Relationship between peritoneal solute transport and dialysate inflammatory markers in peritoneal dialysis patients: A cross-sectional study / Relación entre el transporte de soluto peritoneal y los marcadores inflamatorios del diálisis peritoneal en pacientes en diálisis peritoneal: un estudio transversal

Background: The associated factors of peritoneal small solute transport was not fully understood. This research aimed to investigate the connection between dialysate inflammatory markers (e.g. macrophage migration inhibitory factor, MIF) in peritoneal dialysis (PD) effluent and peritoneal solute transport rate (PSTR) properties. Subjects and design: A total of 80 stable PD patients in the First ShaoYang Hospital were enrolled in present study. Overnight PD effluent and serum inflammatory markers including MIF, MCP-1, VEGF, IL-6, TNFα and TGFβ were detected. Pearson correlation analysis and Logistic regression was performed to determine the risk factors for the increased PSTR. Results: A trend toward increased values of MIF, MCP-1 and IL-6 in PD effluent was observed in subjects with high PSTR when compared with those with low PSTR. The Pearson correlation test showed that D/P Cr exhibited positive correlations with dialysis effluent MIF (r=0.32, p=0.01), MCP-1 (r=0.47, p=0.01), IL-6 (r=0.48, p=0.01). Conversely, no significant correlation was found between D/P Cr and TGF-β (r=0.04, p=0.70), TNF-ɑ (r=0.22, p=0.05), VEGF (r=0.02, p=0.86) and serum inflammatory markers. In the unadjusted regression analysis, dialysis effluent MIF (OR 2.41), MCP-1 (OR 1.72), IL-6 (OR 1.55) were associated with high PSTR condition. Multivariate logistic regression analysis showed that the adjusted odds ratios (OR) of dialysis effluent MIF for high PSTR were 2.47 in all subjects (p=0.03). Conclusion: Elevated MIF, MCP-1 and IL-6 levels in PD effluent were associated with increased PSTR. Elevated dialysis effluent MIF levels was an independent risk factor for high PSTR in subjects with PD treatment. (AU)

Antecedentes: Los factores asociados del transporte peritoneal de pequeños solutos no se conocen completamente. Esta investigación tuvo como objetivo investigar la conexión entre los marcadores inflamatorios del dializado (por ejemplo, el factor inhibidor de la migración de macrófagos [MIF]) en el efluente de diálisis peritoneal (DP) y las propiedades de la tasa de transporte de solutos peritoneal (PSTR). Sujetos y diseño: Se incluyó un total de 80 pacientes con DP estable en el primer Hospital de Shaoyang. Se detectaron efluentes de DP nocturnos y marcadores inflamatorios séricos, incluyendo MIF, MCP-1, VEGF, IL-6, TNF -ɑ, TGF -β. Se realizó un análisis de correlación de Pearson y regresión logística para determinar los factores de riesgo para la PSTR aumentada. Resultados: Se observó una tendencia hacia valores incrementados de MIF, MCP-1 e IL-6 en el efluente de DP en sujetos con PSTR alta, en comparación con aquellos con PSTR baja. La prueba de correlación de Pearson mostró que D/Pcr exhibe correlaciones positivas con el MIF del efluente diálisis (r = 0,32, p = 0,01), MCP-1 (r = 0,47, p = 0,01), IL-6 (r = 0,48, p = 0,01). Por el contrario, no se encontró una correlación significativa entre D/Pcr y TGF-β (r = 0,04, p = 0,70), TNF-ɑ (r = 0,22, p = 0,05), VEGF (r = 0,02, p = 0,86) y marcadores séricos de inflamación. En el análisis de regresión no ajustado, el MIF del efluente diálisis (OR 2,41), la MCP-1 (OR 1,72), la IL-6 (OR 1,55) se asociaron con una PSTR elevada. El análisis de regresión logística multivariante mostró que las odds ratios (OR) ajustadas del MIF del efluente diálisis para PSTR alta fueron de 2,47 en todos los sujetos (p = 0,03). Conclusión: Los niveles elevados de MIF, MCP-1 y IL-6 en el efluente de DP se asociaron con un aumento de la PSTR. Los niveles elevados del MIF del efluente diálisis fueron un factor de riesgo independiente para PSTR elevada en sujetos tratados con DP. (AU)

Neuroinflammatory CSF biomarkers MIF, sTREM1, and sTREM2 show dynamic expression profiles in Alzheimer's disease.

BACKGROUND: There is a need for novel fluid biomarkers tracking neuroinflammatory responses in Alzheimer's disease (AD). Our recent cerebrospinal fluid (CSF) proteomics study revealed that migration inhibitory factor (MIF) and soluble triggering receptor expressed on myeloid cells 1 (sTREM1) increased along the AD continuum. We aimed to assess the potential use of these proteins, in addition to sTREM2, as CSF biomarkers to monitor inflammatory processes in AD. METHODS: We included cognitively unimpaired controls (n = 67, 63 ± 9 years, 24% females, all amyloid negative), patients with mild cognitive impairment (MCI; n = 92, 65 ± 7 years, 47% females, 65% amyloid positive), AD (n = 38, 67 ± 6 years, 8% females, all amyloid positive), and DLB (n = 50, 67 ± 6 years, 5% females, 54% amyloid positive). MIF, sTREM1, and sTREM2 levels were measured by validated immunoassays. Differences in protein levels between groups were tested with analysis of covariance (corrected for age and sex). Spearman correlation analysis was performed to evaluate the association between these neuroinflammatory markers with AD-CSF biomarkers (Aß42, tTau, pTau) and mini-mental state examination (MMSE) scores. RESULTS: MIF levels were increased in MCI (p < 0.01), AD (p < 0.05), and DLB (p > 0.05) compared to controls. Levels of sTREM1 were specifically increased in AD compared to controls (p < 0.01), MCI (p < 0.05), and DLB patients (p > 0.05), while sTREM2 levels were increased specifically in MCI compared to all other groups (all p < 0.001). Neuroinflammatory proteins were highly correlated with CSF pTau levels (MIF: all groups; sTREM1: MCI, AD and DLB; sTREM2: controls, MCI and DLB). Correlations with MMSE scores were observed in specific clinical groups (MIF in controls, sTREM1 in AD, and sTREM2 in DLB). CONCLUSION: Inflammatory-related proteins show diverse expression profiles along different AD stages, with increased protein levels in the MCI stage (MIF and sTREM2) and AD stage (MIF and sTREM1). The associations of these inflammatory markers primarily with CSF pTau levels indicate an intertwined relationship between tau pathology and inflammation. These neuroinflammatory markers might be useful in clinical trials to capture dynamics in inflammatory responses or monitor drug-target engagement of inflammatory modulators.

Microphysiological head and neck cancer model identifies novel role of lymphatically secreted monocyte migration inhibitory factor in cancer cell migration and metabolism.

Regional metastasis of head and neck cancer (HNC) is prevalent (approximately 50% of patients at diagnosis), yet the underlying drivers and mechanisms of lymphatic spread remain unclear. The complex tumor microenvironment (TME) of HNC plays a crucial role in disease maintenance and progression; however, the contribution of the lymphatics remains underexplored. We created a primary patient cell derived microphysiological system that incorporates cancer-associated-fibroblasts from patients with HNC alongside a HNC tumor spheroid and a lymphatic microvessel to create an in vitro TME platform to investigate metastasis. Screening of soluble factor signaling identified novel secretion of macrophage migration inhibitory factor (MIF) by lymphatic endothelial cells conditioned in the TME. Importantly, we also observed patient-to-patient heterogeneity in cancer cell migration similar to the heterogeneity observed in clinical disease. Optical metabolic imaging at the single cell level identified a distinct metabolic profile of migratory versus non-migratory HNC cells in a microenvironment dependent manner. Additionally, we report a unique role of MIF in increasing HNC reliance on glycolysis over oxidative phosphorylation. This multicellular, microfluidic platform expands the tools available to explore HNC biology in vitro through multiple orthogonal outputs and establishes a system with enough resolution to visualize and quantify patient-to-patient heterogeneity.

Migration inhibitory factor and cluster of differentiation 74-mediated dendritic cell apoptosis exacerbates acute acetaminophen-induced liver injury.

INTRODUCTION: We investigated the role of macrophage migration inhibitory factor (MIF) on dendritic cells (DC) during acetaminophen (APAP)-induced acute liver injury (ALI) in mice. METHODS: First, we randomly divided the mice into experimental (ALI model) and control groups, then intraperitoneally injected 600 mg/kg of APAP or phosphate-buffered saline, respectively. Then, we collected liver tissue and serum samples to evaluate liver inflammation using serum alanine aminotransferase level and hematoxylin and eosin (H&E) staining of liver tissues. Flow cytometry was used to identify changes in the quantity and percentage of DCs, as well as the expression of cluster of differentiation (CD) 74 and other apoptosis-related markers in the liver. Next, we randomly divided the mice into APAP-vehicles, APAP-bone marrow-derived dendritic cells (BMDCs), APAP-MIF, APAP-IgG (isotype immunoglobin G antibody) groups (four mice per group), after APAP injection, we injected control extracts, BMDCs, mouse recombinant MIF antibodies, or IgG antibodies into the tail vein. Lastly, the severity of the liver injury and the number of DCs were assessed. RESULTS: The APAP-induced ALI mice had increased hepatic MIF expression but significantly lower amounts of hepatic DCs and apoptotic DCs than healthy mice; CD74 expression on the HDCs also increased markedly. Supplementing APAP-induced ALI mice with BMDCs or MIF antibodies significantly increased the number of hepatic DCs compared with the control mice, alleviating liver damage. CONCLUSION: The MIF/CD74 signaling pathway may mediate hepatic DC apoptosis and promote liver damage.

Preclinical Evaluation of ON203, A Novel Bioengineered mAb Targeting Oxidized Macrophage Migration Inhibitory Factor as an Anticancer Therapeutic.

High levels of macrophage migration inhibitory factor (MIF) in patients with cancer are associated with poor prognosis. Its redox-dependent conformational isoform, termed oxidized MIF (oxMIF), is a promising tumor target due to its selective occurrence in tumor lesions and at inflammatory sites. A first-generation anti-oxMIF mAb, imalumab, was investigated in clinical trials in patients with advanced solid tumors, where it was well tolerated and showed signs of efficacy. However, imalumab has a short half-life in humans, increased aggregation propensity, and an unfavorable pharmacokinetic profile. Here, we aimed to optimize imalumab by improving its physicochemical characteristics and boosting its effector functions. Point mutations introduced into the variable regions reduced hydrophobicity and the antibodies' aggregation potential, and increased plasma half-life and tumor accumulation in vivo, while retaining affinity and specificity to oxMIF. The introduction of mutations into the Fc region known to increase antibody-dependent cellular cytotoxicity resulted in enhanced effector functions of the novel antibodies in vitro, whereas reduced cytokine release from human peripheral blood mononuclear cells in the absence of target antigen by the engineered anti-oxMIF mAb ON203 versus imalumab reveals a favorable in vitro safety profile. In vivo, ON203 mAb demonstrated superior efficacy over imalumab in both prophylactic and established prostate cancer (PC3) mouse xenograft models. In summary, our data highlight the potential of the second-generation anti-oxMIF mAb ON203 as a promising immunotherapy for patients with solid tumors, warranting clinical evaluation.

Tumor-associated macrophages facilitate oral squamous cell carcinomas migration and invasion by MIF/NLRP3/IL-1ß circuit: A crosstalk interrupted by melatonin.

Invasion and migration are significant challenges for treatment of oral squamous cell carcinomas (OSCCs). Tumor-associated macrophages (TAMs) interact with cancer cells and are involved in tumor progression. Our recent study demonstrated that melatonin inhibits OSCC invasion and migration; however, the mechanism by which melatonin influences crosstalk between TAMs and OSCCs is poorly understood. In this study, a co-culture system was established to explore the interactions between human monocytic cells (THP-1 cells) and human tongue squamous cell carcinoma cells (SCC-15 cells). The results were verified using monocyte-derived macrophages (MDMs) isolated and differentiated from primary peripheral blood mononuclear cells. In vivo, assays were performed to confirm the anticancer effects of melatonin. SCC-15 cells co-cultured with THP-1 cells or MDMs exhibited increased migration and invasion, which was reversed by melatonin. Co-culture also increased the expression of macrophage migration inhibitory factor (MIF), CD40, CD163 and IL-1ß, and these changes were also reversed by melatonin. Moreover, IL-1ß secretion in THP-1 cells was MIF- and NLR family pyrin domain-containing 3 (NLRP3)-dependent, and treated with IL-1ß enhanced the invasion and migration of SCC-15 cells. Furthermore, melatonin treatment significantly decreased tumor volumes and weights, and tumors from mice treated with melatonin had lower levels of MIF, NLRP3, and IL-1ß than tumor from control mice. These results demonstrate that macrophages facilitate the progression of OSCCs by promoting the MIF/NLRP3/IL-1ß signaling axis, which can be interrupted by melatonin. Therefore, melatonin could act as an alternative anticancer agent for OSCCs by targeting this signaling axis.

Macrophage Migration Inhibitory Factor Promotes Expression of Matrix Metalloproteinases 1 and 3 in Spinal Cord Astrocytes following Gecko Tail Amputation.

BACKGROUND: The matrix metalloproteinases (MMPs) are zinc-dependent endopeptidases that play a variety of physiological and pathological roles in development, remodeling of tissues and diseases, mainly through degradation of various components of the extracellular matrix (ECM). Particularly, the MMPs have increasingly been found to mediate neuropathology following spinal cord injury (SCI). Proinflammatory mediators are potent activators of the MMPs. However, how the spinal cord regenerative vertebrates circumvent MMPs-mediated neuropathogenesis following SCI remains unclear. METHODS: Following the establishment of gecko tail amputation model, the correlation of MMP-1 (gMMP-1) and MMP-3 (gMMP-3) expression with that of macrophage migration inhibitory factor in gecko (gMIF) was assayed by RT-PCR, Western blot and immunohistochemistry. Transcriptome sequencing of primary astrocytes was performed to analyze the intracellular signal transduction of macrophage migration inhibitory factor (MIF). The effects of MMP-1 and MMP-3 induced by MIF on astrocyte migration were assessed by transwell migration assay. RESULTS: The expression of gMIF significantly increased at lesion site of the injured cord, in parallel with those of gMMP-1 and gMMP-3 in the gecko astrocytes (gAS). Transcriptome sequencing and in vitro cell model revealed that gMIF efficiently promoted the expression of gMMP-1 and gMMP-3 in gAS, which in turn contributed to the migration of gAS. Inhibition of gMIF activity following gecko SCI remarkably attenuated astrocytic expression of the two MMPs, and further influenced gecko tail regeneration. CONCLUSIONS: Gecko SCI following tail amputation promoted production of gMIF, which induced the expression of gMMP-1 and gMMP-3 in gAS. The gMIF-mediated gMMP-1 and gMMP-3 expression was involved in gAS migration and successful tail regeneration.

Tumor Necrosis Factor α-Dependent Lung Inflammation Promotes the Progression of Lung Adenocarcinoma Originating From Alveolar Type II Cells by Upregulating MIF-CD74.

Lung adenocarcinoma is the most common type of lung cancer. We recently reported that inflammation-driven lung adenocarcinoma (IDLA) originates from alveolar type (AT)-II cells, which depend on major histocompatibility complex (MHC) class II to promote the expansion of regulatory T cells. The MHC class II-associated invariant chain (CD74) binds to the macrophage migration inhibitory factor (MIF), which is associated with promoting tumor growth and invasion. However, the role of MIF-CD74 in the progression of lung adenocarcinoma and the underlying mechanisms remain unclear. We aimed to explore the role of MIF-CD74 in the progression of lung adenocarcinoma and elucidate the mechanisms by which tumor necrosis (TNF)-α-mediated inflammation regulates CD74 and MIF expression in IDLA. In human lung adenocarcinoma, CD74 was upregulated on the surface of tumor cells originating from AT-II cells, which correlated positively with lymph node metastasis, tumor origin/nodal involvement/metastasis stage, and TNF-α expression. MIF interaction with CD74 promoted the proliferation and migration of A549 and H1299 cells in vitro. Using a urethane-induced IDLA mouse model, we observed that CD74 was upregulated in tumor cells and macrophages. MIF expression was upregulated in macrophages in IDLA. Blocking TNF-α-dependent inflammation downregulated CD74 expression in tumor cells and CD74 and MIF expression in macrophages in IDLA. Conditioned medium from A549 cells or activated mouse AT-II cells upregulated MIF in macrophages by secreting TNF-α. TNF-α-dependent lung inflammation contributes to the progression of lung adenocarcinoma by upregulating CD74 and MIF expression, and AT-II cells upregulate MIF expression in macrophages by secreting TNF-α. This study provides novel insights into the function of CD74 in the progression of IDLA.

Macrophage Migration Inhibitory Factor in Psoroptes ovis: Molecular Characterization and Potential Role in Eosinophil Accumulation of Skin in Rabbit and Its Implication in the Host-Parasite Interaction.

Psoroptes ovis, a common surface-living mite of domestic and wild animals worldwide, results in huge economic losses and serious welfare issues in the animal industry. P. ovis infestation rapidly causes massive eosinophil infiltration in skin lesions, and increasing research revealed that eosinophils might play an important role in the pathogenesis of P. ovis infestation. Intradermal injection of P. ovis antigen invoked massive eosinophil infiltration, suggesting that this mite should contain some relative molecules involved in eosinophil accumulation in the skin. However, these active molecules have not yet been identified. Herein, we identified macrophage migration inhibitor factor (MIF) in P. ovis (PsoMIF) using bioinformatics and molecular biology methods. Sequence analyses revealed that PsoMIF appeared with high similarity to the topology of monomer and trimer formation with host MIF (RMSD = 0.28 angstroms and 2.826 angstroms, respectively) but with differences in tautomerase and thiol-protein oxidoreductase active sites. Reverse transcription PCR analysis (qRT-PCR) results showed that PsoMIF was expressed throughout all the developmental stages of P. ovis, particularly with the highest expression in female mites. Immunolocalization revealed that MIF protein located in the ovary and oviduct of female mites and also localized throughout the stratum spinosum, stratum granulosum, and even basal layers of the epidermis in skin lesions caused by P. ovis. rPsoMIF significantly upregulated eosinophil-related gene expression both in vitro (PBMC: CCL5, CCL11; HaCaT: IL-3, IL-4, IL-5, CCL5, CCL11) and in vivo (rabbit: IL-5, CCL5, CCL11, P-selectin, ICAM-1). Moreover, rPsoMIF could induce cutaneous eosinophil accumulation in a rabbit model and increased the vascular permeability in a mouse model. Our findings indicated that PsoMIF served as one of the key molecules contributing to skin eosinophil accumulation in P. ovis infection of rabbits.

[Effect of Trichomonas vaginalis macrophage migration inhibitory factor on THP-1 macrophages].

OBJECTIVE: To investigate the effect of Trichomonas vaginalis macrophage migration inhibitory factor (TvMIF) on THP-1 macrophages. METHODS: Recombinant TvMIF protein was prokaryotic expressed and purified, and endotoxin was removed after identification. Following exposure to TvMIF at concentrations of 0, 1, 5, 10, 50 and 100 ng/mL, the cytotoxicity of the recombinant TvMIF protein to THP-1 macrophages was tested using cell counting kit (CCK)-8 assay, and the apoptosis of THP-1 macrophages and reactive oxygen species (ROS) were detected using flow cytometry. The relative expression of nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3), caspase-1, interleukin-1ß (IL-1ß) and IL-18 genes was quantified using real-time fluorescent quantitative PCR (qPCR) assay, and the expression of caspase-1, NLRP3, gasdermin D (GSDMD), gasdermin D N-terminal (GSDMD-NT) and pro-IL-1ß proteins were determined using Western blotting assay. RESULTS: Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) displayed successful expression and purification of the recombinant TvMIF protein with a molecular weight of 15.5 kDa, and the endotoxin activity assay showed the successful removal of endotoxin in the recombinant TvMIF protein (endotoxin concentration < 0.1 EU/mL), which was feasible for the subsequent studies on protein functions. Flow cytometry revealed that the recombinant TvMIF protein at a concentration of 10 ng/mL and less promoted the apoptosis of THP-1 macrophages, and the highest apoptotic rate of THP-1 macrophages was seen following exposure to the recombinant TvMIF protein at a concentration of 5 ng/mL, while the recombinant TvMIF protein at concentrations of 50 and100 ng/mL inhibited the apoptosis of THP-1 macrophages. Exposure to the recombinant TvMIF protein at a concentration 1 ng/mL resulted in increased ROS levels in THP-1 macrophages. qPCR assay quantified significantly elevated caspase-1, NLRP3, IL-18 and IL-1ß expression in THP-1 macrophages 8 hours post-treatment with the recombinant TvMIF protein at a concentration 1 ng/mL, and Western blotting determined increased caspase-1, NLRP3, pro-IL-1ß, GSDMD and GSDMD-NT protein expression in THP-1 macrophages following exposure to the recombinant TvMIF protein at a concentration 1 ng/mL. Pretreatment with MCC950 significantly reduced GSDMD and GSDMD-NT protein expression. CONCLUSIONS: High-concentration recombinant TvMIF protein inhibits macrophage apoptosis, while low-concentration recombinant TvMIF protein activates NLRP3 inflammasome and promotes macrophage pyroptosis.

Acetate attenuates hyperoxaluria-induced kidney injury by inhibiting macrophage infiltration via the miR-493-3p/MIF axis.

Hyperoxaluria is well known to cause renal injury and end-stage kidney disease. Previous studies suggested that acetate treatment may improve the renal function in hyperoxaluria rat model. However, its underlying mechanisms remain largely unknown. Using an ethylene glycol (EG)-induced hyperoxaluria rat model, we find the oral administration of 5% acetate reduced the elevated serum creatinine, urea, and protected against hyperoxaluria-induced renal injury and fibrosis with less infiltrated macrophages in the kidney. Treatment of acetate in renal tubular epithelial cells in vitro decrease the macrophages recruitment which might have reduced the oxalate-induced renal tubular cells injury. Mechanism dissection suggests that acetate enhanced acetylation of Histone H3 in renal tubular cells and promoted expression of miR-493-3p by increasing H3K9 and H3K27 acetylation at its promoter region. The miR-493-3p can suppress the expression of macrophage migration inhibitory factor (MIF), thus inhibiting the macrophages recruitment and reduced oxalate-induced renal tubular cells injury. Importantly, results from the in vivo rat model also demonstrate that the effects of acetate against renal injury were weakened after blocking the miR-493-3p by antagomir treatment. Together, these results suggest that acetate treatment ameliorates the hyperoxaluria-induced renal injury via inhibiting macrophages infiltration with change of the miR-493-3p/MIF signals. Acetate could be a new therapeutic approach for the treatment of oxalate nephropathy.

Effect and mechanisms of dexmedetomidine combined with macrophage migration inhibitory factor inhibition on the expression of inflammatory factors and AMPK in mice.

Reperfusion after acute myocardial infarction can cause ischemia/reperfusion (I/R) injury, which not only impedes restoration of the functions of tissues and organs but may also aggravate structural tissue and organ damage and dysfunction, worsening the patient's condition. Thus, the mechanisms that underpin myocardial I/R injury need to be better understood. We aimed to examine the effect of dexmedetomidine on macrophage migration inhibitory factor (MIF) in cardiomyocytes from mice with myocardial I/R injury and to explore the mechanistic role of adenosine 5'-monophosphate-activated protein kinase (AMPK) signaling in this process. Myocardial I/R injury was induced in mice. The expression of serum inflammatory factors, reactive oxygen species (ROS), adenosine triphosphate (ATP), and AMPK pathway-related proteins, as well as myocardial tissue structure and cell apoptosis rate, were compared between mice with I/R injury only; mice with I/R injury treated with dexmedetomidine, ISO-1 (MIF inhibitor), or both; and sham-operated mice. Dexmedetomidine reduced serum interleukin (IL)-6 and tumor necrosis factor-α concentrations and increased IL-10 concentration in mice with I/R injury. Moreover, dexmedetomidine reduced myocardial tissue ROS content and apoptosis rate and increased ATP content and MIF expression. MIF inhibition using ISO-1 reversed the protective effect of dexmedetomidine on myocardial I/R injury and reduced AMPK phosphorylation. Dexmedetomidine reduces the inflammatory response in mice with I/R injury and improves adverse symptoms, and its mechanism of action may be related to the MIF-AMPK pathway.

MIF Variant rs755622 Is Associated with Severe Crohn's Disease and Better Response to Anti-TNF Adalimumab Therapy.

Crohn's disease (CD), rheumatoid arthritis, psoriatic arthritis and other inflammatory diseases comprise a group of chronic diseases with immune-mediated pathogenesis which share common pathological pathways, as well as treatment strategies including anti-TNF biologic therapy. However, the response rate to anti-TNF therapy among those diseases varies, and approximately one third of patients do not respond. Since pharmacogenetic studies for anti-TNF therapy have been more frequent for other related diseases and are rare in CD, the aim of our study was to further explore markers associated with anti-TNF response in other inflammatory diseases in Slovenian CD patients treated with the anti-TNF drug adalimumab (ADA). We enrolled 102 CD patients on ADA, for which the response was defined after 4, 12, 20 and 30 weeks of treatment, using an IBDQ questionnaire and blood CRP value. We genotyped 41 SNPs significantly associated with response to anti-TNF treatment in other diseases. We found novel pharmacogenetic association between SNP rs755622 in the gene MIF (macrophage migration inhibitory factor) and SNP rs3740691 in the gene ARFGAP2 in CD patients treated with ADA. The strongest and most consistent association with treatment response was found for the variant rs2275913 in gene IL17A (p = 9.73 × 10-3).

MIF inhibition alleviates vitiligo progression by suppressing CD8+ T cell activation and proliferation.

In vitiligo, autoreactive CD8+ T cells have been established as the main culprit considering its pathogenic role in mediating epidermal melanocyte-specific destruction. Macrophage migration inhibitory factor (MIF) is a pleiotropic molecule that plays a central role in various immune processes including the activation and proliferation of T cells; but whether MIF is intertwined in vitiligo development and progression and its involvement in aberrantly activated CD8+ T cells remains ill-defined. In this study, we found that MIF was overabundant in vitiligo patients and a mouse model for human vitiligo. Additionally, inhibiting MIF ameliorated the disease progression in vitiligo mice, which manifested as less infiltration of CD8+ T cells and more retention of epidermal melanocytes in the tail skin. More importantly, in vitro experiments indicated that MIF-inhibition suppressed the activation and proliferation of CD8+ T cells from the lymph nodes of vitiligo mice, and the effect extended to CD8+ T cells in peripheral blood mononuclear cells of vitiligo patients. Finally, CD8+ T cells derived from MIF-inhibited vitiligo mice also exhibited an impaired capacity for activation and proliferation. Taken together, our results show that MIF might be clinically targetable in vitiligo treatment, and its inhibition might ameliorate vitiligo progression by suppressing autoreactive CD8+ T cell activation and proliferation. © 2023 The Pathological Society of Great Britain and Ireland.

Detection of a glutathionyl-carbonylated group (GS-CO-) on D-dopachrome tautomerase with preferential binding of GS-CO- to MIF proteins in rat livers damaged by carbon tetrachloride.

Liver damage has been induced in animal experiments using carbon tetrachloride (CCl4), a potent hepatotoxin. CCl4 is activated by cytochrome P450 2E1, which results in the formation of various metabolites including phosgene. Although D-dopachrome tautomerase (DDT) is abundant in the liver, its role currently remains unclear. The biological activity of DDT, for which the N-terminal proline is a key site, has been detected in various tissues. We herein incidentally detected a 333 Da modification to the N-terminal proline of DDT in rat livers damaged by CCl4. We identified that this modification as glutathionyl carbonylated group, which was formed by condensation of phosgene and reduced glutathione (GSH). We examined other glutathionyl-carbonylated proteins using two dimensional-polyacrylamide gel electrophoresis, mass spectrometry, and Western blotting for GSH, and detected only one glutathionyl-carbonylated protein, macrophage migration inhibitory factor (MIF). DDT belongs to the MIF family of proteins, and amino acid sequence identity between DDT and MIF is 33%. We concluded that MIF family proteins are major targets for glutathionyl carbonylation.

Pathways linking aging and atheroprotection in Mif-deficient atherosclerotic mice.

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.

Impact of Steroid on Macrophage Migration Inhibitory Factor During and After Cardiopulmonary Bypass.

Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine released in response to glucocorticoids, which counter-regulates the effects of glucocorticoids. This study was performed to determine the impact of steroids on the expression of MIF and other pro- and anti-inflammatory cytokines during and after cardiopulmonary bypass (CPB). Twenty adult patients (10 men, 64 ± 8 years old) who underwent elective cardiac surgery by CPB were given either 2000 mg (group-H, n = 10) or 500 mg of methylprednisolone (group-L, n = 10) during CPB. The serum concentrations of MIF, interleukin (IL)-1ß, IL-8, IL-10, and tumor necrosis factor-alpha (TNF-α) were measured at eight time points until 36 hours after skin closure. The early postoperative course was uneventful for all patients. There were no significant differences in duration of operation, CPB, or aortic cross-clamping (AXC) between the two groups. MIF and IL-10 levels peaked just after the conclusion of CPB and decreased gradually thereafter. IL-1ß, IL-8, and TNF-α were undetectable throughout the study period. There were no significant differences in MIF or IL-10 levels between the two groups. Peak levels of MIF in all patients were significantly correlated with the duration of CPB and AXC, whereas no such correlation was observed for IL-10. MIF or IL-10 levels were significantly elevated during and after CPB, but there were no differences between the two doses of steroid administration. Both steroid doses sufficiently suppressed proinflammatory cytokines. MIF better reflected the invasiveness of the operation than IL-10.

Synthesis and evaluation of a new class of MIF-inhibitors in activated macrophage cells and in experimental septic shock in mice.

Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine with enzymatic activities. Anti-inflammatory effects of MIF enzyme inhibitors indicate a link between its cytokine- and catalytic activities. Herein the synthesis, docking, and bioactivity of substituted benzylidene-1-indanone and -1-tetralone derivatives as MIF-tautomerase inhibitors is reported. Many of these substituted benzylidene-1-tetralones and -indan-1-ones were potent MIF-tautomerase inhibitors (IC50 < 10 µmol/L), and the most potent inhibitors were the 1-indanone derivatives 16 and 20. Some of these compounds acted as selective enolase or ketonase inhibitors. In addition, compounds 16, 20, 26, 37 and 61 efficiently inhibited NO, TNFα and IL-6 production in lipopolysaccharide-induced macrophages. Compound 20, 37 and 61 also inhibited ROS generation, and compound 26 and 37 abolished activation of NF-κB. Compound 37 significantly augmented hypothermia induced by high dose of lipopolysaccharide in mice. The possible mechanisms of action were explored using molecular modelling and docking, as well as molecular dynamics simulations.

Distinct patterns of serum and urine macrophage migration inhibitory factor kinetics predict death in sepsis: a prospective, observational clinical study.

Macrophage migration inhibitory factor (MIF) has been considered as a biomarker in sepsis, however the predictive value of the pattern of its kinetics in the serum and in the urine has remained unclarified. It is also unclear whether the kinetics of MIF are different between males and females. We conducted a single-center prospective, observational study with repeated measurements of MIF in serum and urine on days 0, 2, and 4 from admission to the intensive care unit (ICU) in 50 adult septic patients. We found that in patients who died within 90 days, there was an increase in serum MIF level from day 0 to 4, whereas in the survivors there was rather a decrease (p = 0.018). The kinetics were sex-dependent as the same difference in the pattern was present in males (p = 0.014), but not in females (p = 0.418). We also found that urine MIF was markedly lower in patients who died than in survivors of sepsis (p < 0.050). Urine MIF levels did not show temporal changes: there was no meaningful difference between day 0 and 4. These results suggest that kinetics of serum MIF during the initial days from ICU admission can predict death, especially in male patients. Additionally, lower urine MIF levels can also indicate death without showing meaningful temporal kinetics.

Myeloid-derived MIF drives RIPK1-mediated cerebromicrovascular endothelial cell death to exacerbate ischemic brain injury.

Macrophage migration inhibitory factor (MIF) is a multifaced protein that plays important roles in multiple inflammatory conditions. However, the role of MIF in endothelial cell (EC) death under inflammatory condition remains largely unknown. Here we show that MIF actively promotes receptor-interacting protein kinase 1 (RIPK1)-mediated cell death under oxygen-glucose deprivation condition. MIF expression is induced by surgical trauma in peripheral myeloid cells both in perioperative humans and mice. We demonstrate that MIF-loaded myeloid cells induced by peripheral surgery adhere to the brain ECs after distal middle cerebral artery occlusion (dMCAO) and exacerbate the blood-brain barrier (BBB) disruption. Genetic depletion of myeloid-derived MIF in perioperative ischemic stroke (PIS) mice with MCAO following a surgical insult leads to significant reduction in ECs apoptosis and necroptosis and the associated BBB disruption. The adoptive transfer of peripheral blood mononuclear cells (PBMC) from surgical MIFΔLyz2 mice to wild-type (WT) MCAO mice also shows reduced ECs apoptosis and necroptosis compared to the transfer of PBMC from surgical MIFf  l/f  l mice to MCAO recipients. The genetic inhibition of RIPK1 also attenuates BBB disruption and ECs death compared to that of WT mice in PIS. The administration of MIF inhibitor (ISO-1) and RIPK1 inhibitor (Nec-1s) can both reduce the brain EC death and neurological deficits following PIS. We conclude that myeloid-derived MIF promotes ECs apoptosis and necroptosis through RIPK1 kinase-dependent pathway. The above findings may provide insights into the mechanism as how peripheral inflammation promotes the pathology in central nervous system.