Production Of Migration Inhibitory Factor Research Articles

HIF-1α promotes astrocytic production of macrophage migration inhibitory factor following spinal cord injury.

Macrophage migration inhibitory factor (MIF) is an important mediator of neuropathology in various central nervous system (CNS) diseases. However, little is known about its inducers for production from the nerve cells, as well as the underlying regulatory mechanism. Injury-induced HIF-1α has been shown to exacerbate neuroinflammation by activating multiple downstream target molecules. It is postulated that HIF-1α is involved in the regulation of MIF following spinal cord injury (SCI). SCI model of Sprague-Dawley rats was established by cord contusion at T8-T10. The dynamic changes of HIF-1α and MIF protein levels at lesion site of rat spinal cord were determined by Western blot. The specific cell types of HIF-1α and MIF expression were examined by immunostaining. Primary astrocytes were isolated from the spinal cord, cultured and stimulated with various agonist or inhibitor of HIF-1α for analysis of HIF-1α-mediated expression of MIF. Luciferase report assay was used to determine the relationship between HIF-1α and MIF. The Basso, Beattie, and Bresnahan (BBB) locomotor scale was used to assess the locomotor function following SCI. The protein levels of HIF-1α and MIF at lesion site were significantly elevated by SCI. Immunofluorescence demonstrated that both HIF-1α and MIF were abundantly expressed in the astrocytes of the spinal cord. By using various agonists or inhibitors of HIF-1α, it was shown that HIF-1α sufficiently induced astrocytic production of MIF. Mechanistically, HIF-1α promoted MIF expression through interaction with MIF promoter. Inhibition of HIF-1α activity using specific inhibitor markedly reduced the protein levels of MIF at lesion site following SCI, which in turn favored for the functional recovery. SCI-induced activation of HIF-1α is able to promote MIF production from astrocytes. Our results have provided new clues for SCI-induced production of DAMPs, which may be helpful for clinical treatment of neuroinflammation.

Clinical Cases of Bacterial and Fungal Sensitization in Patients with Coxarthrosis and Gonarthrosis depending on Blood Group Differences

Aim: To analyze bacterial and fungal sensitization in persons that have ability to absorb anti-A and anti-B antibodies incompatible with blood group type. Material and methods: 8 people had A blood group, 2 persons had B blood group, 22 persons showed O blood group type. 9 patients with O blood group and 2 patients with B blood group absorbed anti-A antibody. 2 patients with O blood group and 5 patients with A blood group absorbed anti-B antibody. 7 persons with 0 blood group without absorbing activity of anti-A, 4 persons with O blood group and 3 persons with A blood group without anti-B absorbing activity served as control groups. Leukocyte migration inhibition reaction was performed to estimate bacterial and fungal sensitization. Results: Persons with anti-A absorbing ability were characterized by increased antibodies, sensitization to Penicillium, Candida Albicans, Pseudomonas aeruginosa, E. Coli and Clebsiella pneumoniae, increased CIC as compared to those without anti-A absorbing ability. Patients with anti-B absorbing ability showed decreased spontaneous leukocyte migration inhibitory factor production, sensitization to Streptococcus pneumoniae and Staphylococcus aureus by cell type, sensitization to Candida Lusitaniae by humoral type. Persons with anti-A absorbing ability as compared to the persons with anti-B absorbing ability showed sensitization to Penicillium, Candida Albicans, while the persons with anti-B sensitization revealed sensitization to Streptococcus pyogenes. During incubation of erythrocytes with the heated polyclonal anti-A (and anti-B) and complement at room temperature the erythrocytes of the persons with corresponding anti-A (anti-B) absorbance were increased in size and agglutinated by sides. Conclusion: Anti-A and anti-B absorbing activity of erythrocytes is associated with different fungal and bacterial sensitization.

CXCR4/MIF axis amplifies tumor growth and epithelial-mesenchymal interaction in non-small cell lung cancer

Overexpression of C-X-C chemokine receptor type 4 (CXCR4) has been shown in several cancers, including non-small cell lung cancer (NSCLC) and is linked to early metastasis and worse prognosis. The crosstalk between cancer cells and tumor stroma promotes the growth and metastasis and CXCR4 signaling is a key element of this crosstalk.To test the effects of CXCR4 overexpression (CXCR4-OE), we transduced the human NSCLC cell line A549 by using a lentiviral vector. A 3D cell culture model showed generations of tumorspheres and the effects derived by the co-culturing of lung fibroblasts. Using a xenograft mouse model, we also studied the effects of CXCR4-OE in pulmonary cell engraftment and tumor burden in vivo.Our data indicate that CXCR4-OE leads to increased tumorsphere formation and epithelial-mesenchymal transition (EMT). CXCR4-OE by A549 cells resulted in a significant increase in the production of the CXCR4-ligand macrophage migration inhibitory factor (MIF) compared to those transduced with an empty vector (EV) or in which the CXCR4 expression was deleted (KO). In our in vitro system, we did not detect any production of the canonical CXCR4 ligand CXCL12. Autocrine MIF production and CXCR4 signaling are part of a self-perpetuating loop that amplifies tumor growth and EMT. Co-culture with lung fibroblasts further increased tumorsphere formation, partially driven by an increase in IL-6 production. When A549 cells were injected into murine lungs, we observed more abundant and significantly larger tumor lesions in recipients of CXCR4-OE A549 cells compared to those receiving EV or KO cells, consistent with our in vitro findings. Treatment of mice with the MIF antagonist ISO-1 resulted in significantly less tumor burden.In conclusion, our data highlight the role of the CXCR4-OE/MIF/IL-6 axis in epithelial mesenchymal crosstalk and NSCLC progression.

Secretome-Mediated Interactions with Intestinal Epithelial Cells: A Role for Secretome Components from Lactobacillus rhamnosus R0011 in the Attenuation of Salmonella enterica Serovar Typhimurium Secretome and TNF-α-Induced Proinflammatory Responses.

Recent evidence suggests that lactic acid bacteria communicate with host cells via secretome components to influence immune responses but less is known about gut-pathogen secretomes, impact of lactic acid bacteria secretomes on host-pathogen interactions, and the mechanisms underlying these interactions. Genome-wide microarrays and cytokine profiling were used to interrogate the impact of the Lactobacillus rhamnosus R0011 secretome (LrS) on TNF-α and Salmonella enterica subsp. enterica serovar Typhimurium secretome (STS)-induced outcomes in human intestinal epithelial cells. The LrS attenuated both TNF-α- and STS-induced gene expression involved in NF-κB and MAPK activation, as well as expression of genes involved in other immune-related signaling pathways. Specifically, the LrS induced the expression of dual specificity phosphatase 1 (DUSP1), activating transcription factor 3 (ATF3), and tribbles pseudokinase 3 (TRIB3), negative regulators of innate immune signaling, in HT-29 intestinal epithelial cells challenged with TNF-α or STS. TNF-α- and STS-induced acetylation of H3 and H4 histones was attenuated by the LrS, as was the production of TNF-α- and STS-induced proinflammatory cytokines and chemokines. Interestingly, the LrS induced production of macrophage migration inhibitory factor (MIF), a cytokine involved in host-microbe interactions at the gut interface. We propose that the LrS attenuates proinflammatory mediator expression through increased transcription of negative regulators of innate immune activity and changes in global H3 and H4 histone acetylation. To our knowledge, these findings provide novel insights into the complex multifaceted mechanisms of action behind secretome-mediated interdomain communication at the gut-mucosal interface.

Immunological aspects of carbohydrate and lipid metabolism in patients with periprosthetic infection and osteomyelitis

A study of carbohydrate and lipid metabolism and immunological examination of patients with periprosthetic infection after knee and hip joint endoprosthetics (n = 16) and osteomyelitis (n = 20) was conducted. The patients with periprosthetic infection were characterized by anemia, eosinophilia, impaired glucose tolerance, cholestasis, predisposition to elevated atherogenesis, increased circulating immune complexes, autoimmune granulocytotoxic antibodies, immunoglobulins IgM, IgG, IgA and sensitization to synovial membrane and Staphylococcus aureus. The patients with osteomyelitis did not develop increased glucose and cholesterine level, but showed increased aminotransferases, thymol test, chondroitinsulfates, decrease of calcium and production of the leukocyte migration inhibition factor (LIF), both non-specific and to the antigens of cartilage tissue and sensitization to Streptococcus and E. coli. In patients with infections of the musculoskeletal system, glucose metabolism disturbances have been associated with the presence of autoimmune lymphocytotoxic antibodies, reduced production of LiF, both non-specific and to the antigens of synovial membrane. Atherogenesis was associated with increased circulating immune complexes, autoimmune lymphocytotoxic and granulocytotoxic antibodies, decreased LIF and sensitization to connective tissue antigens and pathogenic microorganisms, especially to Streptococcus and Proteus. Anemia was associated with increased autoimmune lymphocytotoxic antibodies, delayed-type sensitization to bone and cartilage tissue, to Staphylococcus and sensitization by accelerated type to E. coli and Proteus. Physicians should analyse immunological data while treating and monitoring the patients with connective tissue infections and disturbances of glucose and cholesterine metabolism.

Biphasic MIF and SDF1 expression during podocyte injury promote CD44-mediated glomerular parietal cell migration in focal segmental glomerulosclerosis.

Glomerular parietal epithelial cell (PEC) activation, as revealed by de novo expression of CD44 and cell migration toward the injured filtration barrier, is a hallmark of podocyte injury-driven focal segmental glomerulosclerosis (FSGS). However, the signaling pathway that mediates activation of PECs in response to podocyte injury is unknown. The present study focused on CD44 signaling, particularly the roles of two CD44-related chemokines, migration inhibitory factor (MIF) and stromal cell-derived factor 1 (SDF1), and their common receptor, chemokine (C-X-C motif) receptor 4 (CXCR4), in the NEP25/LMB2 mouse podocyte-toxin model of FSGS. In the early phase of the disease, CD44-positive PECs were locally evident on the opposite side of the intact glomerular tuft and subsequently increased in the vicinity of synechiae with podocyte loss. Expression of MIF and SDF1 was first increased in injured podocytes and subsequently transferred to activated PECs expressing CD44 and CXCR4. In an immortalized mouse PEC (mPEC) line, recombinant MIF and SDF1 (rMIF and rSDF1, respectively) individually increased CD44 and CXCR4 mRNA and protein levels. rMIF and rSDF1 stimulated endogenous MIF and SDF1 production. rMIF- and rSDF1-induced mPEC migration was suppressed by CD44 siRNA. However, MIF and SDF1 inhibitors failed to show any impact on proteinuria, podocyte number, and CD44 expression in NEP25/LMB2 mice. Our data suggest that injured podocytes upregulate MIF and SDF1 that stimulate CD44 expression and CD44-mediated migration, which is enhanced by endogenous MIF and SDF1 in PECs. This biphasic expression pattern of the chemokine-CD44 axis in podocytes and PECs may be a novel mechanism of "podocyte-PEC cross-talk" signaling underlying podocyte injury-driven FSGS.

GENE-26. HOST GENETIC VARIATIONS IN MACROPHAGE MIGRATION INHIBITOR FACTOR CONFER WORSE PROGNOSIS IN GLIOBLASTOMA

Abstract Genetic and systemic prognostic factors have been identified in glioblastoma (GBM) that correlate with survival, but patient-specific genomic risk factors that impact prognosis or response to immunotherapies have not been elucidated. GBM promotes an immune suppressive microenvironment via many mechanisms including the production of macrophage migration inhibitory factor (MIF) by GBM cancer stem cells, which activates myeloid derived suppressor cells (MDSCs). Increased circulating and intracranial MDSCs as well as intratumoral MIF expression in GBM patients portend a worse overall prognosis. Endogenous MIF expression is dependent on two genetic microsatellite loci: single nucleotide polymorphisms (SNPs) and CATT repeats within the promoter. To determine whether these genetic variations were linked to GBM development and/or prognosis, we assessed peripheral nucleated blood from 520 GBM patients for the MIF SNPs and CATT repeats. MIF microsatellite frequencies were similar between the normal population and GBM patients indicating loci variability was not a risk factor for GBM development. However, newly diagnosed IDH wild-type GBM patients with a minor allele SNP who received standard of care therapy had a 3.1 month shorter progression free survival (PFS), and a 4.3 month shorter overall survival (OS) when compared to patients with two major allele SNPs. This association was seen also with the CATT-repeat analyses. Furthermore, in a multivariate analysis for PFS that included age, sex, Karnofsky performance status, MGMT methylation status, 1p/19q co-deletion, and SNP status as covariates, only age and SNP status were independently associated with shorter PFS. Taken together, patients with variant MIF microsatellite loci experienced shorter PFS and decreased OS compared to those with the most common loci. These results are currently being validated in a separate 1700 patient cohort with the intent of understanding how MIF expression influences myeloid populations within the GBM microenvironment and then developing novel peripheral GBM screening markers for aggressiveness.

Macrophage migration inhibitory factor derived from spinal cord is involved in activation of macrophages following gecko tail amputation.

Macrophages and their initiation of acute inflammation have been defined to be functionally important in tissue repair and regeneration. In injury-induced production of macrophage migration inhibitory factor (MIF), which has been described as a pleiotropic protein that participates in multiple cellular and biologic processes, it is unknown whether it is involved in the regulation of macrophage events during the epimorphic regeneration. In the model of gecko tail amputation, the protein levels of gecko MIF (gMIF) have been determined to be significantly increased in the nerve cells of the spinal cord in association with the recruitment of macrophages to the lesion site. gMIF has been shown to interact with the CD74 receptor to promote the migration of macrophages through activation of Ras homolog gene family member A and to trigger inflammatory responses through MAPK signaling pathways. The determination of microsphere phagocytosis also indicated that gMIF could enhance macrophage phagocytosis. gMIF-mediated recruitment and activation of macrophages have been found to be necessary for gecko tail regeneration, as evidenced by the depletion of macrophages using clodronate liposomes. The results present a novel function of MIF during the epimorphic regeneration, which is beneficial for insights into its pleiotropic property.-Wang, Y., Wei, S., Song, H., Zhang, X., Wang, W., Du, N., Song, T., Liang, H., Chen, X., Wang, Y. Macrophage migration inhibitory factor derived from spinal cord is involved in activation of macrophages following gecko tail amputation.

Artificial shear stress effects on leukocytes at a biomaterial interface.

Medical devices, such as ventricular assist devices (VADs), introduce both foreign materials and artificial shear stress to the circulatory system. The effects these have on leukocytes and the immune response are not well understood. Understanding how these two elements combine to affect leukocytes may reveal why some patients are susceptible to recurrent device-related infections and provide insight into the development of pump thrombosis. Biomaterials-DLC: diamond-like carbon-coated stainless steel; Sap: single-crystal sapphire; and Ti: titanium alloy (Ti6 Al4 V) were attached to the parallel plates of a rheometer. Whole human blood was left between the two discs for 5minutes at +37°C with or without the application of shear stress (0s-1 or 1000s-1 ). Blood was removed and used for complete blood cell counts, flow cytometry (leukocyte activation, cell death, microparticle generation, phagocytic ability, and reactive oxygen species [ROS] production), and the production of pro-inflammatory cytokines. L-selectin expression on monocytes was decreased when blood was exposed to the biomaterials both with and without shear. Applying shear stress to blood on a Sap and Ti surface led to activation of neutrophils shown as decreased L-selectin expression. Sap and Ti blunted the LPS-stimulated macrophage migration inhibitory factor (MIF) production, most notably when sheared on Ti. The biomaterials used here have been shown to activate leukocytes in a static environment. The introduction of shear appears to exacerbate this activation. Interestingly, a widely accepted biocompatible material (Ti) utilized in many different types of devices has the capacity for immune cell activation and inhibition of MIF secretion when combined with shear stress. These findings contribute to our understanding of the contribution of biomaterials and shear stress to recurrent infections and vulnerability to sepsis in some VAD patients as well as pump thrombosis.

Endogenous biological factors modulated by substrate stiffness regulate endothelial differentiation of mesenchymal stem cells.

During the process of tissue regeneration facilitated by stem cells, physical properties of a scaffold affect behavior and activities of the cell. To enhance differentiation of human mesenchymal stem cells (MSCs) into endothelial-like cells (ELCs), we used electrospun fibrous substrates with different stiffness to enhance the differentiation. A simple method of annealing with different lengths of treatment time was employed to modulate stiffness of electrospun fibrous substrates without changing their chemistry. We seeded MSCs on substrates with different stiffness to study how stiffness of a culture substrate affects differentiation of MSCs into ELCs. Results of RT-PCR and western blotting revealed that stiffer substrates with the average surface modulus of 7.82 MPa induced differentiated MSCs to express more VEGF, CD31, and vWF mRNA transcripts and proteins than softer ones with that of 3.8 or 1.44 MPa. We also found that the production of macrophage migration inhibitory factor (MIF) in ELCs was increased with substrate stiffness. After silencing MIF mRNA, MSCs during differentiation showed lower expression levels of VEGF, CD31, and vWF than control cells whereas VEGF-silenced and control cells expressed comparable levels of MIF, indicating that MIF is an upstream molecule regulating VEGF in the mechanism. Our findings provide new insight into how stiffness of a culture substrate regulates differentiation of MSCs into ELCs. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 1595-1603, 2018.

Proteomic Analysis Shows Constitutive Secretion of MIF and p53-associated Activity of COX-2−/− Lung Fibroblasts

The differential expression of two closelyassociated cyclooxygenase isozymes, COX-1 and COX-2, exhibited functions beyond eicosanoid metabolism. We hypothesized that COX-1 or COX-2 knockout lung fibroblasts may display altered protein profiles which may allow us to further differentiate the functional roles of these isozymes at the molecular level. Proteomic analysis shows constitutive production of macrophage migration inhibitory factor (MIF) in lung fibroblasts derived from COX-2−/− but not wild-type (WT) or COX-1−/− mice. MIF was spontaneously released in high levels into the extracellular milieu of COX2−/− fibroblasts seemingly from the preformed intracellular stores, with no change in the basal gene expression of MIF. The secretion and regulation of MIF in COX-2−/− was “prostaglandin-independent.” GO analysis showed that concurrent with upregulation of MIF, there is a significant surge in expression of genes related to fibroblast growth, FK506 binding proteins, and isomerase activity in COX-2−/− cells. Furthermore, COX-2−/− fibroblasts also exhibit a significant increase in transcriptional activity of various regulators, antagonists, and co-modulators of p53, as well as in the expression of oncogenes and related transcripts. Integrative Oncogenomics Cancer Browser (IntroGen) analysis shows downregulation of COX-2 and amplification of MIF and/or p53 activity during development of glioblastomas, ependymoma, and colon adenomas. These data indicate the functional role of the MIF-COX-p53 axis in inflammation and cancer at the genomic and proteomic levels in COX-2-ablated cells. This systematic analysis not only shows the proinflammatory state but also unveils a molecular signature of a pro-oncogenic state of COX-1 in COX-2 ablated cells.

Artesunate inhibits type I interferon-induced production of macrophage migration inhibitory factor in patients with systemic lupus erythematosus.

Macrophage migration inhibitory factor (MIF) is a key regulator of both atherosclerosis and systemic lupus erythematosus (SLE), yet factors leading to its overproduction remain unclear. To explore regulation of MIF in SLE, we studied effects and potential mechanisms of type I interferon (IFN) and artesunate (ART), an antimalarial agent extracted from Chinese herbs, on levels of MIF. Serum and peripheral blood cells from SLE patients and healthy controls were measured for MIF levels by ELISA and type I IFN-inducible gene expressions by real-time PCR, respectively, and assessed for associations by Spearman correlation. ART was added to human umbilical vein endothelial cell (HUVEC) cultures with or without prior IFNα-1b stimulation and to SLE peripheral blood mononuclear cell (PBMC) cultures. Protein levels of STATs and phosphorylated (p-) STATs in HUVECs were determined by Western blotting. Serum MIF levels were elevated in SLE patients and positively associated with disease activity (r = 0.86, p < 0.0001), accumulated damage (r = 0.34, p < 0.05), and IFN scores in SLE PBMCs (r = 0.74, p = 0.0002). The addition of IFNα-1b promoted MIF production in a time- and dose-dependent manner in HUVEC cultures. ART could inhibit expressions of IFN-inducible genes (LY6E and ISG15) in both HUVEC and SLE PBMC cultures, and suppress MIF production and over-expression of p-STAT1, but not p-STAT3 or STAT5, induced by IFNα-1b stimulation. IFNγ-induced expression of p-STAT1 in HUVECs was not inhibited by ART. MIF could be regulated by type I IFN in SLE patients. ART counteracts the effect of IFNα to inhibit MIF production by blocking STAT1 phosphorylation and thus may have therapeutic potential for SLE-associated atherosclerosis.

Circulation: Clinical Summaries.

Circulation: Clinical Summaries.

HIF-α/MIF and NF-κB/IL-6 Axes Contribute to the Recruitment of CD11b+Gr-1+ Myeloid Cells in Hypoxic Microenvironment of HNSCC

CD11b+Gr-1+ myeloid cells have gained much attention due to their roles in tumor immunity suppression as well as promotion of angiogenesis, invasion, and metastases. However, the mechanisms by which CD11b+Gr-1+ myeloid cells recruit to the tumor site have not been well clarified. In the present study, we showed that hypoxia could stimulate the migration of CD11b+Gr-1+ myeloid cells through increased production of macrophage migration inhibitory factor (MIF) and interleukin-6 (IL-6) by head and neck squamous cell carcinoma (HNSCC) cells. Hypoxia-inducible factor-1α (HIF-1α)- and HIF-2α-dependent MIF regulated chemotaxis, differentiation, and pro-angiogenic function of CD11b+Gr-1+ myeloid cells through binding to CD74/CXCR2, and CD74/CXCR4 complexes, and then activating p38/mitogen-activated protein kinase (MAPK) and phosphatidylinositide 3-kinases (PI3K)/AKT signaling pathways. Knockdown (KD) of HIF-1α and HIF-2α in HNSCC cells decreased MIF level but failed to inhibit the CD11b+Gr-1+ myeloid cell migration, because HIF-1α/2α KD enhanced nuclear factor κB (NF-κB) activity that increased IL-6 secretion. Simultaneously blocking NF-κB and HIF-1α/HIF-2α had better inhibitory effect on CD11b+Gr-1+ myeloid cell recruitment in the hypoxic zone than individually silencing HIF-1α/2α or NF-κB. In conclusion, the interaction between HIF-α/MIF and NF-κB/IL-6 axes plays an important role in the hypoxia-induced accumulation of CD11b+Gr-1+ myeloid cells and tumor growth in HNSCC.

Abstract 1547: Tumor-derived macrophage Migration Inhibitory Factor and interleukin-6 cooperated in hypoxic accumulation of CD11b+Gr-1+ myeloid cells.

Abstract Recently, myeloid derived suppressor cells (MDSCs) have gained much attention due to their role in tumor immunity suppression as well as promotion of angiogenesis, tumor cell invasion, and metastases. However, the mechanisms that regulate MDSCs recruitment have not been well clarified. Here, we demonstrated that hypoxic tumor microenvironment stimulate the recruitment of MDSCs through increased production of macrophage migration inhibitory factor (MIF) and interleukin-6 (IL-6) by head and neck squamous cell carcinoma (HNSCC) cells. Knock-down (KD) of HIF-1/2α in HNSCC cells decreased MIF level but did not inhibit the MDSCs migration, because HIF-1/2α KD increased NF-κB activity which further enhanced IL-6 secretion. In vivo administration of neutralizing IL-6 antibody and inhibitors against MIF and NF-κB decreased MDSCs accumulation, accompanied by reduced micro-vessel density (MVD), lymphatic vessel density (LVD), and xenograft tumor growth. The supplementation of MDSCs increased MVD, LVD, and xenograft tumor growth. In conclusion, hypoxic HNSCC cells may stimulate the accumulation of MDSCs through secreting MIF and IL-6. This process is mediated by interactions between NF-κB and HIF-1/2α. Strategies targeting hypoxic tumor microenvironment may represent attractive approaches to HNSCC therapy. [G Zhu and Y Tang contributed equally to this paper.] Citation Format: Guiquan Zhu, Yaling Tang, Ling Li, Xinhua Liang. Tumor-derived macrophage Migration Inhibitory Factor and interleukin-6 cooperated in hypoxic accumulation of CD11b+Gr-1+ myeloid cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1547. doi:10.1158/1538-7445.AM2013-1547

Evaluation of Macrophage Migration Inhibitory Factor Level in Sera of Patients with Alopecia Areata

Backgroundalopecia areata(AA) is a common recurrent non‐scarring type of hair loss. the pathogenesis of AA is still uncertain. The immune system has been implicated in its pathogenesis and certain immunomodulatory cytokines play a role in its development. Migration inhibitory factor(MIF) was reported to the first lymphokine which prevents the random migration of macrophages. A recent finding demonstrated that MIF innitiates inflammation and immune response by stimulating the secretion of several pro‐inflammatory cytokines. As regards skin diseases, MIF production by peripheral blood mononuclear cells has been reported to be markedly increased in patients with atopic dermatitis.The aim of the present work was to measure the serum MIF concentration in patients with AA and in health indivduals.Subjects and methodsThe present study was carried out on 30 patients with AA and 15 healthy controls. The serum level of MIF was measured by enzyme linked immunosorbent assay (ELISA).Resultsthe mean level of MIF in AA patients was 6.535ng/ml+3.51(mean+SD), while in healthy individuals was 1.89ng/ml+0.91(mean+SD)with statistical sinificant difference (p‐0.016).ConclusionMean serum MIF in patients with AA was higher than that in healthy subjects indicating that MIF has a role in the etiopathogenesis of AA

Macrophage migration inhibitory factor gene polymorphism is not associated with pemphigus vulgaris in Iranian patients

Promoter polymorphisms of the macrophage migration inhibitory factor gene are associated with increased production of macrophage migration inhibitory factor. Elevated levels of macrophage migration inhibitory factor have been observed in the sera of patients with pemphigus vulgaris. More than this, macrophage migration inhibitory factor promoter gene polymorphism has been found to confer increased risk of susceptibility to chronic inflammatory diseases. We investigated whether there is an association between promoter polymorphism of the macrophage migration inhibitory factor gene and pemphigus vulgaris. One hundred and six patients with pemphigus vulgaris, and a control panel of one hundred healthy volunteers were genotyped for a single nucleotide polymorphism identified in the 5'-flanking region at the position -173 of the gene, using polymerase chain reaction-restriction fragment length analysis. We found a notably high prevalence of C/C genotype in our nation but no significant difference was observed between patients and controls. The result of this study using a large and well documented trial of patients showed that macrophage migration inhibitory factor -173G-C polymorphism is not associated with pemphigus vulgaris; but as the role of macrophage migration inhibitory factor in the inflammatory process has not been delineated in detail and the prevalence of C/C genotype is notably higher in our nation, this finding merits more consideration.

CD74 and macrophage migration inhibitory factor as therapeutic targets in gastric cancer.

To investigate the relationship and molecular features of CD74/macrophage migration inhibitory factor (MIF)/Toll-like receptor 4 (TLR4) in gastric cancer. CD74, MIF and TLR4 expression in the paraffin-embedded sections of gastric cancer from 120 patients were detected by immunohistochemical staining. Knock down of CD74 expression in gastric cancer cell line MKN-45 was performed by lentivirus transduction and detected by Western blotting. MKN-45 cell proliferation assay under the stimulants was measured by the cell counting kit 8 (CCK8) assay and MIF concentration in the culture medium was detected by enzyme-linked immunosorbent assay. Surface staining of CD74 in the MKN-45 cell line under the stimulation of lipopolysaccharide (LPS) was measured by flow cytometry. MIF, CD74 and TLR4 co-localization in the MKN-45 cell line was performed by the immunoprecipitation. CD74, MIF and TLR4 were found to be expressed in gastric cancer and increased significantly in the advanced stage, and were also associated with lymph node metastasis. Correlation analysis revealed that CD74 was positively correlated with MIF (r = 0.2367, P < 0.01) and both proteins were also associated with TLR4 (r = 0.4414, r = 0.5001, respectively, P < 0.01). LPS can significantly promote MKN-45 cell proliferation (3.027 ± 0.388 vs 4.201 ± 0.092, P < 0.05), induce MIF production (54.333 ± 2.906 pg/mL vs 29.667 ± 3.180 pg/mL, P < 0.01) and cell surface expression of CD74 (75.6% ± 4.046% vs 9.4% ± 0.964%, P < 0.01) at LPS concentration of 1 μg/mL compared to medium control. Knockdown of CD74 or using anti-CD74 and MIF antagonist ISO-1 significantly reduced LPS-induced MKN-45 cell proliferation (4.201 ± 0.092 vs 3.337 ± 0.087, 4.534 ± 0.222 vs 3.368 ± 0.290, 4.058 ± 0.292 vs 2.934 ± 0.197, respectively, P < 0.01). MIF, CD74 and TLR4 could co-localize in the MKN-45 cell line. Upregulation of MIF, CD74 and TLR4 are associated with increasing clinical stage and provide an opportunity as novel gastric cancer chemoprevention and/or treatment strategy.

Macrophage migration inhibitory factor activates cyclooxygenase 2–prostaglandin E 2 in cultured spinal microglia

In our previous study, peripheral inflammatory stimulation evoked production of macrophage migration inhibitory factor (MIF) in the spinal cord and found spinal microglia are the major source of MIF in this context. Given the contribution of the activated-microglia to the inflammatory neuropathy plus the role for upregulated COX 2 expression and PGE 2 production in the severity of clinical manifestations of these neuroinflammatory conditions, we herein tested the hypothesis that in vitro MIF stimulation to spinal microglia could result in an activation of COX 2–PGE 2 system by MIF–CD74 interaction. We found MIF played roles in evoking COX 2 mRNA and protein expression in a dose-dependent manner correspondingly in changes in PGE 2 level in the cultured rat microglia, but these changes could be inhibited by genetic deletion of CD74. Finally, MIF-induced COX 2–PGE 2 activation could be blocked by selective inhibitors of p44/p42 and p38 MAPKs. These data highlight MIF/CD74 interaction induces upregulation of COX 2 expression and PGE 2 secretion in primary rodent microglia, and further this effect is associated with downstream activation of p38 and p44/p42 signaling cascades, and favors the role of MIF as a novel pathway for microglia-associated neuroinflammation.

Kinsenoside Isolated from Anoectochilus Formosanus Suppresses LPS-Stimulated Inflammatory Reactions in Macrophages and Endotoxin Shock in Mice

In the present study, we reported that kinsenoside, a major component of Anoectochilus formosanus, inhibited inflammatory reactions in mouse peritoneal lavage macrophages and protects mice from endotoxin shock. In LPS-stimulated mouse peritoneal lavage macrophages, kinsenoside inhibited the inflammatory mediators, such as nitric oxide, TNF-[alpha], IL-1[beta], monocyte chemoattractant protein 1, and macrophage migration inhibitory factor production. Furthermore, kinsenoside decreased the formation of a nuclear factor [kappa]B-DNA complex and nuclear p65 and p50 protein levels. Kinsenoside inhibited nuclear factor [kappa]B translocation through both I[kappa]B[alpha]-dependent and -independent pathway. In contrast, it stimulated anti-inflammatory cytokine IL-10 generation and enhanced the mRNA expression of IL-10 and suppressor of cytokine signaling 3 in the same cells induced by LPS. In an animal model, both pretreatment and posttreatment of kinsenoside increased the survival rate of ICR mice challenged by LPS (80 mg/kg, i.p.). Pretreatment with kinsenoside decreased serum levels of TNF-[alpha], IL-1[beta], IL-10, monocyte chemoattractant protein 1, and migration inhibitory factor at 1 h after sublethal dose of LPS (40 mg/kg, i.p.) in mice. In contrast, kinsenoside enhanced serum IL-10 level at 24 h after LPS injection in mice. In conclusion, kinsenoside inhibited the production of inflammatory mediators and enhanced anti-inflammatory cytokine generation. Therefore, kinsenoside can alleviate acute inflammatory hazards.