Study On The Mechanism Of Synovial Inflammation Of Osteoarthritis By Toll - Like Receptor Pathway

Objective To study the expression of IL - 1 beta in joint fluid of patients with knee osteoarthirities, analysising the correlation between TCM syndrome type and the expression of IL-1 beta, seeking objective standards for syndrome differentiation of knee OA. To detect the toll-like receptor 4 (TLR4) transcription and expression level in synovial membrane of different stages of OA and find out TLR4 cytology of OA and synovitis. To observe the effects of cinnamaldehyde on the intervention of TLR4 / NF - kB pathways in LPS-induced synovial membrane cell and explore the Tongluozhitong treatment mechanism in the treatment of knee OA.Method1.According to OA diagnostic criteria,42 patients with OA were collected in this study.All patients were from the third affiliated hospital of Beijing medical university orthopaedic clinic.Information of the patient’s age, gender, knee joint X-ray Kellgren - Lawrence (K - L) radiological grading, Lysholm knee OA patients’ joint function Score, Visual Analogue Scale,expression of IL - 1 beta in joint fluid was recorded.Looking for specific indicators to guide syndrome differentiation of Chinese medicine in OA diagnosis.2.80 190-210g adult Wsiatarrats were divided into normal control group, control group, model group (2、4、6week modules) using the method of Hulth to established mild, moderate and severe knee OA model. With observing the cartilage morphological changes to determine the extent of lesion.Using Real - time PCR and Western blot to observe the transcriptional expression of TLR4 and NF - kB in OA model at different disease stage,analysis the sense of expression of TLR4 / NF - kB in OA synovial cells and inflammatory response.3. Cultivation knee joint synovial cells of rats, take 3-5 generations of synovial cell induced by LPS of 10 mg/L concentration, using Elisa to detect quantitative expression of IL - 1 beta in the supernatant fluid of synovial cells, to explore the intervention effect of cinnamicaldehyde in TLR4 signaling pathways.Results:The first partFinally,42 cases were selected in this study.On the TCM syndrome type distribution, the number of cases with liver and kidney deficiency syndrome, phlegm and blood stasis resistance higher than the other two two groups.The X-ray grade of patients with liver and kidney insufficiency, blood stasis, compared with that of spleen and kidney insufficiency,wet joints was statistical difference (p<0.05); The X-ray grade difference between patients with liver and kidney deficiency, phlegm and blood stasis resistance hollows and spleen kidney, wet joints was significant (p<0.01).The function of the patients with liver and kidney insufficiency, the relative stasis score better than the other two groups (p<0.05); The function score of patients with spleen and kidney deficiency,wet joints was higher than patients with liver and kidney deficiency syndrome, phlegm and blood stasis resistance syndrome, but the difference was not statistically significant (p>0.05). Between groups of different syndrome types in patients with pain score had significant difference (p< 0.05).The scale of pain of patients with liver and kidney insufficiency, blood stasis is lower. The difference of scale of pain of between patients with spleen and kidney deficiency,wet joints and liver and kidney deficiency, phlegm and blood stasis resistance increased significantly, but no statistical differences in score. IL - 1 beta content of patients with liver and kidney insufficiency, the relative stasis and spleen and kidney deficiency, wet joints was significantly higher in patients with liver and kidney deficiency, phlegm and blood stasis resistance; IL - 1 beta content in the two groups of joint fluid were higher than in liver and kidney insufficiency, the relative stasis.TLR4 expression between three groups of the patients with the single factor analysis of variance, the results showed very statistical difference (F=15.91, p< 0.01).Patients with liver and kidney deficiency, relative stasis,TLR4 expression level difference was significant higner than other two groups(P<0.05);There was statistical difference (P<0.01) in TLR4 expression level between patients with liver and kidney deficiency, the channels stasis group and liver and kidney deficiency, phlegm and blood stasis resistance syndrome.The second partWith prolong the time of moduling, the synovial tissue of rats TLR4 mRNA expression gradually decreased, and were significantly higher than that of normal group (P<0.05); When modeling after 2weeks,TLR4 mRNA expression was the highest. With building the extension of time, the synovial tissue of rats NF-κB mRNA expression increased dramatically, and were significantly higher than that of normal group (P<0.05);When building 4 w, NF-κB mRNA expression significantly increased than before.The third partControl group supernatant inmanufacturing IL - 1 beta levels did not reach the minimum detectable concentration;LPS group supernatant IL - 1 beta level higher than that of cinnamic aldehyde intervention group, there was statistical difference (p<0.05).And compraring with control group, LPS group and cinnamic aldehyde TLR4 expression was obviously increased, there was significant difference (p<0.01); TLR4 expression level of Synovial cells of rats after cinnamic aldehyde intervention has decreased, the difference was statistically significant (p<0.05).Conclusion X-ray grading of knee OA, IL-1β expression level of joint fluid has obvious differences between the different syndrome types, thus may serve as reference to guide Chinese medicine dialectical.With the increase of pain, knee OA patients were significantly reduced on the function, the pain will affect patient activity ability.In patients with liver and kidney deficiency, channels stasis syndrome showed X-ray grading level, joint activity function, pain evaluation of knee OA was in the late stage,which was the most serious illness.Classic Hulth method can successfully simulate the rat knee OA disease processes; There was significant correlation between different degree of OA synovial inflammation lesion and the expression of TLR4 and NF-kB.In order to further explore the targeted therapy for OA, made effective treatment, controlling progression of OA and judging prognosis of OA provide experimental parameters.Cinnamicaldehyde can inhibit the synovial cells of rats induced by LPS rise in TLR4 protein expression,and the synovial inflammation, reduced the secretion of inflammatory mediators by synovial tissue.