The Effects Of Ultra-high Molecular Weight Polyethylene On HMGB1 Production In RAW264.7 And Underlying Mechanisms

Objectives:This study constructed the inflammatory reaction model which the wear particles(WP) stimulate macrophage, to observe the inflammatory factor of the inflammatory induced by the ultra-high molecular polyethylene wear particles(UHMWPE-WP), especially the late inflammatory protein high mobility group protein B1(HMGB1) expression and possible mechanism, to provide the theoretical basis and experimental basis of the clinical research for the artificial joint aseptic loosening.Methods: Part1: the model construction of wear particle induced macrophage inflammatory reaction buildingThe cells were divided into three groups: Control Group, LPS Group, WP Group.1. The appropriate concentration of LPS and WP were analyzed by MTT. 2. The content of the inflammatory mediators in the supernatant fluid were detected by ELISA.Part2: the impact of wear particle on the macrophage inflammatory and the possible mechanismThe cells were divided into three groups: Control Group, WP Group, WP+HMGB1 A-box Group, HMGB1 A-box Group, LPS Group.1. The content of the inflammatory mediators in the supernatant fluid were detected by ELISA. 2. The expression of HMGB1 was detected by ELISA and Western Blot. 3. The phosphorylation of NF-κB p65 was detected by Western Blot.Results:Part1: the model construction of wear particle induced macrophage inflammatory reaction building1. The resuls of MTT showed that the appropriate concentration of LPS was 1 μg/m L; he appropriate concentration of LPS was 250 ng/m L. 2. The content of the inflammatory mediators in the supernatant fluid were detected by ELISA. Compared with the concentration in the blank group, the concentration of TNF-α and IL-1, IL-6 in the model group has significant difference(P<0.05).It proved that the model is builded successfully, the WP induce the inflammation.Part2: the impact of wear particle on the macrophage inflammatory and the possible mechanism1. The content of the inflammatory mediators in the supernatant fluid were detected by ELISA. Compared with the concentration in the blank group, the concentration of TNF-α and IL-1, IL-6 in the model group has significant difference(P<0.05). After the HMGB1 A-box using in the model, the degree of the inflammatory response is reduced obviously. 2. The content of the HMGB1 in the supernatant fluid were detected by ELISA. Compared with the concentration in the blank group, the concentration of HMGB1 in the model group has significant difference(P<0.05). After the HMGB1 A-box using in the model, the expression of the HMGB1 is reduced obviously. 3. The influence of the WP on the NF-κB p65 phosphorylation is detected by Western Blot. Compared with the blank group, we found that the activation degree of NF-κB p-65 increase significantly.Conclusions: The mice mononuclear macrophage can be induced to produce inflammation by theUHMWPE-WP, raising the expression of the mice mononuclear macrophages HMGB1 protein, the mechanism may be related to the NF-κB signaling pathway.