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.

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 regulates proliferation of gastric cancer cells via the PI3K/Akt pathway

AIM:To investigate the effects of macrophage migration inhibitory factor (MIF) on proliferation of human gastric cancer MGC-803 cells and expression of cyclin D1 and p27Kip1 in them,and further determine whether the effects are related to the PI3K/Akt signal transduction pathway. METHODS:Gastric cancer MGC-803 cells were cultured and then treated with 50 μg/L recombinant human MIF (rhMIF) with and without a PI3K inhibitor,LY294002 (25 μmol/L). MTT assay was used to detect the prolifer-ation of MGC-803 cells. Cell cycle was detected by flow cytometry. Expression of cyclin D1 and p27Kip1 mRNA was by reverse transcription-polymerase chain reaction. Protein expression of phosphorylated Akt (p-Akt),Akt,cyclin D1 and p27Kip1 was examined by immunocyto-chemistry and Western blotting. RESULTS:rhMIF signifi cantly stimulated the prolifera-tion of MGC-803 cells and cell cycle progression from G1 phase to S phase in a concentration-and time-de-pendent manner. After the MGC-803 cells were treated with rhMIF for 24 h,the expression of cyclin D1 was signifi cantly up-regulated compared with the cells not treated with rhMIF at both mRNA and protein levels(0.97 ± 0.02 vs 0.74 ± 0.01,P = 0.002; 0.98 ± 0.05 vs 0.69 ± 0.04,P = 0.003). The p27Kip1 was down-regulated but only statistically significant at the protein level. rhMIF significantly increased the expression of p-Akt,which reached the peak at 30 min,but did not affect the expression of Akt. However,LY294002 inhibited all the effects of rhMIF.CONCLUSION:Macrophage MIF increases the proliferation of gastric cancer cells,induces the expression of cyclin D1 at the transcriptional level and inhibits the expression of p27Kip1 at the post-transcriptional level via the PI3K/Akt pathway.

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.

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.

Upregulation of macrophage migration inhibitory factor and calgizzarin by androgen in TM4 mouse Sertoli cells

Aim： To identify proteins induced by androgen in Sertoli cells during spermatogenesis. Methods： We analyzed protein profiles in TM4 Sertoli cells treated with dihydrotestosterone （DHT） using surface enhanced laser desorption ionization time-of-flight mass spectrometry （SELDI-TOF-MS）. Results： We found increases in the expression of a 5.0-kDa protein at 15 min, an 11.3-kDa protein at 24 h and 4.3 kDa, 5.7 kDa, 5.8 kDa, 9.95 kDa and 9.98 kDa proteins at 48 h after the treatment. In contrast, the expression of 6.3 kDa and 8.6 kDa proteins decreased at 30 min, and 4.9 kDa, 5.0 kDa, 12.4 kDa and 19.8 kDa proteins at 48 h after the treatment. The ll.3-kDa protein was identified as macrophage migration inhibitory factor （MIF） known to having various functions. The 9.98-kDa protein was identified as calgizzarin related to calcium channels. The timing of their expression suggests that MIF and calgizzarin are involved in late regulation of spermatogenesis in Sertoli cells by androgen. Conclusion： MIF and calgizzarin are two important androgen-responsive proteins produced by Sertoli cells and they might play a role in regulating spermatogenesis.

Effects of leukemia inhibitory factor on endogenous neural stem cell proliferation and glycoprotein-130 expression in a mouse model of cerebral infarction

Leukemia inhibitory factor （LIF） has been shown to promote proliferation of endogenous neural stem cells. In this study, we treated mice with cerebral infarction using LIF to investigate whether the LIF receptor subunit glycoprotein （gp）130 is involved in neuroprotection. After LIF treatment, the motor function of model mice was significantly improved. Immunofluorescence histochemistry showed increased numbers of endogenous neural stem cells surrounding the infarct foci. Western blot analysis revealed that gp130 expression was significantly decreased surrounding the infarcted foci. Results demonstrated that LIF promoted the proliferation of endogenous neural stem cells by inhibiting gp130 protein expression.

Effects of leukemia inhibitory factor and basic fibroblast growth factor on free radicals and endogenous stem cell proliferation in a mouse model of cerebral infarction

The present study established a mouse model of cerebral infarction by middle cerebral artery occlusion, and monitored the effect of 25 tJg/kg leukemia inhibitory factor and （or） basic fibroblast growth factor administration 2 hours after model establishment. Results showed that following administration, the number of endogenous neural stem cells in the infarct area significantly increased, malondialdehyde content in brain tissue homogenates significantly decreased, nitric oxide content, glutathione peroxidase and superoxide dismutase activity significantly elevated, and mouse motor function significantly improved as confirmed by the rotarod and bar grab tests. In particular, the effect of leukemia inhibitory factor in combination with basic fibroblast growth factor was the most significant. Results indicate that leukemia inhibitory factor and basic fibroblast growth factor can improve the microenvironment after cerebral infarction by altering free radical levels, improving the quantity of endogenous neural stem cells, and promoting neurological function of mice with cerebral infarction.

CD74 and macrophage migration inhibitory factor as therapeutic targets in gastric cancer

AIM:To investigate the relationship and molecular features of CD74/macrophage migration inhibitory factor(MIF)/Toll-like receptor 4(TLR4) in gastric cancer.METHODS:CD74,MIF and TLR4 expression in the paraffin-embedded sections of gastric cancer from 120 patients were detected by immunohistochemical staining.Knock down of CD74 expression in gastric cancer cell line MKN-45 was performed by lentivirus transduction and detected by Western blotting.MKN-45 cell proliferation assay under the stimulants was measured by the cell counting kit 8(CCK8) assay and MIF concentration in the culture medium was detected by enzymelinked immunosorbent assay.Surface staining of CD74 in the MKN-45 cell line under the stimulation of lipopolysaccharide(LPS) was measured by flow cytometry.MIF,CD74 and TLR4 co-localization in the MKN-45 cell line was performed by the immunoprecipitation.RESULTS:CD74,MIF and TLR4 were found to be expressed in gastric cancer and increased significantly in the advanced stage,and were also associated with lymph node metastasis.Correlation analysis revealed that CD74 was positively correlated with MIF(r = 0.2367,P < 0.01) and both proteins were also associated with TLR4(r = 0.4414,r = 0.5001,respectively,P < 0.01).LPS can significantly promote MKN-45 cell proliferation(3.027 ± 0.388 vs 4.201 ± 0.092,P < 0.05),induce MIF production(54.333 ± 2.906 pg/mL vs 29.667 ± 3.180 pg/mL,P < 0.01) and cell surface expression of CD74(75.6% ± 4.046%vs 9.4% ± 0.964%,P < 0.01) at LPS concentration of 1 μg/mL compared to medium control.Knockdown of CD74 or using antiCD74 and MIF antagonist ISO-1 significantly reduced LPS-induced MKN-45 cell proliferation(4.201 ± 0.092 vs 3.337 ± 0.087,4.534 ± 0.222 vs 3.368 ± 0.290,4.058 ± 0.292vs 2.934 ± 0.197,respectively,P < 0.01).MIF,CD74 and TLR4 could co-localize in the MKN-45 cell line.CONCLUSION:Upregulation of MIF,CD74 and TLR4 are associated with increasing clinical stage and provide an opportunity as novel gastric cancer chemoprevention and/or treatment strategy.

Macrophage migration inhibitory factor facilitates astrocytic production of the CCL2 chemokine following spinal cord injury

Spinal cord injury causes accumulation of a large number of leukocytes at the lesion site where they contribute to excessive inflammation.Overproduced chemokines are responsible for the migratory process of the leukocytes,but the regulatory mechanism underlying the production of chemokines from resident cells of the spinal cord has not been fully elucidated.We examined the protein levels of macrophage migration inhibitory factor and chemokine C-C motif chemokine ligand 2 in a spinal cord contusion model at different time points following spinal cord injury.The elevation of macrophage migration inhibitory factor at the lesion site coincided with the increase of chemokine C-C motif chemokine ligand 2 abundance in astrocytes.Stimulation of primary cultured astrocytes with different concentrations of macrophage migration inhibitory factor recombinant protein induced chemokine C-C motif chemokine ligand 2 production from the cells,and the macrophage migration inhibitory factor inhibitor 4-iodo-6-phenylpyrimidine attenuated the stimulatory effect.Further investigation into the underlying mechanism on macrophage migration inhibitory factor-mediated astrocytic production of chemokine C-C motif chemokine ligand 2 revealed that macrophage migration inhibitory factor activated intracellular JNK signaling through binding with CD74 receptor.Administration of the macrophage migration inhibitory factor inhibitor 4-iodo-6-phenylpyrimidine following spinal cord injury resulted in the reduction of chemokine C-C motif chemokine ligand 2-recruited microglia/macrophages at the lesion site and remarkably improved the hindlimb locomotor function of rats.Our results have provided insights into the functions of astrocyte-activated chemokines in the recruitment of leukocytes and may be beneficial to develop interventions targeting chemokine C-C motif chemokine ligand 2 for neuroinflammation after spinal cord injury.

Gene knockout or inhibition of macrophage migration inhibitory factor alleviates lipopolysaccharide-induced liver injury via inhibiting inflammatory response

Background:Liver injury is one of the most common complications during sepsis.Macrophage migration inhibitory factor(MIF)is an important proinflammatory cytokine.This study explored the role of MIF in the lipopolysaccharide(LPS)-induced liver injury through genetically manipulated mouse strains.Methods:The model of LPS-induced liver injury was established in wild-type and Mif-knockout C57/BL6 mice.Serum levels of alanine aminotransferase(ALT),aspartate aminotransferase(AST),and total bilirubin(TBil)were detected,and the expressions of MIF,tumor necrosis factor-α(TNF-α)and interleukin-1β(IL-1β)were measured.Liver histopathology was conducted to assess liver injury.Moreover,the inhibitions of MIF with(S,R)-3-(4-hydroxyphenyl)-4,5-dihydro-5-isoxazole acetic acid methyl ester(ISO-1)and 4-iodo-6-phenylpyrimidine(4-IPP)were used to evaluate their therapeutic potential of liver injury.Results:Compared with wild-type mice,the liver function indices and inflammation factors presented no significant difference in the Mif-/-mice.After 72 h of the LPS-induced liver injury,serum levels of ALT,AST,and TBil as well as TNF-αand IL-1βwere significantly increased,but the knockout of Mif attenuated liver injury and inflammatory response.In liver tissue,m RNA levels of TNF-α,IL-1βand NF-κB p65 were remarkably elevated in LPS-induced liver injury,while the knockout of Mif reduced these levels.Moreover,in LPS-induced liver injury,the inhibitions of MIF with ISO-1 and 4-IPP alleviated liver injury and slightly attenuated inflammatory response.Importantly,compared to mice with LPS-induced liver injury,Mif knockout or MIF inhibitions significantly prolonged the survival of the mice.Conclusions:In LPS-induced liver injury,the knockout of Mif or MIF inhibitions alleviated liver injury and slightly attenuated inflammatory response,thereby prolonged the survival of the mice.Targeting MIF may be an important strategy to protect the liver from injury during sepsis.

D-dopachrome tautomerase from Japanese sea bass(Lateolabrax japonicus) is a chemokine-like cytokine and functional homolog of macrophage migration inhibitory factor

D-dopachrome tautomerase(DDT),a member of the macrophage migration inhibitory factor(MIF)protein superfamily,is a newly described cytokine with chemokine-like characteristics.However,research on fish DDT remains limited.In this study,we identified a DDT homolog(LjDDT)from the Japanese sea bass,Lateolabrax japonicus.Sequence analysis showed that LjDDT had typical sequence features of known DDT and MIF homologs and was most closely related to DDT of rock bream(Oplegnathus fasciatus).LjDDT transcripts were detected in all tested tissues of healthy Japanese sea bass,with the highest expression found in the liver.Upon infection with Vibrio harveyi,LjDDT transcripts were significantly down-regulated in the three tested tissues,including the liver,spleen,and head kidney.Recombinant LjDDT(rLjDDT)and the corresponding antibody(anti-rLjDDT)were subsequently prepared.The administration of 100μg/g anti-rLjDDT had a statistically significant protective effect on the survival of V.harveyi-infected fish.Moreover,rLjDDT was able to induce the migration of monocytes/macrophages(MO/MФ)and lymphocytes both in vitro and in vivo,but without significant influence on the migration of neutrophils.rLjDDT exhibited chemotactic activity for lipopolysaccharide(LPS)-stimulated M1-type MO/MΦin vitro,but not for cAMP-stimulated M2-type MO/MΦ.Furthermore,the knockdown of LjCD74,but not LjCXCR4,significantly down-regulated the rLjDDT-enhanced migration of MO/MΦand relieved the rLjMIF-inhibited migration of MO/MΦ.These results indicate that LjCD74 may be the major chemotactic receptor of LjDDT and LjMIF in Japanese sea bass MO/MΦ.Combined rLjDDT+rLjMIF treatment had no significant effect on the migration of MsiRNA,LjCD74si-,or LjCXCR4sitreated MO/MΦcompared to the control group,suggesting that the roles of LjDDT and LjMIF may be antagonistic.In conclusion,our study demonstrates for the first time that DDT may play a role in the immune responses of fish against bacterial infection through chemotactic recruitment of MO/MΦvia mediation of CD74 as an antagonist of MIF.

Association of the macrophage migration inhibitory factor promoter polymorphisms with benign lymphoepithelial lesion of lacrimal gland

AIM： To identify the association of the macrophage migration inhibitory factor （MIF） gene polymorphism with the susceptibility of benign lymphoepithelial lesions （BLEL） of the lacrimal gland. METHODS： A total of 40 BLEL of lacrimal gland cases were matched with 40 healthy subjects （HS）. Extraction the plasma and whole blood DNA of patients of lacrimal gland BLEL and HS. Elisa and polymerase chain reaction was used to determine in plasma contents of MIF and MIF gene SNP-173G〉C and STR -794 CATT（8） polymorphism, respectively. RESULTS： The MIF levels in plasma were significantly higher in patients with lacrimal gland BI.EL versus HS （P〈0.001）. The -173 G〉C MIF polymorphism was significantly associated with lacrimal gland BLEL, with a significantly higher frequency of the C allele in lacrimal gland BLEL patients compared with HS （OR=2.38, 95% C1=1.07-5.31, P=0.032）, and the -173 C/x is more frequent in patients than in HS, P=0.037. Besides, we found that the carriage rate of the MIF -173C/x is associated with higher plasma levels of MIF in the BLEI. of lacrimal gland. CONCLUSION： MIF -173G/C variants play an insidious role in susceptibility of BLEL of lacrimal gland. Otherwise,there is no statistically significant correlation exists between MIF-794 CATT （） and BLEL of lacrimal gland.

Expression of Leukemia Inhibitory Factor in Airway Epithelial Tissue of Asthmatic Rats

In order to investigate the expression of leukemia inhibitory factor （LIF） in airway epithelial tissues of normal and asthmatic rats, the influence of dexamethasone and the role of LIF in pathogenesis of asthma, 30 Sprague-Dawley （SD） rats were randomly divided into 3 groups （10 for each group）： normal group, asthma model group, and dexamethasone-interfered group. In asthma model group and dexamethasone-interfered group, asthma rat models were established by intraperitoneal （i.p.） injection of 10% ovalbumin （OVA） and challenge with 1% OVA via inhalation. Rats in dexamethasone-interfered group were pretreated with dexamethasone （2 mg/kg, i.p） 30 min before each challenge. The expression of LIF protein in lung was detected by immunohistochemistry. The results showed that LIF protein was mainly expressed in cytoplasm of bronchial epithelial cells. The expression of LIF protein in the airway epithelial tissue of asthma model group was significantly higher than that in normal group and dexamethasone-interfered group （P〈0.01）, but there was no significant difference between normal group and dexamethasone-interfered group （P〉0.05）. It was concluded that the expression of LIF was increased significantly in the airway epithelial tissue of the asthma rats, and dexamethasone could down-regulate the expression of LIF. It was suggested that LIF might play an important role in the pathogenesis of asthma as an inflammation regulator.

Macrophage migration inhibitory factor decreased T-type Ca^(2+) channel current through activating Src

Aims T-type Ca2+ current(ICaT)plays an important role in the pathogenesis of atrial fibrillation（AF）.The present study sought to investigate the role of Macrophage migration inhibitory factor（MIF）,a pleiotropic cytokine,in the regulation of T-type Ca2+ channel in atrium myocytes.Methods We used whole-cell voltage-clamp technique and biochemical assays to study the regulation and expression of ICa,T in mouse atrium myocytes（HL-1 cells）.Results Serum MIF concentrations was slightly increased in patients with AF compared to sinus rhythm（SR） controls.In cultured HL-1 cells, significant amounts of MIF were produced in response to hydrogen peroxide（H2O2）,but not AngiotensinⅡ（AngⅡ）. Mouse recombinant MIF（rMIF）（20 or 40 nM,24 h） suppressed peak ICa,T by-38%and-60%in a concentration-dependent manner,impaired the voltage-dependent activation of ICa,T,and down-regulated of TCC alG mRNA.Src inhibitors genistein and PPl significantly enhanced ICaT.The depression of ICa,T induced by rMIF could be reversed by genistein and PP1.Conclusions MIFis involved in the pathogenesis of AF,probably by decreasing ICa,T through impairment of the channel function and activation of c-Src kinases in atrium myocytes.

Moxibustion of Zusanli(ST36)and Shenshu(BL23)alleviates the inflammation of rheumatoid arthritis in rats through regulating macrophage migration inhibitory factor/glucocorticoids signaling

OBJECTIVE:To test the hypothesis that moxibustion may inhibit rheumatoid arthritis(RA)synovial inflammation by regulating the expression of macrophage migration inhibitory factor(MIF)/glucocorticoids(GCs).METHODS:Fifty male Sprague-Dawley rats were randomly divided into five groups(n=10 each):blank Control(CON)group,RA Model(RA)group,Moxibustion(MOX)group,MIF inhibitor(S,R)-3-(4-hydroxyphenyl)-4,5-dihydro-5-isoxazole acetic acid methyl ester(ISO-1)group,and Moxibustion+MIF inhibitor ISO-1(MOX+ISO-1)group.Rats in the ISO-1 group and ISO-1+MOX group were intraperitoneally injected with the inhibitor ISO-1.The rats in the RA group,ISO-1 group,MOX group,and ISO-1+MOX group were injected with Freund's complete adjuvant(FCA)in the right hind footpad to establish an experimental RA rat model.In the MOX group and MOX+ISO-1 group,rats were treated with Moxa.The thickness of the footpads of the rats in each group was measured at three-time points before,after modeling and after moxibustion treatment.The contents of serum MIF,corticosterone(CORT),tumor necrosis factor-α(TNF-α)and interleukin-1β(IL-1β)were detected by enzyme-linked immunosorbent assay;and the contents of synovial MIF were detected by Western blot.Hematoxylin-eosin(HE)staining method was used to observe the pathological changes of synovial tissue under a section light microscope,and pathological scoring was performed according to the grading standard of the degree of synovial tissue disease.RESULTS:Moxibustion was found to reduce the level of MIF and alleviate inflammation in RA rats in this study.In addition,after inhibiting the expression of MIF,the level of CORT increased,and the level of TNF-α decreased.Treating RA rats with inhibited MIF by moxibustion,the level of CORT was almost unchanged,but the level of TNF-α further decreased.The correlation analysis data suggested that MIF was positively related to the expression of TNF-α and negatively correlated with the expression of CORT.CONCLUSION:Reducing MIF to increase CORT and decrease TNF-α by moxibustion treatment in RA.MIF may be a factor for moxibustion to regulate the expression of CORT,but the expression of TNF-α is due to the incomplete regulation of the MIF.This study added to the body of evidence pointing to moxibustion's antiinflammatory mechanism in the treatment of RA.

Pathogenesis of coxsackievirus B3-induced myocarditis： role of macrophage migration inhibitory factor

Background Macrophage migration inhibitory factor （MIF） is an upstream regulator in immune and inflammatory responses.However,its role in viral myocarditis remains unknown.In this study,we investigated the role of the MIF in coxsackievirus B3 （CVB3）-induced myocarditis.Methods Mice were randomized into two groups receiving either Eagle＇s minimal essential medium （EMEM,control group） or virus solution （infected group）.Subsets of mice in the infected group were sacrificed on days 3,7,14 and 28 after inoculation.Expression of MIF was detected using an enzyme-linked immunosorbent assay （ELISA）,reverse transcription polymerase chain reaction and immunohistochemistry.A neutralizing antibody （Ab） to MIF was injected intraperitoneally from day 0 to 7 after inoculation.Disease severity was estimated by histopathology of the heart and by the heart weight to body weight ratio,and the interleukin-1β （IL-1β） and tumor necrosis factor α （TNF-α） in the myocardium were measured by ELISA on day 14.Results The serum MIF concentration and expression levels of myocardial MIF mRNA and protein were significantly elevated in mice on days 7 and 14 post-infection.The survival rate was markedly higher and disease severity was obviously less in mice treated with anti-MIF Ab.Furthermore,MIF blockade significantly decreased the IL-1β and TNF-α in the myocarditic heart.Conclusion These results demonstrate that MIF is an important naturally occurring inflammatory cytokine in CVB3-induced myocarditis,and anti-MIF Ab may lessen the inflammatory response.

The effect of Helicobacter pylori infection on expression of macrophage migration inhibitory factor by T cells and macrophages in gastric mucosa

Background Macrophage migration inhibitory factor (MIF) plays a pivotal role in inflammatory and immune-mediated diseases. However, its molecular function and role in gastrointestinal diseases has rarely been studied and thus warrants an in-depth investigation. This study was designed and conducted to determine MIF expression in Helicobacter pylori (H. pylori)-induced gastritis and the effect of H. pylori on MIF expression in monocytes in vitro. Methods Gastric specimens of 62 patients with chronic gastritis were obtained through endoscopic biopsies. Both gastric antrum and body were examined for histopathologic changes. Positive H. pylori was determined through rapid urease test and histopathological examination. A patient was classified as H. pylori positive if both tests showed positive results. The updated Sydney System was employed to assess the severity and activity of gastric inflammation. Double immunoassaying for MIF/T-cells (CD45RO) and MIF/macrophage (KP1), as well as in situ hybridization for the expression of MIF mRNA were used for the current analysis. THP-1, a monocyte cell line, was co-incubated with H. pylori strains (ATCC26695) and subsequently examined for the expression of MIF protein and mRNA by enzyme linked immunosorbent assay and retrospective transcription-polymerase chain reaction, respectively. Results Among 62 patients with chronic gastritis, significant increase in total T-cells, MIF+ T-cells, total macrophages, MIF+ macrophages and MIF mRNA+ cells was observed in 42 H. pylori positive patients compared to H. pylori negative patients. Moreover, the increase of the MIF mRNA+ cells was highly correlated with the severity of the disease(number of MIF mRNA+cells/mm2, mild: 2834±382, moderate: 3569±123, severe: 3881±118, P<0.01). In vitro results showed that the expression of MIF protein and mRNA in monocytes was significantly increased after incubation with H. pylori strains.Conclusions Overexpression of MIF is common in H. pylori-induced gastric inflammation, which suggests MIF may play an important role in the initiation and development of this disease.

Serum macrophage migration inhibitory factor as a potential biomarker to evaluate therapeutic response in patients with allergic asthma:an exploratory study

Background:Previous studies have shown that macrophage migration inhibitory factor(MIF)is involved in the pathogenesis of asthma.This study aimed to investigate whether serum MIF reflects a therapeutic response in allergic asthma.Methods:We enrolled 30 asthmatic patients with mild-to-moderate exacerbations and 20 healthy controls,analyzing the parameter levels of serum MIF,serum total immunoglobulin E(tIgE),peripheral blood eosinophil percentage(EOS%),and fractional exhaled nitric oxide(FeNO).Lung function indices were used to identify disease severity and therapeutic response.Results:Our study showed that all measured parameters in patients were at higher levels than those of controls.After one week of treatment,most parameter levels decreased significantly except for serum tIgE.Furthermore,we found that serum MIF positively correlated with EOS%as well as FeNO,but negatively correlated with lung function indices.Receiver operator characteristic(ROC)curve analysis indicated that among the parameters,serum MIF exhibited a higher capacity to evaluate therapeutic response.The area under the curve(AUC)of MIF was 0.931,with a sensitivity of 0.967 and a specificity of 0.800.Conclusions:Our results suggested that serum MIF may serve as a potential biomarker for evaluating therapeutic response in allergic asthma with mild-to-moderate exacerbations.

Macrophage migration inhibitory factor enhances neoplastic cell invasion by inducing the expression of matrix metalloproteinase 9 and interleukin-8 in nasopharyngeal carcinoma cell lines

Background Nasopharyngeal carcinoma (NPC) shows highly invasive and metastatic features. This study aims to investigate macrophage migration inhibitory factor (MIF)-induced invasion of NPC cells in vitro and the effects on matrix metalloproteinases (MMPs) and interleukin-8 (IL-8),and to study the mechanism of tumor cell invasion and metastasis in the early stage of NPC. Methods Two nasopharyngeal carcinoma cell lines,CNE-1 and CNE-2,were adopted in this study. The NPC cell invasion and migration were evaluated by microinvasion assay. The variation of expression percentages of MMP2- or MMP9-positive cells was detected by flow cytometry in two cell lines with or without MIF treatment. Western blotting and RT-PCR were used to assay the protein and mRNA expressions of MMP2 and MMP9. The IL-8 concentration secreted by NPC cells was compared with the cells with different treatments using ELISA. Results After treating with MIF for 48 hours,the cell numbers of CNE-1 and CNE-2 which went through the 8-μm filter membrane were increased. Compared with non-MIF treated NPC cells,significant difference could be found both in CNE-1( P =0.005) and CNE-2 cells ( P =0.001) . The percentages of MMP9-positive cells were significantly increased in both CNE-1 [from (28.5±2.5)% to (82.4±3.5)%, P =0.001] and CNE-2 [from (32.8±3.5)% to (86.1±1.6)%, P =0.002]. The relative intensity of MMP9 protein expression was also enhanced in both cell lines (CNE-1: from 83.1±6.0 to 242.9±22.9, P =0.002;CNE-2: from 84.4±4.3 to 278.9±29.7, P =0.003). Correspondingly,the increased MMP9 mRNA expression level was significantly detectable in both cell lines.The concentration of IL-8 in the supernatant of CNE-2 was higher [(1201.8±593.3) pg/ml] after treatment. It was also remarkably higher than that in the supernatant of CNE-2 without treatment ( P =0.026). However,there was no significant difference in the concentration variation of IL-8 in CNE-1 ( P =0.581), while the IL-8 mRNA level was only enhanced in CNE-2. Conclusions MIF can induce potent invasion of NPC cell lines in vitro , and the infiltrating lymphocytes in NPC might be responsible for the invasion and metastasis of tumor cells. MIF cytokine which is secreted by these infiltrating lymphocytes might contribute to the invasion as well as metastasis of NPC in the early stages by induction of MMP9 and IL-8 in an indirect pathway.