Study On The Effect Of CXCR4 In Acute Lung Injury Caused By Lipopolysaccharide

Part IExpression and effect of CXCR4 in rat lung tissue in acute lung injury caused by endotoxin[Objective]By detecting the expression of CXCR4 and pro-inflammatory cytokine in rat lung tissue and analyzing their relationship at the condition of lipopolysaccharide (LPS), to investigate the effect of CXCR4 in acute lung injury caused by endotoxin. [Methods]Spraque-Dawley rats were divided into 5 groups: (1) contral group, (2) group 2h, (3) group 4h, (3) group 8h, (3) group 12h. By injecting lipopolysaccharide (LPS) into caudal vein, the model of acute lung injury was duplicated. Two, four, eight and twelve hours later, the expression of CXCR4, SDF-1α, TNF-α, IL-1β and IL-18 were detected through the methods of immunohistochemistry, Western Blot and RT-PCR respectively. [Results]CXCR4 and SDF-1α exist in normal lung tissue. In acute lung injury caused by LPS, the expression of CXCR4 and SDF-1α increased at RNA and protein level with the same trends of TNF-α, IL-1β and IL-18.[Conclusion]LPS can promote the expression of CXCR4 and SDF-1α in rat lung tissue and cause the release of lots of pro-inflammatory cytokines, further more, results in lung injury. CXCR4 and SDF-1α take important roles in acute lung injury caused by LPS. Part IIThe expression and effect of CXCR4 in rat alveolar macrophage caused by lipopolysaccharide[Objective]By detecting the expression of CXCR4 and pro-inflammatory cytokine in rat alveolar macrophage cultured in vitro and analyzing their relationship at the condition of LPS and/or AMD3100, to investigate the effect of CXCR4 in the course of LPS activating pro-inflammatory cytokine system. [Methods]Alveolar macrophages isolated and cultured in vitro were grouped at random.(1) Stimulating alveolar macrophages with LPS only.(2) Stimulating alveolar macrophages with AMD3100 firstly. Thirty minutes later, stimulating alveolar macrophages with LPS.(3) Stimulating alveolar macrophages with AMD3100 and LPS together at the same time.(4) Stimulating alveolar macrophages with LPS firstly. Thirty minutes later, stimulating alveolar macrophages with AMD3100.After the stimulation of LPS, the expression of CXCR4 and the concentrations of TNF-α and IL-6 in culture fluid were detected through the methods of Western Blot, immunohistochemistry and RT-PCR respectively. [Results]At different times after stimulating alveolar macrophages with LPS only, contrasting to normal control groups, CXCR4 evidently increased at RNA and protein level. So did concentrations of TNF-α and IL-6 in culture fluid. AMD3100 decreased concentrations of TNF-α and IL-6 in culture fluid, especially in Group AMD3100 preconditioning, but had no effect on the expression of CXCR4. [Conclusion]LPS can promote the expression of CXCR4 in rat alveolar macrophages and cause the release of lots of pro-inflammatory cytokines. AMD3100 has no effect on the expression of CXCR4 but can inhibit the release of pro-inflammatory cytokine. CXCR4 may take an important role in the course of lipopolysaccharide recognition and signal transduction. Part IIIThe effect of CXCR4 on Lipopolysaccharide-tolerence[Objective]By detecting the expression of CXCR4 and pro-inflammatory cytokine in rat lung tissue and alveolar macrophage cultured in vitro and analyzing their relationship at the condition of LPS preconditioning, to investigate the effect of CXCR4 in the course of acute lung injury caused by lipopolysaccharide. [Methods] 1. The expression of CXCR4 in rat lung tissue at the condition of LPS preconditioningSpraque-Dawley rats were grouped at random.(1) Group LPS: By injecting LPS into caudal vein, the model of acute lung injury was duplicated.(2) Group LPS preconditioning: LPS was injected into rat abdominal cavity firstly. Twenty-four hours later, Do it again. After another 72 hours, LPS was injected into caudal vein.At the different time (2, 4, 8, 12hour) after LPS injected into caudal vein, the expression of CXCR4, TNF-α, IL-1β and IL-18 in lung tissue were detected through the methods of immunohistochemistry, Western Blot and RT-PCR respectively. 2. The expression of CXCR4 in rat alveolar macrophage cultured in vitro at thecondition of LPS preconditioningAlveolar macrophages isolated and cultured in vitro were grouped at random.(1) Group LPS: LPS stimulate alveolar macrophages only one times.(2) Group LPS preconditioning: Low dose LPS precondition alveolar macrophages for 12 hours, then high dose LPS stimulate alveolar macrophages.At the different time (2, 4, 6, 8 hours) after LPS stimulsting, the expression of CXCR4, TNF-α, IL-1β and IL-18 in alveolar macrophages were detected through the methods of immunohistochemistry, Western Blot and RT-PCR respectively. [Results]The expression of CXCR4, TNF-α, IL-1β and IL-18 in lung tissue and alveolar macrophages were decreased in Group LPS preconditioning contrasting to Group LPS. [Conclusion]LPS preconditioning can alleviate acute lung injury caused by endotoxin. In this course, the down-regulation of expression level of CXCR4 may take an important role.