Increased Levels of Macrophage Migration Inhibitory Factor in Patients with Familial Mediterranean Fever

Objective: To determine the level of macrophage migration inhibitory factor (MIF), its relationship with Mediterranean fever (MEFV) gene mutations and oxidative stress in familial Mediterranean fever (FMF).Methods: Fifty one unrelated attack free FMF patients (24 M and 27 F, 32.8±8.7 years) and 30 healthy controls (16 M and 14 F, 32.7±7 years) were included in the study. Serum MIF, total oxidant status (TOS) and total anti-oxidant status (TAS) were studied.Results: Age, sex distribution, anthropometrical indices, smoking status, serum lipids and TAS concentrations were similar between the patients and controls. However; erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), MIF, and TOS were significantly higher in the patients'' group compared with healthy subjects. MIF, TOS and TAS levels were not different between patients with or without M694V mutations.Conclusion: We found increased concentrations of MIF in patients with FMF. Increased MIF levels were significantly correlated with oxidative stress and in regression analysis MIF concentrations were independent from the inflammatory activity as assessed by ESR and CRP. M694V mutations seem no effect on MIF and oxidative stress.     

Increased Expression of Macrophage Migration Inhibitory Factor and DJ-1 contribute to Cell Invasion and Metastasis of Nasopharyngeal Carcinoma

Background and aim: Both macrophage migration inhibitory factor (MIF) and DJ-1 protein have been shown to relate with cell invasion and metastasis in tumors. However, the role of DJ-1 in invasion and metastasis of nasopharyngeal carcinoma (NPC) and its relation to MIF expression in NPC are not fully understood. The aim of present study is to determine whether or not MIF and DJ-1 are correlated with tumor invasion and influence a worse outcome in NPC, as well as its related mechanism.Methods: 125 cases of NPC and 45 normal tissues of nasopharynx were collected. The expression of MIF and DJ-1 in tissue microarray was evaluated by immunohistochemical staining. Correlation between immunostainings and clinicopathological parameters, as well as the follow-up data of patients, was analyzed statistically. The association of MIF and DJ-1 with cell invasion and migration in NPC cell line were evaluated by small interfering RNA (siRNA) transfection, invasion assay and Western blotting.Results: MIF and DJ-1 staining was diffused and strong in tumor cells, whereas they were generally weaker and less common in normal lining epithelia of nasopharynx. High MIF expression in tumor cells (71.2%, 89/125 cases) were significantly associated with advanced clinical stage, lymph node metastasis, and worse prognosis of NPC patients. High expression of DJ-1 (75.2%, 94/125 cases) were closely correlated to lymph node metastasis and MIF high-expression. Only MIF high expression (P = 0.010) and lymph node metastasis (P = 0.004) emerged as strong independent prognostic factors for overall survival of NPC patients. In vitro, down-regulated expression of DJ-1 in NPC cell lines by siRNA was observed to reduce cell migration and invasion potential, however, exogenous MIF promoted cells invasion.Conclusions: The data provided evidence that increased expression of MIF and DJ-1 induced cell invasion and metastasis of NPC, supporting the idea that MIF and DJ-1 may play important roles as regulators in the progression of NPC.     

Epitope Mapping of Monoclonal Antibody 1B9 Against Plasmodium falciparum-Derived Macrophage Migration Inhibitory Factor

A homologue of mammalian macrophage migration inhibitory factor (MIF) has been identified in Plasmodium falciparum (PfMIF). This parasite-derived cytokine and its antiserum were detected in the circulation of patients with malaria. Using a monoclonal antibody, designated mAb1B9, against PfMIF, we performed biopanning using two phage display peptide libraries to screen for the main sequence of the epitope recognized by the antibody. We then expressed a series of truncated peptides in order to identify the precise sequence of the epitope. The epitope recognized by mAb1B9 is 22 amino acids long and has the following sequence: ³⁶LGYIMSNYDYQKNLRFGGSNEA⁵⁷. Western analysis showed that the residues ³⁶LG³⁷ and ⁵²G differentiated PfMIF from the rodent malaria parasite-derived MIFs, and the residues ⁴³Y and ⁴⁸NL⁴⁹ differentiated PfMIF from P. vivax- and P. knowlesi-derived MIFs. The precise identification of this epitope, the first identified for PfMIF, will increase the specificity of the sandwich ELISA assays used to evaluate patients with malaria. These results indicate that mAb1B9 is useful for investigating the function of PfMIF in immune responses to malaria. Both the epitope and the monoclonal antibody against it will be valuable tools in epidemiological studies concerning this P. falciparum-derived cytokine.     

Chemical Characterization of Macrophage Cytotoxicity Factor,Macrophage Migration Inhibitory Factor,T-Helper Cell-Replacing Factor and Colony-Stimulating Factor from Culture Supernatants of Concanavalin A-Stimulated Murine Spleen Cells

Supernatants from Concanavalin A-stimulated murine spleen cells were subjected to hydrophobic interaction chromatography on phenyl-Sepharose. Macrophage cytotoxicity factor (MCF), macrophage migration inhibitory factor (MIF), T-helper cell-replacing factor (TRF) and colony-stimulating factor (CSF) were bound at high ionic strength and were released stepwise at low ionic strength. CSF thus could be separated from MCF, MIF and TRF and the bulk of other proteins. Chromatography of pools containing MCF, MIF and TRF on Sephadex did not lead to a separation of the three activities which were all found in a molecular weight range of 25.000-55.000. Isoelectric focusing of these pools in pH range from 4 to 9 gave two peaks for MCF at pH 8.2 and 7.2, whereas MIF activity focused from pH 4.5 to 5.5. TRF activity was found in a single sharp peak at pH 5.3. The results demonstrate that the four biological activities can be distinguished on a chemical basis and are accessible for purification and chemical characterization.     

Mannose Binding Lectin and Macrophage Migration Inhibitory Factor Gene Polymorphisms in Turkish Children with Cardiomyopathy: No Association with MBL2 Codon 54 A/B Genotype, but an Association between MIF -173 CC Genotype

Myocardial inflammation is one of the commonest mechanisms in cardiomyopathy (CMP). Mannose binding lectin (MBL) is a key molecule in innate immunity, while macrophage migration inhibitory factor (MIF) is a constitutive element of the host defenses. We investigated the possible association between polymorphisms of MBL2 and MIF genes and CMP in Turkish children. Twenty-children with CMP and 30 healthy controls were analyzed for codon 54 A/B polymorphism in MBL, and -173 G/C polymorphism in MIF genes by using PCR-RFLP methods. No significant difference was found between genotypes and alleles of MBL2 gene codon 54 A/B polymorphism in patients and controls (p>0.05). However, serum uric acid levels was found higher in dilated CMP patients with AA genotype. Frequency of MIF -173 CC genotype was significantly higher in patients (p<0.05), and sodium levels were higher in patients with MIF -173 CC genotype. This study is the first to investigate the MBL and MIF gene polymorphisms in Turkish children with CMP. We conclude that CC genotype of MIF (-173) polymorphism may be a risk factor for CMP patients. However, further studies with larger samples are needed to address the exact role of this polymorphism in CMP.     

VEGF和MMP-2的表达对肝细胞癌侵袭转移的影响

采用免疫组化 ABC法对 5 0例肝细胞癌手术切除标本的血管内皮生长因子 (VEGF)和基质金属蛋白酶- 2 (MMP- 2 )表达进行检测 ,以探讨 VEGF及 MMP- 2在肝细胞癌中的表达与其复发、转移的关系。结果发现 VEGF和 MMP- 2在肝细胞癌组织中的阳性表达率分别为 86 % (43/5 0 )和 6 0 % (30 /5 0 ) ,在正常肝组织中分别为 5 3.3%(16 /30 )和 30 % (9/30 ) ,癌组织与正常组织间存在着显著差异 (P<0 .0 5 ) ;VEGF和 MMP- 2的阳性表达率与肝细胞癌的转移和包膜形成相关 ,在肝细胞癌生长、侵袭和转移过程中起着重要的作用 ,对预测肝细胞癌复发、转移具有重要的意义     

心绞痛患者血浆C反应蛋白及巨噬细胞移动抑制因子的变化

目的通过检测稳定型及不稳定型心绞痛患者血浆高敏c反应蛋白（hsCRP）及巨噬细胞移动抑制因子（MIF），了解hsCRP及MIF在其中的变化。方法61例稳定型心绞痛患者、69例不稳定型心绞痛患者和55例健康志愿者。以ELISA法检测分析其血浆中高敏C反应蛋白及巨噬细胞移动抑制因子的变化。结果不稳定型心绞痛组患者血浆hsCRP水平（12．93±3．67）mg／L及MIF水平（23．60±6．31）μg／L高于稳定型心绞痛患者[hsCRP（4．37±1．28）mg／L，MIF（9．89±1．04）μg／L]及对照组[hsCRP（3．51±1．47）mg／L，MIF（9．58±1．25）μg／L]，差异有统计学意义（P均〈0．05）；而稳定型心绞痛患者及对照组hsCRP及MIF水平比较差异则无统计学意义（P〉0．05）。结论不稳定型心绞痛患者其hsCRP及MIF水平较健康对照组及稳定型心绞痛患者均有明显升高，对hsCRP及MIF检测可能有助于鉴别高危冠心病患者。     

Reprint of: The non-mammalian MIF superfamily

Macrophage migration inhibitory factor (MIF) was first described as a cytokine 50 years ago, and emerged in mammals as a pleiotropic protein with pro-inflammatory, chemotactic, and growth-promoting activities. In addition, MIF has gained substantial attention as a pivotal upstream mediator of innate and adaptive immune responses and with pathologic roles in several diseases. Of less importance in mammals is an intrinsic but non-physiologic enzymatic activity that points to MIF's evolution from an ancient defense molecule. Therefore, it is not surprising that mif-like genes also have been found across a range of different organisms including bacteria, plants, protozoa, helminths, molluscs, arthropods, fish, amphibians and birds. While Genebank analysis identifying mif-like genes across species is extensive, contained herein is an overview of the non-mammalian MIF-like proteins that have been most well studied experimentally. For many of these organisms, MIF contributes to an innate defense system or plays a role in development. For parasitic organisms however, MIF appears to function as a virulence factor aiding in the establishment or persistence of infection by modulating the host immune response. Consequently, a combined targeting of both parasitic and host MIF could lead to more effective treatment strategies for parasitic diseases of socioeconomic importance.     

糖尿病肾病合并冠心病患者C反应蛋白及巨噬细胞移动抑制因子的改变

目的通过检测糖尿病肾病合并冠心病患者血浆高敏C反应蛋白（hsCRP）及巨噬细胞移动抑制因子（MIF），了解hsCRP及MIF在糖尿病肾病合并冠心病患者中的变化，协助临床决策。方法收入64例糖尿病肾病合并冠心病患者及57例糖尿病肾病但无明确冠心病依据的患者，同时收入54例健康志愿者，以ELISA法检测分析其血浆中高敏C反应蛋白及巨噬细胞移动抑制因子的变化。结果糖尿病肾病合并冠心病组患者血浆hsCRP水平（9．93±2．58）mg／L及MIF水平（23．61±6．64）μg／L高于无明确冠心病依据的患者[hsCRP（7．37±1．32）mg／L，MIF（15．56±3．70）μg／L]，差异有统计学意义（P均〈0．05），两组患者血浆hsCRP及MIF水平又均显著高于对照组[hsCRP（3．51±2．00）mg／L，MIF（9．57±1．25）μg／L]。结论检测糖尿病肾病患者血浆hsCRP及MIF，可能有助于鉴别糖尿病肾病患者有无合并冠心病的危险。     

Localization of Matrix Metalloproteinase (MMP)-9 in Lung Tissue of a Murine Model of Allergic Asthma

MMP-9 (gelatinase B) is recognized in chronic obstructive pulmonary disease (COPD) and now asthma as playing a central role in matrix degradation in injury, as well as contributing to the remodeling process. The increasing focus on MMP-9 in human and animal research supports the need for a reliable immunostain in lung tissue. However, MMP-9 immunostaining in murine systems is hampered by several factors. First, many of the anti-human antibodies do not readily cross-react with murine MMP-9 despite the high degree of conservation between human and murine MMP-9. Secondly, the availability of detailed protocols is limited. Lung MMP-9 immunostaining is further complicated by technical issues such as edge effect, availability of positive and negative controls, antigen retrieval, staining specificity, and the need to achieve a delicate balance of primary and secondary antibody concentrations, and colorimetric reagents which will allow visualization of specific cell expression in highly delicate lung tissue, while also demonstrating adequate uptake in (extra-pulmonary) tissue controls. We describe a detailed method for immunostaining MMP-9 in mouse lung paraffin-embedded tissue utilizing human ovary as a control since MMP-9 is known to be over-expressed in human ovarian carcinomas.     

Blockade of macrophage migration inhibitory factor (MIF) prevents the antigen-induced response in a murine model of allergic airway inflammation

Objective and Design: The role of macrophage migration inhibitory factor (MIF), a proinflammatory cytokine, was tested using a mouse asthma model. Materials: One hundred and four male BALB/c mice were used in this study. Treatment: Mice were actively sensitized with an intraperitoneal injection of ovalbumin (OVA) and challenged with repeated nebulization of 1 w/v% OVA. Polyclonal anti-MIF antibody was intraperitoneally injected at 10 mg/kg during the antigen challenge period. Methods: Bronchoalveolar lavage (BAL) was performed 8 h after the last challenge. Airway hyperresponsiveness to inhaled methacholine was measured 24 h after the last challenge. Results: Antigen challenge to immunized mice induced increase in inflammatory cells and concentration of Th2 cytokines in BAL fluid (BALF), and caused the development of airway hyperresponsiveness. Anti-MIF antibody significantly decreased the numbers of inflammatory cells including macrophages, eosinophils, lymphocytes and neutrophils in BALF from OVA-challenged mice. Prednisolone decreased the numbers of eosinophils, lymphocytes and neutrophils but not macrophages. Anti-MIF antibody reduced airway hyperresponsiveness. Anti-MIF antibody affected neither the cytokine levels in BALF nor the IgE levels in serum. Conclusion: MIF was involved in the antigen-induced inflammatory cell accumulation in the lung and airway hyperresponsiveness without affecting immune responses. Received 17 November 2005; returned for revision 14 June 2006; accepted by M. Katori 25 July 2006     

Glucocorticoids and macrophage migration inhibitory factor (MIF) are neuroendocrine modulators of inflammation and neuropathic pain after spinal cord injury

Traumatic spinal cord injury (SCI) activates the hypothalamic–pituitary–adrenal (HPA) axis, a potent neuroendocrine regulator of stress and inflammation. SCI also elicits a profound and sustained intraspinal and systemic inflammatory response. Together, stress hormones and inflammatory mediators will affect the growth and survival of neural and non-neural cells and ultimately neurologic recovery after SCI. Glucocorticoids (GCs) are endogenous anti-inflammatory steroids that are synthesized in response to stress or injury, in part to regulate inflammation. Exogenous synthetic GCs are often used for similar purposes in various diseases; however, their safety and efficacy in pre-clinical and clinical SCI is controversial. The relatively recent discovery that macrophage migration inhibitory factor (MIF) is produced throughout the body and can override the anti-inflammatory effects of GCs may provide unique insight to the importance of endogenous and exogenous GCs after SCI. Here, we review both GCs and MIF and discuss the potential relevance of their interactions after SCI, especially their role in regulating maladaptive mechanisms of plasticity and repair that may contribute to the onset and maintenance of neuropathic pain.     

巨噬细胞移动抑制因子、高敏C-反应蛋白与颈动脉粥样硬化和急性脑梗死的关系

目的探讨巨噬细胞移动抑制因子、高敏C-反应蛋白与颈动脉粥样硬化和急性脑梗死的关系。方法103例首次发病的脑梗死患者和40例健康体检者，应用酶联免疫双抗体夹心法测定血清MIF浓度，免疫散射比浊法测定血清hs—CRP浓度。应用颈动脉超声检测颈动脉内膜状况，并对急性脑梗死患者进行神经功能缺损评分。结果急性脑梗死患者血清MIF和hs—CRP浓度显著高于正常对照组（P〈0．01）。不稳定斑块组（混合斑块组、软斑组）血清MIF和hs-CRP浓度显著高于稳定斑块组（硬化斑块组）和内膜粗糙组（P〈0．01）。血清MIF和hs—CRP浓度分别与急性脑梗死患者神经功能缺损评分呈正相关。结论急性脑梗死患者血清MIF和hs-CRP水平可以反映颈动脉斑块的性质和稳定性，是临床了解脑梗死严重程度的重要指标。     

Production of Five Human Lymphokines (GranulocyteMacrophage Colony Stimulating Factor,Interferon-γ, Interleukin 2, Macrophage Cytotoxicity Factor and Macrophage Migration Inhibitory Factor) from Con A Stimulated Lymphocyte Cultures in Bioreactors

Cultivation parameters for the production of five lymphokines, granulocyte-macrophage colony stimulating factor (GM-CSF), interferon-y (IFN-γ), interleukin 2 (IL-2), macrophage cytotoxicity factor (MCF), and macrophage migration inhibitory factor (MIF) from human spleen cells or peripheral blood lymphocytes were optimized. Cultivation was done in bioreactors containing up to 200 ml of medium, usually serum-free. The reactors were equipped with surface aeration facilities, stirrers and oxygen electrodes.Whereas stirring speed alone did not influence the yields of lymphokines, good aeration was especially beneficial for high IL-2 yields. However, all lymphokines were also produced under anaerobic conditions. The concentration of the mitogen concanavalin A was mainly critical for optimal IL-2 release. Optimal cell concentrations varied from 5 x 10₆/ml (for GM-GSF and MCF) to 10 x 10₆/ml (for IL-2 and IFN-γ). It was possible to increase the yields of individual lymphokines 3 to 10-fold per batch of lymphocytes by a reinduction procedure which involved a change of medium and mitogen every 24 hrs. Reinduction was possible up to 4 times, especially when serum was present in the culture media.     

Partial MHC class II constructs inhibit MIF/CD74 binding and downstream effects

MIF and its receptor, CD74, are pivotal regulators of the immune system. Here, we demonstrate for the first time that partial MHC class II constructs comprised of linked β1α1 domains with covalently attached antigenic peptides (also referred to as recombinant T‐cell receptor ligands — RTLs) can inhibit MIF activity by not only blocking the binding of rhMIF to immunopurified CD74, but also downregulating CD74 cell‐surface expression. This bifunctional inhibition of MIF/CD74 interactions blocked downstream MIF effects, including enhanced secretion of proinflammatory cytokines, anti‐apoptotic activity, and inhibition of random migration that all contribute to the reversal of clinical and histological signs of EAE. Moreover, we demonstrate that enhanced CD74 cell‐surface expression on monocytes in mice with EAE and subjects with multiple sclerosis can be downregulated by humanized RTLs, resulting in reduced MIF binding to the cells. Thus, binding of partial MHC complexes to CD74 blocks both the accessibility and availability of CD74 for MIF binding and downstream inflammatory activity.     

Macrophage migration inhibitory factor (MIF) inhibitor,Z-590 suppresses cartilage destruction in adjuvant-induced arthritis via inhibition of macrophage inflammatory activation

Background: Macrophage migration inhibitory factor (MIF) is a pleiotropic pro-inflammatory mediator that is involved in the progression of rheumatoid arthritis (RA). Previously, we demonstrated a small molecule compound 3-[(biphenyl-4-ylcarbonyl) carbamothioyl] amino benzoic acid (Z-590) could inhibit MIF activity with docking-based virtual screening and experimental evaluation.

Methods: The LPS activated RAW264.7 macrophage cells were used to determine the anti-inflammatory effects of Z-590 in vitro. A rat adjuvant-induced arthritis (AIA) model was used to determine the anti-arthritic effects of Z-590 in vivo.

Results: MIF inhibitor Z-590 significantly inhibited the production of NO, TNF-α and IL-6 in LPS-activated RAW 264.7 macrophage cells and markedly inhibited LPS-induced expression of TNF-α, IL-6, inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). Z-590 also significantly reduced paw edema, serum level of TNF-α, IL-6 and spleen index in the adjuvant-induced arthritis (AIA) rat model. Furthermore, Z-590 markedly ameliorated joint inflammation and articular cartilage damage in AIA rat model.

Conclusion: MIF inhibitor Z-590 possesses potent anti-arthritic activity through suppression of macrophage activation, and could be a potential therapeutic treatment for RA.     

巨噬细胞移动抑制因子在大肠黏膜癌变患者中表达增强

目的 探讨巨噬细胞移动抑制因子(MIF)表达与大肠癌临床病理因素的关系.方法 采用免疫组化法测定大肠组织中MIF的表达.ELISA测定血清中MIF水平.结果 MIF在正常大肠黏膜、大肠腺瘤、大肠癌中阳性表达强度和阳性表达率依次增高,3组之间存在显著性差异(P<0.001);组织中MIF表达强度与血清中MIF水平呈正相关性;MIF的表达与大肠癌的分化程度、淋巴结转移、肝转移有关.结论 MIF可能在大肠癌的发生和进展中起着重要的作用.