| Objective: To investigate the function and mechanism of TLR9-MyD88-TRAF6-IRF5 signaling pathway and its signaling molecules in systemic lupus erythematosus(SLE).Method: 38 SLE patients were divided into inactive group(21)and active group(17)according to the SLEDAI score.Real time quantitative PCR(RT-qPCR)was used to detected the expressions of TLR9?MyD88?TRAF6?IRF5 mRNA in 38 SLE patients and 19 healthy volunteers,ELISA was used to determine IFN-??IL-6?IL-12 in the serum of all subjects.In addition,separated and cultuered the B cell of the whole blood from 4 SLE patients and 4 volunteers by Magnetic Beads.After 3 days of culture,the cell suspension was divided into two wells which named treatment group and blank control group respectively.TLR9 antagonist was added to the treatment group,TLR9 blank control was added to the blank control group,then continued culturing for another 24 hours.RT-qPCR was used to detected the expressions of TLR9?MyD88?TRAF6?IRF5 in B cell of two groups.ELISA was used to detected the IL-6?IL-10 levels in the culture supernatant.Result: 1.The expressions of TLR9-MyD88-TRAF6-IRF5 singal pathway in PBMC(1)The levels of TLR9?TRAF6?IRF5 mRNA in active SLE group were higher than that in inactive group and healthy group,the inactive group were higher than that in healthy group,the differences between the three groups were statistically significant(p < 0.05).The expression of MyD88 was higher in inactive group than that in healthy group and active group,the difference between the active group and inactive group was not statistically significant(p>0.05)while the difference between inactive group and healthy control was statistically statistically significant(p<0.05).The levels of IL-6?IL-10 in active group were higher than in inactive group and healthy group,the levels in the inactive group were higher than the healthy group,the differences between the three groups were statistically significant(p<0.05).The level of IFN-? in active group was higher than in inactive group and healthy group(p<0.05),while there was not any differences between the inactive group and healthy group(p>0.5).(2)The correlation beween the molecules of TLR9-MyD88-TRAF6-IRF5 signal pathway and active index of SLEIn SLE group,the expression of TLR9 was positive correlate with the MyD88?TRAF6?IRF5?IFN-??IL-6?IL-10?dsDNA?SLEDAI,and it was negative correlate with C3 ? C4.The differences were statistically significant(p<0.05).There were no any correlations of MyD88 and TRAF6?IRF5?IFN-??IL-6?IL-10?SLEDAI?dsDNA(p>0.05).The expression of TRAF6 was positive correlate with IRF5?IFN-??SLEDAI?dsDNA,the differences were statistically significant(p?0.05),There was no differences between the correlations of TRAF6 and IL-10?IL-6?C3?C4.IRF5 was positive correlate with IFN-??IL-6?IL-10?SLEDAI?dsDNA and negative with C3,the differences were statistically significant(p<0.05).There was no difference between IRF5 and C4.(3)The statistical analysis of the clinical manifestations of SLE related to signal molecules in TLR9-MyD88-TRAF6-IRF5 signaling pathwayIn the SLE group,the expressions of TLR9?TRAF6 and IRF5 in patients with arthritis were higher than those without arthritis,and the differences were statistically significant(p<0.05).While the expression of MyD88 was lower in patients with arthritis than patients without arthritis,and the difference was statistically significant(p<0.05).There were no significant differences in the expression levels of TLR9?TRAF6 and IRF5 in patients with facial erythema and patients without facial erythema(p<0.05),while the expression of MyD88 in patients with facial erythema had lower level and the difference was statistically significant(p<0.05).2.The expressions of the molecules of TLR9-MyD88-TRAF6-IRF5 in B cell separated from the whole blood after adding TLR9 antagonist or TLR9 blank control when cultured in three day(1)After separating B cells from the whole blood by magnetic beads,the purity of B cells was more than 95%.(2)In vitro,the expressions of TLR9?MyD88?TRAF6 and IRF5 on B cells in SLE patients were higher than thoes in healthy controls,and the differences were statistically significant(p<0.05).(3)After treating B cell with TLR9 antagonist or TLR9 blank control in vitro,the expressions of TLR9?MyD88?TRAF6 and IRF5 were lower in the treated group than in the blank control group,and the differences were statistically significant(p<0.05).For the same molecule,the difference of the expressions in the blank control group and the treatment group was greater in the healthy control group than in the SLE group,and the differences were statistically significant(p<0.05).(4)The expressions of IL-6 and IL-10 in the supernatant of B cell of SLE patients were lower than that in healthy controls(p<0.05).The differences of the blank control group and the treatment group were larger in the healthy control group than in the SLE group(p<0.05).Conclusion: 1.TLR9-MyD88-TRAF6-IRF5 signal pathway maybe take participate in disease progression of SLE,and it may related to the disease activity.These pathway may become one of the indicators for judging disease activity.2.This signal pathway may be abnormally tolerated on B ells in SLE patients... |
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