Objective To explore the mechanism by which gentiopicroside (GPS) prevents macrophage-mediated hepatic fibrosis by regulating the macrophage migration inhibitory factor (MIF)-secreted phosphoprotein 1 (SPP1) signaling pathway in hepatic stellate cells. Methods LX-2 cells were divided into control group, transforming growth factor β(TGF-β) group, and TGF-β combined with GPS (25, 50, 100, 150 μmol/mL) groups. Cell proliferation was detected by EDU assay, cell invasion was assessed by Transwell^TM assay, and the protein expressions of α-smooth muscle actin (α-SMA) and type I collagen (COL1A1) were measured by Western blot. M1-type macrophage-conditioned medium (M1-CM) was used to treat LX-2 cells in the TGF-β group and TGF-β combined with GPS group. The concentrations of inducible nitric oxide synthase (iNOS) and arginase 1 (Arg1) in the cell supernatant, as well as cell proliferation, invasion ability, and the expressions of α-SMA and COL1A1 were detected. Bioinformatics analysis was performed to identify the target intersections of GPS, hepatic fibrosis, and macrophage-related genes. Drug affinity responsive target stability (DARTS) experiments and Western blot were used to verify the regulatory effect of GPS on MIF. Furthermore, LX-2 cells were divided into control group, TGF-β group, TGF-β combined with M2-CM group, TGF-β and oe-NC combined with M2-CM group, and TGF-β and oe-MIF combined with M2-CM group to analyze the concentrations of iNOS and Arg1 in the cell supernatant, as well as changes in cell proliferation, invasion, and the expressions of α-SMA and COL1A1. LX-2 cells were also divided into control group, TGF-β group, TGF-β combined with oe-NC group, TGF-β combined with oe-MIF group, and TGF-β and oe-MIF combined with GPS group to determine the protein expressions of MIF and SPP1 by Western blot. A rat model of hepatic fibrosis was constructed to explore the potential therapeutic effects of GPS on hepatic fibrosis in vivo. Results Compared with the control group, the proliferation and invasion abilities of LX-2 cells in the TGF-β group were increased, and the protein expressions of α-SMA and COL1A1 were enhanced. GPS intervention inhibited the proliferation and invasion of LX-2 cells under TGF-β conditions and reduced the expressions of α-SMA and COL1A1. Compared with the control group, the concentration of iNOS in the cell supernatant of the TGF-β group was upregulated, while the concentration of Arg1 was decreased. M1-CM treatment further increased the concentration of iNOS, decreased the concentration of Arg1, and promoted cell proliferation and invasion, as well as upregulated the expressions of α-SMA and COL1A1 on the basis of TGF-β intervention. However, GPS could reverse the effects of M1-CM intervention. Bioinformatics analysis revealed that MIF was one of the target intersections of GPS, hepatic fibrosis, and macrophage-related genes, and GPS could target and inhibit its expression. Compared with the TGF-β group, after M2-CM intervention, the concentration of iNOS in the cell supernatant decreased, the concentration of Arg1 increased, the proliferation and invasion abilities of LX-2 cells were reduced, and the expressions of α-SMA and COL1A1 were weakened. However, overexpression of MIF reversed the effects of M2-CM intervention. Western blot results showed that compared with the control group, the protein expressions of MIF and SPP1 were enhanced in the TGF-β group. Overexpression of MIF further enhanced the expressions of MIF and SPP1, while GPS intervention inhibited the expressions of MIF and SPP1. In the animal experiment, GPS intervention treatment alleviated liver injury in rats with hepatic fibrosis and inhibited the expressions of MIF and SPP1, as well as α-SMA and COL1A1 in liver tissue. Conclusion GPS may prevent macrophage-mediated hepatic fibrosis by inhibiting the MIF-SPP1 signaling pathway in hepatic stellate cells.
Hepatic Stellate Cells/metabolism* ; Signal Transduction/drug effects* ; Macrophage Migration-Inhibitory Factors/genetics* ; Liver Cirrhosis/prevention & control* ; Macrophages/drug effects* ; Iridoid Glucosides/pharmacology* ; Humans ; Cell Proliferation/drug effects* ; Animals ; Cell Line ; Collagen Type I/metabolism* ; Collagen Type I, alpha 1 Chain ; Intramolecular Oxidoreductases/genetics* ; Rats ; Transforming Growth Factor beta/pharmacology* ; Actins/metabolism*

OBJECTIVES: Keloids are fibrotic skin disorders characterized by excessive collagen deposition and a high recurrence rate, closely associated with inflammatory mediators. However, existing epidemiological studies are limited by confounding factors and reverse causality, making it difficult to establish causation. This study aims to investigate the causal relationship between circulating cytokines and keloids using Mendelian randomization analysis. METHODS: Significant single nucleotide polymorphisms (SNPs) associated with circulating cytokines (exposures) and keloids (outcomes) were extracted from genome-wide association study (GWAS) summary datasets. Eligible SNPs were selected as instrumental variables (IVs). Exposure data were derived from a cytokine GWAS including 8 293 Finnish participants, and outcome data from a keloid GWAS based on the UK Biobank. The inverse-variance weighted (IVW) method served as the primary analytical approach to estimate causal effects, supplemented by weighted median (WME), MR-Egger regression, and other sensitivity analyses. Horizontal pleiotropy was assessed using MR-Egger regression and the MR pleiotropy residual sum and outlier (MR-PRESSO) test, while Cochran's Q test evaluated heterogeneity. Leave-one-out analysis was used to verify robustness and consistency. A reverse MR analysis was also conducted, with keloid as the exposure and cytokines as outcomes, to rule out reverse causation. RESULTS: IVW analysis identified significant positive causal associations between two cytokines and keloids-macrophage migration inhibitory factor (MIF) [odds ratio (OR)=2.081, 95% confidence interval (CI) 1.219 to 3.552, P=0.007] and monocyte chemoattractant protein-1 (MCP-1) (OR=1.673, 95% CI 1.036 to 2.701, P=0.035). Conversely, stem cell factor (SCF) showed a negative causal relationship with keloids (OR=0.518, 95% CI 0.269 to 0.998, P=0.049). Results from the MR-Egger and weighted median analyses were consistent with IVW findings. No evidence of horizontal pleiotropy was observed (P>0.05). Except for interleukin-6 (P=0.014), no heterogeneity was detected in other cytokines. Leave-one-out analysis further confirmed the robustness of the causal associations. In reverse MR analysis, keloids were causally related only to β-nerve growth factor (beta-NGF) (OR=1.048, 95% CI 1.002 to 1.095, P=0.039), with no heterogeneity or pleiotropy detected in most cytokines (P>0.05). CONCLUSIONS MIF and MCP-1 exhibit positive causal associations with keloid formation, while SCF shows a negative causal relationship. These findings provide new evidence for the causal involvement of inflammatory cytokines in keloid pathogenesis and offer potential molecular targets for developing novel keloid therapies.
Humans ; Keloid/blood* ; Mendelian Randomization Analysis ; Cytokines/genetics* ; Polymorphism, Single Nucleotide ; Genome-Wide Association Study ; Chemokine CCL2/genetics* ; Interleukin-6/genetics* ; Macrophage Migration-Inhibitory Factors/genetics* ; Male ; Stem Cell Factor/blood* ; Female ; Intramolecular Oxidoreductases

OBJECTIVE: Pneumoconiosis, a lung disease caused by irreversible fibrosis, represents a significant public health burden. This study investigates the causal relationships between gut microbiota, gene methylation, gene expression, protein levels, and pneumoconiosis using a multi-omics approach and Mendelian randomization (MR). METHODS: We analyzed gut microbiota data from MiBioGen and Esteban et al. to assess their potential causal effects on pneumoconiosis subtypes (asbestosis, silicosis, and inorganic pneumoconiosis) using conventional and summary-data-based MR (SMR). Gene methylation and expression data from Genotype-Tissue Expression and eQTLGen, along with protein level data from deCODE and UK Biobank Pharma Proteomics Project, were examined in relation to pneumoconiosis data from FinnGen. To validate our findings, we assessed self-measured gut flora from a pneumoconiosis cohort and performed fine mapping, drug prediction, molecular docking, and Phenome-Wide Association Studies to explore relevant phenotypes of key genes. RESULTS: Three core gut microorganisms were identified: Romboutsia ( OR = 0.249) as a protective factor against silicosis, Pasteurellaceae ( OR = 3.207) and Haemophilus parainfluenzae ( OR = 2.343) as risk factors for inorganic pneumoconiosis. Additionally, mapping and quantitative trait loci analyses revealed that the genes VIM, STX8, and MIF were significantly associated with pneumoconiosis risk. CONCLUSIONS This multi-omics study highlights the associations between gut microbiota and key genes ( VIM, STX8, MIF) with pneumoconiosis, offering insights into potential therapeutic targets and personalized treatment strategies.
Humans ; Male ; East Asian People/genetics* ; Europe ; Gastrointestinal Microbiome ; Lung ; Macrophage Migration-Inhibitory Factors/metabolism* ; Mendelian Randomization Analysis ; Multiomics ; Pneumoconiosis/microbiology* ; Intramolecular Oxidoreductases

OBJECTIVES: This research was performed to explore the effect of macrophage migration inhibitory factor (MIF) on the apoptosis of bone marrow mesenchymal stem cells (BMSCs) in ischemia and hypoxia environments. METHODS: The cell viability of BMSCs incubated under hypoxia/ischemia (H/I) conditions with or without pretreatment with MIF or triglycidyl isocyanurate (TGIC) was detected using cell counting kit-8 (CCK-8) analysis. Plasmids containing long noncoding RNA (lncRNA) maternally expressed gene 3 (MEG3) or β-catenin small interfering RNA (siRNA) were used to overexpress or downregulate the corresponding gene, and the p53 signaling pathway was activated by pretreatment with TGIC. The influences of MIF, overexpression of lncRNA MEG3, activation of the p53 signaling pathway, and silencing of β-catenin on H/I-induced apoptosis of BMSCs were revealed by western blotting, flow cytometry, and terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick-end labeling (TUNEL) staining. RESULTS: From the results of CCK-8 assay, western blotting, and flow cytometry, pretreatment with MIF significantly decreased the H/I-induced apoptosis of BMSCs. This effect was inhibited when lncRNA MEG3 was overexpressed by plasmids containing MEG3. The p53 signaling pathway was activated by TGIC, and β-catenin was silenced by siRNA. From western blot results, the expression levels of β-catenin in the nucleus and phosphorylated p53 (p-p53) were downregulated and upregulated, respectively, when the lncRNA MEG3 was overexpressed. Through flow cytometry, MIF was also shown to significantly alleviate the increased reactive oxygen species (ROS) level of BMSCs caused by H/I. CONCLUSIONS In summary, we conclude that MIF protected BMSCs from H/I-induced apoptosis by downregulating the lncRNA MEG3/p53 signaling pathway, activating the Wnt/β-catenin signaling pathway, and decreasing ROS levels.
Humans ; RNA, Long Noncoding/metabolism* ; Macrophage Migration-Inhibitory Factors/metabolism* ; beta Catenin/metabolism* ; Reactive Oxygen Species/metabolism* ; Sincalide/metabolism* ; Tumor Suppressor Protein p53/metabolism* ; Apoptosis ; Mesenchymal Stem Cells ; Wnt Signaling Pathway/genetics* ; RNA, Small Interfering/metabolism* ; Hypoxia/metabolism* ; Ischemia ; Bone Marrow Cells

OBJECTIVETo investigate the inhibitory effect and underlying mechanism of total saponins from Paris polyphylla var. yunnanensis on the proliferation of salivary adenoid cystic carcinoma ACC-83 cells.

METHODSIn vitro cell culture was performed. The proliferation of ACC-83 cells treated with different concentrations (5, 10, 20, 40, 60, 80, 100 μg·mL⁻¹) of total saponins from Paris polyphylla var. yunnanensis was observed using CCK-8 assay. Meanwhile, the apoptosis of ACC-83 cells treated with different concentrations (25, 50, 100 μg·mL⁻¹) of the total saponins was observed using flow cytometry. The expression levels of macrophage migration inhibitory factor (MIF) and CD74 were measured using Western blot and reverse transcription-polymerase chain reaction.

RESULTSThe total saponins from Paris polyphylla var. yunnanensis induced apoptosis and expressed dose-effect relationship. ACC-83 cells expressed MIF and CD74, and the total saponins suppressed MIF and CD74 expression in ACC-83 cells.

CONCLUSIONSThe total saponins from Paris polyphylla var. yunnanensis can significantly inhibit the proliferation, suppress MIF and CD74 expression, and promote apoptosis in ACC-83 cells. This study provides a theoretical basis for the treatment of salivary adenoid cystic carcinoma using Paris polyphylla var. yunnanensis.
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Carcinoma, Adenoid Cystic ; drug therapy ; Humans ; Intramolecular Oxidoreductases ; Liliaceae ; Macrophage Migration-Inhibitory Factors ; Rhizome ; Salivary Gland Neoplasms ; drug therapy ; Saponins ; therapeutic use

Objective To investigate the effect of 15-Deoxy-△(12,14)-prostaglandin J2 (15 d-PGJ2) on the expression of macrophage migration inhibitory factor (MIF) and its underlying mechanism in J774A.1. Methods The murine monocyte/macrophage cell line J774A.1 were divided into six groups:lipopolysaccharide (LPS) group,incubated with 1 μg/ml LPS for 1 h;normal control group,incubated with PBS for 1 h;negative control group,incubated with 5 μmol/L 15 d-PGJ2 for 1 h;15 d-PGJ2 group,incubated with 5 μmol/L 15 d-PGJ2 for 1 h followed by 1 μg/ml LPS for 1 h;GW9662 group,incubated with 5 μmol/L 15 d-PGJ2 for 1 h following GW9662 10 μmol/L for 1 h,and then incubated with 1 μg/ml LPS for 1 h;and Vehicle group,control of GW9662,GW9662 was replaced by its solvent DMSO. The expression of MIF was detected via immunofluorescence and agarose gel electrophoresis. RT-qPCR and Western blotting were used to test whether 15 d-PGJ2 could regulate mRNA and protein expression of MIF in J774A.1 upon LPS challenge. The effect of peroxisome proliferator-activated receptor-γ (PPAR-γ) antagonist GW9662 on the regulation of MIF by 15 d-PGJ2 was observed. The effects of 15 d-PGJ2 on the nuclear translocation of PPAR-γ upon LPS challenge were detected via high content screening analysis. Results MIF DNA and protein expressions were detected in J774A.1. MIF mRNA expression was up-regulated (1.75±0.09,P=0.037) when challenged with LPS and 15 d-PGJ2 inhibited its upregulation (0.84±0.08,P=0.026) in J774A.1. The protein level was consistent with the mRNA level. PPAR-γ antagonist GW9662 reversed the effect of 15 d-PGJ2 (mRNA,1.48±0.06,P=0.016;protein,1.28). Furthermore,nuclear translocation of PPAR-γ was regulated by 15 d-PGJ2 in J774A.1 upon LPS challenge(1.39±0.02 vs. 1.01±0.03,P=0.003). Conclusion 15 d-PGJ2 may down-regulate the MIF expression in J774A.1 in a PPAR-γ-dependent manner.
Anilides ; pharmacology ; Animals ; Cell Line ; Intramolecular Oxidoreductases ; metabolism ; Lipopolysaccharides ; Macrophage Migration-Inhibitory Factors ; metabolism ; Mice ; Monocytes ; drug effects ; PPAR gamma ; antagonists & inhibitors ; Prostaglandin D2 ; analogs & derivatives ; pharmacology

OBJECTIVETo investigate the interaction of single nucleotide polymorphisms of macrophage migration inhibitory factor (MIF) gene -173G/C and glutathione peroxidase 1(GPX1) gene 594C/T polymorphisms and high-fat diet in ulcerative colitis (UC).

METHODSThe genetic polymorphisms of MIF -173G/C and GPX1 594C/T were determined with a polymorphism-polymerase chain reaction (PCR)-endonuclease method in peripheral blood leukocytes derived from 1500 UC cases and 1500 healthy controls.

RESULTSThe frequencies of MIF -173CC and GPX1 594TT were 55.60% and 55.73% in the UC cases and 16.67% and 16.47% in the healthy controls, respectively. Statistical tests also showed a significant difference in the frequencies between the two groups (P<0.01; P<0.01, respectively). Individuals carrying MIF -173CC also had a significantly higher risk of UC compared with those with MIF -173GG (OR=6.8662, 95%CI: 4.5384-9.6158). Individuals carrying GPX1 594TT had a high risk of UC (OR=7.0854, 95%CI: 4.4702-10.5283). Combined analysis showed that the percentages of MIF -173CC/GPX1 594TT in the UC and control groups were 31.00% and 2.73%, respectively (P<0.01). Individuals carrying MIF -173CC/GPX1 594TT had a high risk of UC (OR=49.0113, 95%CI: 31.7364-61.8205). The high-fat diet rate of the case group was significantly higher than that of the control group (OR=3.3248, 95%CI: 1.9461-5.0193, P<0.01), and statistic analysis suggested an interaction between high-fat diet and MIF -173CC and GPX1 594TT which increase risk of UC (γ =6.9293; γ =6.9942).

CONCLUSIONMIF -173CC and GPX1 594TT and high-fat diet are the risk factors for UC, and the significant interactions between genetic polymorphisms of MIF -173G/C, GPX1 594C/T and high-fat diet may increase the risk for UC.

Case-Control Studies ; Colitis, Ulcerative ; enzymology ; genetics ; metabolism ; psychology ; Diet, High-Fat ; adverse effects ; Dietary Fats ; metabolism ; Feeding Behavior ; Female ; Gene-Environment Interaction ; Genetic Predisposition to Disease ; Glutathione Peroxidase ; genetics ; Humans ; Intramolecular Oxidoreductases ; genetics ; Macrophage Migration-Inhibitory Factors ; genetics ; Male ; Polymorphism, Single Nucleotide ; Risk Factors

Macrophage migration inhibitory factor (MIF) is originally identified in the culture medium of activated T lymphocytes as a soluble factor that inhibits the random migration of macrophages. MIF is now recognized as a multipotent cytokine involved in the regulation of immune and inf lammatory responses. In rheumatoid arthritis (RA), MIF promotes inf lammatory responses by inducing proinflammatory cytokines and tissue-degrading molecules, promoting the proliferation and survival of synovial fibroblasts, stimulating neutrophil chemotaxis, and regulating angiogenesis and osteoclast differentiation. Expression of MIF in synovial tissue and synovial fluid levels of MIF are elevated in RA patients. Specifically, MIF levels correlate with RA disease activity and high levels are associated with bone erosion. In animal models of RA, the genetic and therapeutic inhibition of MIF has been shown to control inflammation and bone destruction. Based on the role of MIF in RA pathogenesis, small molecular inhibitors targeting it or its receptor pathways could provide a new therapeutic option for RA patients.
Arthritis, Rheumatoid* ; Chemotaxis ; Cytokines ; Fibroblasts ; Humans ; Inflammation ; Macrophage Migration-Inhibitory Factors ; Macrophages* ; Models, Animal ; Neutrophils ; Osteoclasts ; Synovial Fluid ; T-Lymphocytes

BACKGROUND: Despite evidence from animal and clinical studies showing the detrimental effects of macrophage migration inhibitory factor (MIF) on bone metabolism, there are no clinical studies relating circulating MIF levels to osteoporosis-related phenotypes. This cross-sectional study investigated the association of plasma MIF with bone mineral density (BMD), bone turnover markers (BTMs), and prevalence of osteoporosis in postmenopausal Korean women. METHODS: A total of 246 women not taking any medications or diagnosed with any diseases that could affect bone metabolism were enrolled. BMD values at the lumbar spine, femoral neck, and total femur, and blood levels of MIF and BTMs were measured in all subjects. Osteoporosis was defined by World Health Organization criteria. RESULTS: Before and after adjustment for confounding variables, higher MIF levels were significantly associated with lower BMD values at all measured sites and higher levels of all BTMs. All BMD values and BTMs significantly changed in a dose-dependent fashion across increasing MIF quartile. When participants were divided into two groups according to osteoporosis status, postmenopausal women with osteoporosis demonstrated 24.2% higher plasma MIF levels than those without osteoporosis (P=0.041). The odds ratio per each standard deviation increment of MIF levels for prevalent osteoporosis was 1.32 (95% confidence interval, 1.01 to 1.73). CONCLUSION: This study provides the first epidemiological evidence that higher plasma MIF may be associated with higher risk of osteoporosis resulting from lower bone mass and higher bone turnover rate, and thus it could be a potential biomarker of poor bone health outcomes in postmenopausal women.
Animals ; Bone Density* ; Bone Remodeling* ; Confounding Factors (Epidemiology) ; Cross-Sectional Studies ; Female ; Femur ; Femur Neck ; Humans ; Macrophage Migration-Inhibitory Factors ; Macrophages* ; Metabolism ; Odds Ratio ; Osteoporosis ; Phenotype ; Plasma* ; Prevalence ; Spine ; World Health Organization

OBJECTIVE: To investigate the expression of MIF, NF-κB p65 and IL-1β in the tissue of nasal polyps and normal inferior turbinate, to analyze their relevance, and to explore their role in nasal polyps. METHOD: The infiltrating results of EOS and others inflammatory cells in 48 cases diagnosed as nasal polyps (nasal polyps group) were detected by HE staining, and the expression of MIF, NF-κB p65 and IL-1β were investigated by immunohistochemistry. Twenty-one patients who were performed septoplasty orthotics were included as the control group; the VAS and Lund-Kennedy score were used to evaluate the degree of nasal polyps in patients and the correlation analysis was conducted between the disease severity and the expression levels of this three factors. RESULT: (1) The infiltrating results of EOS and the expression level of MIF, NF-κB p65, IL-1β in nasal polyps group are obviously higher than these in the control group (P < 0.05); Spearman correlation analysis shows that MIF, NF-κb p65 and IL-1β are positively correlated with each other (r = 0.74, 0.66, 0.60, P < 0.05); the nuclear activation rate of NF-κB p65 is positively correlated with MIF, IL-1β (r = 0.67, 0.63, P < 0.05); the infiltration degree of EOS is positively correlated with MIF, IL-1β (r = 0.49, 0.55, P < 0.05), but has no correlation with the NF-κB p65 and its nuclear activation rate. (2) The VAS grade of the nasal polyps group is 8.24 ± 1.72 and the nasal endoscopic examination grade is 8.63 ± 3.81. Spearman correlation analysis shows that the VAS grade is positively correlated with the level of MIF (r = 0.71, P < 0.05), but had no correlation with NF-κB p65, its nuclear activation rate and IL-1β. The nasal endoscopic examination grade is positively correlated with MIF and the nuclear activation rate of NF-κB p65 (r = 0.79, 0.73, P < 0.05), but has no correlation with the level of NF-κB p65 and IL-1β (P > 0.05). CONCLUSION MIF, NF-κB p65 and IL-1β may promote the development of the nasal polyps, and there may exist the IL-1β--NF-κB--MIF approach in nasal polyps; MIF and NF-κB may participate in maintaining physiological function of inferior turbinate and have relations with the lightest sustained inflammation of inferior turbinate. The MIF and NF-κB p65 nuclear activation rate can be used as a standard of the nasal polyp severity and the judgement prognosis.
Humans ; Inflammation ; metabolism ; Interleukin-1beta ; metabolism ; Intramolecular Oxidoreductases ; metabolism ; Macrophage Migration-Inhibitory Factors ; metabolism ; Nasal Polyps ; metabolism ; Transcription Factor RelA ; metabolism