Association of Toll-like receptor 7, CTLA-4 gene polymorphisms with severe asthma / 公共卫生与预防医学

Objective To explore the association of Toll-like receptor 7, CTLA-4 gene polymorphisms and severe asthma. Methods From February 2018 to March 2020, 175 asthma patients admitted to the respiratory department of our hospital were selected as the research subjects (109 cases of mild disease and 66 cases of severe disease), and 248 cases of healthy people who were included in the outpatient physical examination of our hospital during the same period were selected as the normal control group. Toll-like receptor 7 and CTLA-4 gene polymorphisms in the above groups were determined, and the relationship between Toll-like receptor 7 and CTLA-4 polymorphisms and severe asthma was evaluated by calculating the odds ratio (OR) and 95% confidence interval(CI). The relationship between the genotypes of Toll-like receptor 7 and CTLA-4 polymorphisms and severe asthma were evaluated by logistic regression analysis. Results The proportion of TLR7 rs3853839 CC genotype, CTLA-4 rs231725 AA genotype, TLR7 rs3853839 C allele frequency and CTLA-4 rs231725 A allele frequency in severe asthma group and mild asthma group were higher than those in normal control group(P<0.05). The proportion of TLR7 rs3853839 CC genotype, the proportion of CTLA-4 rs231725 AA genotype, the frequency of TLR7 rs3853839 C allele, and the frequency of CTLA-4 rs231725 A allele in the severe asthma group were higher than those in the mild asthma group(P<0.05). TLR7 rs3853839 CC genotype (OR=10.32, 95%CI=5.59-23.89), CTLA-4 rs231725 AA genotype (OR=13.21, 95%CI=3.58-20.25), TLR7 rs3853839 C allele frequency (OR=11.32, 95% CI=4.25-21.14) and CTLA-4 rs231725 A allele frequency (OR=13.24, 95% CI=6.59-20.21) could increase the susceptibility to severe asthma(P<0.05). TLR7 rs3853839CC genotype, TLR7 rs3853839C allele frequency, CTLA-4 rs231725AA genotype and CTLA-4 rs231725A allele frequency were risk factors for severe asthma(P<0.05). Conclusion TLR7 rs3853839 CC genotype, TLR7 rs3853839 C allele frequency, CTLA-4 rs231725 AA genotype and CTLA-4 rs231725 A allele frequency are associated with the occurrence of severe asthma.

TLR7 mediates increased vulnerability to ischemic acute kidney injury in diabetes

SUMMARY OBJECTIVE Diabetes is a risk factor for acute kidney injury (AKI). However, its mechanism of pathogenesis has not been elucidated. The aim of the study was to investigate the role of inflammation and the toll-like receptor 7 (TLR7) in ischemic AKI for diabetes. METHODS A high glucose hypoxia-reoxygenation model of human renal tubular epithelial (HK-2) cells was used to generate AKI induced by ischemia-reperfusion in diabetes. The activity of cells was measured by CCK-8 assay and LDH activity. Inflammatory cytokines were assessed by ELISA. TLR7, MyD88, and NF-κB expressions were examined by western blotting. Apoptosis was evaluated by flow cytometry. RESULTS The high glucose group and low glucose group were subjected to hypoxia-reoxygenation. The low glucose group developed only mild cell damage, apoptosis, and inflammatory response. In contrast, an equivalent hypoxia-reoxygenation injury provoked severe cell damage, apoptosis, and inflammatory response in the high glucose group. Expression of TLR7 and its related proteins were measured in the high glucose group before and after hypoxia-reoxygenation. The high glucose group exhibited more significant increases in TLR7 expression following hypoxia-reoxygenation than the low glucose group. In addition, the expression of TLR7 and its related proteins after hypoxia-reoxygenation were higher in the high glucose group than in the low glucose group. Inhibition of TLR7 provides significant protection against ischemic injury in diabetes. CONCLUSION Our results suggest that diabetes increases the vulnerability to ischemia-induced renal injury. This increased vulnerability originates from a heightened inflammatory response involving the TLR7 signal transduction pathway.

RESUMO OBJETIVO O diabetes é um fator de risco para a lesão renal aguda (LRA). No entanto, seu mecanismo de patogênese não foi elucidado. O objetivo do estudo foi investigar o papel da inflamação e do receptor Toll-like 7 (TLR7) na LRA isquêmica no diabetes. MÉTODOS Um modelo de hipóxia-reoxigenação de células epiteliais tubulares renais humanas (HK-2) na presença de concentrações altas de glicose foi utilizado para gerar LRA induzida por isquemia-reperfusão em diabetes. A atividade das células foi medida pelo ensaio Cell Counting Kit-8 (CCK-8) e pela atividade da lactato desidrogenase (LDH). As citocinas inflamatórias foram avaliadas por ensaio imunoenzimático (Elisa). A expressão de TLR7, do fator de diferenciação mieloide 88 (MyD88) e do fator de transcrição nuclear-κB (NF-κB) foi examinada por Western blotting. A apoptose foi avaliada por citometria de fluxo. RESULTADOS Os grupos glicose alta e glicose baixa foram submetidos à hipóxia-reoxigenação. O grupo de baixa glicose desenvolveu apenas danos celulares ligeiros, apoptose e uma resposta inflamatória. Em contraste, no grupo de alta glicose, uma lesão equivalente de hipóxia-reoxigenação provocou danos celulares graves, apoptose e uma resposta inflamatória. A expressão de TLR7 e suas proteínas relacionadas foi medida no grupo de alta glicose antes e após a hipóxia-reoxigenação. O grupo de alta glicose exibiu maiores aumentos na expressão de TLR7 após hipóxia-reoxigenação do que o grupo de baixa glicose. Além disso, a expressão de TLR7 e suas proteínas relacionadas após a hipóxia-reoxigenação foi maior no grupo com alto nível de glicose do que no grupo com baixo nível de glicose. A inibição do TLR7 fornece proteção significativa contra a lesão isquêmica no diabetes. CONCLUSÃO Nossos resultados sugerem que o diabetes aumenta a vulnerabilidade à lesão renal induzida por isquemia. Essa vulnerabilidade acrescida tem por origem uma resposta inflamatória aumentada envolvendo a via de transdução de sinal do TLR7.

The association between toll-like receptor 7 single nucleotide polymorphisms and severity of hand, foot and mouth disease caused by enterovirus 71 infection / 中华传染病杂志

Objective To explore the association between toll-like receptor 7 (TLR7) single nucleotide polymorphisms (SNP) and susceptibility of hand, foot and mouth disease caused by enterovirus (EV)71 infection.Methods The genotype of SNP (rs179019 and rs3853839) was determined in 775 EV7l-infected cases (including 439 mild cases and 336 severe cases) and 748 healthy control cases with TaqMan assay.The difference of allele frequencies was compared.The difference of TLR7 mRNA expression in peripheral blood mononuclear cells (PBMCs) derived from different SNP genotype carriers was detected.PBMCs derived from different SNP genotype carriers were stimulated by imiquimod and the TLR7-specific interferon-α(IFN-α) and interleukin-6 (IL-6) secretions were detected.Logistic regression was used to analyze for genotype frequency.Results The frequencies of rs3853839 genotype CC and CG in female patients of severe group were significantly higher than mild group (rs3853839 GC: OR=0.36,95%CI:0.14-0.82, P=0.01;rs3853839 CC: OR=0.19,95%CI:0.11-0.69,P=0.01).In addition, the frequency of rs3853839 genotype C in severe male group was significantly higher compared with that in mild group (OR=0.35,95%CI:0.19-0.63, P=0.01).Female carriers with rs3853839 genotype CC had significantly lower TLR7 mRNA expression than genotype GC and GG (CC vs GG: P=0.005;CC vs GC: P=0.016).Male carriers with rs3853839 genotype C also had significantly lower TLR7 mRNA expression than genotype G (C vs G: P=0.004).After stimulation of imiquimod, the expression of IFN-α (CC vs GG, P=0.001;CC vs GC: P=0.026) and IL-6 productions (CC vs GG: P=0.001;CC vs GC: P=0.011) were significantly lower in female carriers with rs3853839 genotype CC.The same patterns were observed in male carriers with rs3853839 genotype CC (IFN-α: P=0.003;IL-6: P=0.018).Conclusions The rs3853839C allele is the risk factor of severe infection of EV71, which may be due to specific cytokine profiles in rs3853839C allele carriers in children.

Research on Toll-like Receptor 7 Agonist in Liver Diseases / 胃肠病学

As an important protein molecule for innate immunity,Toll-like receptor 7 (TLR7) is also a bridge between innate immunity and adaptive immunity.When TLR7 is activated,it can exert its effect on liver diseases through different signaling pathways.Studies on the role and application of TLR7 agonist in liver diseases are increasing in recent years.This article reviewed the research on TLR7 agonist in liver diseases.

Expression of Toll-like receptors 7 and 9 as well as their regulatory molecules myeloid differentia-tion factor 88 and nuclear factor-κB in peripheral blood mononuclear cells of patients with vitiligo / 中华皮肤科杂志

Objective To compare expression of Toll?like receptors 7 and 9(TLR7, TLR9)as well as their regulatory molecules myeloid differentiation factor 88(MyD88)and nuclear factor?κB(NF?κB)in peripheral blood mononuclear cells(PBMCs)between patients with vitiligo and healthy individuals, and to explore their significance. Methods Flow cytometry was performed to measure expression of TLR7 and TLR9 in PBMCs among 36 patients with vitiligo and 22 healthy controls, and real?time fluorescence?based quantitative PCR(RT?PCR)was conducted to determine mRNA expression of MyD88 and NF?κB in the above blood samples. Results Compared with healthy controls, patients with vitiligo showed higher expression of TLR7 and mRNA expression of MyD88 and NF?κB, but lower expression of TLR9. However, significant differences were only observed in the mRNA expression of NF?κB(t=2.814, P=0.008), but not in the expression of TLR7 and TLR9 or the mRNA expression of MyD88 between patients and controls (t = 1.477, 1.761, 0.058, all P > 0.05). Conclusion NF?κB, as a key signaling molecule of TLR7 and TLR9 regulation pathways, increases obviously in patients with vitiligo, suggesting that NF?κB may be involved in the pathogenesis of vitiligo.

Toll-like receptor 7 involves the injury in acute kidney ischemia/reperfusion of STZ-induced diabetic rats

ABSTRACT PURPOSE: To determine whether Toll-like receptor 7 (TLR7) is the potential targets of prevention or progression in the renal ischemia/reperfusion (I/R) injury of STZ-induced diabetic rats. METHODS: Thirty six Sprague-Dawley rats were randomly arranged to the nondiabetic (ND) or diabetic group (DM), with each group further divided into sham (no I/R injury), I/R (ischemia-reperfusion) and CD (given by Chloroquine) group. Preoperatively, Chloroquine (40 mg/kg, intraperitoneal injection.) was administrated 6 days for treatment group. I/R animals were subjected to 25 min of bilateral renal ischemia. Renal function, histology, apoptosis, cytokines, expression of TLR7, MyD88 and NF-κB were detected. RESULTS: The serum levels of blood urea nitrogen, creatinine, IL-6 and TNF-α, apoptotic tubular epithelial cells, expression of TLR7, MyD88 and NF-κB were significantly increased in DM+I/R group, compared with ND+I/R group (p<0.05). All these changes were further improved by TLR7 inhibition Chloroquine except Paller scores (p<0.05). CONCLUSION: Toll-like receptor 7 inhibition attenuates the acute renal ischemia/reperfusion injury of STZ-induced diabetic in SD rats.

Effect of Toll-like receptor 7 agonist on JNK signaling pathway during liver injury in septic mice / 中华麻醉学杂志

Objective To evaluate the effect of Toll-like receptor 7 (TLR7) agonist on c-Jun Nterminal kinase (JNK) signaling pathway during liver injury in the septic mice.Methods One hundred eighty pathogen-free adult male C57BL/6 mice,aged 10-14 weeks,weighing 20-26 g,were randomly divided into 3 groups (n=60 each) using a random number table:sham operation group (group S);sepsis group (group Sep);TLR7 agonist group (group GDQ).Sepsis was induced by cecum ligation and puncture.In group GDQ,TLR7 agonist 1.5 g/kg was injected intraperitoneally at 24 h before establishment of the model.At 6,12 and 24 h after operation,10 mice in each group were sacrificed,and the livers were removed to detect the expression of interleukin-6 (IL-6) and IL-10 by Western blot.The expression of JNK was determined by immuno-histochemistry,and the histopathologic changes of livers were examined with a light microscope at 24 h after operation.The survival of mice was observed at 14 days after operation,and the 14-day survival rates were calculated.Results Compared with group S,the 14-day survival rates were significantly decreased,and the expression of IL-6,IL-10 and JNK was significantly up-regulated at 6,12 and 24 h after operation in Sep and GDQ groups (P＜0.05).Compared with group Sep,the 14-day survival rates were significantly increased,and the expression of IL-6,IL-10 and JNK was significantly down-regulated at 6,12 and 24 h after operation in group GDQ (P＜0.05).The pathological changes of livers were significantly attenuated in group GDQ as compared with group Sep.Conclusion TLR7 agonist can reduce the liver injury through blocking the JNK signaling pathway in the septic mice.

Investigation for anti-tumor effects of CIK cells induced by TLR7 agonist instead of IFN-γ / 中国免疫学杂志

Objective:To investigate the immune effects of CIK cells induced by Toll like receptor 7 agonist (Tlr7a) instead of IFN-γon killing lymphoma cells in vitro .Methods: Mononuclear cells were isolated from healthy human peripheral blood .CIK were induced by Tlr7a in vitro instead of IFN-γ.Two groups were divided as follows:CIK group,Tlr7a-CIK group.Then the main investigation on immune effects included immune phenotype was detected respectively , and cytotoxicity of the effectors was analyzed .Results: In Tlr7a-CIK group,the amount of CD56+cells was more than CIK group (P<0.05),and the cytotoxicity was also stronger (P<0.05). Conclusion:Tlr7a instead of IFN-γcould promote the immune effects of CIK cells on killing tumor cells in vitro .

JMJD3 participates in activation and apoptosis of IFN-αand TLR7-induced B cells / 中国免疫学杂志

Objective:To explore the effect of histone demethylase JMJD3 on B cell activation and apoptosis.Methods:B cells were sorted and purified from the peripheral blood of healthy people and SLE patients by using magnetic bead.After B cells were treated with IFN-αor R848 or IFN-α+R848,the percentages of CD86+B cells,CD69+B cells,CD86+Annexin V+B cells and CD69+Annexin V+B cells were detected by flow cytometry.The expression of JMJD3 was detected by Real Time PCR and Western blot.Results:The purity of sorted B cells was up to 95%.IFN-αenhanced both the activation and apoptosis and the JMJD3 expression of TLR7-activated B cells.The expression of JMJD3 was dependent on MAPK signal pathway,but not the NF-κB signaling pathway.Moreover,JMJD3 was highly expressed in B cells of peripheral blood from SLE patients compared to those from healthy people.Furthermore,JMJD3 inhibitors could inhibit the activation and apoptosis of IFN-αand R848 activated B cells.Conclusion:JMJD3 participated in the activation and apoptosis of IFN-αand TLR7-induced B cells, suggesting JMJD3 inhibitors may possess therapeutic effect for alleviating symptom of SLE.

mRNA expressions of Toll-like receptor 3, 7 and type Ⅰ interferons in peripheral blood of patients with chronic hepatitis C / 中华临床感染病杂志

Objective To investigate mRNA expressions of Toll-like receptors (TLR3 and TLR7)and type Ⅰ interferons (IFN-α and IFN-β) in peripheral blood mononuclear cells (PBMCs) of patients with chronic hepatitis C (CHC).Methods Thirty patients with CHC and 30 healthy controls were collected from Renmin Hospital of Wuhan University during September 2013 and May 2014.Liver function was detected using enzyme-linked immunosorbent assay (ELISA).mRNA expressions of TLR3,TLR7,IFN-α and IFN-β were detected by real-time quantitative PCR,and their relative expression values were obtained by 2-△△Ctmethod.The t test was performed for the comparison of mean differences between CHC patients and healthy controls,and Pearson analysis was used to analyze the correlations between TLR mRNA and IFN mRNA,liver function,HCV RNA load.Results Taking mRNA expressions in healthy control group as 1,the relative mRNA expressions of TLR3,TLR7,IFN-α and IFN-β in CHC group were 0.086,0.633,0.145 and 0.423 respectively (t =4.25,2.39,2.37 and 2.80,all P ＜ 0.01).Pearson correlation analysis showed that mRNA expressions of TLR3 and TLR7 were positively correlated with that of IFN-β (r =0.381 and 0.487,all P ＜ 0.05),but not correlated with IFN-α mRNA expression,HCV RNA load,ALT and AST (all P ＞ 0.05).Conclusion The mRNA expressions of TLR3,TLR7 and IFN-α,IFN-β decrease in CHC patients,and the mRNA expressions of TLR3 and TLR7 are positively correlated with that of IFN-β,indicating that increasing the expression of TLRs might be a new strategy in CHC treatment.

The TLR7 agonist Imiquimod promote the immunogenicity of msenchymal stem cells

BACKGROUND: Mesenchymal stem cells (MSCs) are considered the best candidate in stem cells therapy due to their multipotent differentiation ability, low expression of co-stimulatory molecules (CD80, CD86, CD34 and HLA-II) and immunosuppression effects on in vivo immune responses. MSCs were now widely used in clinical trials but received no encourage results. The major problem was the fate of engrafted MSCs in vivo could not be defined. Some studies indicated that MSCs could induce immune response and result in the damage and rejection of MSCs. As toll like receptors (TLRs) are important in inducing of immune responses, in this study we study the role of TLR7 in mediating the immune status of MSCs isolated from umbilical cord. RESULTS: Our results indicated that TLR7 agonist Imiquimod could increase the proliferation of PBMC isolated from healthy human volunteers and release of lactate dehydrogenase (LDH) in supernatant from PBMC-UCMSCs co-culture system. Flow cytometry and quantitative PCR also confirmed the regulated expression of surface co-stimulatory molecules and pro-inflammatory genes (IL-6, IL-8, IL-12, TGF-β and TNF-α). And the down-regulation expression of stem cell markers also confirmed the loss of stemness of UCMSCs. We also found that the osteo-differentiation ability of UCMSCs was enhanced in the presence of Imiquimod. CONCLUSION: To our knowledge, this is the first report that activation of TLR7 pathway increases the immunogenicity of UCMSCs. Extensive researches have now been conducted to study whether the change of immune status will be help in tumor rejection based on the tumor-tropism of MSCs.

Mast cells generate cysteinyl leukotrienes and interferon- as well as evince impaired IgE-dependent degranulation upon TLR7 engagement.

Mast cells are numerous at anatomical sites close to external environment, virtually at the portals of infection. A few data indicated that these cells express cytoplasmic Toll-like receptors (TLRs) recognizing virus-derived molecules. Accordingly, mast cells could participate in anti-viral defense or/and in viral-related diseases. However, data concerning the influence of viruses on mast cell activity are limited. Thus, the aim of our study was to determine mast cell response to TLR7 ligand, i.e. resiquimod (R848), a synthetic mimic of viral ssRNA. Since mast cells play a central role in allergic reactions the effect of TLR7 agonist was also investigated on FcεRI-dependent mast cell response. Experiments were carried out in vitro on freshly isolated fully mature rat peritoneal mast cells. Mast cells exhibit constitutive TLR7 molecule expression and its up-regulation after the agonist challenge. TLR7-mediated mast cell stimulation resulted in cysteinyl leukotriene (cysLT) and interferon (IFN)-β synthesis, whereas no histamine and CXCL8 secretion was stated. Moreover, mast cell priming with TLR7 ligand caused the reduction in anti-IgE-induced histamine release. The results suggest that ssRNA viruses could directly activate mast cells to alter their phenotype and to release of potent proinflammatory mediators or indirectly modulate IgE-dependent allergic processes.

The expression and clinical significance of toll-like receptor (TLR) 4 and TLR7 on renal tissue in children with pri-mary nephrotic syndrome / 临床儿科杂志

Objective To study the expression and clinical significance of toll-like receptor (TLR) 4 and TLR7 in primary nephrotic syndrome (PNS) of different pathological types in children. Methods Renal expressions of TLR4 and TLR7 were amined and analyzed retrospectively in renal biopsy specimens from 110 PNS patients and 21 healthy controls by immunohisto-chemical method. According to the renal pathologic classification of PNS, the TLR4 and TLR7 expression levels in PNS of dif-ferent types were compared. Results Compared with the renal tissue of healthy controls, the expression level of TLR4 on renal tissue of PNS patients was significantly increased (P<0.01). Among MN, MsPGN and FSGS, the highest expression of TLR4 was observed in MCD (P<0.01). Compared with the renal tissue of healthy controls, the expression level of TLR7 in re-nal tissue of PNS patients was also significantly increased (P<0.01). Among MCD, MN and FSGS, the highest expression of TLR7 was observed in MsPGN (P<0.01). Conclusions TLR4 and TLR7 expression levels are increased in renal tissue of PNS patients and the expression levels may be correlated with renal pathological types.

Association of TLR7 gene copy number variations and systemic lupus erythematosus in Han population / 中华风湿病学杂志

Objective To examine the variation in TLR7 gene copy number of patients with systemic lupus erythematosus (SLE) in Han population,and investigate the relationship between TLR7 gene copy number variations and clinical phenotypes of SLE.Methods Determination of gene copy number of TLR7 was achieved by AccuCopyTM multiple gene copy number detection method in 337 cases of Han SLE patients and 338 healthy controls.According to the clinical phenotype stratification,all cases were divided into lupus nephritis and non-lupus nephritis group,the hematological involvement and non-hematological involvement group,anti-Smith antibody positive and negative group.The data were analyzed by non-parametric rank test.Results Based on sex,there was no significant difference in the variations in TLR7 gene copy number between in female SLE patients and female healthy controls (Z=-1.175,P=0.240).There was also no difference in male group (Z=-1.085,P=0.278).Comparing gene copy numbers variation based on the presence or absence of lupus nephritis,hematological involvement,and anti-Smith antibody,there was no statistical significant difference in female SLE patients(Z=-0.888,P=0.375; Z=-1.085,P=0.278; Z=-0.529,P=0.597).There was no difference in variation in male SLE patients,neither (Z=-0.460,P=0.646; Z=-0.340,P=0.733;Z=-0.158,P=0.874).Conclusion The variations in TLR7 gene copy number are not correlated with SLE and clinical phenotypes of SLE in Han population.

Expressions of TLR7, MyD88 and TRAF6 in peripheral blood from patients with condyloma acuminatum / 中华皮肤科杂志

Objective To explore the role of Toll-like receptor 7 (TLR7) and related signal transduction molecules in mechanisms underlying human papilloma virus (HPV) infection and condyloma acuminatum (CA) recurrence.Methods Peripheral blood was obtained from 30 healthy controls,35 patients with primary CA,32 patiens with recurrent CA and 30 patients with recurrent CA treated by imiquimod and ALA-PDT.Two-color flow cytometric analysis was used to detect TLR7 expression in different peripheral blood T cell subsets,Western blot to determine the expression levels of an adapter protein myeloid differentiation primary response gene 88 (MyD88),and signaling molecules including tumor necrosis factor receptor-associated factor (TRAF6),phosphatidylinositol3-kinase (PI3K),protein kinase B (AKT),p42/44 and nuclear factor-kappa B (NF-κB) in peripheral blood CD3+ T cells,from these subjects.Statistical analysis was carried out by Student's t test using the software SPSS 13.0.Results TLR7 was expressed in peripheral blood CD3+CD4+ T cells and CD3+CD8+ T cells from the healthy controls.Compared with the healthy controls,the patients with primary and recurrent CA showed no significant changes in TLR7 expression in CD3+CD8+ T cells,but a statistical increase in TLR7 expression in CD3+CD4+ T cells (23.3％ ± 8.4％ and 32.8％ ± 8.9％ vs.12.6％ ± 6.3％,t =4.72,10.76,both P ＜ 0.01).Decreased expression of TLR7 in CD3+CD4+ T cells was observed in patients treated with imiquimod and ALAPDT compared with the patients with recurrent CA (20.3％ ± 5.7％ vs.32.8％ ±8.9％,t =5.41,P ＜ 0.01).The protein expressions of MyD88,TRAF6,PI3K,p42/44 and NF-κB were significantly increased in CD3 + T cells,while the AKT protein expression experienced no significant changes in patients with primary or recurrent CA compared with the healthy controls.A significant decrease was observed in the protein expression of MyD88,TRAF6,p42/44 and NF-κB in the treated patients compared with those with recurrent CA.Conclusions TLR7,which is highly expressed in peripheral blood CD3+CD4+ T cells in patients with CA,may take part in the host immune response against HPV and serve as a recognition receptor for HPV infection.

Expressions of Toll-like receptor 7 and 9 mRNA in peripheral blood mononuclear cells from patients with vitiligo / 中华皮肤科杂志

ObjectiveTo detect the expressions of Toll-like receptor(TLR) 7 and 9 mRNA in peripheral blood mononuclear cells(PBMCs) from patients with vitiligo and their significance.Methods Real-time fluorescence-based quantitative reverse transcription-PCR was performed to detect the expressions of TLR7 and TLR9 mRNA in PBMCs from 50 patients with vitiligo and 25 normal human controls.Two-sample t test was conducted by using SPSS 11.5 software to compare the expression difference of TLR7 and TLR9 mRNA between the patients and controls.ResultsThe mRNA expression level (the ratio of the absolute copy number of a target gene to a control gene) of TLR7 and TLR9 in PBMCs were significantly higher in patients with vitiligo than in the normal human controls(0.85 ± 1.90 vs.0.44 ± 1.18,P ＜ 0.05; 0.94 ± 2.25 vs.0.11 ±0.31,P ＜ 0.01 ).No significant difference was observed in the expression level of TLR7 or TLR9 mRNA in PBMCs between patients with stable and active vitiligo,or between patients with localized and generalized vitiligo(both P ＞ 0.05).ConclusionsThe expression level of TLR7 and TLR9 mRNA is elevated in PBMCs from patients with vitiligo,which may be involved in the pathogenesis of vitiligo.

A critical amino acid in Toll-like receptor 7 TIR domain impacting its binding with MyD88 / 第二军医大学学报

Objective To study the mechanism of Toll-like receptor 7 (TLR7) signal transduction by identifying the critical amino acid in TLR7 Toll/Interleukin-1 receptor (TIR) domain. Methods We carried out a series of truncating mutagenesis and site-direct mutagenesis to obtain TLR7 mutants. These TLR7 mutants were transfected into HEK293T cells. NF-κB signal pathway activation was determined by the reporter gene assay. Immunoprecipitation and immunoblotting were employed to determine the binding between TLR7 and MyD88. Results We identified a RXR signal motif with two arginines at position 1004 and 1006, which plays a significant role in TLR7 signal transduction. We further demonstrated that only the arginine at position 1004 could affect the binding between TLR7 and MyD88. Conclusion We conclude that the arginine at position 1004 of human TLR7 is critical for its binding with MyD88 in signal transduction.

Antagonism of geniposide on Toll-like receptor 7/nuclear factor-κB signaling pathways in cells with influenza A virus infection / 中华微生物学和免疫学杂志

Objective To observe how geniposide as an anti-inflammatory agent through inhibition Toll-like receptor 7/nuclear factor-κB signaling pathways activation, as well as TNF-α and IL-6 release infectioned by influenza virus. Methods Epithelial cells was exposed to human influenza viruses A/Gui/81/23(H3N2) infection for 2 h before treatment with geniposide for 24 h. NF-κB responsive element luciferase reportor gene was transfected and dual luciferase cis-reporting systems was used to assay the transcriptional activity of NF-κB under the stimulated circumstance of influenza virus infection. The phosphory level and nuclear transposition of NF-κB was observed by fluorescence inverted microscope. RT-PCR was used to detect the gene transcription level of TLR7, TNF-α and IL-6. Results The relative luciferase reporter assay of NF-κB was apparently improved by influenza virus infection. But geniposide significantly repressed the relative value of luciferase. The phosphorylation level and rate for nuclear transposition of NF-κB was apparently improved by influenza A virus infection observed by fluorescence inverted microscope. But geniposide significantly repressed the phosphorylation level and rate for nuclear transposition. RT-PCR showed upregulation of TLR7 and pre-inflammatory markers TNF-α and IL-6 in A549 cells infected by influenza virus, geniposide had a significant effect on the expression of TLR7 and inflammatory markers TNF-α and IL-6 after treated with influenza virus. Conclusion Geniposide as an antiinflammatory agent antagonized influenza A virus infection through inhibiting Toll-like receptor 7/nuclear factor-κB signaling pathways activation, as well as on the downregulation of the downstream inflammatory markers target gene expression TNF-α and IL-6.

The expression and clinical significance of Toll-like receptor 7 and 9 in peripheral blood B cells of patients with systemic lupus erythematosus / 中华风湿病学杂志

Objective The aim of this study is to examine the expressions of Toll like receptor (TLR) 7 and TLR9 in the peripheral blood B lymphocytes of SLE patients and to analyze the correlation between TLR7/9 and clinical parameters. Methods lntracellular expression of TLR7/9 in the peripheral blood CD19+Blymphocytes was analyzed in 50 SLE patients and 30 healthy controls by flow cytometry. The difference of intracellular TLR7/9 expression levels in two groups was compared. Furthermore,the correlation between TLR7/9 expression and clinical parameters such as ESR, CRP, complement 3 (C3), complement 4 (CA), the level of serum IgG, anti-double stranded DNA antibody, anti-nuclear antibodies, SLEDAI score and urine protein excretion level, were analyzed. Results Compared with healthy subjects, the proportion of B cells expressing TLR7 and TLR9 was higher among SLE patients. Positive correlation was observed between TLR7 expression levels and clinical measurement of the SLEDAI and ESR. Negative correlation was observed between TLR7 expression levels and serum C3 levels. Positive correlation was observed between TLR9 expression levels and SLEDAI scores. Negative correlation was observed between TLR9 expression levels and serum C3 levels. Conclusion TLR7 and TLR9 expression is increased in the peripheral blood B cells of SLE patients, and correlates well with clinical parameters.