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As a receptor mediating the transmembrane signal transduction in the innate immunity, Toll-like receptor 4 (TLR4) is a bridge between innate immunity and required immunity, and plays an important role when signal-transducing of some cells are activated. Recent reports show that TLR4 expresses in the different glial cells and strongly links to the innate immune activation and inflammatory response in the central nervous system (CNS). TLR4 plays a key role in the processes of brain damage by infection of the CNS, stroke, cerebral hemorrhage and trauma. In this review, we concentrate on recent findings regarding the progress of function and mechanism of TLR4 in the processes of the CNS damage in various diseases.
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AIM: To investigate the expression of heat shock protein 60(HSP60) and Toll-like receptor 4 transduction system in mouse cardiac transplantation. METHODS: The mouse cervical heart transplantation model was established. The animals were divided into control group (the donor and recipient were all C57BL/6 mice) and experimental group (the donor was BALB/c mice and recipient was C57BL/6 mice). The heart and blood were collected for study at 3 d and 7 d. The pathological analysis of the hearts was performed. The levels of cytokines in the serum were determined using ELISA. The expression of HSP60, TLR4, MyD88 and NF-κB in the myocardium was determined by immunohistochemistry and Western blotting. RESULTS: The expression levels of HSP60, TLR4, MyD88 and NF-κB were higher in experimental group than those in control group. Severe rejection was observed in experimental group, whereas no distinct rejection in control group was found. The cytokines (TNF-α, IFN-γ, IL-12) increased significantly in experimental group as compared to those in control group. CONCLUSION: HSP60 increases significantly after heart transplantation, which may activate Toll-like receptor 4 transduction system in a MyD88-dependent pathway and promote allograft rejection. Regulation of HSP60 signal transduction may be a novel way for treating allograft rejection.
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AIM: To investigate the effect of lipopolysaccharide (LPS) on the mRNA expression of Toll-like receptor 4 (TLR4) and CD14 in lens epithelial cells (LECs).METHODS: The cultured LECs were incubated with defferent concentrations of LPS for defferent times.The mRNA expression of TLR4 and CD14 in LECs were detected by reverse transcription polymerase chain reaction (RT-PCR).RESULTS: The mRNA expression of TLR4 in the LECs treated with LPS at concentrations of 50 μg/L, 100 μg/L, 200 μg/L, 500 μg/L or 1 000 μg/L was higher than that in the LECs without LPS treatment (P<0.01), respectively.The mRNA expression of TLR4 in the LECs incubated with 100 μg/L LPS for 24 h, 48 h or 72 h was higher than that in LECs without LPS treatment at same time points (P<0.01).The mRNA expression of CD14 in the LECs incubated with LPS at concentration of 100 μg/L for 24 h was also higher than that in the LECs without LPS treatment (P<0.01).CONCLUSION: LPS enhances the mRNA expression of TLR4 and CD14 in LECs, which may be related to intraocular response, as well as to the formation and development of after-cataract.
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AIM: To establish an animal model of experimental autoimmune myocarditis (EAM) in BALB/c mice and to investigate the expression and significance of Toll-like receptor 3 in mouse EAM. METHODS: BALB/c mice were immunized with cardiac myosin extracted from porcine ventricular myocardium covered by complete freund's adjuvant (CFA) on 0 d and 7 d, then divided into immunized with CFA only. Serum and myocardium samples were collected at 14 d and 21 d after the first immunization. HE staining was used to identify the areas of inflammation. The myosin IgG antibody was examined by indirect ELISA assay. The changes of TLR3 protein and mRNA expression in myocardial tissue were measured by immunohistochemistry and real time-PCR. RESULTS: Compared to control group, immunohistochemistry results showed that there was positive expression of TLR3 in the myocardium of mice with EAM and the mRNA of TLR3 were more than 20 times (P<0.05). The expression of interferon beta mRNA in EAM group was more than 14 times as many as basal expression, that of tumor necrosis factor alpha was more than 18 times (P<0.05). CONCLUSION: The expression of Toll-like receptor 3 in myocardium is up-regulated in experimental autoimmune myocarditis. The inflammatory response to cardiac myosin may associate with the TLR3 signal transduction pathway.
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AIM:To investigate the expression of TLR2 and TLR4 in hepatocellular carcinoma(HCC),and to analyze their correlations to clinicopathologic features of HCC.METHODS:The protein and mRNA levels of TLR2 and TLR4 in HCC and para-tumor tissue were determined by immunohistochemistry and real-time fluorescence quantitative PCR(RFQ-PCR).RESULTS:The protein and mRNA levels of TLR2 and TLR4 in HCC were lower than those in para-tumor tissue(P
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AIM:To study the influence of stimulation by LPS and CD40 ligandization in vitro on the TLR4-MD2 expression and IL-12 production in dendritic cells (DCs) modified by sCD40 gene and provide the experimental clues of inducing dornor-specific immune tolerance.METHODS: Plasmid pEGFP-N1/sCD40 and pEGFP-N1 was transfected into DC2.4 cell line with lipofectamine. After 6 h of treatment with LPS and anti-CD40mAb, the expression of TLR4-MD2 on DCs was determined with flow cytometry (FCM) and RT-PCR. IL-12p70 protein was detected by ELISA.RESULTS: LPS treatment of DCs down-regulated surface expression of TLR4-MD2, LPS treatment along with anti-CD40mAb significantly up-regulated TLR4-MD2 surface expression. CD40 ligandization did not affect TLR4-MD2 mRNA expression in DCs but partly increased its low level induced by LPS and markly enhanced IL-12p70 secretion after LPS stimulation. DCs modified by sCD40 gene inhibited the above effect.CONCLUSION: After treatment with LPS and anti-CD40mAb, DCs modified by sCD40 gene down-regulate surface expression of TLR4-MD2 and IL-12p70 secretion decreases significantly, which might be linked with the interruption of TLR4-MD2 transportation from cytoplasm.
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AIM: To investigate the effects of transforming growth factor ?1 (TGF-?1) on murine-derived dendritic cells (DC). METHODS: Murine bone marrow cells were cultured with GM-CSF and TGF-?1 to develop TGF ?-DC. Then they were stimulated by lipopolysaccharide (LPS). Their phenotypes were assessed by flow cytometry (FCM). The allogeneic stimulating capacity of DC was measured by mixed lymphocyte reaction (MLR) using BrdU ELISA method. IL-12 p70 protein was detected by ELISA and the expressions of Toll like receptor 4 (TLR4) on DCs were measured by semi-quantitative RT-PCR and FCM. RESULTS: Compared to immature DC (imDC) cultured with GM-CSF alone, the expressions of CD_80, CD_86, I-A~b and CD_40 in TGF ?-DC were lower. The TGF ?-DC was resistant to maturation by LPS. Maturation resistance was evident from a failure to up-regulate CMs, to stimulate larger T cell proliferation and to increase secretion of IL-12 p70. Down-regulation of TLR4 expression on TGF ?-DC was also found. CONCLUSION: TGF-?1 inhibits the expression of co-stimulatory molecules on DC. It is resistant to maturation stimulus (LPS) and might be linked with TLR4 down-regulation.
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AIM: To investigate the signaling mechanisms involved in the priming effects of lipopolysaccharide(LPS) on heat killed Staphylococcus aureus(HKSa)-induced nitric oxide(NO) production in macrophages.METHODS: Murine macrophage RAW264.7 was used in the experiment.Griess reagent was used to measure the content of nitrite in culture medium.Real-time PCR and Western blot was utilized to examine the mRNA and protein levels of toll-like receptor 2(TLR2),respectively.Dual luciferase reporter assay was used to assess the transcriptional activity of nuclear factor of activated T cells(NF-AT).RESULTS: The RAW264.7 cells pretreated with LPS for 24 h significantly enhanced NO production induced by HKSa,suggesting that LPS primed the macrophages and increased the reactivity of the cells to HKSa.LPS increased the mRNA and protein levels of TLR2 in a dose-dependent manner in RAW264.7 cells.The RAW264.7 cells pretreated with LPS enhanced NO production induced by peptidoglycan,one of the specific ligand of TLR2.The priming effect of LPS on HKSa-induced NO production was partly blocked by TLR2 neutralizing antibody.LPS significantly enhanced the transcriptional activity of NF-AT,which was inhibited by BAPTA/AM(a cell-permeable cytosolic calcium chelator) and cyclosporine A(CsA,an inhibitor for calcineurin).Both BAPTA/AM and CsA inhibited the priming effect of LPS on HKSa-induced NO production in RAW264.7 cells.CONCLUSION: The present study confirms the priming effect of LPS on the reactivity of RAW264.7 cells to HKSa.Pattern recognition receptor TLR2 and calcium/calcineurin/NF-AT signaling pathway may be involved in the priming process initiated by LPS.
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AIM:To investigate the effects of ox-LDL on TLR2 and TLR4 expression and production of TNF-?,IL-10,IL-12,NO and MDA in macrophages and to observe intervention effect of GW1929 in above procedure.METHODS:The mouse peritoneal macrophages were pretreated with ox-LDL(50 mg/L,100 mg/L)and GW1929(20 ?mol/L)respectively for 24 h.The concentrations of MDA,NO-2/NO-3,TNF-?,IL-10 and IL-12 in the culture fluid were detected.Flow cytometry was used to observe TLR2 and TLR4 expressions after the mouse peritoneal macrophages were pretreated with ox-LDL(50 mg/L)and GW1929(20 ?mol/L)respectively for 6 h,12 h,and 24 h.RESULTS:The concentrations of MDA,NO-2/NO-3,TNF-? and IL-10 in ox-LDL(50 mg/L,100 mg/L)group were higher than those in control and GW1929 group obviously,but the concentrations of above index in ox-LDL(50 mg/L,100 mg/L)+GW1929 group were lower than those in ox-LDL(50 mg/L,100 mg/L)group apparently.No IL-12 in every group was detected.Expressions of TLR-2 in ox-LDL+GW1929(6 h,12 h,24 h)group were lower than those in ox-LDL(6 h,12 h,24 h)group respectively.TLR-4 expressions in ox-LDL+GW1929(12 h)were lower than those in ox-LDL(12 h)apparently.CONCLUSION:ox-LDL up-regulates TLR2 and TLR4 expressions and promotes the production of ROX,NO,TNF-? and IL-10 in macrophages.GW1929 is capable of inhibiting the above ox-LDL effects.
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0.05) was observed. The protein expressions of TLR-4, NF-?B p65, HSP70 and ICAM-1mRNA in IR group were significantly increased as compared to C group and PD group, while those expressions in PD group were evidently higher than those in C group (all P