CO-EXPRESSION OF MACROPHAGE COLONY-STIMULATING FACTOR WITH ITS RECEPTOR IN HUMAN HEPATOMA CELLS AND ITS POTENTIAL ROLES

Objective: To investigate the potential role of macrophage colony-stimulating factor (M-CSF) and macrophage colony-stimulating factor receptor (M-CSF-R) on the growth of human hepatoma cells. Methods: Specimens of different origin, including tissues of human hepatocellular carcinoma (HCC), human fetal liver (FL) and normal liver (NL), the hepatoma cell lines, as well as the peripheral blood mononuclear cells (PBMC) from patients with HCC or liver metastatic tumor (LMT), were used to detect the expression levels of M-CSF and M-CSF-R by ABC immunohistochemistry staining and reverse transcription polymerase chain reaction methods the expression levels of M-CSF and M-CSF-R. Influence of monoclonal antibody against M-CSF (B5) or M-CSF-R (RE2) on proliferation ability of hepatoma cell linesin vitro was also studied. Results: The results showed that hepatoma tissues produced elevated levels of both M-CSF and M-CSF-R compared with those of fetal liver (P<0.001). The M-CSF/M-CSF-R expression levels of PBMC from hepatoma patients were higher than those of LMT patients (P<0.01,P<0.05) and the normal people (P<0.001). The hepatoma cell lines showed strong positive for M-CSF and M-CSF-R production. Both B5 and RE2 displayed a dose-dependent inhibitory effect on the growth and proliferation of hepatoma cells. Conclusion: The study indicates a co-expression model for M-CSF-R in hepatoma cells, suggesting an involvement of M-CSF/M-CSF-R in growth signaling of those malignant cells. The M-CSF/M-CSF-R seems to function through an autonomy mechanism in human hepatoma.

Granulocyte-macrophage colony-stimulating factor （GM-CSF） and T-cell responses： what we do and don＇t know

Granulocyte-macrophage colony-stimulating factor （GM-CSF） is an important hematopoietic growth factor and immune modulator. GM-CSF also has profound effects on the functional activities of various circulating leukocytes. It is produced by a variety of cell types including T cells, macrophages, endothelial cells and fibroblasts upon receiving immune stimuli. Although GM-CSF is produced locally, it can act in a paracrine fashion to recruit circulating neutrophils, monocytes and lymphocytes to enhance their functions in host defense. Recent intensive investigations are centered on the application of GM-CSF as an immune adjuvant for its ability to increase dendritic cell （DC） maturation and function as well as macrophage activity. It is used clinically to treat neutropenia in cancer patients undergoing chemotherapy, in AIDS patients during therapy, and in patients after bone marrow transplantation. Interestingly, the hematopoietic system of GM-CSF-deficient mice appears to be normal; the most significant changes are in some specific T cell responses. Although molecular cloning of GM-CSF was carried out using cDNA library oft cells and it is well known that the T cells produce GM-CSF after activation, there is a lack of systematic investigation of this cytokine in production by T cells and its effect on T cell function. In this article, we will focus mainly on the immunobiology of GM-CSF in T cells.

Preclinical evaluation of herpes simplex virus armed with granulocyte-macrophage colony-stimulating factor in pancreatic carcinoma

AIM: To investigate the therapeutic efficacy and mechanisms of action of oncolytic-herpes-simplex-virus encoding granulocyte-macrophage colony-stimulating factor(HSVGM-CSF) in pancreatic carcinoma.METHODS: Tumor blocks were homogenized in a sterile grinder in saline.The homogenate was injected into the right armpit of each mouse.After vaccination,the mice were randomly assigned into four groups: a control group,a high dose HSVGM-CSFgroup [1 × 107plaque forming units(pfu)/tumor],a medium dose HSVGM-CSF group(5 × 106pfu/tumor) and a low dose HSVGM-CSF group(5 × 105pfu/tumor).After initiation of drug administration,body weights and tumor diameters were measured every 3 d.Fifteen days later,after decapitation of the animal by cervical dislocation,each tumor was isolated,weighed and stored in 10% formaldehyde solution.The drug effectiveness was evaluated according to the weight,volume and relative volume change of each tumor.Furthermore,GM-CSF protein levels in serum were assayed by enzyme-linked immunosorbent assays at 1,2,3 and 4 d after injection of HSVGM-CSF.RESULTS: Injection of the recombinant mouse HSV encoding GM-CSF resulted in a significant reduction in tumor growth compared to the control group,and dosedependent effects were observed: the relative tumor proliferation rates of the low dose,medium dose and high dose groups on 15 d after injection were 45.5%,55.2% and 65.5%,respectively.The inhibition rates of the tumor weights of the low,middle,and high dose groups were 41.4%,46.7% and 50.5%,respectively.Furthermore,the production of GM-CSF was significantly increased in the mice infected with HSVGM-CSF.The increase in the GM-CSF level was more pronounced in the high dose group compared to the other two dose groups.CONCLUSION: Our study provides the first evidence that HSVGM-CSFcould inhibit the growth of pancreatic cancer.The enhanced GM-CSF expression might be responsible for the phenomenon.

DETERMINATION OF SERUM SOLUBLE MACROPHAGE COLONY-STIMULATING FACTOR RECEPTOR LEVELS IN PATIENTS WITH HEMATOLOGICAL DISEASES

Objective: To investigate the serum levels of soluble macrophage colony-stimulating factor receptor (M-CSFsR) in normal subjects and patients with hematological diseases and its clinical implications in hematological diseases. Methods: The concentration of M-CSFsR was determined by ELISA. The serum M-CSFsR was identified and characterized by immunoprecipitation and Western blotting. Results: The mean serum level of M-CSFsR of 123 normal individuals was 0.48 ng/ml ± 0.41 ng/ml. Immunoprecipitation and Western blotting assay revealed a ～ 90kD band of serum M-CSFsR. The mean serum M-CSFsR level of 60 patients with acute lymphoblastic leukemia (ALL), 36 patients with acute myeloblastic leukemia (AML), 13 patients with myelodysplastic syndrome (MDS) and 42 patients with aplastic anemia (AA) .were 0.22 ng/ml±0.23 ng/ml, 0.17 ng/ml±0.16 ng/ml, 0.19 ng/ml±0.16 ng/ml and 0.23 ng/ml±0.21 ng/ml, respectively, which were significantly lower than that of normal subjects (P=0.002 ,P<0.0001,P<0.0001 andP<0.0001). The mean serum M-CSFsR level of 51 idiopathic thrombocytopenic purpura (ITP) patients was significantly higher than that of normal subjects (2.05 ng/ml±2.75 ng/ml,P<0.0001). Conclusion: The serum M-CSFsR levels of patients with ALL, AML, MDS and AA were significantly lower, while the level of patients with ITP was significantly higher than that of normal individuals. Patients with severe ITP (platelet count<30×l09/L) had the highest M-CSFsR level. It suggested that the abnormal levels of serum M-CSFsR may associate with some hematological diseases and may contribute to the pathological process.

Impaired granulocyte-macrophage colony-stimulating factor bioactivity accelerates surgical recurrence in ileal crohn's disease

AIM To examine the relationship between elevated granulocyte-macrophage colony-stimulating factor(GMCSF) auto-antibodies(Ab) level and time to surgical recurrence after initial surgery for Crohn's disease(CD). METHODS We reviewed 412 charts from a clinical database at tertiary academic hospital. Patients included in the study had ileal or ileocolonic CD and surgical resection of small bowel or ileocecal region for management of disease. Serum samples were analyzed for serological assays including GM-CSF cytokine, GM-CSF Ab, ASCA Ig G and Ig A, and genetic markers including SNPs rs2066843, rs2066844, rs2066845, rs2076756 and rs2066847 in NOD2, rs2241880 in ATG16 L1, and rs13361189 in IRGM. Cox proportional-hazards models were used to assess the predictors of surgical recurrence.RESULTS Ninety six percent of patients underwent initial ileocecal resection(ICR) or ileal resection(IR) and subsequently 40% of patients required a second ICR/IR for CD. GMCSF Ab level was elevated at a median of 3.81 mcg/mL. Factors predicting faster time to a second surgery included elevated GM-CSF Ab [hazard ratio(HR) 3.52, 95%CI: 1.45-8.53, P = 0.005] and elevated GM-CSF cytokine(HR = 2.48, 95%CI: 1.31-4.70, P = 0.005). Factors predicting longer duration between first and second surgery included use of Immunomodulators(HR = 0.49, 95%CI: 0.31-0.77, P = 0.002), the interaction effect of low GM-CSF Ab levels and smoking(HR = 0.60, 95%CI: 0.45-0.81, P = 0.001) and the interaction effect of low GM-CSF cytokine levels and ATG16 L1(HR = 0.65, 95%CI: 0.49-0.88, P = 0.006).CONCLUSION GM-CSF bioavailability plays a critical role in maintaining intestinal homeostasis. Decreased bioavailability coupled with the genetic risk markers and/or smoking results in aggressive CD behavior.

Granulocyte-macrophage colony-stimulating factor protects mice against hepatocellular carcinoma by ameliorating intestinal dysbiosis and attenuating inflammation

BACKGROUND Hepatocellular carcinoma(HCC)is the third leading cause of cancer mortality worldwide.The gut microbiota can help maintain healthy metabolism and immunity.Granulocyte-macrophage colony-stimulating factor(GM-CSF)is a critical factor in promoting health and homeostasis;it promotes intestinal immunity,stimulates bone marrow precursors to generate macrophage colonies,and enhances the antibacterial and antitumor activity of circulating monocytes.As such,GM-CSF may protect against HCC development by regulating immunity as well as intestinal microecology.AIM To investigate the impact of GM-CSF on the gut microbiome and metabolic characteristics of HCC.METHODS Thirty-six male BALB/c nude mice were divided into three groups:Control(n=10),HCC(n=13),and HCC+GM-CSF(GM-CSF overexpression,n=13).We utilized HCC cells to establish orthotopic transplantation tumor models of HCC with normal and over-expressing GM-CSF.Liver injury,immune inflammatory function and intestinal barrier function were evaluated.The fecal microbiome and metabolome were studied using 16S rRNA absolute quantification sequencing and gas chromatography-mass spectrometry.RESULTS GM-CSF overexpression significantly affected the gut microbiome of mice with HCC and resulted in a high abundance of organisms of the genera Roseburia,Blautia and Butyricimonass,along with a significant reduction in Prevotella,Parabacteroides,Anaerotruncus,Streptococcus,Clostridium,and Mucispirillum.Likewise,GM-CSF overexpression resulted in a substantial increase in fecal biotin and oleic acid levels,along with a prominent decrease in the fecal succinic acid,adenosine,fumaric acid,lipoic acid,and maleic acid levels.Correlation analysis revealed that the intestinal microbiota and fecal metabolites induced by GM-CSF were primarily involved in pathways related to reducing the inflammatory response,biotin metabolism,and intestinal barrier dysfunction.CONCLUSION GM-CSF can protect against HCC development by regulating immunity and modulating the abundance of specific intestinal microorganisms and their metabolites.This study provides new insights into the therapeutic approaches for HCC.

CONSTRUCTION OF EUKARYOTIC EXPRESSION VECTOR WITH GRANULOCYTE-MACROPHAGE COLONY-STIMULATING FACTOR GENE

Objective: To construct the eukaryotic expression vector that express human granulocyte-macrophage colony-stimulating factor (hGM-CSF) gene for making highly express in mammalian cells. Methods: Extract totally RNA from the induced human fetal lung (HFL) cell line. HGM-CSF cDNA was obtained by reverse transcription-polymerase chain reaction (RT-PCR), and then directionally subcloned into the HindIII and EcoRI site on the pcDNA3.1 plasmid, which was controlled by the CMV promoter, to form the recombinant expressing vector pcDNA3.1-GM-CSF. Results: The PCR amplification was identified and the sequence was analyzed, the results showed that hGM-CSF was properly inserted into the vector and the sequence was correct.

Granulocyte-macrophage colony-stimulating factor-transfected bone marrow stromal cells for the treatment of ischemic stroke

Adult, male, Sprague-Dawley rats were injected with granulocyte-macrophage colony-stimulating factor-transfected bone marrow stromal cells （GM-CSF-BMSCs） into the ischemic boundary zone at 24 hours after onset of middle cerebral artery occlusion. Results showed reduced infarct volume, decreased number of apoptotic cells, improved neurological functions, increased angiogenic factor expression, and increased vascular density in the ischemic boundary zone in rats that underwent GM-CSF-BMSCs transplantation compared with the BMSCs group. Experimental findings suggested that GM-CSF-BMSCs could serve as a potential therapeutic strategy for ischemic stroke and are superior to BMSCs alone.

Macrophage colony-stimulating factor expressed in non-cancer tissues provides predictive powers for recurrence in hepatocellular carcinoma

AIM To investigate the role of macrophage colony-stimulating factor(M-CSF) in patients with hepatocellular carcinoma(HCC) after surgery. METHODS Expression of M-CSF, distribution of M2 macrophages(Mφs), and angiogenesis were assessed in the liver, including tumors and peritumoral liver tissues. The prognostic power of these factors was assessed. Mouse isolated hepatic Mφs or monocytes were cultured with media containing M-CSF. The concentration of vascular endothelial growth factor(VEGF) in media was assessed. Furthermore, the role of the M-CSF-matured hepatic Mφs on proliferation of the vascular endothelial cell(VEC) was investigated. RESULTS A strong correlation between the expressions of M-CSF and CD163 was observed in the peritumoral area. Also, groups with high density of M-CSF, CD163 or CD31 showed a significantly shorter time to recurrence(TTR) than low density groups. Multivariate analysis revealedthe expression of M-CSF or hepatic M2Mφs in the peritumoral area as the most crucial factor responsible for shorter TTR. Moreover, the expression of M-CSF and hepatic M2Mφs in the peritumoral area had better predictable power of overall survival. Values of VEGF in culture media were significantly greater in the hepatic Mφs compared with the monocytes. Proliferation of the VEC was greatest in the cells co-cultured with hepatic Mφs when M-CSF was present in media.CONCLUSION M-CSF increases hepatocarcinogenesis, most likely by enhancing an angiogenic factor derived from hepatic Mφ and could be a useful target for therapy against HCC.

Mudskipper interleukin-34 modulates the functions of monocytes/macrophages via the colony-stimulating factor-1 receptor 1

Interleukin-34(IL-34)is a novel cytokine that plays an important role in innate immunity and inflammatory processes by binding to the colonystimulating factor-1 receptor(CSF-1R).However,information on the function of IL-34 in fish remains limited.In the present study,we identified an IL-34 homolog from mudskippers(Boleophthalmus pectinirostris).In silico analysis showed that the mudskipper IL-34(BpIL-34)was similar to other known IL-34 variants in sequence and structure and was most closely related to an orange-spotted grouper(Epinephelus coioides)homolog.BpIL-34 transcripts were constitutively expressed in various tissues,with the highest level of expression found in the brain.Edwardsiella tarda infection significantly up-regulated the mRNA expression of BpIL-34 in the mudskipper tissues.The recombinant mature BpIL-34 peptide(rBpIL-34)was purified and used to produce anti-rBpIL-34 IgG.Western blot analysis combined with PNGase F digestion revealed that native BpIL-34 in monocytes/macrophages(MOs/MФs)was N-glycosylated.In vitro,rBpIL-34 treatment enhanced the phagocytotic and bactericidal activity of mudskipper MOs/MФs,as well as the mRNA expression of pro-inflammatory cytokines like tumor necrosis factorα(BpTNF-α)and BpIL-1βin these cells.Furthermore,the knockdown of mudskipper CSF-1R1(BpCSF-1R1),but not mudskipper BpCSF-1R2,significantly inhibited the rBpIL-34-mediated enhanced effect on MO/MФfunction.In conclusion,our results indicate that mudskipper BpIL-34 modulates the functions of MOs/MФs via BpCSF-1R1.

Antiviral Activity of Silkworm Expressed Recombinant Human Macrophage Colony-Stimulating Factor in Vitro and Its Mechanism

Cells, pretreated with the recombinant human macrophage colony-stimulating factor (rhM-CSF) expressed in silkworm larvae, were inoculated with several viruses to observe the effect of rhM-CSF on viral replication. The results showed that in cultures of fibroblast derived from human fetal skin-muslce tissues infected with the viruses (including VSV, rhinovirus, influenza virus type A3, HSV-1, HSV-2, adenovirus type 6 and type 11), rhM-CSF inhibited the virus-induced cytopathy, which defered or relieved the cytophathy and that in the cells derived from breast feeding rabbit kidney infected with HSV-1, rhM-CSF reduced titer of the virus in a rhM-CSF dose-dependent pattern,in which rhM-CSF with the dose of 1×106 U/L made the viral titer drop dwon over 200 times. This antiviral activity of rhM-CSF could be partially neutralized by anti-IFN-βMcAb, but not by antiTNF-α, anti-IFN-α, or anti-IFA-γ McAb, indicating the mechanism of the antiviral activity of MCSF is related to the induction of IFN-β.

The roles of macrophage migration inhibitory factor in retinal diseases

Macrophage migration inhibitory factor(MIF),a multifunctional cytokine,is secreted by various cells and participates in inflammatory reactions,including innate and adaptive immunity.There are some evidences that MIF is involved in many vitreoretinal diseases.For example,MIF can exacerbate many types of uveitis;measurements of MIF levels can be used to monitor the effectiveness of uveitis treatment.MIF also alleviates trauma-induced and glaucoma-induced optic nerve damage.Furthermore,MIF is critical for retinal/choroidal neovascularization,especially complex neovascularization.MIF exacerbates retinal degeneration;thus,anti-MIF therapy may help to mitigate retinal degeneration.MIF protects uveal melanoma from attacks by natural killer cells.The mechanism underlying the effects of MIF in these diseases has been demonstrated:it binds to cluster of differentiation 74,inhibits the c-Jun N-terminal kinase pathway,and triggers mitogen-activated protein kinases,extracellular signal-regulated kinase-1/2,and the phosphoinositide-3-kinase/Akt pathway.MIF also upregulates Toll-like receptor 4 and activates the nuclear factor kappa-B signaling pathway.This review focuses on the structure and function of MIF and its receptors,including the effects of MIF on uveal inflammation,retinal degeneration,optic neuropathy,retinal/choroidal neovascularization,and uveal melanoma.

Neutrophil peptide 1 accelerates the clearance of degenerative axons during Wallerian degeneration by activating macrophages after peripheral nerve crush injury

Macrophages play an important role in peripheral nerve regeneration,but the specific mechanism of regeneration is still unclear.Our preliminary findings indicated that neutrophil peptide 1 is an innate immune peptide closely involved in peripheral nerve regeneration.However,the mechanism by which neutrophil peptide 1 enhances nerve regeneration remains unclear.This study was designed to investigate the relationship between neutrophil peptide 1 and macrophages in vivo and in vitro in peripheral nerve crush injury.The functions of RAW 264.7 cells we re elucidated by Cell Counting Kit-8 assay,flow cytometry,migration assays,phagocytosis assays,immunohistochemistry and enzyme-linked immunosorbent assay.Axonal debris phagocytosis was observed using the CUBIC(Clear,Unobstructed Brain/Body Imaging Cocktails and Computational analysis)optical clearing technique during Wallerian degeneration.Macrophage inflammatory factor expression in different polarization states was detected using a protein chip.The results showed that neutrophil peptide 1 promoted the prolife ration,migration and phagocytosis of macrophages,and CD206 expression on the surfa ce of macrophages,indicating M2 polarization.The axonal debris clearance rate during Wallerian degeneration was enhanced after neutrophil peptide 1 intervention.Neutrophil peptide 1 also downregulated inflammatory factors interleukin-1α,-6,-12,and tumor necrosis factor-αin invo and in vitro.Thus,the results suggest that neutrophil peptide 1 activates macrophages and accelerates Wallerian degeneration,which may be one mechanism by which neutrophil peptide 1 enhances peripheral nerve regeneration.

IMMUNOHISTOCHEMICAL OBSERVATION OF MACROPHAGE COLONY STIMULATING FACTOR AND ITS RECEPTOR IN BREAST CANCER AND HEPATOMA TISSUES

Objective: To study the potential role of cellular macrophage colony-stimulating factor (cM-CSF) and cellular macrophage colony-stimulating factor receptor (cM-CSF-R) with breast cancer and hepatoma and search the way for clinical application. Methods: Frozen surgical specimens from 48 breast cancer patients, including 29 cases of histological grade II and 19 eases of grade III, and 16 hepatoma patients were investigated by Avidin Biotin Complex (ABC) immunohistochemical assay with anti-M-CSF monoclonal antibody (Mab) and anti-M-CSF-R Mab. Pathohistological examination was performed as well. Results: cM-CSF and cM-CSF-R were detected in tested specimens. The expression levels of cM-CSF and cM-CSF-R in grade III group were higher than in grade II group and more higher than control group hyperplasia of breast. Hepatoma tissues also showed higher expression level of cM-CSF and cM-CSF-R than normal adult and fetal liver. Conclusion: Breast cancer and hepatoma tissues presented higher expression levels of cM-CSF and cM-CSF-R than control and expression level might be related with tumor’s process.

Brain-derived neurotrophic factor mediates macrophage migration inhibitory factor to protect neurons against oxygen-glucose deprivation

Macrophage migration inhibitory factor(MIF)is a chemokine that plays an essential role in immune system function.Previous studies suggested that MIF protects neurons in ischemic conditions.However,few studies are reported on the role of MIF in neurological recovery after ischemic stroke.The purpose of this study is to identify the molecular mechanism of neuroprotection mediated by MIF.Human neuroblastoma cells were incubated in Dulbecco’s modified Eagle’s medium under oxygen-glucose deprivation(OGD)for 4 hours and then returned to normal aerobic environment for reperfusion(OGD/R).30 ng/mL MIF recombinant(30 ng/mL)or ISO-1(MIF antagonist;50μM)was administered to human neuroblastoma cells.Then cell cultures were assigned to one of four groups:control,OGD/R,OGD/R with MIF,OGD/R with ISO-1.Cell viability was analyzed using WST-1 assay.Expression levels of brain-derived neurotrophic factor(BDNF),microtubule-associated protein 2(MAP2),Caspase-3,Bcl2,and Bax were detected by western blot assay and immunocytochemistry in each group to measure apoptotic activity.WST-1 assay results revealed that compared to the OGD/R group,cell survival rate was significantly higher in the OGD/R with MIF group and lower in the OGD/R with ISO-1 group.Western blot assay and immunocytochemistry results revealed that expression levels of BDNF,Bcl2,and MAP2 were significantly higher,and expression levels of Caspase-3 and Bax were significantly lower in the MIF group than in the OGD/R group.Expression levels of BDNF,Bcl2,and MAP2 were significantly lower,and expression levels of Caspase-3 and Bax were significantly higher in the ISO-1 group than in the OGD/R group.MIF administration promoted neuronal cell survival and induced high expression levels of BDNF,MAP2,and Bcl2(anti-apoptosis)and low expression levels of Caspase-3 and Bax(pro-apoptosis)in an OGD/R model.These results suggest that MIF administration is effective for inducing expression of BDNF and leads to neuroprotection of neuronal cells against hypoxic injury.

Macrophage migration inhibitory factor as a potential prognostic factor in gastric cancer

AIM:To investigate macrophage migration inhibitory factor(MIF) expression and its clinical relevance in gastric cancer,and effects of MIF knockdown on proliferation of gastric cancer cells. METHODS:Tissue microarray containing 117 samples of gastric cancer and adjacent non-cancer normal tissues was studied for MIF expression by immunohistochemistry(IHC) semiquantitatively,and the association of MIF expression with clinical parameters was analyzed. MIF expression in gastric cancer cell lines was detected by reverse transcriptionpolymerase chain reaction(RT-PCR) and Western blot. Two pairs of si RNA targeting the MIF gene(MIF si-1 and MIF si-2) and one pair of scrambled si RNA as a negative control(NC) were designed and chemically synthesized. All si RNAs were transiently transfected in AGS cells with OligofectamineTM to knock down the MIF expression,with the NC group and mock group(OligofectamineTM alone) as controls. At 24,48,and 72 h after transfection,MIF m RNA was analyzed by RTPCR,and MIF and proliferating cell nuclear antigen(PCNA) proteins were detected by Western blot.The proliferative rate of AGS cells was assessed by methylthiazolyl tetrazolium(MTT) assay and colony forming assay.RESULTS:The tissue microarray was informative for IHC staining,in which the MIF expression in gastric cancer tissues was higher than that in adjacent noncancer normal tissues(P < 0.001),and high level of MIF was related to poor tumor differentiation,advanced T stage,advanced tumor stage,lymph node metastasis,and poor patient survival(P < 0.05 for all). After si RNA transfection,MIF m RNA was measured by real-time PCR,and MIF protein and PCNA were assessed by Western blot analysis. We found that compared to the NC group and mock group,MIF expression was knocked down successfully in gastric cancer cells,and PCNA expression was downregulated with MIF knockdown as well. The cell counts and the doubling times were assayed by MTT 4 d after transfection,and colonies formed were assayed by colony forming assay 10 d after transfection; all these showed significant changes in gastric cancer cells transfected with specific si RNA compared with the control si RNA and mock groups(P < 0.001 for all).CONCLUSION:MIF could be of prognostic value in gastric cancer and might be a potential target for small-molecule therapy.

Early treadmill exercise increases macrophage migration inhibitory factor expression after cerebral ischemia/reperfusion

The neuroprotective function of macrophage migration inhibitory factor(MIF) in ischemic stroke was rarely evaluated.This study aimed to investigate the effects of early treadmill exercise on recovery from ischemic stroke and to determine whether these effects are associated with the expression levels of MIF and brain-derived neurotrophic factor(BDNF) in the ischemic area.A total of 40 male Sprague-Dawley rats were randomly assigned to the ischemia and exercise group [middle cerebral artery occlusion(MCAO)-Ex,n = 10),ischemia and sedentary group(MCAO-St,n = 10),sham-surgery and exercise group(Sham-Ex,n= 10),or sham-surgery and sedentary group(Sham-St,n = 10).The MCAO-Ex and MCAO-St groups were subjected to MCAO for 60 minutes,whereas the Sham-Ex and Sham-St groups were subjected to an identical operation without MCAO.Rats in the MCAO-Ex and Sham-Ex groups then ran on a treadmill for 30 minutes once a day for 5 consecutive days.After reperfusion,the hanging time tested by the wire hang test was longer and the relative fractional anisotropy determined by MRI was higher in the peri-infarct region of the MCAO-Ex group compared with the MCAO-St group.The expression levels of MIF and BDNF in the peri-infarct region were upregulated in the MCAO-Ex group.Increased MIF and BDNF levels were positively correlated with relative fractional anisotropy changes in the peri-infarct region.There was no significant difference in the levels of MIF and BDNF in the peri-infarct region between the Sham-Ex and Sham-St groups.Our study demonstrated that early exercise(initiated 48 hours after the MCAO) could improve motor and neuronal recovery after ischemic stroke.Furthermore,the increased levels of MIF and BDNF in the peri-infarct region(penumbra) may be one of the mechanisms of enhanced neurological function recovery.All experiments were approved by the Institutional Animal Care and Use Committee in Asan Medical Center in South Korea(2016-12-126).

Serum and ascites levels of macrophage migration inhibitory factor, TNF-α and IL-6 in patients with chronic virus hepatitis B and hepatitis cirrhosis

Objective: To study the potential role of macrophage migration inhibitory factor (MIF), tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in the development of chronic virus hepatitis B (CH) and hepatitis cirrhosis (HC). Methods: The serum concentrations of MIF, TNF-α and IL-6 in 18 patients with chronic virus hepatitis B and in 14 patients with hepatitis cirrhosis without as- citic fluid, and the serum and ascites cytokine con- centrations in 22 HC patients with ascitic fluid were detected by enzyme linked immunity sorbed assay. Results: The cytokine concentrations of the patients were significantly higher than those of the controls. The serum levels of MIF, TNF-α and IL-6 of the 22 patients with ascitic fluid were higer than those of 14 HC patients without ascites. In the 18 patients with CH, the serum cytokine concentrations were the low- est. The serum cytokine concentrations of the 22 HC patients with ascites were significantly higher than those of the 14 HC patients without ascites (P< 0. 01). Their serum cytokine concentrations were sig- nificantly higher than those in the 18 patients with CH (P<0. 01). The concentration of IL-6 in ascites was the highest among all the groups. The serum le- vels of MIF, TNF-α and IL-6 are correlated with al- anine aminotransferase (ALT) in the patients with CH, but not in those with HC with or without asci- tes. Conclusions: These results indicated that MIF, TNF- α and IL-6 may participate in the pathological process of CH and cirrhosis, that IL-6 seems to play an important role in ascites formation, and that se- rum levels of MIF, TNF-α and IL-6 appear to reflect the severity of tissue injury in HBV disease.

Influence of granulocyte-macrophage colonystimulating factor and tumor necrosis factor on anti-hepatoma activities of human dendritic cells

INTRODUCTIONDendritic cells (DCs) play a key regulatory role inantitumor immunity,especially in its immuneaccessory role via MHC-Ⅰ molecules.We haverecently reported that DCs were able to enhance thekilling activity of Lymphokine and PHA activatedkiller (LPAK) cells in vitro.In the presentstudy,we evaluated the effects of GM-CSF andTNF upon antitumor activities of freshly

Cloning and mRNA expression of macrophage migration inhibitory factor (MIF) gene of large yellow croaker (P seudosciaena crocea)

Mammalian macrophage migration inhibitory factor （MIF） plays an important role as an indispensable mediator in the pathogenesis of inflammatory disease like septicemia, but little is known about the role of MIF homologue in fish septicemia. The authors have cloned the MIF homologue in large yellow croaker Pseudosciaena crocea （LycMIF） using RACE approach. The full-length cDNA of LycMIF was 634 bases and contained an ORF of 345 bases encoding a protein of 115 amino acid residues. As demonstrated by RT-PCR and QRT-PCR assay, MIF mRNAs were constitutively expressed in 11 selected tissues and were abundant in brain and liver. Moreover, the LycMIF transcripts in the liver and head kidney were responsive to bacteria infection and could be significantly up-regulated. Our results provide the first direct evidence that fish MIF was implicated in pathogenesis of fish vibrosis and play an important role in response to bacteria infection.