Macrophage Migration Inhibitory Factor Protein Expression Research Articles

A small-molecule allele-selective transcriptional inhibitor of the MIF immune susceptibility locus

Functional variants of the gene for the cytokine macrophage migration inhibitory factor (MIF) are defined by a 4-nucleotide promoter microsatellite (-794 CATT5-8, rs5844572) and confer risk for autoimmune, infectious, and oncologic diseases. We describe herein the discovery of a prototypic, small molecule inhibitor of MIF transcription with selectivity for high microsatellite repeat number and correspondingly high gene expression. Utilizing a high-throughput luminescent proximity screen, we identify 1-carbomethoxy-5-formyl-4,6,8-trihydroxyphenazine (CMFT) to inhibit the functional interaction between the transcription factor ICBP90 (a.k.a. UHRF1) and the MIF -794 CATT5-8 promoter microsatellite. CMFT inhibits MIF mRNA expression in a -794 CATT5-8 length-dependent manner with an IC50 of 470 nM, and preferentially reduces ICBP90-dependent MIF mRNA and protein expression in high-genotypic versus low-genotypic MIF – expressing macrophages. RNA expression analysis also showed CMFT to downregulate MIF-dependent, inflammatory gene expression with little evidence of off-target metabolic toxicity. These findings provide proof-of-concept for advancing the pharmacogenomic development of precision-based MIF inhibitors for diverse autoimmune and inflammatory conditions.

Downregulation of PDCD4 through STAT3/ATF6/autophagy mediates MIF-induced PASMCs proliferation/migration and vascular remodeling

To address the molecular mechanisms underlying macrophage migration inhibitory factor (MIF) induced pulmonary artery smooth muscle cells (PASMCs) proliferation, migration and vascular remodeling in pulmonary hypertension (PH), primary cultured rat PASMCs and monocrotaline (MCT)-induced rats with PH were applied in the present study. The results showed that MIF increased signal transducer and activator of transcription 3 (STAT3) phosphorylation, and then stimulated activating transcription factor 6 (ATF6) activation, subsequently triggered autophagy activation, which further led to programmed cell death factor 4 (PDCD4) lysosomal degradation, and eventually promoted PASMCs proliferation/migration. In lung tissues of MCT rats, MIF protein expression was elevated, phosphorylation of STAT3 and activation of ATF6 were increased, activation of autophagy was evident, and reduction of PDCD4 was observed. Intervention with MIF inhibitor 4-Iodo-6-phenylpyrimidine (4-IPP), ATF6 blocker melatonin or autophagy inhibitor chloroquine, confirmed the in vitro interaction among MIF, STAT3, ATF6, autophagy and PDCD4 in MCT induced rats with PH. Targeting MIF/STAT3/ATF6/autophagy/PDCD4 axis effectively prevented the development of PH by suppressing PASMCs proliferation and vascular remodeling. In conclusions, we demonstrate that MIF activates the STAT3/ATF6/autophagy cascade and then degrades PDCD4 leading to PASMCs proliferation/migration and pulmonary vascular remodeling, suggesting that intervention this axis might have potential value in management of PH.

Macrophage migration inhibitory factor promotes glucocorticoid resistance of neutrophilic inflammation in a murine model of severe asthma

BackgroundSevere neutrophilic asthma is resistant to treatment with glucocorticoids. The immunomodulatory protein macrophage migration inhibitory factor (MIF) promotes neutrophil recruitment to the lung and antagonises responses to glucocorticoids. We hypothesised...

Clinicopathological significance of the EMT-related proteins and their interrelationships in prostate cancer. An immunohistochemical study.

The chronic inflammation influences a microenvironment, where as a result of losing control over tissue homeostatic mechanisms, the carcinogenesis process may be induced. Inflammatory response cells can secrete a number of factors that support both initiation and progression of cancer and also they may consequently induct an epithelial-mesenchymal transition (EMT), the process responsible for development of distant metastasis. Macrophage migration inhibitory factor (MIF) acts as a pro-inflammatory cytokine that is considered as a link between chronic inflammation and tumor development. MIF can function as a modulator of important cancer-related genes expression, as well as an activator of signaling pathways that promotes the development of prostate cancer. The study was performed on FFPE tissues resected from patients who underwent radical prostatectomy. To investigate the relationship of studied proteins with involvement in tumor progression and initiation of epithelial-to-mesenchymal transition (EMT) process, we selected clinicopathological parameters related to tumor progression. Immunohistochemical analyses of MIF, SOX-4, β-catenin and E-cadherin were performed on TMA slides. We found a statistically significant correlation of overall β-catenin expression with the both lymph node metastasis (p<0.001) and presence of angioinvasion (p = 0.012). Membrane β-catenin expression was associated with distant metastasis (p = 0.021). In turn, nuclear MIF was correlated with lymph node metastasis (p = 0.003). The positive protein-protein correlations have been shown between the total β-catenin protein expression level with level of nuclear SOX-4 protein expression (r = 0.27; p<0.05) as well as negative correlation of β-catenin expression with level of nuclear MIF protein expression (r = -0.23; p<0.05). Our results seem promising and strongly highlight the potential role of MIF in development of nodal metastases as well as may confirm an involvement of β-catenin in disease spread in case of prostate cancer.

MIF promoter polymorphism increases peripheral blood expression levels, contributing to increased susceptibility and poor prognosis in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the most common malignancies in the world. The etiology and pathogenesis of HCC remain unclear. Macrophage migration inhibitory factor (MIF) plays a critical role in the pathogenesis of hepatocellular carcinoma. The association between MIF polymorphisms and its expression level in HCC has rarely been demonstrated. In the present study, the peripheral blood of 202 patients with HCC (HCC group), 242 patients with chronic hepatitis B (CHB group), 215 patients with liver cirrhosis (LC group) and 227 healthy volunteers (normal group) were collected, DNA was extracted and the target fragment of MIF gene was amplified using PCR. The products were then sequenced, and the expression levels of MIF protein were tested using ELISA. The results showed that the MIF rs755622 polymorphism was associated with an increased susceptibility and metastasis of HCC, and that the genotypes GC and CC were associated with poor prognosis of HCC. Compared with the normal, CHB and LC groups, the expression of MIF in the peripheral blood of the HCC group was significantly increased, and the high expression was associated with to poor prognosis. In the HCC group, MIF protein levels for genotypes GC and CC were increased compared with those of genotype GG. The current study indicated that the MIF rs755622 polymorphism is associated with susceptibility and metastasis of HCC, and that the GC and CC genotypes may be indicators of poor prognosis, which may be ascribed to the MIF rs755622 polymorphism leading to elevated MIF protein expression in peripheral blood.

Macrophage migration inhibitory factor may play a protective role in osteoarthritis

BackgroundOsteoarthritis (OA) is the most prevalent form of arthritis and the major cause of disability and overall diminution of quality of life in the elderly population. Currently there is no cure for OA, partly due to the large gaps in our understanding of its underlying molecular and cellular mechanisms. Macrophage migration inhibitory factor (MIF) is a procytokine that mediates pleiotropic inflammatory effects in inflammatory diseases such as rheumatoid arthritis (RA) and ankylosing spondylitis (AS). However, data on the role of MIF in OA is limited with conflicting results. We undertook this study to investigate the role of MIF in OA by examining MIF genotype, mRNA expression, and protein levels in the Newfoundland Osteoarthritis Study.MethodsOne hundred nineteen end-stage knee/hip OA patients, 16 RA patients, and 113 healthy controls were included in the study. Two polymorphisms in the MIF gene, rs755622, and -794 CATT5-8, were genotyped using polymerase chain reaction–restriction fragment length polymorphism (PCR-RFLP) and PCR followed by automated capillary electrophoresis, respectively. MIF mRNA levels in articular cartilage and subchondral bone were measured by quantitative polymerase chain reaction. Plasma concentrations of MIF, tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and interleukin-1 beta (IL-1β) were measured by enzyme-linked immunosorbent assay.Resultsrs755622 and -794 CATT5-8 genotypes were not associated with MIF mRNA or protein levels or OA (all p ≥ 0.19). MIF mRNA level in cartilage was lower in OA patients than in controls (p = 0.028) and RA patients (p = 0.004), while the levels in bone were comparable between OA patients and controls (p = 0.165). MIF protein level in plasma was lower in OA patients than in controls (p = 3.01 × 10−10), while the levels of TNF-α, IL-6 and IL-1β in plasma were all significantly higher in OA patients than in controls (all p ≤ 0.0007). Multivariable logistic regression showed lower MIF and higher IL-1β protein levels in plasma were independently associated with OA (OR per SD increase = 0.10 and 8.08; 95% CI = 0.04–0.19 and 4.42–16.82, respectively), but TNF-α and IL-6 became non-significant.ConclusionsReduced MIF mRNA and protein expression in OA patients suggested MIF might have a protective role in OA and could serve as a biomarker to differentiate OA from other joint disorders.

Inhibition of Macrophage Migration Inhibitory Factor Protects against Inflammation through a Toll-like Receptor-Related Pathway after Diffuse Axonal Injury in Rats.

Objective We have previously demonstrated that inflammation induced by toll-like receptors (TLRs) 2/4 exert cerebral deleterious effects after diffuse axonal injury (DAI); however, the underlying mechanisms are not fully understood. Macrophage migration inhibitory factor (MIF) is a multifunctional cytokine involved in inflammatory responses. The purpose of this study was to investigate the role of MIF in inflammation induced by TLRs in the cortices of DAI rats. Methods The rat DAI model was established by head rotational acceleration and confirmed by β-APP, HE, and silver staining. MIF protein expression at 3 h, 6 h, 12 h, 1 d, and 3 d after DAI was measured by western blot. The localization of MIF was measured by immunofluorescence. MIF antagonist ISO-1 was intracerebroventricularly injected to inhibit MIF. Neuronal and axonal injury and glial responses were assessed by TUNEL, immunohistochemistry, and TEM. Expression of TLR2, TLR4, ERK, phospho-ERK, NF-κB, and phospho-NF-κB was examined by western blot. The level of IL-1β, IL-6, and TNF-α was measured by ELISA. Results MIF expression was significantly increased, peaking at 1 day after DAI, and MIF was mainly localized in microglial cells and neurons. ISO-1 suppressed neuronal apoptosis, axonal injury, and glial responses and decreased the expression of downstream signaling molecules related to TLR2/4, including ERK, phospho-ERK, NF-κB, phospho-NF-κB, IL-1β, IL-6, and TNF-α. Conclusion MIF was involved in the neuronal and axonal damage through a TLR-related pathway following DAI.

The small molecule macrophage migration inhibitory factor antagonist MIF098, inhibits pulmonary hypertension associated with murine SLE

Pulmonary arterial hypertension (PAH) is a severe complication of systemic lupus erythematosus (SLE), with unclear etiopathogenesis. We evaluated the role of macrophage migration inhibitory factor (MIF), which has been implicated in idiopathic pulmonary hypertension (PH), in SLE-associated PAH. Circulating MIF was measured in SLE patients, SLE-PAH patients, and healthy donors. In situ pulmonary artery MIF protein expression was determined in spontaneous SLE mice (MRL/lpr) and hypoxia-induced C57BL/6J mice. Daily MIF098 was administered to C57BL/6J mice, and these mice were maintained in a hypoxic chamber for 4 weeks. The right ventricular systolic pressure (RVSP) and pathological characteristics of the pulmonary artery (PA), such as hyperproliferation, muscularization, and fibrosis were then measured in each group of mice. Data were also obtained in vitro using pulmonary smooth muscle cells (PASMC) challenged with platelet-derived growth factor (PDGF)-BB or 1% O2 hypoxia. As a result, circulating MIF was elevated in SLE-PAH patients compared with SLE patients or healthy donors. Higher RVSP SLE mice produced more MIF protein than lower RVSP SLE mice in the pulmonary artery. MIF098 decreased RVSP and inhibited distal pulmonary artery hyperproliferation, muscularization, and collagen deposition in hypoxia challenged mice. In addition, MIF098 inhibited PASMC proliferation and migration by regulating mitogen-activated protein kinase/extracellular signal-regulated kinase 1/2 (MAPK/ERK1/2) signal- and cell-cycle-related proteins. MIF098 also reduced collagen synthesis by inhibiting the TGFβ1/Smad2/Smad3 pathway in cell-based experiments. In conclusion, MIF may serve as a biomarker and a therapeutic target of SLE-associated PAH. Pharmacologic MIF antagonism may be an effective means to ameliorate SLE-PAH.

The expression of macrophage migration inhibitory factor and intercellular adhesion molecule-1 in rats with periodontitis and atherosclerosis

ObjectiveThe aim of this study was to explore the role of macrophage migration inhibitory factor (MIF) and intercellular adhesion molecule (ICAM)-1 in rats with periodontitis and atherosclerosis. MethodsTwenty-four male Sprague-Dawley rats were randomly divided into 4 groups: control group (C), periodontitis group (P), atherosclerosis group (AS), and periodontitis plus atherosclerosis group (P + AS). The levels of MIF and ICAM-1 in serum were detected by an enzyme-linked immunosorbent assay (ELISA). The protein expression of MIF and ICAM-1 in the carotid artery tissues was examined by immunohistochemical staining. ResultsThe results of the ELISA showed that the serum level of MIF in the P + AS group (59.40 ± 3.92 ng/mL) was significantly higher than that in the C group (42.93 ± 2.63 ng/mL), the P group (45.57 ± 2.59 ng/mL) and the AS group (50.88 ± 4.20 ng/mL) (P < 0.05). Similarly, the ICAM-1 level in the P + AS group (6.77 ± 1.47 ng/mL) was much higher than that in the C (1.33 ± 0.25 ng/mL), P (3.99 ± 0.44 ng/mL) and AS groups (4.19 ± 0.89 ng/mL) (P < 0.05). Furthermore, the expression of MIF, as assessed by immunohistochemical staining, was significantly higher in the artery tissues of the P + AS group than in the tissues of the other three groups (P < 0.05). However, there was no significant difference in the protein level of ICAM-1 between the P + AS and AS groups, where its expression was much stronger than in the C and the P groups. ConclusionOur results indicate that there is a close association between periodontitis and atherosclerosis. MIF and ICAM-1 may play a role in the development of atherosclerosis and periodontitis.

Macrophage migration inhibitory factor antagonist (S,R)3‑(4‑hydroxyphenyl)‑4,5‑dihydro‑5‑isoxazole acetic acid methyl ester attenuates inflammation and lung injury in rats with acute pancreatitis in pregnancy.

Macrophage migration inhibitory factor (MIF) is an inflammatory cytokine involved in many acute and chronic inflammatory diseases. However, its role in acute lung injury associated with acute pancreatitis in pregnancy (APIP) has not yet been elucidated. The present study was undertaken to clarify the effect and potential mechanism of MIF antagonist (S,R)3-(4-hydroxyphenyl)-4,5-dihydro-5-isoxazole acetic acid methyl ester (ISO-1) in the development of acute lung injury in rats with APIP. Eighteen late-gestation SD rats were randomly assigned to three groups: Sham operation (SO) group, APIP group, and ISO-1 group. All the rats were sacrificed 6 h after modeling. The severity of pancreatitis was evaluated by serum amylase (AMY), lipase (LIPA), tumor necrosis factor (TNF)-α, interleukin (IL)-1β and IL-6 and assessing the histopathological score. Lung injury was determined by performing histology and inflammatory cell infiltration investigations. Western blot analysis was used to detect the protein expression of MIF, phosphorylated and total P38 and nuclear factor-κB (NF-κB) protein in lungs. The results showed that MIF was upregulated in the lung of APIP rats. Compared with APIP group, the intervention of ISO-1 alleviated the pathological injury of the pancreas and lungs, decreased serum AMY and LIPA, attenuated serum concentrations of TNF-α, IL-1β, and IL-6, reduced the number of MPO-positive cells in the lung and inhibited the activation of P38MAPK and NF-κB. These results suggest that MIF is activated in lung injury induced by APIP. Furhtermore, the present findings indicate that the MIF antagonist ISO-1 has a protective effect on lung injury and inflammation, which may be associated with deactivating the P38MAPK and NF-κB signaling pathway.

Expression of macrophage migration inhibitory factor gene in placenta tissue and its correlation with gestational diabetes mellitus

Objective: To investigate the relationship between the expression of macrophage migration inhibitory factor (MIF) gene and the incidence of gestational diabetes mellitus (GDM) in Chinese Han population in northern China. Method: This study was consisted of 126 GDM women and 150 cases of healthy subjects who came from the obstetrics department of Maternity Hospital of Qingdao University and Maternity and Child Care Hospital of Donggang District of Rizhao. The protein expression of MIF in placenta tissue was detected by Western blot method. The blood glucose, insulin levels and other clinical physicochemical index were tested. The differences of MIF, fasting blood-glucose (FBG), fasting insulin (FIN) and the homeostasis model assessment of insulin resistance (HOMA-IR) were compared between the two groups of pregnant women. Result: The level of MIF protein in GDM group (0.85±0.10) was higher than that in healthy pregnant group (0.12±0.09), with significant difference (P<0.001). Insulin resistance index in GDM group were higher than that in healthy pregnant group, with significant difference (P<0.001). Conclusion: The expression of MIF in placental tissues of GDM women was increased and correlated with insulin resistance in GDM patients, suggesting that MIF may play an important role in the occurrence and development of GDM.

Hypoxia Drives AML Proliferation in the Bone Marrow Microenvironment Via Macrophage Inhibitory Factor

Abstract Introduction Hypoxia is an important component of the bone marrow microenvironment and the hematopoietic stem cell niche. Studies have shown that hypoxia contributes to the development and maintenance of acute myeloid leukemia (AML) cells within the bone marrow microenvironment. Hypoxia is principally maintained by members of the hypoxia-inducible factor (HIF), in particular HIF1a and its target genes, including MIF. We have previously shown that AML cells express constitutively high macrophage migration inhibitory factor (MIF) which drives IL-8 expression by the BM-MSC which in turn supports AML cell survival and proliferation. The aim of the present study is to determine if there is a connection between the role of hypoxia in regulating AML survival and MIF survival signals. Furthermore we investigate the role BM-MSC in regulating the hypoxic response. Methods Primary AML and BM-MSC were isolated from AML patients following informed consent and under approval from the UK National Research Ethics Service (LRCEref07/H0310/146). AML cell lines and primary AML blasts were cultured under normoxic (20% oxygen) or hypoxic conditions (1% oxygen) for 4 - 24 hours, mRNA expression of MIF, HIF1a, VEGF and IL-8 were determined by RT-PCR. MIF and IL-8 protein was determined using target specific ELISA. HIF1a protein expression was determined by western blotting. Hypoxia-mimetic agents, cobalt chloride (CoCl2) and desferrioxamine (DFO) were used. Cell proliferation was determines using CellTiter Glo and trypan blue exclusion. CFU-assays were performed using complete methylcellulose media. Results To determine If MIF is regulated by HIF1a in AML cells, we mimicked hypoxic conditions using CoCl2 and DFO in AML cells. Both CoCl2 and DFO upregulate MIF transcription and protein expression in OCI-AML3 cell lines and in primary AML blasts. Moreover, hypoxia increases both MIF mRNA expression and MIF chemokine expression compared to normoxic conditions. Lentiviral mediated knockdown of MIF in AML cells show significantly reduced cell proliferation and colony formation in methylcellulose media. Recombinant MIF induced interleukin-8 in AML blasts and the MIF inhibitor blocked MIF induced IL-8 release. Lentiviral mediated KD of HIF1a decreased MIF expression in human AML cells and a significant reduced their proliferative capacity. Finally we found that hypoxia increased MIF in AML blasts which was further increased when in co-culture with BM-MSC. Conclusions The results reported here suggest that hypoxia significantly affects the expression of the survival cytokine MIF in AML blasts. Furthermore, we show that BM-MSC regulate HIF1a induced MIF expression in AML under hypoxic conditions. We propose this hypoxia regulated HIF1a/MIF axis is essential to blast survival in the bone marrow niche. Disclosures Rushworth: Infinity Pharmaceuticals: Research Funding.

Expression pattern of miR-451 and its target MIF (macrophage migration inhibitory factor) in colorectal cancer

AimsTo investigate the expression pattern of microRNA-451 (miR-451) in patients with colorectal carcinoma and correlate with the expression of its target gene MIF (macrophage migration inhibitory factor).MethodsMatched cancer and non-cancer...

Tanshinone IIA pretreatment attenuates ischemia/reperfusion-induced renal injury

Tanshinone IIA is a chemical compound extracted from the root of traditional Chinese herb Salvia miltiorrhiza Bunge. Tanshinone IIA has been suggested to possess anti-inflammatory activity and antioxidizing capability. Recently, accumulating results have indicated the antitumor activity of tanshinone IIA; thus, it has attracted increasing attention. In addition, tanshinone IIA has been indicated to attenuate ischemia/reperfusion induced renal injury (I/RIRI); however, little is known regarding the underlying mechanisms involved in this process. In the present study an I/RIRI rat model was used to analyze the effects of tanshinone IIA on myeloperoxidase (MPO), TNF-α and IL-6 activities using ELISA kits. Furthermore, macrophage migration inhibitory factor (MIF), cleaved caspase-3, B-cell lymphoma 2 (Bcl-2) and p38 mitogen-activated protein kinase (MAPK) protein expression levels were evaluated using western blot analysis. The results indicated that tanshinone IIA protected renal function in I/RIRI rats. ELISA demonstrated that tanshinone IIA significantly reduced MIF, TNF-α and IL-6 activities in I/RIRI rats. Western blot analysis showed that tanshinone IIA significantly suppressed MIF, cleaved caspase-3 and p38 MAPK protein expression levels in I/RIRI rats. The present results suggest that tanshinone IIA pretreatment attenuates I/RIRI via the downregulation of MPO expression, inflammation, MIF, cleaved caspase-3 and p38 MAPK.

Expression of macrophage migration inhibitory factor (MIF) in bovine oviducts is higher in the postovulatory phase than during the oestrus and luteal phase.

Whether macrophage migration inhibitory factor (MIF) in the bovine oviduct is important for early embryogenesis has not been well substantiated. The aim of the present study was to test the hypothesis that bovine oviduct expresses higher levels of MIF during the post-ovulation phase. Both ampullary and isthmic samples were collected from Japanese black heifers during oestrus (Day 0; n=5), postovulation (Day 3; n=6) and luteal phase (Days 9-12; n=5). MIF mRNA and protein were extracted from the ampullary and isthmic samples and their levels measured by real-time polymerase chain reaction and western blot analysis respectively. Fluorescent immunohistochemistry was performed on frozen ampullary and isthmic sections using antibodies against MIF. MIF mRNA and protein expression was higher in the postovulatory phase than during oestrus and the luteal phase (P<0.05). Fluorescent immunohistochemistry confirmed that in all phases of the oestrous cycle evaluated, the primary site of MIF expression in the ampulla and isthmus was the tunica mucosa. In conclusion, the bovine ampulla and isthmus have higher MIF expression during the postovulatory phase. Further studies are needed to clarify the role of MIF in bovine oviducts.

Transient transfection of macrophage migration inhibitory factor small interfering RNA disrupts the biological behavior of oral squamous carcinoma cells.

Macrophage migration inhibitory factor (MIF) is closely associated with tumorigenesis. The present study aimed to investigate the effects of MIF on the proliferation, migration and colony formation of oral squamous cell carcinoma (OSCC), and to quantify the protein expression levels of MIF in OSCC tissue samples. Firstly, small interfering (si) RNA was used to knock down the gene expression of MIF in Tca8113, HN5 and SCC25 OSCC cells. Secondly, proliferation, migration and colony formation of the OSCC cells were determined by MTT, transmigration and colony formation assays, respectively. Western blotting was performed to detect changes in the protein expression levels of the epithelial mesenchymal transition markers, Twist-related protein 1 (Twist1), matrix metalloproteinase (MMP)-2 and MMP-9. Finally, immunohistochemistry was used to examine the protein expression of MIF in OSCC tissue samples. The results demonstrated that siRNA against MIF significantly downregulated the expression levels of MIF in all OSCC cells, and decreased their proliferation and migration ability. Colony formation ability was also inhibited in the OSCC cells following transfection with MIF siRNA. Furthermore, western blotting demonstrated that the protein expression of Twist1 was decreased similarly to those of MIF. The protein expression of MMP-2 revealed no change, whereas that of MMP-9 decreased. The protein expression of MIF was detected in OSCC tissue samples with staining predominantly located in the cell membrane and cytoplasm. The present study demonstrated that MIF may be important in the pathogenesis and progression of OSCC, and indicated its potential therapeutic value.

Macrophage Migration Inhibitory Factor in Acute Adipose Tissue Inflammation.

Macrophage migration inhibitory factor (MIF) is a pleiotropic cytokine and has been implicated in inflammatory diseases. However, little is known about the regulation of MIF in adipose tissue and its impact on wound healing. The aim of this study was to investigate MIF expression in inflamed adipose and determine its role in inflammatory cell recruitment and wound healing. Adipose tissue was harvested from subcutaneous adipose tissue layers of 24 healthy subjects and from adipose tissue adjacent to acutely inflamed wounds of 21 patients undergoing wound debridement. MIF protein and mRNA expression were measured by ELISA and RT-PCR. Cell-specific MIF expression was visualized by immunohistochemistry. The functional role of MIF in cell recruitment was investigated by a chemotaxis assay and by flow cytometry of labeled macrophages that were injected into Mif –/–and wildtype mice. Wound healing was evaluated by an in vitro scratch assay on human fibroblast monolayers. MIF protein levels of native adipose tissue and supernatants from acutely inflamed wounds were significantly elevated when compared to healthy controls. MIF mRNA expression was increased in acutely inflamed adipose tissue indicating the activation of MIF gene transcription in response to adipose tissue inflammation. MIF is expressed in mature adipocytes and in infiltrated macrophages. Peripheral blood mononuclear cell migration was significantly increased towards supernatants derived from inflamed adipose tissue. This effect was partially abrogated by MIF-neutralizing antibodies. Moreover, when compared to wildtype mice, Mif –/–mice showed reduced infiltration of labeled macrophages into LPS-stimulated epididymal fat pads in vivo. Finally, MIF antibodies partially neutralized the detrimental effect of MIF on fibroblast wound healing. Our results indicate that increased MIF expression and rapid activation of the MIF gene in fat tissue adjacent to acute wound healing disorders may play a role in cell recruitment to the site of inflammation and wound healing.

Association between macrophage migration inhibitory factor in the endometrium and estrogen in endometriosis.

Recent studies have shown that macrophage migration inhibitory factor (MIF) has a possible role in endometriosis-related pain and infertility, yet it has not been explored whether the mRNA level of MIF is altered in endometrial tissues from patients with endometriosis. The aim of the present study was to compare the expression of MIF in endometrial tissues from women with and without endometriosis, and to analyze the association between endometrial MIF expression and 17β-estradiol (E2). The protein and mRNA expression of MIF in the human endometrial tissue was assessed by western blotting and reverse transcription-polymerase chain reaction analysis, respectively. The MIF expression of women with endometriosis was found to be significantly higher than that of the controls. A positive correlation was noted between the serum E2 level and MIF expression. In endometrial cells from women with endometriosis, the level of E2-induced MIF upregulation was significantly higher than that in cells from women without endometriosis. In conclusion, this study demonstrated a significant increase in MIF expression in the endometrial tissues of women with endometriosis and an association between MIF expression and E2 level. MIF expression in endometrial cells from patients with endometriosis showed an increased sensitivity to stimulation by E2.

The expression of microRNA-451 in human endometriotic lesions is inversely related to that of macrophage migration inhibitory factor (MIF) and regulates MIF expression and modulation of epithelial cell survival.

What is the role of microRNA-451 (miR-451) in human endometriotic tissue? miR451 expression was elevated in endometriotic lesion tissue. MiR451 modulated the expression of macrophage migration inhibitory factor and limited cell survival. microRNAs are post-transcriptional regulators of gene expression which have been reported to be mis-expressed in endometriotic tissue. The exact pattern of expression and role of miR451 in endometriosis is currently unknown. Thirty women with endometriosis are included in the study. Matched eutopic (N = 30) and endometriotic lesion tissue (N = 43) were collected. miR-451, macrophage migration inhibitory factor (MIF), cyclin E1 (CCNE) and phosphatase and tensin homolog (PTEN) mRNA expression were examined by quantitative real-time (qRT)-PCR while MIF protein expression was evaluated by western blot analysis. miR-451 regulation of MIF in vitro translation was confirmed by 3'untranslated region (UTR) reporter assays and western blot analysis. The effect of miR-451 on cell survival was assessed using a human endometrial epithelial cell line (HES). Compared with eutopic endometrium, both MIF mRNA and protein were significantly (P < 0.05) decreased in endometriotic lesions and this was associated with a significant (P < 0.05) increase in miR-451 expression. Transfection of HES cells with luciferase reporter constructs for MIF revealed that miR-451 specifically bound to the 3'UTR to regulate expression. Further, forced expression of miR-451 induced a significant (P < 0.05) down-regulation of both MIF mRNA and protein in HES cells which was associated with a significant (P < 0.05) reduction in cell survival. Inhibition of MIF using a specific antagonist verified that reduction of MIF contributes to HES cell survival. miR-451 and MIF expression were only examined in tissue from women with endometriosis. Our data support the hypothesis that miR-451 is elevated in endometriotic tissue and, through regulating MIF expression, may function to limit endometriotic lesion cell survival. This study was funded by the National Institutes of Health/NICHD by grant NIH HD069043 to W.B.N. The authors have no competing interests.

Expression of macrophage migration-inhibitory factor in duodenal ulcer and its relation to Helicobacter pylori infection.

The aim of this study was to examine the expression of macrophage migration-inhibitory factor (MIF) in duodenal ulcer epithelial cells and its relation to Helicobacter pylori (Hp) infection, and to discuss the pathogenic roles of MIF expression and Hp infection in duodenal ulcer. MIF protein and mRNA expression was examined in samples from patients with duodenal ulcer with and without Hp infection (N = 40 each, experimental group), and in normal duodenal bulb mucosal tissue (N = 40, control group) using immunohistochemistry and in situ hybridization. Patients without Hp infection received routine treatment, and treatment was provided to the patients positive for Hp to eradicate Hp infection. Hp and MIF expression levels before treatment and after the ulcer had been cured were compared. The positive rates of MIF protein and mRNA in patients with Hp infection before treatment were 67.5 and 65%, respectively, and were 18.9 and 21.6% in the 37 patients from whom Hp was eliminated. These were statistically different both before and after treatment compared with controls (P < 0.05). In the patients without Hp infection, the positive rates of MIF protein and mRNA expression before (45 and 47.5%, respectively) and after (32.5 and 30%) treatment were not significantly different (P > 0.05). The results of this study suggested that MIF is related to the development of duodenal ulcer, and that the presence of Hp is closely related with the expression of MIF in the duodenal mucosa and the development of duodenal ulcer.