Receptor For Migration Inhibitory Factor Research Articles

Macrophage migration inhibitory factor receptor CD74 expression is associated with expansion and differentiation of effector T cells in COVID-19 patients.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused millions of COVID-19 cases and deaths worldwide. Severity of pulmonary pathologies and poor prognosis were reported to be associated with the activation non-virus-specific bystander T cells. In addition, high concentrations of the macrophage migration inhibitory factor (MIF) were found in serum of COVID-19 patients. We hypothesized that these two pathogenic factors might be related and analyzed the expression of receptors for MIF on T cells in COVID-19. T cells from PBMCs of hospitalized patients with mild and severe COVID-19 were characterized. A significantly higher proportion of CD4+ and CD8+ T cells from COVID-19 patients expressed CD74 on the cell surface compared to healthy controls. To induce intracellular signaling upon MIF binding, CD74 forms complexes with CD44, CXCR2, or CXCR4. The vast majority of CD74+ T cells expressed CD44, whereas expression of CXCR2 and CXCR4 was low in controls but increased upon SARS-CoV-2 infection. Hence, T cells in COVID-19 patients express receptors that render them responsive to MIF. A detailed analysis of CD74+ T cell populations revealed that most of them had a central memory phenotype early in infection, while cells with an effector and effector memory phenotype arose later during infection. Furthermore, CD74+ T cells produced more cytotoxic molecules and proliferation markers. Our data provide new insights into the MIF receptor and co-receptor repertoire of bystander T cells in COVID-19 and uncovers a novel and potentially druggable aspect of the immunological footprint of SARS-CoV-2.

MIF in the cerebrospinal fluid is decreased during relapsing-remitting while increased in secondary progressive multiple sclerosis

BackgroundMacrophage migration inhibitory factor (MIF) is involved in the function of both the innate and adaptive immune systems and in neuroprotection and has recently been implicated in multiple sclerosis (MS). ObjectivesDetermination of MIF levels in the cerebrospinal fluid (CSF) of patients with distinct subtypes of MS and the cellular localization of MIF in human brain tissue. MethodsThe levels of MIF were investigated in CSF from patients with clinically isolated syndrome (CIS) (n = 26), relapsing-remitting MS (RRMS) (n = 22), secondary progressive MS (SPMS) (n = 19), and healthy controls (HCs) (n = 24), using ELISA. The effect of disease-modifying therapies in the RRMS and SPMS cohorts were examined. Cellular distribution of MIF in the human brain was studied using immunochemistry and the newly available OligoInternode database. ResultsMIF was significantly decreased in treatment-naïve CIS and RRMS patients compared to HCs but was elevated in SPMS. Interestingly, MIF levels were sex-dependent and significantly lower in women with CIS and RRMS. MIF expression in the human brain was localized to neurons, astrocytes, pericytes, and oligo5 oligodendrocytes but not in microglia. ConclusionThe finding that MIF was decreased in newly diagnosed CIS and RRMS patients but was high in patients with SPMS may suggest that MIF levels in CSF are regulated by local MIF receptor expression that affects the overall MIF signaling in the brain and may represent a protective mechanism that eventually fails.

Soluble cluster of differentiation 74 regulates lung inflammation through the nuclear factor-κB signaling pathway

The soluble form of the migration inhibitory factor receptor cluster of differentiation 74 (sCD74) has previously been shown to be elevated during the development of acute lung injury (ALI) in mice. However, the biological role of increased sCD74 in ALI remans poorly understood. Synthesized recombinant sCD74 protein was administered to mice intratracheally. Pro-inflammatory genes in lung tissue and the inflammatory factors in bronchoalveolar lavage fluid (BALF) were analyzed using RT-PCR and ELISA, respectively. Additionally, RAW264.7, A549, and human umbilical vein endothelial cells (HUVEC) were treated with sCD74, and the resulting pro-inflammatory factor protein and gene expression levels were analyzed in the supernatants and cell lysates. Meanwhile, the level of nuclear factor (NF)-κB components in cell lysates after stimulating macrophages with sCD74 was also assessed. After administration of 0.5 mg/kg body weight sCD74 to mice, the expression of Tnfa, Mip2, and Il6 increased in lung tissues after 2 h by 2.1-, 3.4-, and 2.8-fold, respectively. Moreover, the BALF concentrations of TNF-α and MIP-2 reached maximal levels of 560 and 107 pg/mL at 8 h compared to those in the saline group, respectively. Similarly, TNFA, MIP2, and IL6 expression increased by 4.0-, 11.8-, and 2.6-fold, respectively, 2 h after stimulating macrophages with 1000 ng/mL sCD74. The levels of phospho-IκB and phospho-p65 were also significantly increased in the cytoplasm and nucleus of macrophages following sCD74 stimulation. Taken together, these results suggest that sCD74 is a critical cellular factor involved in the lung acute inflammatory response through nuclear factor-κB signaling.

Glioblastoma Myeloid-Derived Suppressor Cell Subsets Express Differential Macrophage Migration Inhibitory Factor Receptor Profiles That Can Be Targeted to Reduce Immune Suppression.

The application of tumor immunotherapy to glioblastoma (GBM) is limited by an unprecedented degree of immune suppression due to factors that include high numbers of immune suppressive myeloid cells, the blood brain barrier, and T cell sequestration to the bone marrow. We previously identified an increase in immune suppressive myeloid-derived suppressor cells (MDSCs) in GBM patients, which correlated with poor prognosis and was dependent on macrophage migration inhibitory factor (MIF). Here we examine the MIF signaling axis in detail in murine MDSC models, GBM-educated MDSCs and human GBM. We found that the monocytic subset of MDSCs (M-MDSCs) expressed high levels of the MIF cognate receptor CD74 and was localized in the tumor microenvironment. In contrast, granulocytic MDSCs (G-MDSCs) expressed high levels of the MIF non-cognate receptor CXCR2 and showed minimal accumulation in the tumor microenvironment. Furthermore, targeting M-MDSCs with Ibudilast, a brain penetrant MIF-CD74 interaction inhibitor, reduced MDSC function and enhanced CD8 T cell activity in the tumor microenvironment. These findings demonstrate the MDSC subsets differentially express MIF receptors and may be leveraged for specific MDSC targeting.

Macrophage migration inhibitory factor regulates integrin-β1 and cyclin D1 expression via ERK pathway in podocytes

AimsMacrophage migration inhibitory factor (MIF) is found to increase in proliferative glomerulonephritis. MIF binds to the MIF receptor (CD74) that activates MAP kinase (ERK and p38). Integrins and cyclinD1 regulate cell proliferation, differentiation and adhesion. This study evaluates whether MIF can regulate integrin-β1/cyclin D1 expression and cell adhesion of podocytes. Main methodsExpression of integrin-β1 mRNA/protein and cyclin D1 mRNA under stimulation of MIF was evaluated by real-time PCR and Western blotting. MIF receptor (CD74) and MAP kinase under MIF treatment were examined to determine which pathway regulated integrin-β1 and cyclin D1 expression. Cell adhesion was evaluated under MIF treatment and/or anti-integrin-β1 antibody by cell adhesion assay. Key findingsProtein levels of integrin-β1 were up-regulated under MIF treatment in a dosage-dependent manner. CD74 protein levels were not changed after MIF treatment. Integrin-β1 and cyclin D1 mRNA levels were up-regulated after MIF 100 ng/ml treatment. ERK inhibitor U0126 reduced MIF-induced the increase in integrin-β1 mRNA and protein expression following MIF stimulation. However, p38 inhibitor SB 203580 did not inhibit MIF-induced increase in integrin-β1 mRNA and protein expression following MIF stimulation. MIF-induced increase in cyclin D1 mRNA level also was inhibited only by U0126 following MIF stimulation. Podocyte adhesion was increased after MIF treatment, but, anti-integrin-β1 antibody decreased MIF-enhanced podocyte adhesion. SignificanceMIF increases integrin-β1 and cyclin D1 expression through the ERK pathway in podocytes, and the up-regulated expression of integrin-β1 increases podocyte adhesion. These results provide further understanding for the role of MIF in developing proliferative glomerulonephritis.

Macrophage Migration Inhibitory Factor Receptor, CD74, is Overexpressed in Human and Baboon ( Papio Anubis) Endometriotic Lesions and Modulates Endometriotic Epithelial Cell Survival and Interleukin 8 Expression.

CD74 is the primary receptor for macrophage migration inhibitory factor (MIF). Although expression of MIF has been described in endometriotic lesions, the cellular localization and function of the MIF receptor, CD74, are poorly understood. To further explore the role of CD74 in the pathophysiology of endometriosis, we utilized specimens from women with diagnostically confirmed endometriosis, women with no signs or symptoms of endometriosis (controls), and 8 baboons with experimentally induced endometriosis. Compared to eutopic endometrium from women with endometriosis, CD74 transcript expression was significantly increased in endometriotic lesion tissue. Similarly, cellular expression of CD74 was significantly greater in ectopic lesion tissue compared to paired eutopic endometrium, which both expressed greater CD74 expression compared to eutopic endometrium from control patients. Localization of CD74 was predominant to epithelial cells of ectopic and matched eutopic endometrium and was not influenced by the stage of the menstrual cycle. Eutopic endometrium from control patients did not express detectable levels of CD74 protein by immunohistochemistry. This pattern of expression and CD74 protein localization could be recapitulated in endometriotic lesion tissue from baboons with experimentally induced disease. Transfection of the endometriotic epithelial cell lines, 12Z with CD74 short hairpin RNA (shRNA), resulted in a significant decrease in CD74 protein expression, which was associated with a significant reduction in cellular proliferation as well as the expression of the prosurvival cytokine interleukin 8. Together, these data support the hypothesis that CD74 is elevated in endometriotic lesion tissue and may contribute to the pathogenesis of endometriosis by promoting cell survival.

Abstract 3560: Targeting the MIF receptor CD74 in bladder cancer tumorigenesis

Abstract Macrophage migration inhibitory factor (MIF) is a pleiotropic cytokine that can stimulate cellular proliferation in multiple tumors. Our previous data indicates genetic deletion of MIF results in T1 stage arrest in the N-Butyl-N-(4-hydroxybutyl) nitrosamine (BBN) induced bladder cancer model. Similarly, pharmacological inhibition of MIF results in lower stage tumors and reduced bladder weights when given therapeutically. CD74 is a human leukocyte antigen class II trafficking and processing protein that also functions as an extracellular receptor for MIF. Additionally, CD74 complexes with CD44 to act as the cognate MIF receptor complex, and upregulation of CD74 has been noted in high grade bladder cancer. We hypothesized CD74 would be upregulated in malignant bladder cancer cell lines, and knockout of CD74 would reduce bladder cancer tumorigenesis in the murine BBN model. The objectives of this study were to determine the effect of CD74 knockout on development and progression of urothelial cell carcinoma; as well as evaluate expression of the putative MIF signaling axis in BCa cell lines. BCa cell lines HTB-9, HTB-5, RT-4, T24 and HT-1376 and benign urothelial UROtsa cells were analyzed for MIF, CD74, and CD44 expression via quantitative PCR or western blot to assess expression of the MIF signaling axis. CD74 and CD44 expression by q-PCR were dramatically elevated in BCa cell lines vs. benign cell lines. Protein levels of CD74 and CD44 were elevated in multiple cell lines as well. MIF expression levels did not change between cell lines. To ascertain differences in BCa tumorigeneis, WT C57Bl/6J mice or CD74-/- mice were given 0.05% BBN in the drinking water for 22 weeks. Bladders were weighed and evaluated for pathologic staging by histology. After exposure to BBN, bladders from CD74-/- mice had a 33% reduction in weight vs. WT bladders (p=.062). Pathological analysis revealed lower stage tumors in bladders of CD74-/- mice (33% incidence; 6.7% pTa, 0% pTis, 6.7% pT1, 20% pT2, 6.7% pT3) as compared to bladders of WT mice (81.3% incidence; 0% pTa, 6.3% pTis, 25% pT1, 12.5% pT2, 37.5% pT3), which was a statistically significant reduction in tumor stage. Lower tumor stage correlated significantly with the CD74-/- genotype (p=0.02, Fisher's Exact Test). These data suggest CD74 likely mediates a portion of the effects of MIF in BCa, raising the possibility of both CD74 dependent, and independent MIF signaling pathways. Citation Format: Justin Penticuff, Sambantham Shanmugam, Joshua Warrick, Brian Chase, Erika Abbott, Benjamin L. Woolbright, John A. Taylor. Targeting the MIF receptor CD74 in bladder cancer tumorigenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3560.

Invariant Chain Complexes and Clusters as Platforms for MIF Signaling.

Invariant chain (Ii/CD74) has been identified as a surface receptor for migration inhibitory factor (MIF). Most cells that express Ii also synthesize major histocompatibility complex class II (MHC II) molecules, which depend on Ii as a chaperone and a targeting factor. The assembly of nonameric complexes consisting of one Ii trimer and three MHC II molecules (each of which is a heterodimer) has been regarded as a prerequisite for efficient delivery to the cell surface. Due to rapid endocytosis, however, only low levels of Ii-MHC II complexes are displayed on the cell surface of professional antigen presenting cells and very little free Ii trimers. The association of Ii and MHC II has been reported to block the interaction with MIF, thus questioning the role of surface Ii as a receptor for MIF on MHC II-expressing cells. Recent work offers a potential solution to this conundrum: Many Ii-complexes at the cell surface appear to be under-saturated with MHC II, leaving unoccupied Ii subunits as potential binding sites for MIF. Some of this work also sheds light on novel aspects of signal transduction by Ii-bound MIF in B-lymphocytes: membrane raft association of Ii-MHC II complexes enables MIF to target Ii-MHC II to antigen-clustered B-cell-receptors (BCR) and to foster BCR-driven signaling and intracellular trafficking.

Macrophage Migration Inhibitory Factor Receptor CD74 Mediates Alphavirus‐Induced Arthritis and Myositis in Murine Models of Alphavirus Infection

ObjectiveArthrogenic alphaviruses such as Ross River virus (RRV) and chikungunya virus (CHIKV) circulate worldwide. This virus class causes debilitating illnesses that are characterized by arthritis, arthralgia, and myalgia. In previous studies, we identified macrophage migration inhibitory factor (MIF) as a critical inflammatory factor in the pathogenesis of alphaviral diseases. The present study was undertaken to characterize the role of CD74, a cell surface receptor of MIF, in both RRV‐ and CHIKV‐induced alphavirus arthritides.MethodsMouse models of RRV and CHIKV infection were used to investigate the immunopathogenesis of arthritic alphavirus infection. The role of CD74 was assessed using histologic analysis, real‐time polymerase chain reaction, flow cytometry, and plaque assay.ResultsIn comparison to wild‐type mice, CD74−/− mice developed only mild clinical features and had low levels of tissue damage. Leukocyte infiltration, characterized predominantly by inflammatory monocytes and natural killer cells, was substantially reduced in the infected tissue of CD74−/− mice, but production of proinflammatory cytokines and chemokines was not decreased. CD74 deficiency was associated with increased monocyte apoptosis, but had no effect on monocyte migratory capacity. Consistent with these findings, alphaviral infection resulted in a dose‐dependent up‐regulation of CD74 expression in human peripheral blood mononuclear cells, and serum MIF levels were significantly elevated in patients with RRV or CHIKV infection.ConclusionCD74 appears to regulate immune responses to alphaviral infection through its effects on cellular recruitment and survival. These findings suggest that both MIF and CD74 play a critical role in mediating alphaviral disease, and blocking these factors with novel therapeutic agents could substantially ameliorate the pathologic manifestations.

Lack of macrophage migration inhibitory factor in mice does not affect hallmarks of the inflammatory/immune response during the first week after stroke

BackgroundMacrophage migration inhibitory factor (MIF) has been proposed to play a detrimental role in stroke. We recently showed that MIF promotes neuronal death and aggravates neurological deficits during the first week after experimental stroke, in mice. Since MIF regulates tissue inflammation, we studied the putative role of MIF in post-stroke inflammation.MethodsWe subjected C57BL/6 mice, Mif-/- (MIF-KO) or Mif+/+ (WT), to a transient occlusion of the right middle cerebral artery (tMCAo) or sham-surgery. We studied MIF expression, GFAP expression and the number of CD74-positive cells in the ischemic brain hemisphere 7 days after tMCAo using primarily immunohistochemistry. We determined IFN-γ, IL-2, IL-4, IL-5, IL-10, IL-12, KC/CXCL-1 and TNF-α protein levels in the brain (48 h after surgery) and serum (48 h and 7 days after surgery) by a multiplex immunoassay.ResultsWe observed that MIF accumulates in neurons and astrocytes of the peri-infarct region, as well as in microglia/macrophages of the infarct core up to 7 days after stroke. Among the inflammatory mediators analyzed, we found a significant increase in cerebral IL-12 and KC levels after tMCAo, in comparison to sham-surgery. Importantly, the deletion of Mif did not significantly affect the levels of the cytokines evaluated, in the brain or serum. Moreover, the spleen weight 48 h and 7 days subsequent to tMCAo was similar in WT and MIF-KO mice. Finally, the extent of GFAP immunoreactivity and the number of MIF receptor (CD74)-positive cells within the ischemic brain hemisphere did not differ significantly between WT and MIF-KO mice subjected to tMCAo.ConclusionsWe conclude that MIF does not affect major components of the inflammatory/immune response during the first week after experimental stroke. Based on present and previous evidence, we propose that the deleterious MIF-mediated effects in stroke depend primarily on an intraneuronal and/or interneuronal action.

Further evidence for increased macrophage migration inhibitory factor expression in prostate cancer

BackgroundMacrophage migration inhibitory factor (MIF) is a cytokine associated with prostate cancer, based on histologic evidence and circulating (serum) levels.Recent studies from another laboratory failed to document these results. This study's aims were to extend and confirm our previous data, as well as to define possible mechanisms for the discrepant results. Additional aims were to examine MIF expression, as well as the location of MIF's receptor, CD74, in human prostatic adenocarcinoma compared to matched benign prostate.MethodsMIF amounts were determined in random serum samples remaining following routine PSA screening by ELISA. Native, denaturing and reducing polyacrylamide gels and Western blot analyses determined the MIF form in serum. Prostate tissue arrays were processed for MIF in situ hybridization and immunohistochemistry for MIF and CD74. MIF released into culture medium from normal epithelial, LNCaP and PC-3 cells was detected by Western blot analysis.ResultsMedian serum MIF amounts were significantly elevated in prostate cancer patients (5.87 ± 3.91 ng/ml; ± interquartile range; n = 115) compared with patients with no documented diagnosis of prostate cancer (2.19 ± 2.65 ng/ml; n = 158). ELISA diluent reagents that included bovine serum albumin (BSA) significantly reduced MIF serum detection (p < 0.01). MIF mRNA was localized to prostatic epithelium in all samples, but cancer showed statistically greater MIF expression. MIF and its receptor (CD74) were localized to prostatic epithelium. Increased secreted MIF was detected in culture medium from prostate cancer cell lines (LNCaP and PC-3).ConclusionIncreased serum MIF was associated with prostate cancer. Diluent reagents that included BSA resulted in MIF serum immunoassay interference. In addition, significant amounts of complexed MIF (180 kDa under denaturing conditions by Western blot) found in the serum do not bind to the MIF capture antibody. Increased MIF mRNA expression was observed in prostatic adenocarcinoma compared to benign tissue from matched samples, supporting our earlier finding of increased MIF gene expression in prostate cancer.

Macrophage glycolipid receptors for human migration inhibitory factor (MIF): Differentiated HL-60 cells exhibit MIF responsiveness and express surface glycolipids which both bind MIF and convert nonresponsive cells to responsiveness

The human promyelocytic leukemia line HL-60 when treated with a phorbol diester (TPA) differentiates into cells (HL60-TPA) that respond to human migration inhibitory factor (MIF). Unresponsive HL-60 cells became responsive to MIF when preincubated with a glycolipid-enriched preparation extracted from HL60-TPA cells, human monocytes, human macrophage-like (U937) cell line, or with the purified glycolipid receptor for MIF from guinea pig peritoneal macrophages. Human blood monocytes exhibited an increased response to MIF when preincubated with glycolipids from HL60-TPA and U937 cells but not from HL-60 cells, Finally, glycolipids from HL60-TPA cells but not from HL-60 cells were able to reversibly bind MIF when covalently coupled to agarose. These studies suggest that TPA induces the differentiation of HL-60 cells into MIF-responsive cells through the expression of a glycolipid receptor for MIF.

Agglutination of murine and guinea pig peritoneal cells by alpha-L-fucose-binding lectin: Evonymus europaea.

Among lectins from Lotus tetragonolobus, Ulex europaeus I and Evonymus europaea, agglutinating cells with blood group H determinants containing L-fucose alpha 1 leads to 2-linked to subterminal D-galactose, only the last lectin agglutinates thioglycolate- and paraffin oil-stimulated murine and guinea pig peritoneal exudate cells (PEC). The agglutination is inhibited by specific inhibitors of Evonymus lectin only: lacto-N-fucopentaose I and lactose. These results suggest the presence of a determinant on the surface of PEC, containing L-fucose alpha 1-linked at the nonreducing end which is different from blood group H determinants. Nonstimulated murine peritoneal cells (PC) are not agglutinated by the lectin but become agglutinable after neuraminidase treatment. Unstimulated guinea pig PC from different animals are agglutinated to a different extent by the same lectin concentration and show increased agglutinability after neuraminidase digestion. These results show that receptor for Evonymus lectin also exists on the nonstimulated PC but access to it is hindered by sialic acid. Trypsin- and pronase-digested PEC show increased agglutinability with Evonymus lectin. These results suggest that the lectin receptor is a glycolipid. Since alpha-linked L-fucose has been suggested as a part of the macrophage receptor for migration inhibitory factor in the guinea pig (Remold, J. Exp. Med. 1973. 138: 1065), the effect of Evonymus europaea lectin on the migration of PEC was studied. It was found that lectin inhibits the migration of PEC in the capillary tube assay up to 80%.

Glycolipid affinity purification of migration inhibitory factor.

Migration inhibitory factor (MIF) is an immune mediator secreted by primed lymphocytes on in vitro challenge by antigen or mitogen, which elicits various physiological responses from its target cell, the macrophage1,2. Previous studies suggest that membrane glycoconjugates have an important role in regulating the interaction between MIF and the macrophage. A macrophage glycolipid functions as a putative receptor for MIF as evidenced by its ability to enhance the macrophage response to MIF3,4. This enhancing ability of glycolipids requires the presence of sialic acid5 and fucose6; these carbohydrate residues are also required on the macrophage surface for the induction of an intact response by MIF5,7. Furthermore, it has been reported4 that incubating a preparation of mixed gangliosides from bovine brain with lymphocyte supernatants abolished their MIF and macrophage activating factor (MAF) activities. Taken together, these results are evidence that a glycolipid serves as a membrane receptor for MIF. We predict that glycolipids which enhance the macrophage response to MIF, also bind MIF with subsequent elution of biological activity, and present here evidence in support of this notion. We also demonstrate that glycolipid covalently bound to agarose provides a novel means of affinity purification of MIF.

Isolation of a guinea pig macrophage glycolipid with the properties of the putative migration inhibitory factor receptor.

We have previously established that preincubation of guinea pig macrophages with crude preparations of macrophage glycolipids enhances their response to a mediator of cellular immunity, migration inhibitory factor. This biologically active moiety found in the extracted material was attributed to a single glycolipid component. This component was purified to homogeneity by phase partition, ion exchange, adsorption, and thin layer chromatography. It migrates in the ganglioside region on thin layer plates, contains sialic acid and sphingosine. There is no apparent identity with any of the common gangliosides possessing the gangliotetraose structure. Homogeneity was demonstrated by rechromatography in four additional solvent systems. These results are discussed in the context of their implications for a deeper understanding of the putative receptor function of this glycolipid.

Role of sialic acid in the macrophage glycolipid receptor for MIF.

Acidic glycolipids from guinea pig macrophages enhance the response of macrophages to migration inhibitory factor (MIF), suggesting a role of glycolipid receptors for this lymphocyte mediator. Neuraminidase treatment of these glycolipids results in the loss of their biologic activity. This activity remains intact after incubation of the glycolipids with beta-galactosidase. In order to investigate whether sialic acid is essential for the macrophage's response to MIF, macrophages were incubated with neuraminidase. Neuraminidase treatment of peritoneal exudate cells results in the abrogation of macrophage responsiveness to MIF. Other exoglycosidases such as beta-galactosidase and beta-glucosidase had no effect upon the macrophage response. The effect of neuraminidase was found to be reversible within 18 hr. These experiments suggest that macrophage glycolipids containing sialic acid are components of the macrophage receptor for MIF.

Further characterization of the putative glycolipid receptor for mif: Role of fucose associated with an acidic glycolipid

Water soluble glycolipids were extracted from guinea pig macrophages. These glycolipids, when incubated with macrophages, augment the cells' response to migration inhibitory factor. The glycolipids were fractionated by diethylaminoethyl-Sephadex ion exchange chromatography into neutral and acidic fractions. Only the acidic glycolipid fraction was able to enhance the responsiveness of macrophages to migration inhibitory factor. Additional studies indicate that the enhancing activity of these glycolipid preparations can be abrogated by the removal of terminal fucose residues with α-L-fucosidase. The possibility that fucose functions as an essential component of a macrophage glycolipid receptor for migration inhibitory factor is discussed.

Evidence for a surface receptor for human migration inhibitory factor on a B-lymphoid cell line

Reception by PGLC-33H target cells for the migration inhibitory factor (MIF) produced by this established line has been investigated by pulse time and temperature dependence, MIF absorption, and abrogation by trypsinization. PGLC-33H supernatants containing MIF were concentrated 5× with Carbowax and dialyzed against serum free RPMI-1640 before use. Prior to standard capillary migration assay a minimum 30 min pulse of MIF at 37 °C is required for significant migration inhibition (MI > 20%). No significant MI is observed when cells are pulsed at 4 °C for up to 2 hr. Preincubation with PGLC-33H for 1 hr at 37 °C reduces activity of supernatants from 38 to 13% MI; at 4 °C to 27% MI. Trypsinization of target cells for 30 min at 25 °C abrogates response to MIF (43 to −14% MI). Trypsinized cells did not reduce activity of supernatants. MIF activity is abolished (32 to 3% MI) in samples preincubated with supernatants of the trypsinized cells inactivated with serum. These data suggest that cells from the human B-lymphoid cell line PGLC-33H have a surface receptor for human MIF.

Requirement for alpha-L-fucose on the macrophage membrane receptor for MIF.

alpha-L-fucose abolishes the activity of guinea pig migration inhibitory factor (MIF) on the macrophages. Other sugars such as alpha-D-glucose, beta-D-galactose, alpha-L-rhamnose, methyl-alpha-D-mannoside, and N-acetyl-beta-D-glucosamine had no effect. Theabolition of MIF activity by alpha-L-fucose was reversible. When macrophages were incubated with alpha-L-fucosidase, a glycosidase which splits terminal alpha-L-fucose from oligosaccharides, the macrophages no longer responded to MIF. On the other hand, MIF incubated with alpha-L-fucosidase was still active. These experiments strongly suggest that alpha-L-fucose comprises an essential part of a macrophage membrane receptor for MIF.

Mechanism of action of migration inhibitory factor (MIF). I. Evidence for a receptor for MIF present on the peritoneal macrophage but not on the alveolar macrophage.

The initial interaction between migration inhibitory factor (MIF) and the guinea pig alveolar and peritoneal macrophage was studied. MIF-containing supernatants were generated from sensitized lymph node lymphocytes obtained from guinea pigs immunized with bovine gamma globulin in complete Freund's adjuvant. MIF-containing supernatants were markedly inhibitory for the migration of the peritoneal macrophage but had no effect on the alveolar macrophage. A linear relationship was observed between per cent inhibition of migration and serial twofold dilution of supernatant. Reexpressed in arbitrary MIF units, this relationship reflects a dose-response relationship with saturation characteristics. Pulse exposure of peritoneal macrophages to MIF resulted in adsorption of MIF onto both viable and nonviable cells with corresponding depletion of supernatant MIF. The alveolar macrophage did not adsorb MIF. Pulse adsorption of MIF onto the peritoneal macrophage is dependent on time, temperature, and cell number. Pretreatment of the cells with proteolytic enzyme prevents the adsorption of MIF while leaving migration unaffected. These observations support the existence of a specific cell surface receptor for MIF. The existence of such a receptor provides selectivity of immune modulation of macrophage populations by lymphocytes in delayed hypersensitivity reactions.