Total Glucosides of Paeony Improve Renal Injury in Mice with Systemic Lupus Erythematosus by Regulating TLR9/MyD88/NF-κB Pathway / 中国实验方剂学杂志

ObjectiveTo investigate the intervention effect of total glucosides of paeony （TGP） on the renal injury of MRL/lpr mice based on the Toll-like receptor 9 （TLR9）/myeloid differentiation factor 88 （MyD88）/nuclear transcription factor-κB （NF-κB） signaling pathway and explore the immunological mechanism of TGP in preventing and treating systemic lupus erythematosus （SLE）. MethodMRL/lpr female mice of SPF grade were randomly divided into a model group， a dexamethasone group （0.15 g·kg-1）， and high- （0.078 g·kg-1） and low-dose （0.039 g·kg-1） TGP groups， and female C57BL/6J mice were assigned to a blank group， with 7 mice in each group. Mice in each group were treated with corresponding drugs or normal saline by gavage at the same time every day. After 4 weeks， samples were collected. The kidney and spleen were weighed， and the organ index was calculated. Serum creatinine （SCr） and blood urea nitrogen （BUN） levels in each group were detected by biochemical assay. Hematoxylin-eosin （HE） staining was used to observe the histopathological changes in the kidney. The degree of renal fibrosis was evaluated by Masson staining. The serum levels of interleukin （IL）-2， interferon （IFN）-α， IL-4， and anti-nuclear antibody （ANA） were detected by enzyme-linked immunosorbent assay （ELISA）. The mRNA expression of TLR9， MyD88， and NF-κB p65 in renal tissues was detected by real-time quantitative polymerase chain reaction （Real-time PCR）. The protein expression of TLR9 and NF-κB p65 in renal tissues was detected by immunofluorescence. The protein expression of TLR9， MyD88， and NF-κB p65 in renal and spleen tissues was tested by Western blot. ResultCompared with the blank group， the model group showed increased SCr， BUN， spleen index， and kidney index （P<0.05）， deteriorated pathological injury and fibrosis in renal tissues， elevated serum levels of IFN-α， IL-4， and ANA， decreased level of IL-2 （P<0.05）， and up-regulated TLR9， MyD88， and NF-κB p65 mRNA and protein levels in the kidney and spleen （P<0.05）. Compared with the model group， the TGP groups displayed reduced SCr， BUN， spleen index， and kidney index （P<0.05）， relieved pathological damage and fibrosis in renal tissues， decreased serum levels of IFN-α， IL-4， and ANA （P<0.05）， increased level of IL-2， and declining mRNA and protein expression levels of TLR9， MyD88， and NF-κB p65 in the kidney and spleen （P<0.05）. ConclusionTGP may inhibit the expression of downstream inflammatory factors to regulate immunity and resist SLE-induced renal injury by regulating the TLR9/MyD88/NF-κB signaling pathway.

Expression changes of TLR-9 and MyD88 in retina of mice following optic nerve injury / 国际眼科杂志(Guoji Yanke Zazhi)

@#AIM: To investigate the expression changes of Toll like recepter 9(TLR-9)and myeloid differentiation factor 88(MyD88)in retina of mice following optic nerve injury(ONI).<p>METHODS: There were 36 male 8-week-old C57BL/6J mice randomly divided into 6 groups: blank control(no treatment), ONI 1d group(materials were taken at 1d after optic nerve injury), ONI 3d group(materials were taken at 3d after optic nerve injury), ONI 5d group(materials were taken at 5d after optic nerve injury), ONI 7d group(materials were taken at 7d after optic nerve injury), ONI 14d group(materials were taken at 14d after optic nerve injury). The mice optic nerve model was made by optic nerve gripping, and the mRNA and protein levels of Toll like recepter 9 and myeloid differentiation factor 88 in each retinal were measured by RT-qPCR and Western-blot.<p>RESULTS: The mRNA and protein levels of Toll like recepter 9 and myeloid differentiation factor 88 in the retina of ONI 1d group were not significantly different from those of the blank control group(<i>P</i>>0.05), the mRNA and protein levels of TLR-9 and MyD88 in the retina of ONI 3d group, ONI 5d group, ONI 7d group and ONI 14d group were significantly increased compared with the blank control group, and the differences were statistically significant(<i>P</i><0.01). Compared with the blank control group, the mRNA and protein levels of TLR-9 and MyD88 in the retina of mice began to increase at ONI 3d(<i>P</i><0.01), peaked at ONI 5d(<i>P</i><0.001), and gradually decreased at ONI 7d(<i>P</i><0.01).<p>CONCLUSION: Optic nerve injury can activate the expression of TLR-9 and MyD88 in mice retina. TLR-9 and MyD88 may play an essential role in the process of optic nerve injury.

The role of Toll-like receptors in chemoradiotherapy-induced gastrointestinal mucositis / 口腔疾病防治

@#Mucositis is a common gastrointestinal complication in cancer patients undergoing chemoradiotherapy, including oral mucositis and gastrointestinal mucositis, with clinical manifestations of oral ulcers, vomiting, diarrhea and pain that seriously reduce the quality of life of patients and even affect anticancer therapy. Toll-like receptor (TLR) are important receptors involved in innate immunity and in the development of chemoradiation-induced mucositis by mediating the effect between microorganisms and the host. A comprehensive understanding of the role of TLR in mucositis is helpful to guide the prevention and treatment of mucositis. This paper reviews the available studies on TLR and mucositis. The results of the literature review indicate that different TLR have different roles in chemoradiation-induced mucositis: TLR2 is an important receptor in the inflammatory cascade of chemoradiation-induced mucositis; TLR4 activation can increase gastrointestinal mucosal inflammation and lead to oral epithelial ulceration; TLR5 agonists can reduce the degree of radiation-induced mucositis damage; and antagonizing or knocking out TLR9 can reduce chemoradiation-induced gastrointestinal mucositis. However, no TLR agonists or inhibitors have yet been applied in clinical practice, and additional studies are needed to explore the role of different TLR in mucositis in the future to provide a reference for the precise prevention and treatment of chemoradiation-induced mucositis.

Construction of TLR9 gene knockout mice with preliminary phenotype identification / 中国药理学通报

Aim To construct TLR9 gene knockout mouse model and preliminarily identify the phenotypes. Methods The TLR9 knockout mice were established by CRISPR/Cas9 system. The genotype of knockout mice was identified by capillary gel electrophoresis, while the PCR product was sequenced to analyse the knockout efficiency. The mRNA and protein expression level of TLR9 were determined by qRT-PCR, Western blot and immunohistochemistry. Genetic traits, body weight and blood routine changes were also measured. Pathological changes of mouse tissues were observed by hematoxylin-eosin (HE) staining. Results PCR and sequencing results showed that the stable TLR9 gene knockout mice were generated. The TLR9 gene expres-sions of knockout mice in spleen and liver tissues were significantly lower than those of wild-type mice. The growth and breeding of TLR9 knockout mice were normal , as well as all indexes of body weight and peripheral blood routine. The histomorphological characteristics of TLR9 knockout mice showed no significant difference compare to wild-type mice. Conclusions The TLR9 knockout mice are successfully established, providing an experimental animal model for studying the biological functions and regulatory mechanisms of the TLR9.

Role of TLR 9 expression in maternal peripheral blood and placenta in intrauterine transmission of HBV / 中华流行病学杂志

Objective@#To explore the role of TLR 9 in intrauterine transmission of hepatitis B virus (HBV) through blood pathway and placenta.@*Methods@#Epidemiological investigation was carried out in 290 HBsAg positive parturients and 45 normal parturients (control group) in Northwest Women and Children Hospital of Shaanxi Province. Enzyme-linked immunosorbent assay (ELISA) was used to detect five serological makers of hepatitis B and TLR 9 levels in peripheral blood of pregnant women and newborns. HBV DNA was detected by real-time fluorescence quantitative PCR. Detection of TLR 9 expression in placenta by immunohistochemical method. A case-control study was conducted to analyze the difference of TLR 9 levels in placenta and peripheral blood of HBsAg- positive pregnant women with intrauterine transmission of HBV.@*Results@#The incidence of dominant HBV infection (DBI), occult HBV infection (OBI) and intrauterine transmission of HBV were 9.28% (27/291), 40.21% (117/291) and 49.48% (144/291) respectively. (1) The level of TLR 9 in peripheral blood of HBsAg-positive parturients, non-HBV intrauterine transmission (NBIT) group and OBI group were significantly lower than that of control group (P<0.001). The level of TLR 9 in DBI group was significantly higher than those in NBIT group and OBI group (P=0.000). (2) The TLR 9 level in HBeAg-negative group was significantly lower than that in HBeAg-positive parturients in OBI group (P=0.01). (3) With the increased severity of intrauterine transmission of HBV in each HBV DNA load group, the TLR 9 level in maternal peripheral blood increased significantly (P<0.05). (4) With the increased severity of intrauterine transmission of HBV, the levels of TLR 9 increased significantly in antiviral therapy, immunoglobulin injection and non-hepatitis B vaccine groups (P<0.05). (5) The expression of TLR 9 in placenta tissues with DBI group was significantly higher than that in OBI group and NBIT group (P<0.05).@*Conclusions@#HBV can inhibit the secretion of TLR 9 in parturient to some extent, but HBeAg can stimulate the secretion of TLR 9. However, with the increased severity of intrauterine transmission of HBV, the level of TLR 9 in parturients is increased by intra-group cross-differentiation. Therefore, TLR 9 is not an independent marker for screening and grouping, but it can be used as an reference indicator for the monitoring and management of HBsAg-positive parturients.

Expressions and correlations of TLR2 ,TLR4 ,TLR7 and TLR9 in eosinophil-enriched blood cells from patients with allergic rhinitis / 西安交通大学学报(医学版)

Objective To investigate expressions and correlations of TLR2,TLR4,TLR7 and TLR9 in eosinophil-enriched cell populations from patients with allergic rhinitis (AR),and elucidate their roles in AR. Methods Peripheral venous blood samples were collected from healthy controls (HCs)and AR patients,and then incubated with crude extracts of Artemisia pollen,dust mite,and Platanus pollen,respectively.Levels of TLR2 , TLR4,TLR7 and TLR9 in blood eosinophil-enriched cells were detected by flow cytometry.Correlations between TLR2+,TLR4+,TLR7+and TLR9+eosinophils were analyzed by SPSS.Results Levels of TLR2+eosinophils from patients with AR were reduced by 4%,mean fluorescence intensity (MFI)of TLR4+eosinophil was elevated by 20%,and TLR7+eosinophils increased up to 4.8 folds compared with HCs when cultured with medium only (P＜0.05).Artemisia pollen extracts induced approximately 7 .8 % of increase in TLR2+eosinophils from AR patients.In addition,correlations between TLR2+and TLR4+eosinophils,TLR2+and TLR7+eosinophils,and TLR7+and TLR9+eosinophils were -0.670 (P＜0.01),-0.430 (P＜0.05)and 0.446 (P＜0.05),respectively. However,allergens had few effects on TLR2,TLR4,TLR7 and TLR9 expressions in HCs.Conclusion Eosinophil-derived TLR2 ,TLR4 and TLR7 are likely to play a key role in AR.TLR2 ,TLR4 and TLR7 might become the potential targets for AR treatment.

Toll-like receptors in B cell activation and STAT3 signal pathway / 国际儿科学杂志

Toll-like receptors(TLR) are widely exist in antigen presenting cells as a kind of pattem recognition receptor involved in the recognition of molecular structures specific for microbial pathogens,and have an important effect on innate and adaptive immune responses.There are 10 types of TLR in human beings which can be grouped into two main categories:cell surface receptors and receptors localized in the endosome.Among all the TLRs,TLR7 and TLR9,which expressed widely in human B cells,are important to B cells' functions.Two signal transduction pathways of TLR including MyD88 pathway and non-MyD88 pathway associate with B cells activation,proliferation,differentiation and antibody secretion.TLR7 and TLR9's stimulus,such as CpG-containing oligodeoxynucleotides(ODN) can activate memory B cells and naive B cells,promote cells proliferation,plasma cells generation,cytokine secretion and protect them from apoptosis.More than that,TLR7 and TLR9 can stimulate the production of IgG and IgM,make antibody shift to IgG2a and block the production of IgG1 and IgE.Interestingly,TLR-MyD88 pathway is recently confirmed to have connection with STAT3 signal pathway.However,the mutation of STAT3 will cause autosomal dominant hyper-IgE syndrome (HIES),a primary immunodeficiency characterized by elevated IgE levels,eczema,recurrent infections,pneumonia as well as multi-system symptoms.Therefore,the pathogenesis of HIES maybe related to the inhibition of TLR-MyD88-STAT3 signal pathway.As a result,we write a review about TLR in B cell activation and STAT3 signal pathway,and discuss their connection between HIES.

Protective effect of CpG-ODN conditioning on hypoxic/ischemicbrain damage in neonatal rats / 中国药理学通报

Aim To study the therapeutic effect of CpG-ODN, an agonist of Toll-like receptor 9 (TLR9), on hypoxic/ischemic encephapathy in neonatal rats and investigate the mechanisms.Methods Fifty healthy 7-day-old neonatal Wistar rats (in either gender, weighing 12~17g) were randomly divided into sham operation group, HIBD group, and CpG-ODN low group(0.35 mL·kg-1), CpG-ODN middle group(1.40 mL·kg-1), CpG-ODN high group(5.60 mL·kg-1).The neurological function was scored after 48h operation;ten rats of each group was executed respectively and brains tissue was taken;HE staining was used to observe the brain pathological changes.Western blot assay was used to detect the expressions of TLR9 and phosphor-p38 mitogen-activated protein kinases(p-p38 MAPK), and enzyme linked immunosorbent assay (ELISA) method was adopted to detect TNF-α expression.Results The CpG-ODN low, middle group were improved in impairment significantly compared with the HIBD group, and the brain pathological change was lessened, while the CpG-ODN high group was impaired significantly compared with the HIBD group (P<0.05), and brain pathological change was sharpened.Western blot showed the up-regulation in TLR9 and p-p38 MAPK and a significant increase of the expression of TNF-α in the brain tissue in CpG-ODN group with statistical difference in HIBD group and sham operation group(P<0.05).Conclusions The neuro-behavioral score and nervous system function can be improved and the hypoxic/ischemic brain damage can be reduced in neonatal rats in the CpG-ODN low, middle group.The protective mechanisms may be suitably via activating p38 MAPK signaling pathway to promote p38 MAPK phosphory1ation and up-regulation of the expression of TNF-α in the brain tissue of rats.

Study on the relationship between the expression of toll-like receptor 9-mRNA and inflammatory factors in patients with acute pancreatitis / 临床外科杂志

Objective To investigate the correlation between the expression of Toll-like receptor 9(TLR9)-mRNA and inflammatory factors in patients with acute pancreatitis(AP).Methods A total of 40 patients with AP diagnosed in our hospital from February 2016 to September 2016 in 24 h were enrolled.EDTAK2 anticoagulant venous blood was collected on day 1,3,5.And pancreatic elastase,proinflammatory cytokines and anti-inflammatory cytokines were detected.Peripheral blood monouclear cells were isolated and the mRNA expression of TLR9 was tested by polymerase chain reaction.The change of TLR9-mRNA in 5 days was analyzed.Results The level of TLR9-mRNA in peripheral blood mononuclear cells of patients with AP at 1,3,5 days were significantly higher than that in the control group,and the difference was statistically significant (P ＜ 0.05).Changes of the TLR9-mRNA expression in peripheral blood mononuclear cells in patients with AP was positively correlated with the levels of pancreatic elastase and proinflammatory cytokines,and the difference was statistically significant (P ＜ 0.05).Conclusion The expression of TLR9-mRNA in peripheral blood mononuclear macrophages was closely related to the up-regulation of inflammatory cytokines in AP patients,suggesting that TLR9 may mediate the occurrence and development of AP.To investigate the correlation between the expression of TLR9-mTLR9-mRNA and inflammatory factors in patients with AP.Methods A total of 40 patients with AP diagnosed in our hospital from February 2016 to September 2016 in 24h were selected.EDTAK2 anticoagulant venous blood was collected on day 1,3,5.And Pancreatic elastase,proinflammatory cytokines and anti-inflammatory cytokines were also detected.EDTAK2 anticoagulant venous blood was collected,tested the above indicators and used them as the benchmark level of the corresponding indicators.Results The levels of TLR9-mRNA in peripheral blood mononuclear cells of patients with AP at 1,3,5 days were significantly higher than in the control group,and the difference was statistically significant (P ＜ 0.05).Changes of TLR9-mRNA expression in peripheral blood mononuclear cells in patients with AP was positively correlated with pancreatic elastase and proinflammatory cytokine,and the difference was statistically significant (P ＜ 0.05).Conclusion The expression of TLR9-mRNA in peripheral blood mononuclear macrophages was closely related to the up regulation of inflammatory cytokines in AP patients,suggesting that TLR9 may mediate the occurrence and development of AP.

Research on influence of TLR9 on pancreatic cancer nude mouse model growth and resistance to chemotherapy / 重庆医学

Objective To observe the effects of TLR9 on the nude mouse transplanted tumor growth of human pancreanc cancer and its drug resistance.Methods The nude mouse transplated tumor of human pancreatic cancer PANC-1 was established and randomly divided into 6 groups for conducting the experiment:sterile normal saline group,TLR9 agonist,TLR9 inhibitor group,gemcitabine group,TLR9 inhibitor plus gemcitabine Bin group,TLR9 agonist plus gemcitabine.The tumor size and growth situation were recorded by the vernier caliper.The immunohistochemical method was used to detect tumor TLR9 receptor expression.The tumor growth,metastasis and paracancerous nssue invasion situation were observed by the magnetic resonance imaging (MRI).Results The volume and growth speed of resected tumor mass in the gemcitabine group,TLR9 agonist + gemcitabine group,TLR9 inhibitor plus gemcitabine group was significantly smaller than those in other groups (P＜0.05),which in the TLR9 agonist + gemcitabine group were significantly greater than those in the TLR9 inhibitor plus gemcitabine group and gemcitabine group (P＜0.05),the difference between the TLR9 inhibitor plus gemcitabine group and gemcitabine group had statistical significance (P＜0.05),while the difference among the TLR9 agonist group,TLR9 inhibitor group and normal saline group had no stastistical significance (P＞0.05).The tumor in mice at 7 weeks after planting showed oval shape with clear boundary by MRI observation,no obvious metastais and paracancerous invasion were seen in paracancerous nssues no statistically significant,5 weeks,6 weeks after planting,seven weeks mice observed in MRI,the tumor into an,state clearly that the transfer of the surrounding tissue,no significant vascular invasion,heart,liver,kidney disease.The TLR9 expression on the surface of tumor tissue was detected and identified.Conclusion Pancreatic cancer nude mouse transplated tumor has definitely positive expression of TLR9,TLR9 activation can significantly decrease the sensitivity of pancreatic cancer to chemotherapy,increases the drug resistance of tumor,on contrary promotes the tumor growth.

Rab5a promotes expression of pro-inflammatory cytokines and type Ⅰ IFN in CpG induced macrophages / 中国免疫学杂志

Objective:Using the macrophage cell lines RAW264.7 stably expressing Rab5a and its dominant negative mutant Rab5aN133I as models to analyze the effect and the mechanism of Rab 5a,Rab5aN133I on CpG-induced production of pro-inflammatory cytokines and type Ⅰ IFN.Methods: The eukaryotic expression vectors of Rab5a and Rab5aN133I were transfected into RAW264.7 cells by liposome,and screened with G418.The G418-resistant colonies were obtained and amplified.The transformed cell lines were i-dentified by RT-PCR,Real time-PCR and Western blot.The production of cytokines were measured after transformed cell lines of Rab5a and Rab5aN133I was stimulation with CpG for 8 h.Results: Rab5a expression in transfected cells was significantly higher than the control cell group (P<0.05).Overexpression of Rab5a significantly promoted the production of TNF -α,IL1-β(P<0.01) and IFN-β( P<0.05) in CpG stimulated RAW264.7.The production of cytokines was restored in Rab 5aN133I transfected cell line.Conclusion:Rab 5a promotes CpG-induced pro-inflammatory cytokines and typeⅠIFN in macrophages,it may be act as a positive regulator in TLR9 signaling pathway.

Evaluation of Toll-like, chemokine, and integrin receptors on monocytes and neutrophils from peripheral blood of septic patients and their correlation with clinical outcomes

Recognition of pathogens is performed by specific receptors in cells of the innate immune system, which may undergo modulation during the continuum of clinical manifestations of sepsis. Monocytes and neutrophils play a key role in host defense by sensing and destroying microorganisms. This study aimed to evaluate the expression of CD14 receptors on monocytes; CD66b and CXCR2 receptors on neutrophils; and TLR2, TLR4, TLR5, TLR9, and CD11b receptors on both cell types of septic patients. Seventy-seven septic patients (SP) and 40 healthy volunteers (HV) were included in the study, and blood samples were collected on day zero (D0) and after 7 days of therapy (D7). Evaluation of the cellular receptors was carried out by flow cytometry. Expression of CD14 on monocytes and of CD11b and CXCR2 on neutrophils from SP was lower than that from HV. Conversely, expression of TLR5 on monocytes and neutrophils was higher in SP compared with HV. Expression of TLR2 on the surface of neutrophils and that of TLR5 on monocytes and neutrophils of SP was lower at D7 than at D0. In addition, SP who survived showed reduced expression of TLR2 and TLR4 on the surface of neutrophils at D7 compared to D0. Expression of CXCR2 for surviving patients was higher at follow-up compared to baseline. We conclude that expression of recognition and cell signaling receptors is differentially regulated between SP and HV depending on the receptor being evaluated.

Effects total glucosides paeony on downregulating the expression ERKI/2, TLR4 and TRL9 in rats with nonalcoholic fatty liver disease / 中国药学杂志

OBJECTIVE: To investigate total glucosides paeony (TGP) on the expression phosphorylated extracellelar signal regulated kinase 1/2(p-ERK1/2), Toll-like receptors 4(TLR4) and Toll-like receptors 9(TLR9) in rats with nonalcoholic fatty liver disease (NAFLD) induced by fructose and high-fat feed. METHODS: SD rats were divided into normal group and test groups. The rats in test groups were fed with fructose and high-fat feed for 10 weeks totally to induce the test model. After 6 weeks the model was established, the rats were divided into four groups randomly, the model group (NAFLD), the metformin group (Met, 200 mg · kg-1), the low-dose TGP group(TGP-L, 100 mg · kg-1) and the high-dose TGP group(TGP-H, 200 mg · kg-1) (n=10). Four weeks later all the rats were killed and checked the indexes such as serum fasting blood glucose(FBG), fasting insulin(Fins), insulin sensitivity index (ISI), total cholesterol (TC), low-density lipoprotein (LDL-C), high-density lipoprotein (HDL-C), triglyceride (TG), free fatty acids (FFA), alanine aminotransferase (ALT), aspartate aminotransferase (AST), glutathione S-transferase (GST) and liver index. The expression p-ERK1/2, TLR4 and TLR9 were inspected by Western-blot. RESULTS: Compared with the normal group, the rats in test groups were with the high levels serum FBG, insulin, TC, LDL-C, TG, FFA, ALT, AST, GST, liver index, p-ERK1/2, TLR4 and TLR9 (P 0.05). CONCLUSION: By downregulating the expression ERK1/2, TLR4 and TRL9, TGP can improve abnormal glucose and lipid metabolism and insulin resistance, enhance insulin sensitivity and ameliorate liver function in rats with NAFLD induced by fructose and high-fat feed.

Single Nucleotide Polymorphisms of Toll-Like Receptor 7 and Toll-Like Receptor 9 in Hepatitis C Virus Infection Patients from Central China

PURPOSE: To analyze the correlation of polymorphisms of toll-like receptor 7 (TLR7) (rs179009) and toll-like receptor 9 (TLR9) (rs187084) in hepatitis C virus (HCV) infections in the Han population. MATERIALS AND METHODS: The genotypes of TLR7IVS2-151 in HCV infection were detected by Sanger sequencing using polymerase chain reaction-restriction fragment length polymorphism to determine the TLR9 T-1486C single nucleotide polymorphisms (SNP) for all enrolled patients. RESULTS: We found no significant difference between males with spontaneous clearance of HCV versus those chronically infected [chi2=2.71, p=0.10, odd ratios (OR)=0.58, 95% confidence interval (CI) 0.31-1.11]. However, significant differences were found for the distribution of TLR7 (rs179009) in females (chi2=9.46, p=0.01). In females, a significant difference was also found between chronic hepatitis C and those with spontaneous clearance of HCV in terms of TLR7 IVS2-151G/A allele frequencies (chi2=9.50, p=0.00, OR=0.46, 95% CI 0.28-0.75). In HCV-infected patients, no significant association was found between the frequency of TLR9 genotypes and alleles. CONCLUSION: The site of TLR7 IVS2-151 (rs179009) G/A may be a factor for susceptibility of chronic HCV in the female Han population. TLR9T-1486C (rs18084) SNP may not play a major role in HCV infection. However, individual risk profiles for HCV infection did vary by sex and this relationship should be further investigated.

Effect of PvMSP1 on differentiation,maturation and function of dendritic cells / 中国血吸虫病防治杂志

Objective To investigate the effects of Plasmodium vivax merozoite surface protein 1(PvMSP1)on differentia-tion,maturation and function of dendritic cells(DC)and the mechanisms of PvMSP1 on the activation of DC via toll like receptors (TLR). Methods DCs were incubated with different doses of PvMSP1(1.0,10.0,100.0μg/ml)in vitro. The changes of CD83, CD86,and HLA-DR on DC were detected by flow cytometry(FCM);the expressions of cytokine IL-10 and IL-12 of DC were mea-sured by ELISA;the expressions of TLR4 and TLR9 mRNA of DC were measured by RT-PCR;the proliferation induction to autol-ogous lymphocytes of DC was measured by MTT. Meanwhile,the untreated DC and LPS inducing DC were as the negative control and positive control,respectively. All the data were analyzed statistically. Results Compared with the untreated DC,the propor-tions of CD83,CD86 and HLA-DR on DC induced by LPS and PvMSP1 increased significantly(all P0.05). In the PvMSP1-treated group,the DC TLR4 mRNA production increased(P0.05);DC stimulated the proliferation of autologous lympho-cytes. Conclusion PvMSP1 enhances DC differentiation and maturation,and the mature DC induced by PvMSP1 has the ability of antigen presenting. The route for PvMSP1 inducing DC maturation might be TLR4 pathway rather than TLR9 pathway.

Kinetic Analysis of CpG-Induced Mouse B Cell Growth and Ig Production

Immune cells express toll-like receptors (TLRs) and respond to molecular patterns of various pathogens. CpG motif in bacterial DNA activates innate and acquired immune systems through binding to TLR9 of immune cells. Several studies reported that CpG can directly regulate B cell activation, differentiation, and Ig production. However, the role of CpG in B cell growth and Ig production is not fully understood. In this study, we analyzed the effect of CpG on the kinetics of mouse B cell viability, proliferation, and Igs production. Overall, CpG enhanced mouse B cell growth and production of Igs in a dose-dependent manner. Unlike LPS, 100 nM CpG (high dose) did not support TGF-beta1-induced IgA and IgG2b production. Moreover, 100 nM CpG treatment abrogated either LPS-induced IgM or LPS/TGF-beta1-induced IgA and IgG2b production, although B cell growth was enhanced by CpG under the same culture conditions. We subsequently found that 10 nM CpG (low dose) is sufficient for B cell growth. Again, 10 nM CpG did not support TGF-beta1-induced IgA production but, interestingly enough, supported RA-induced IgA production. Further, 10 nM CpG, unlike 100 nM, neither abrogated the LPS/TGF-beta1-nor the LPS/RA-induced IgA production. Taken together, these results suggest that dose of CpG is critical in B cell growth and Igs production and the optimal dose of CpG cooperates with LPS in B cell activation and differentiation toward Igs production.

The Role of Plasmacytoid and Myeloid Dendritic Cells in Induction of Asthma in a Mouse Model and the Effect of a TLR9 Agonist on Dendritic Cells

PURPOSE: To determine the role of plasmacytoid dendritic cells (pDC) and myeloid dendritic cells (mDC) in priming effector T cells to induce allergy, and to evaluate the effect of immunostimulatory sequences (ISS, TLR9 agonist) on dendritic cells. METHODS: Cultured mDC and pDC with/without ISS were injected intratracheally into sensitized Balb/C mice. Mice were sacrificed, and then pulmonary function tests, bronchoalveolar lavage (BAL), cell counts, and cytokine levels were evaluated. Migration of dendritic cells was also evaluated after ISS administration. RESULTS: In mice injected with mDC, airway hyperresponsiveness, eosinophil counts, and Th2 cytokine levels in BAL increased with increasing numbers of mDC injected. However, in mice injected with pDC, none of these changed, suggesting poor priming of T cells by pDC. In addition, mDC pulsed with ISS inhibited asthmatic reactions, and ISS administration inhibited migration of DC to the lung. CONCLUSIONS: We suggest that pDC played a limited role in priming T cells in this asthma model and that mDC played a major role in inducing asthma. In addition, ISS inhibited migration of DC to the lung.

Derp2-mutant gene vaccine inhibits airway inflammation and up-regulates Toll-like receptor 9 in an allergic asthmatic mouse model.

Background: A DNA vaccine encoding the whole segment of the Derp2 allergen could prevent allergic airway inflammation in a Derp2 allergen-induced allergic airway inflammation mouse model. Objective: This study investigated the effect of DNA vaccine encoding Derp2-mutant gene in which an IgE epitope was deleted on airway inflammation and the role of TLR9 in the asthmatic mouse model. Methods: A Derp2-mutant DNA vaccine was constructed. Mice were immunized, sensitized and challenged. Airway inflammation, airway hyper reactivity (AHR) and serum antibody were tested. The expression of Toll like receptor9 (TLR9) was detected with western-blot and immunehistochemistry. Results: We demonstrated that the Derp2-mutant-DNA induced IgG2a and inhibited IgE production, inhibited airway allergenic inflammation and AHR. Derp2-mutant-DNA vaccine induce TLR9 expression in lung tissue. Conclusions: The data indicate that allergen DNA vaccine deleted IgE epitope could prevent allergenic airway inflammation, AHR, and upregulate lung TLR9 express.

Advances in DNA recognition receptors / 中国肿瘤生物治疗杂志

DNA, as the material basis of all living cells, triggers innate immune responses through TLR9 and other cytosolic recognition receptors. In recent years, the research progress of TLR9 is mainly manifested by the following four aspects: (1) the determinants of TLR9 interacting with its ligands; (2) the mechanisms and the importance of TLR9 translocation from the endoplasmic reticulum to the endosome; (3) the roles of the endosomal acidification and maturation, and subsequent TLR9 cleavage in TLR9 signal transduction pathway; and (4) the possible mechanisms by which the organism distinguish self DNA from microbial DNA. Meanwhile, a series of experiments on TLR9 antagonists and TLR9 deficient mice confirmed the presence of TLR9-independent cytosolic DNA sensors. So far, three TLR9-independent DNA sensors have been found, and they are DAI, AIM2, and RNA polymerase Ⅲ.

El ADN de Leishmania mexicana activa al macrófago murino e induce aumento en la expresión de TLR9 / Leishmania mexicana DNA activates murine macrophages and increases their TLR9 expression

Antecedentes: Los macrófagos son células de la respuesta inmune que reconocen patrones moleculares asociados a patógenos (PAMP) mediante receptores presentes en la superficie de la célula como en compartimentos intracelulares, como los TLR (toll like receptors). Distintos TLR reconocen ligandos que comparten múltiples patógenos. La unión de TLR con su ligando desencadena una cascada de señalización que termina en la producción de citocinas y moléculas coestimuladoras a través de la translocación de NF-κB al núcleo. Nuestro grupo demostró que el lipofosfoglucano de Leishmania es un ligando de TLR2 que activa células NK. Schieicher y cols.12 informo recientemente la activación de células dendríticas plasmacitoides con ADN genómico de Leishmania infantum a través de TLR9, con alta producción de IFN tipo I. Objetivo: En el presente trabajo exploramos si el ADN de Leishmania mexicana contiene motivos CpG no metilados capaces de activar al macrófago murino derivado de médula ósea, como ha sido descrito anteriormente para motivos CpG no metilados de ADN bacteriano. Resultados y conclusiones: Encontramos que el ADN de Leishmania mexicana posee motivos CpG no metilados que activan macrófagos murinos de la cepa BALB/c, llevando a la producción de citocinas proinflamatorias como TNFα e IL12P40 y a la sobreexpresión del mARN de TLR9.

BACKGROUND: Macrophages are immune system cells that recognize pathogen associated molecular patterns (PAMPs) through receptors that can be located on the cell membrane or in intracellular compartments, such as the TLR (toll like receptors). Different TLRs bind to ligands shared among multiple pathogens. The binding of ligands to TLRs induces a signaling cascade that leads to cytokine and co-stimulatory molecule production due to the nuclear translocation of NF-kappaB. We demonstrated that Leishmania lipophosphoglycan (LPG) is a ligand for TLR2, leading to NK-cell activation. Schieicher et al. recently reported that genomic DNA from Leishmania infantum activates plasmacitoid dendritic cells through TLR9, leading to IFN type I production. OBJECTIVE: In the present study we explored wether Leishmania mexicana DNA contained non-methylated CpG motifs able to activate murine bone marrow derived macrophages, as previously described for bacterial DNA containing CpG motifs. RESULTS AND CONCLUSIONS: We observed that Leishmania mexicana DNA contains non-methylated CpG morifs able ofactivating murine bone marrow derived macrophages, leading to the production of proinflammatory cytokines such as TNFalpha and IL- 12(P40) as well as the over expression of mRNA for TLR9.