Redox-dependent plasticity of oxMIF facilitates its interaction with CD74 and therapeutic antibodies.

MIF is a ubiquitous protein involved in proinflammatory processes, which undergoes an oxidation-driven conformational change to oxidized (ox)MIF. We demonstrate that hypochlorous acid, produced by neutrophil-released myeloperoxidase (MPO) under inflammatory conditions, effectively oxidizes MIF into the oxMIF isoform, which is specifically recognized by the anti-oxMIF therapeutic antibody, ON104. NMR investigation of MIF oxidized by the MPO system revealed increased flexibility throughout the MIF structure, including at several catalytic and allosteric sites. Mass spectrometry of MPO-oxMIF revealed methionines as the primary site of oxidation, whereas Pro2 and Tyr99/100 remained almost unmodified. ELISA, SPR and cell-based assays demonstrated that structural changes caused by MPO-driven oxidation promoted binding of oxMIF to its receptor, CD74, which does not occur with native MIF. These data reveal the environment and modifications that facilitate interactions between MIF and its pro-inflammatory receptor, and a route for therapeutic intervention targeting the oxMIF isoform.

The newly engineered monoclonal antibody ON104, targeting the oxidized Macrophage Migration Inhibitory Factor (oxMIF), ameliorates clinical and histopathological signs of collagen-induced arthritis.

Macrophage Migration Inhibitory Factor (MIF) is a pleiotropic inflammatory cytokine that emerged as a pivotal regulator in the pathogenesis of several autoimmune diseases including rheumatoid arthritis (RA). MIF occurs in two immunologically distinct conformational isoforms, indicated as reduced (redMIF) and oxidized MIF (oxMIF) where the latter exerts disease-related activities. In this study we demonstrate the presence of circulating oxMIF in RA patients and investigate the in vivo effects of an oxMIF-neutralizing antibody in a murine model of RA. By advanced antibody engineering we generated the fully human anti-oxMIF antibody ON104 with abolished effector functions. The therapeutic potential of ON104 was tested in a model of Collagen-Induced Arthritis (CIA) in DBA/1j mice. At disease onset, the mice received ON104 twice a week for three weeks. Clinical symptoms were assessed daily, and histological examinations of the joints were performed at the end of the study. Antibody ON104, specifically targeting human and murine oxMIF, is highly affine and does not elicit effector functions in vitro. The treatment of CIA mice with ON104 profoundly modulated disease progression with marked amelioration of clinical signs of arthritis that was associated with reduced synovial and cartilage damage and reduced F4/80-positive macrophages in the joints. These data prove that oxMIF is a relevant target in a well-known model of human RA and its specific neutralization by the antibody ON104 ameliorates clinical and histological signs of the disease in the so-treated mice. Thus, ON104 represents a new and promising treatment option for RA and possibly other autoimmune diseases.

BMPER is a marker of adipose progenitors and adipocytes and a positive modulator of adipogenesis.

Autocrine and paracrine signaling regulating adipogenesis in white adipose tissue remains largely unclear. Here we used single-cell RNA-sequencing (RNA-seq) and single nuclei RNA-sequencing (snRNA-seq) to identify markers of adipose progenitor cells (APCs) and adipogenic modulators in visceral adipose tissue (VAT) of humans and mice. Our study confirmed the presence of major cellular clusters in humans and mice and established important sex and diet-specific dissimilarities in cell proportions. Here we show that bone morphogenetic protein (BMP)-binding endothelial regulator (BMPER) is a conserved marker for APCs and adipocytes in VAT in humans and mice. Further, BMPER is highly enriched in lineage negative stromal vascular cells and its expression is significantly higher in visceral compared to subcutaneous APCs in mice. BMPER expression and release peaked by day four post-differentiation in 3T3-L1 preadipocytes. We reveal that BMPER is required for adipogenesis both in 3T3-L1 preadipocytes and in mouse APCs. Together, this study identified BMPER as a positive modulator of adipogenesis.

Late-in-life treadmill training rejuvenates autophagy, protein aggregate clearance, and function in mouse hearts.

Protein quality control mechanisms decline during the process of cardiac aging. This enables the accumulation of protein aggregates and damaged organelles that contribute to age-associated cardiac dysfunction. Macroautophagy is the process by which post-mitotic cells such as cardiomyocytes clear defective proteins and organelles. We hypothesized that late-in-life exercise training improves autophagy, protein aggregate clearance, and function that is otherwise dysregulated in hearts from old vs. adult mice. As expected, 24-month-old male C57BL/6J mice (old) exhibited repressed autophagosome formation and protein aggregate accumulation in the heart, systolic and diastolic dysfunction, and reduced exercise capacity vs. 8-month-old (adult) mice (all p < 0.05). To investigate the influence of late-in-life exercise training, additional cohorts of 21-month-old mice did (old-ETR) or did not (old-SED) complete a 3-month progressive resistance treadmill running program. Body composition, exercise capacity, and soleus muscle citrate synthase activity improved in old-ETR vs. old-SED mice at 24 months (all p < 0.05). Importantly, protein expression of autophagy markers indicate trafficking of the autophagosome to the lysosome increased, protein aggregate clearance improved, and overall function was enhanced (all p < 0.05) in hearts from old-ETR vs. old-SED mice. These data provide the first evidence that a physiological intervention initiated late-in-life improves autophagic flux, protein aggregate clearance, and contractile performance in mouse hearts.

Metabolic regulation by PPARγ is required for IL-33-mediated activation of ILC2s in lung and adipose tissue.

Type 2 innate lymphoid cells (ILC2s) play a critical role early in the response to infection by helminths and in the development of allergic reactions. ILC2s are also involved in the physiologic regulation of adipose tissue and its metabolic response to cold shock. We find that the metabolic sensor peroxisome proliferator-activated receptor gamma (PPARγ) is highly expressed in ILC2s of the lung and adipose tissue and increases responsiveness to IL-33. In turn, activation of ILC2 by IL-33 leads to increased expression of PPARγ, a prerequisite for proliferation and expression of the effector cytokines IL-5 and IL-13. In contrast, pharmacological inhibition of PPARγ leads to decreased expression of CD36 and fatty acid uptake, a necessary source of energy for ILC2s and of potential ligands for PPARγ. As a consequence, treatment of mice with a PPARγ antagonist reduces the severity of an ILC2-dependent acute airway inflammation. Together, our results demonstrate the critical role of the metabolic sensor PPARγ for the functions of ILC2s.

[Ischemia-reperfusion injury after kidney transplantation]. / Ischémie reperfusion en transplantation rénale.

Ischemia-reperfusion injury is an inescapable phenomenon in kidney transplantation. It combines lesional processes of biochemical origin associated with oxydative stress and of immunological origin in connection with the recruitment and activation of innate immunity cells. Histological lesions associate acute tubular necrosis and interstitial œdema, which can progress to interstitial fibrosis. The extent of these lesions depends on donor characteristics (age, expanded criteria donor, etc.) and cold ischemia time. In the short term, ischemia-reperfusion results in delayed recovery of graft function. Cold ischemia time also impacts long-term graft survival. Preclinical models, such as murine and porcine models, have furthered understanding of the pathophysiological mechanisms of ischemia-reperfusion injury. Due to its renal anatomical proximity to humans, the porcine model is relevant to assessment of the molecules administered to a donor or recipient, and also of additives to preservation solutions. Different donor resuscitation and graft perfusion strategies can be studied. In humans, prevention of ischemia-reperfusion injury is a research subject as concerns donor conditioning, additive molecules in preservation solutions, graft reperfusion modalities and choice of the molecules administered to the recipient. Pending significant advances in research, the goal is to achieve the shortest possible cold ischemia time.

Identification of a Paracrine Signaling Mechanism Linking CD34high Progenitors to the Regulation of Visceral Fat Expansion and Remodeling.

Accumulation of visceral (VIS) is a predictor of metabolic disorders and insulin resistance. This is due in part to the limited capacity of VIS fat to buffer lipids allowing them to deposit in insulin-sensitive tissues. Mechanisms underlying selective hypertrophic growth and tissue remodeling properties of VIS fat are not well understood. We identified subsets of adipose progenitors (APs) unique to VIS fat with differential Cd34 expression and adipogenic capacity. VIS low (Cd34 low) APs are adipogenic, whereas VIS high (Cd34 high) APs are not. Furthermore, VIS high APs inhibit adipogenic differentiation of SUB and VIS low APs in vitro through the secretion of soluble inhibitory factor(s). The number of VIS high APs increased with adipose tissue expansion, and their abundance in vivo caused hypertrophic growth, fibrosis, inflammation, and metabolic dysfunction. This study unveils the presence of APs unique to VIS fat involved in the paracrine regulation of adipogenesis and tissue remodeling.

Interleukin-1 Family Cytokines: Keystones in Liver Inflammatory Diseases.

The pyrogenic property being the first activity described, members of the interleukin-1 superfamily (IL-1α, IL-1ß, IL-18, and the newest members: IL-33, IL-36, IL-37, and IL-38) are now known to be involved in several inflammatory diseases such as obesity, atherosclerosis, cancer, viral and parasite infections, and auto-inflammatory syndromes as well as liver diseases. Inflammation processes are keystones of chronic liver diseases, of which the etiology may be viral or toxic, as in alcoholic or non-alcoholic liver diseases. Inflammation is also at stake in acute liver failure involving massive necrosis, and in ischemia-reperfusion injury in the setting of liver transplantation. The role of the IL-1 superfamily of cytokines and receptors in liver diseases can be either protective or pro-inflammatory, depending on timing and the environment. Our review provides an overview of current understanding of the IL-1 family members in liver inflammation, highlighting recent key investigations, and therapeutic perspectives. We have tried to apply the concept of trained immunity to liver diseases, based on the role of the members of the IL-1 superfamily, first of all IL-1ß but also IL-18 and IL-33, in modulating innate lymphoid immunity carried by natural killer cells, innate lymphoid cells or innate T-αß lymphocytes.

Autophagy in Adipose Tissue Physiology and Pathophysiology.

Significance: Alterations in adipose tissue function have profound consequences on whole body energy homeostasis because this tissue is central for fat accumulation, energy expenditure, glucose and insulin metabolism, and hormonal regulation. With the obesity reaching epidemic proportions globally, it is important to understand the mechanisms leading to adipose tissue malfunction. Recent Advances: Autophagy has originally been viewed as an adaptive response to cellular stress, but in recent years this process was shown to regulate important cellular processes. In adipose tissue, autophagy is a key regulator of white adipose tissue (WAT) and brown adipose tissue (BAT) adipogenesis, and dysregulated autophagy impairs fat accumulation both in vitro and in vivo. Animal studies have also suggested an important role for autophagy and mitophagy during the transition from beige to white fat. Human studies have provided evidence for altered autophagy in WAT, and these alterations correlated with the degree of insulin resistance. Critical Issues: Despite these important advances in the study of autophagy in adipose tissue, we still do not understand the physiological role of autophagy in mature white and brown adipocytes. Furthermore, several human studies involving autophagy assessment were performed on whole adipose tissue, which complicates the interpretation of the results considering the cellular heterogeneity of this tissue. Future Directions: Future studies will undoubtedly expand our understanding of the role of autophagy in fully differentiated adipocytes, and uncover novel cross-talks between this tissue and other organs in regulating lipid metabolism, redox signaling, energy homeostasis, and insulin sensitivity.

Autophagy Ablation in Adipocytes Induces Insulin Resistance and Reveals Roles for Lipid Peroxide and Nrf2 Signaling in Adipose-Liver Crosstalk.

Autophagy is a homeostatic cellular process involved in the degradation of long-lived or damaged cellular components. The role of autophagy in adipogenesis is well recognized, but its role in mature adipocyte function is largely unknown. We show that the autophagy proteins Atg3 and Atg16L1 are required for proper mitochondrial function in mature adipocytes. In contrast to previous studies, we found that post-developmental ablation of autophagy causes peripheral insulin resistance independently of diet or adiposity. Finally, lack of adipocyte autophagy reveals cross talk between fat and liver, mediated by lipid peroxide-induced Nrf2 signaling. Our data reveal a role for autophagy in preventing lipid peroxide formation and its transfer in insulin-sensitive peripheral tissues.

The Impact of Invariant NKT Cells in Sterile Inflammation: The Possible Contribution of the Alarmin/Cytokine IL-33.

Although the contribution of iNKT cells to induction of sterile inflammation is now well-established, the nature of the endogenous compounds released early after cellular stress or damage that drive their activation and recruitment remains poorly understood. More precisely, iNKT cells have not been described as being reactive to endogenous non-protein damage-associated molecular-pattern molecules (DAMPs). A second subset of DAMPs, called alarmins, are tissue-derived nuclear proteins, constitutively expressed at high levels in epithelial barrier tissues and endothelial barriers. These potent immunostimulants, immediately released after tissue damage, include the alarmin IL-33. This factor has aroused interest due to its singular action as an alarmin during infectious, allergic responses and acute tissue injury, and as a cytokine, contributing to the latter resolutive/repair phase of sterile inflammation. IL-33 targets iNKT cells, inducing their recruitment in an inflammatory state, and amplifying their regulatory and effector functions. In the present review, we introduce the new concept of a biological axis of iNKT cells and IL-33, involved in alerting and controlling the immune cells in experimental models of sterile inflammation. This review will focus on acute organ injury models, especially ischemia-reperfusion injury, in the kidneys, liver and lungs, where iNKT cells and IL-33 have been presumed to mediate and/or control the injury mechanisms, and their potential relevance in human pathophysiology.

Endogenous IL-33 Contributes to Kidney Ischemia-Reperfusion Injury as an Alarmin.

Inflammation is a prominent feature of ischemia-reperfusion injury (IRI), which is characterized by leukocyte infiltration and renal tubular injury. However, signals that initiate these events remain poorly understood. We examined the role of the nuclear alarmin IL-33 in tissue injury and innate immune response triggered by experimental kidney ischemia-reperfusion. In wild-type mice, we found that IL-33 was constitutively expressed throughout the kidney in peritubular and periglomerular spaces, mainly by microvascular endothelial cells, from which it was released immediately during IRI. Compared with wild-type mice, mice lacking IL-33 (IL-33Gt/Gt) exhibited reductions in early tubular cell injury and subsequent renal infiltration of IFN-γ/IL-17A-producing neutrophils, with preservation of renal functions. This protection associated with decreased renal recruitment of myeloid dendritic cells, natural killer (NK) cells, and invariant natural killer T (iNKT) cells, the latter of which were reported as deleterious in IRI. Increases in the level of circulating IL-12, a key IL-33 cofactor, and the expression of ST2, an IL-33-specific receptor, on the surface of iNKT cells preceded the IL-33- and iNKT cell-dependent phase of neutrophil infiltration. Furthermore, IL-33 directly targeted iNKT cells in vitro, inducing IFN-γ and IL-17A production. We propose that endogenous IL-33 is released as an alarmin and contributes to kidney IRI by promoting iNKT cell recruitment and cytokine production, resulting in neutrophil infiltration and activation at the injury site. Our findings show a novel molecular mediator contributing to innate immune cell recruitment induced by renal ischemia-reperfusion and may provide therapeutic insights into AKI associated with renal transplantation.