Absence Of Macrophage Migration Inhibitory Factor Research Articles

Macrophage migration inhibitory factor (MIF) and pregnancy may impact the balance of intestinal cytokines and the development of intestinal pathology caused by Toxoplasma gondii infection

Toxoplasma gondii (T. gondii) is an intracellular parasite responsible for causing toxoplasmosis. When infection occurs during pregnancy, it can produce severe congenital infection with ocular and neurologic damage to the infant. From the oral infection parasite reaches the intestine, causing inflammatory response, damage in tissue architecture and systemic dissemination. Macrophage migration inhibition factor (MIF) is a cytokine secreted from both immune and non-immune cells, including gut epithelial cells. MIF is described to promote inflammatory responses, to be associated in colitis pathogenesis and also to play role in maintaining the intestinal barrier. The aim of the present study was to evaluate the influence of the pregnancy and MIF deficiency on T. gondii infection in the intestinal microenvironment and to address how these factors can impact on the intestinal architecture and local cytokine profile. For this purpose, small intestine of pregnant and non-pregnant C57BL/6 MIF deficient mice (MIF−/−) and Wild-type (WT) orally infected with 5 cysts of ME-49 strain of T. gondii were collected on day 8th of infection. Intestines were processed for morphological and morphometric analyses, parasite quantification and for cytokines mensuration. Our results showed that the absence of MIF and pregnancy caused an increase in T. gondii infection index. T. gondii immunolocalization demonstrated that segments preferentially infected with T. gondii were duodenum and ileum. The infection caused a reduction in the size of the intestinal villi, whereas, infection associated with pregnancy caused an increase in villi size due to edema caused by the infection. Also, the goblet cell number was increased in the ileum of MIF−/− mice, when compared to the corresponding WT group. Analyses of cytokine production in the small intestine showed that MIF was up regulated in the gut of pregnant WT mice due to infection. Also, infection provoked an intense Th1 response that was more exacerbated in pregnant MIF−/− mice. We also detected that the Th2/Treg response was more pronounced in MIF−/− mice. Altogether, our results demonstrated that pregnancy and MIF deficiency interferes in the balance of the intestinal cytokines and favors a Th1-immflamatory profile, which in turn, impact in the development of pathology caused by T. gondii infection in the intestinal microenvironment.

Macrophage Migration Inhibitory Factor (MIF) Is Essential for Type 2 Effector Cell Immunity to an Intestinal Helminth Parasite.

Immunity to intestinal helminths is known to require both innate and adaptive components of the immune system activated along the Type 2 IL-4R/STAT6-dependent pathway. We have found that macrophage migration inhibitory factor (MIF) is essential for the development of effective immunity to the intestinal helminth Heligmosomoides polygyrus, even following vaccination which induces sterile immunity in wild-type mice. A chemical inhibitor of MIF, 4-IPP, was similarly found to compromise anti-parasite immunity. Cellular analyses found that the adaptive arm of the immune response, including IgG1 antibody responses and Th2-derived cytokines, was intact and that Foxp3+ T regulatory cell responses were unaltered in the absence of MIF. However, MIF was found to be an essential cytokine for innate cells, with ablated eosinophilia and ILC2 responses, and delayed recruitment and activation of macrophages to the M2 phenotype (expressing Arginase 1, Chil3, and RELM-α) upon infection of MIF-deficient mice; a macrophage deficit was also seen in wild-type BALB/c mice exposed to 4-IPP. Gene expression analysis of intestinal and lymph node tissues from MIF-deficient and -sufficient infected mice indicated significantly reduced levels of Arl2bp, encoding a factor involved in nuclear localization of STAT3. We further found that STAT3-deficient macrophages expressed less Arginase-1, and that mice lacking STAT3 in the myeloid compartment (LysMCrexSTAT3fl/fl) were unable to reject a secondary infection with H. polygyrus. We thus conclude that in the context of a Type 2 infection, MIF plays a critical role in polarizing macrophages into the protective alternatively-activated phenotype, and that STAT3 signaling may make a previously unrecognized contribution to immunity to helminths.

Mif deficiency promotes adiposity in fructose-fed mice.

The macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine involved in inflammation, regulation of energy metabolism and glucocorticoid action. Chronic low-grade inflammation may be caused by fructose intake, contributing to visceral adipose tissue (VAT) dysfunction. Since MIF is a known antagonist of glucocorticoid signaling, and deregulated glucocorticoid signaling can contribute to lipid metabolism disturbances, we hypothesized that altered MIF signaling might underlie fructose-induced adiposity through glucocorticoid action. We analyzed physiological and biochemical parameters, adipose tissue histology, insulin sensitivity and lipid metabolism in WT and MIF-/- C57Bl/6J mice consuming 20% fructose solution for 9 weeks. Glucocorticoid prereceptor metabolism and glucocorticoid receptor (GR) protein level were examined in VAT, together with the expression of glucocorticoid-target genes involved in lipid metabolism. The expression of adipogenic and lipogenic transcriptional regulators peroxisome proliferator-activated receptor gamma (PPARG) and sterol regulatory element-binding protein 1c (SREBP1c) was also assessed. Results showed disturbed insulin sensitivity in all MIF-/- mice, regardless of the diet. Mice on fructose diet had increased energy intake, but increased visceral adiposity and enlarged adipocytes were observed only in fructose-fed MIF-/- mice. Increased VAT corticosterone level and 11 beta-hydroxysteroid dehydrogenase type 1, hexose-6-phosphate dehydrogenase and GR protein levels were observed in the same animals, together with induced expression of examined lipogenic genes and accumulation of PPARG and SREBP1c. In conclusion, the results showed that dietary fructose was associated with increased visceral adiposity through activation of GR-regulated lipogenic genes, but only in the absence of MIF, which set the state of hyperinsulinemia and insulin resistance.

Macrophage migration inhibitory factor protects from nonmelanoma epidermal tumors by regulating the number of antigen-presenting cells in skin.

The response of the skin to harmful environmental agents is shaped decisively by the status of the immune system. Keratinocytes constitutively express and secrete the chemokine-like mediator, macrophage migration inhibitory factor (MIF), more strongly than dermal fibroblasts, thereby creating a MIF gradient in skin. By using global and epidermis-restricted Mif-knockout (Mif-/- and K14-Cre+/tg; Miffl/fl) mice, we found that MIF both recruits and maintains antigen-presenting cells in the dermis/epidermis. The reduced presence of antigen-presenting cells in the absence of MIF was associated with accelerated and increased formation of nonmelanoma skin tumors during chemical carcinogenesis. Our results demonstrate that MIF is essential for maintaining innate immunity in skin. Loss of keratinocyte-derived MIF leads to a loss of control of epithelial skin tumor formation in chemical skin carcinogenesis, which highlights an unexpected tumor-suppressive activity of MIF in murine skin.-Brocks, T., Fedorchenko, O., Schliermann, N., Stein, A., Moll, U. M., Seegobin, S., Dewor, M., Hallek, M., Marquardt, Y., Fietkau, K., Heise, R., Huth, S., Pfister, H., Bernhagen, J., Bucala, R., Baron, J. M., Fingerle-Rowson, G. Macrophage migration inhibitory factor protects from nonmelanoma epidermal tumors by regulating the number of antigen-presenting cells in skin.

Macrophage migration inhibitory factor inhibits the antiinflammatory effects of glucocorticoids via glucocorticoid-induced leucine zipper.

Glucocorticoids remain a mainstay in the treatment of rheumatoid arthritis (RA). Dose-dependent adverse effects highlight the need for therapies that regulate glucocorticoid sensitivity to enable dosage reduction. Macrophage migration inhibitory factor (MIF) is a proinflammatory protein that has been implicated in the pathogenesis of RA; it impairs glucocorticoid sensitivity via MAPK phosphatase 1 (MKP-1) inhibition. The intracellular protein glucocorticoid-induced leucine zipper (GILZ) mimics the effects of glucocorticoids in models of RA, but whether it represents a target for the modulation of glucocorticoid sensitivity remains unknown. We undertook this study to investigate whether GILZ is involved in the regulation of glucocorticoid sensitivity by MIF. GILZ expression was studied in the presence and absence of MIF, and the role of GILZ in the MIF-dependent regulation of the glucocorticoid sensitivity mediator MKP-1 was studied at the level of expression and function. GILZ expression was significantly inhibited by endogenous MIF, both basally and during responses to glucocorticoid treatment. The effects of MIF on GILZ were dependent on the expression and Akt-induced nuclear translocation of the transcription factor FoxO3A. GILZ was shown to regulate the expression of MKP-1 and consequent MAPK phosphorylation and cytokine release. MIF exerts its effects on MKP-1 expression and MAPK activity through inhibitory effects on GILZ. These findings suggest a previously unsuspected interaction between MIF and GILZ and identify GILZ as a potential target for the therapeutic regulation of glucocorticoid sensitivity.

MIF deficiency does not alter glucose homeostasis or adipose tissue inflammatory cell infiltrates during diet-induced obesity.

ObjectiveCirculating Macrophage Migration Inhibitory Factor (MIF) levels have been shown to positively correlate with body mass index (BMI) in humans. Our objective in this study was to determine the effects of MIF deficiency in a model of high fat diet-induced obesity.Design and MethodsMIF wild type (MIF WT) and MIF deficient (MIF−/−) C57Bl/6J mice were fed a high fat diet (HFD) for up to 15 weeks. Weight and metabolic responses were measured over the course of the disease. Immune cell infiltrates in visceral and subcutaneous adipose tissue were examined by flow cytometry.ResultsThere was no difference in weight gain or adipose tissue mass in MIF−/− mice compared to MIF WT mice. Both groups fed HFD developed glucose intolerance at the same rate and had similar elevations in fasted blood insulin. MDSC abundance was evaluated and showed no MIF-dependent differences. Macrophages were elevated in the visceral adipose tissue of obese mice, but there was no difference between the two groups.ConclusionsWhile high fat diet feeding induced obesity with the expected perturbations in glucose homeostasis and adipose tissue inflammation, the presence or absence of MIF had no effect on any parameter examined.

Macrophage migration inhibitory factor produced by the tumour stroma but not by tumour cells regulates angiogenesis in the B16-F10 melanoma model

Background:Macrophage migration inhibitory factor (MIF) has been proposed as a link between inflammation and tumorigenesis. Despite its potentially broad influence in tumour biology and prevalent expression, the value of MIF as a therapeutic target in cancer remains unclear. We sought to validate MIF in tumour models by achieving a complete inhibition of its expression in tumour cells and in the tumour stroma.Methods:We used MIF shRNA-transduced B16-F10 melanoma cells implanted in wild-type and MIF−/− C57Bl6 mice to investigate the effect of loss of MIF on tumour growth. Cytokine detection and immunohistochemistry (IHC) were used to evaluate tumours ex vivo.Results:Macrophage migration inhibitory factor shRNA inhibited expression of MIF protein by B16-F10 melanoma cells in vitro and in vivo. In vitro, the loss of MIF in this cell line resulted in a decreased response to hypoxia as indicated by reduced expression of VEGF. In vivo the growth of B16-F10 tumours was inhibited by an average of 47% in the MIF−/− mice compared with wild-type but was unaffected by loss of MIF expression by the tumour cells. Immunohistochemistry analysis revealed that microvessel density was decreased in tumours implanted in the MIF−/− mice. Profiling of serum cytokines showed a decrease in pro-angiogenic cytokines in MIF−/− mice.Conclusion:We report that the absence of MIF in the host resulted in slower tumour growth, which was associated with reduced vascularity. While the major contribution of MIF appeared to be in the regulation of angiogenesis, tumour cell-derived MIF played a negligible role in this process.

Abstract 159: Macrophage Migration Inhibitory Factor (MIF) enhances proliferation and expression of tumorigenic cytokines in a bladder cancer cell line

Abstract Background: MIF is a proinflammatory cytokine with regulatory properties over tumor suppressor proteins involved in bladder cancer. We have previously reported that the absence of MIF in transgenic mice or the use of a small molecule oral inhibitor of MIF (CPSI-1306, Cytokine PharmaSciences, Inc, King of Prussia, PA) leads to decreased angiogenesis and invasive potential in the BBN mouse model of bladder cancer. The objective of this study was to evaluate potential pathways of MIF activity in a cell culture model. Methods: High grade, metastatic HTB-5 bladder cancer cells were treated with rhMIF 1-100 ng/ml (CPSI). Measurements included cell counts, proliferation by 3H-thymidine incorporation (TdR), phosphorylated and total ERK by Western analysis and mRNA expression of TNF-α, IL-6, VEGF and CD74 (putative receptor for MIF) by real time PCR. CPSI-1306 (0.5-500 nM) was tested for its ability to decrease ERK phosphorylation. Results: MIF 100 ng/ml increased cell counts 2-fold (p<0.01) on culture days 2-4. MIF increased TdR, normalized to cell numbers, 30% (p<0.01). MIF (1-100 ng/ml) stimulated ERK phosphorylation in a dose dependent manner, which was abrogated by the addition of CPSI-1306 at doses above 5 nM. A specific inhibitor of ERK (PD98059, 50 μm) blocked the MIF stimulated increase in cell number. MIF treatment for 24 hrs increased mRNA expression of the agiogenic factor VEGF and the cytokine TNF-α 2-fold (p<0.01). IL-6 mRNA was unchanged. mRNA levels for the putative MIF receptor, CD74, increased 1.3 fold with treatment (p=0.01). Conclusion: MIF increased ERK phosphorylation, which was blocked by CPSI-1306, a small molecule inhibitor of MIF. MIF also increased cell proliferation via an ERK pathway. These data, along with the increased cytokine expression, provide a potential explanation for the decreased angiogenesis and invasion seen in our in vivo murine models using global knockout of MIF or CPSI-1306. This study is the first to begin to elucidate mechanistic pathways involved in MIFs role in bladder cancer and possible pathways involved in CPSI-1306 inhibition. Funding Source: ACS MRSG-08-270-01-CCE, NIH RO1DK48361 Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 159. doi:1538-7445.AM2012-159

Migratory response of mesenchymal stem cells to macrophage migration inhibitory factor and its antagonist as a function of colony-forming efficiency

Human mesenchymal stem cells (MSCs) are capable of repairing pulmonary disorders, but their efficacy is limited by poor engraftment. A strategy is proposed to augment MSC migration to lung tissue by antagonizing macrophage migration inhibitory factor (MIF), a pro-inflammatory cytokine. Recombinant MIF (85 ng/ml) inhibited in vitro chemokinesis of multipotent MSCs by nearly 50 and 20% for donor preparations with colony-forming efficiencies of 22 +/- 4% and 66 +/- 3%, respectively (P < 0.05). The small-molecule MIF antagonist, (S,R)-3-(4-hydroxyphenyl)-4,5-dihydro-5-isoxazole acetic acid methyl ester (ISO-1, 85 microg/ml), restored MSC migration for all donors to levels found in the absence of MIF. At this concentration, ISO-1 increased migration to conditioned medium from bronchial epithelial cell cultures by >or=3-fold for all donor MSC preparations (P < 0.05). Transcript levels for the MIF receptor, CD74, in MSCs were independent of colony-forming efficiency. These data suggest that MIF and its antagonists may be relevant to the control of MSC homing and efficacy of stem cell therapies in a variety of clinical scenarios.

Macrophage migration inhibitory factor is critical to interleukin‐5‐driven eosinophilopoiesis and tissue eosinophilia triggered by Schistosoma mansoni infection

Macrophage migration inhibitory factor (MIF) participates in the pathogenesis of inflammatory diseases, including asthma, in which it enhances airway hypersensitivity and tissue eosinophilia. Herein, we investigated the role of MIF in eosinophilopoiesis and tissue eosinophilia using Schistosoma mansoni infection. MIF-deficient (Mif(-/-)) mice had similar numbers of adult worms, eggs, and granulomas compared to wild-type mice, but the size of granulomas was strikingly reduced due to smaller numbers of eosinophils. MIF did not affect the acquired response to infection, as Mif(-/-) mice produced normal amounts of Th2 cytokines and IgE. Nevertheless, recombinant MIF (rMIF) behaved as a chemoattractant for eosinophils, what could partially explain the reduced eosinophilia in infected Mif(-/-) mice. Moreover, the percentage of eosinophils was reduced in bone marrows of Mif(-/-) mice chronically infected with S. mansoni compared to wild type. Mif(-/-) had impaired eosinophilopoiesis in response to interleukin (IL)-5 and addition of rMIF to bone marrow cultures from IL-5 transgenic mice enhanced the generation of eosinophils. In the absence of MIF, eosinophil precursors were unable to survive the IL-5-supplemented cell culture, and were ingested by macrophages. Treatment with pancaspase inhibitor z-VAD or rMIF promoted the survival of eosinophil progenitors. Together, these results indicate that MIF participates in IL-5-driven maturation of eosinophils and in tissue eosinophilia associated with S. mansoni infection.

The Role of Macrophage Migration Inhibitory Factor in Mouse Islet Transplantation

Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine produced by many tissues including pancreatic beta-cells. This study investigates the impact of MIF on islet transplantation using MIF knock-out (MIFko) mice. Early islet function, assessed with a syngeneic marginal islet mass transplant model, was enhanced when using MIFko islets (P<0.05 compared with wild-type [WT] controls). This result was supported by increased in vitro resistance of MIFko islets to apoptosis (terminal deoxynucleotide tranferase-mediated dUTP nick-end labeling assay), and by improved glucose metabolism (lower blood glucose levels, reduced glucose areas under curve and higher insulin release during intraperitoneal glucose challenges, and in vitro in the absence of MIF, P<0.01). The beneficial impact of MIFko islets was insufficient to delay allogeneic islet rejection. However, the rejection of WT islet allografts was marginally delayed in MIFko recipients by 6 days when compared with WT recipient (P<0.05). This effect is supported by the lower activity of MIF-deficient macrophages, assessed in vitro and in vivo by cotransplantation of islet/macrophages. Leukocyte infiltration of the graft and donor-specific lymphocyte activity (mixed lymphocyte reaction, interferon gamma ELISPOT) were similar in both groups. These data indicate that targeting MIF has the potential to improve early function after syngeneic islet transplantation, but has only a marginal impact on allogeneic rejection.

Macrophage Migration Inhibitory Factor (MIF) Promotes the Development of Murine Chronic Lymphocytic Leukemia (CLL)

Introduction: Tumor formation results from a complex interplay between genetic/epigenetic alterations, cell cycle dysregulation and promotion by the tumor environment. Stimulation by extracellular survival factors is important for chronic lymphocytic leukemia (CLL), since the leukemic cells undergo spontaneous apoptosis when removed from their normal milieu. Since preliminary experiments demonstrated that macrophage migration inhibitory factor (MIF), a chemokine-like proinflammatory mediator and an intracellular regulator of growth and apoptosis, is overexpressed in human CLL, we investigated whether MIF participates in the pathogenesis of murine CLL.Methods: We studied the role of MIF in CLL by crossing the Eμ-TCL1-transgenic mouse model with MIF knockout (MIF−/−) mice. B-cell-specific overexpression of T cell leukemia-1 (TCL1) leads to accumulation and proliferation of IgM+/CD5+ mature B-cells via activation of AKT. This results in a CLL-like disease with peripheral lymphocytic leukemia, lymphadenopathy, splenomegaly, BM infiltration and premature death after 8–15 months. TCL1+/wtMIF−/− and TCL1+/wtMIF+/+ mice were compared with respect to leukemia development, tumor burden, cytogenetics and survival.Results: The MIF receptors CD74/CD44 and CXCR2 are expressed on murine B-cells. TCL1+/wtMIF+/+ mice exhibited increased numbers of IgM+/CD5+ B-cells already in the preleukemic phase at month 3 and developed overt leukemia (WBC > 20G/l) 3 months earlier than their MIF−/− counterparts (p = 0.02). Leukemia load at 12 months of age as measured by hepatosplenomegaly was increased in TCL1+/wtMIF+/+ animals and lymphatic organs were densely infiltrated by small, mature lymphocytes. The accelerated disease progression in the presence of MIF translated into a median survival which was 60 days shorter than in the absence of MIF (TCL1+/wtMIF+/+ 400 days, TCL1+/wtMIF−/− 460 days, p = 0.04). SKY analysis in leukemic splenocytes yielded various complex genetic aberrations with trisomies (e.g. +15), tetraploidy, translocations and deletions. Overexpression of tp53 due to the presence of an inactivating mutation in the p53 gene was found more frequently in TCL1+/wtMIF+/+ than in TCL1+/wtMIF−/− animals. Although the rates of DNA-damage-induced apoptosis in pre-leukemic and leukemic mice ex vivo were not significantly different between the genotypes, this defect in the p53-dependent apoptosis pathway corresponded with a reduced rate of spontaneous apoptosis in spleens of leukemic TCL1+/wtMIF+/+ animals.Conclusions: Our experience with the Eμ-TCL-1-transgenic mice shows that this model is suitable for the identification of novel regulators of CLL-like disease. We provide genetic proof that MIF acts to promote the early preleukemic and the leukemic phase of TCL1-induced CLL and thereby identify MIF as a novel regulator of CLL pathogenesis. Ongoing efforts are focussing on further characterizing the differences in pathology, the activation of the AKT pathway and cell cycle control between TCL1+/wtMIF−/− and TCL1+/wtMIF+/+ mice.

Treatment of Inflammatory Autoimmune Disease Using a Small Molecule Cytokine Inhibitor

Multiple sclerosis (MS) is an autoimmune disease of the central nervous system (CNS). Macrophage migration inhibitory factor (MIF), a ubiquitous inflammatory cytokine, is elevated in the CSF of MS patients during relapse. We have shown that MIF−/− (KO) mice show markedly suppressed clinical signs of experimental autoimmune encephalomyelitis (EAE) relative to wild type (WT) controls. The purpose of this study was to identify the timing in which MIF is important for progression of disease. Using adoptive transfer, we showed that activated lymphocytes from KO mice could transfer EAE to WT recipients, but that WT cells were unable to transfer EAE to KO recipients, suggesting that the CNS environment was protective in the absence of MIF. We used a small molecule inhibitor of MIF (INH) at two different times to selectively test the induction of EAE in the absence of MIF. Mice receiving INH at Day 0 were not protected, but mice treated after disease onset had less severe EAE. Despite the reduced severity of disease, lymphocytes from INH‐treated mice secreted substantial IFN‐γ and IL‐17. INH‐treated mice had a larger population of CD4+25+foxP3 lymphocytes. These findings suggest that MIF is important for disease progression but is not critical for T cell activation; an inhibitor could be a novel therapeutic approach for treating ongoing MS and other autoimmune diseases. (Supported by NIH grant AI 064320 and NMSS grant RG3272)

Macrophage Migration Inhibitory Factor Induces B Cell Survival by Activation of a CD74-CD44 Receptor Complex

Macrophage migration inhibitory factor (MIF) is an upstream activator of innate immunity that regulates subsequent adaptive responses. It was previously shown that in macrophages, MIF binds to a complex of CD74 and CD44, resulting in initiation of a signaling pathway. In the current study, we investigated the role of MIF in B cell survival. We show that in B lymphocytes, MIF initiates a signaling cascade that involves Syk and Akt, leading to NF-kappaB activation, proliferation, and survival in a CD74- and CD44-dependent manner. Thus, MIF regulates the adaptive immune response by maintaining the mature B cell population.

Estrogen modulates cutaneous wound healing by downregulating macrophage migration inhibitory factor.

Characteristic of both chronic wounds and acute wounds that fail to heal are excessive leukocytosis and reduced matrix deposition. Estrogen is a major regulator of wound repair that can reverse age-related impaired wound healing in human and animal models, characterized by a dampened inflammatory response and increased matrix deposited at the wound site. Macrophage migration inhibitory factor (MIF) is a candidate proinflammatory cytokine involved in the hormonal regulation of inflammation. We demonstrate that MIF is upregulated in a distinct spatial and temporal pattern during wound healing and its expression is markedly elevated in wounds of estrogen-deficient mice as compared with intact animals. Wound-healing studies in mice rendered null for the MIF gene have demonstrated that in the absence of MIF, the excessive inflammation and delayed-healing phenotype associated with reduced estrogen is reversed. Moreover, in vitro assays have shown a striking estrogen-mediated decrease in MIF production by activated murine macrophages, a process involving the estrogen receptor. We suggest that estrogen inhibits the local inflammatory response by downregulating MIF, suggesting a specific target for future therapeutic intervention in impaired wound-healing states.