXCR2 and CXCR4 to promote atherogenic leukocyte recruitment. Here, we hypothesized that chemokine–chemokine interactions extend to ACKs and that MIF forms heterocomplexes with classical chemokines. We tested this hypothesis by using an unbiased chemokine protein array. Platelet chemokine CXCL4L1 (but not its variant CXCL4 or the CXCR2/CXCR4 ligands CXCL8 or CXCL12) was identified as a candidate interactor. MIF/CXCL4L1 complexation was verified by co-immunoprecipitation, surface plasmon-resonance analysis, and microscale thermophoresis, also establishing high-affinity binding. We next determined whether heterocomplex formation modulates inflammatory/atherogenic activities of MIF. Complex formation was observed to inhibit MIF-elicited T-cell chemotaxis as assessed by transwell migration assay and in a 3D-matrix-based live cell-imaging set-up. Heterocomplexation also blocked MIF-triggered migration of microglia in cortical cultures in situ, as well as MIF-mediated monocyte adhesion on aortic endothelial cell monolayers under flow stress conditions. Of note, CXCL4L1 blocked binding of Alexa-MIF to a soluble surrogate of CXCR4 and co-incubation with CXCL4L1 attenuated MIF responses in HEK293-CXCR4 transfectants, indicating that complex formation interferes with MIF/CXCR4 pathways. Because MIF and CXCL4L1 are platelet-derived products, we finally tested their role in platelet activation. Multi-photon microscopy, FLIM-FRET, and proximity-ligation assay visualized heterocomplexes in platelet aggregates and in clinical human thrombus sections obtained from peripheral artery disease (PAD) in patients undergoing thrombectomy. Moreover, heterocomplexes inhibited MIF-stimulated thrombus formation under flow and skewed the lamellipodia phenotype of adhering platelets. Our study establishes a novel molecular interaction that adds to the complexity of the chemokine interactome and chemokine/receptor-network. MIF/CXCL4L1, or more generally, ACK/CXC-motif chemokine heterocomplexes may be target structures that can be exploited to modulate inflammation and thrombosis.