The Role Of Quorum Sensing In The Pulmonary Infection In Rats Due To Pseudomonas Aeruginosa

[Obejective] To investigate the role of quorum sensing in the pulmonary infection in rats due to Pseudomonas aeruginosa. To further elucidate the effect of quorum sensing on the extracelluar virulence factors involved the pathogenesis of Pseudomonas aeruginosa. To explore the mechanism of how quorum sensing systems of Pseudomonas aeruginosa operate during the infectious process.[Method] (l)The animal model of chronic pulmonary infection was establishedby inoculating two different Pseudomonas aeruginosa embedded in minute seaweed algiante beads which was made by an ejection set with an acuminate hole to Sprague-Dawley rats. And two groups of rat model were established. The sterile alginate beads were inoculted to rats as the control group. (2) The in vivo pathogenic effects of the two different Pseudomonas aeruginosa 'which were the wide-type Pseudomonas aeruginosa PAO1 and its double mutant PAO-JP2, in which the signal-generating parts of the quorum sensing systems(lasR-lasI and RhlR-RhlI expressing the autoinducer N-acylhomoserine lactone) are defective were compared based on the successefully established lung infection models. The indicator about bacteriology, pathology, immuology were monitored. The percentage of polymorphonuclear leukocyte(PMN) in peripheral blood white cells was counted with an optic microscope after Wright's staining. The numbers of Pseudomonas aeruginosa in lung were determined by viable counting method(VCM) using log10CFU/g as the measure value. The levels of antibody to Pseudomonas aeruginosa in sera and cytokines including interferon gamma ( DFN-Y ) ,Interleukin-4(IL-4) in lung homogenate was measured by ELISA method. Pathology changes of lung were evaluated by the macroscopic and microscopic graded scores. The IL-4 levels in lung tissues were detected by immunohistochemistry methods.I Results 1 (1) The rat model of chronic pulmonary infection was established asexpected. The bacteria numbers in lung were measured 3,7,14,28 days after infection . No bacterium were detected in the control group. PAO1 and PAO-JP2 were detected from rats of PAO1 and PAO-JP2 infected groups respectively and the number was all higher than 103CFU/g. The pathyological changes showed: 3, 7 days after infection, lung abscess, edema, consolidation, atelectasis can be seen from lungs of both groups. And at optical microscopic, alginate-Pseudomonas aeruginosa caused a pronounced inflammatory reaction with polymorphonuclear cells surrouding a bead. Small microcolonies formed at the periphery of the bead were also seen. 14, 28 days after infection , the consolidation reduced gradually and lung atelectasis, fibrinous adhesions granulomas became the major pathological changes. (2) the difference between two groups of pulmonary infection models : ﹖he infection aspect: the mortality in the PAO1-JP2-infected group(10.59%) was significantly lower than that of the RAO1-infected group(29.8%). In the early stages of infection(3 days after inoculation), no difference was found between the two groups in indicators of pathological changes and bacteriology( log10CFU/g). In middle to later stages of infection(7 to 28 after infection), the bacteriology indicator(log10CFU/g) and the pathological scores in the PAO-JP2 infected group were signicantly lower than that of the PAO1 infected group. (2)The immunology aspect: Compared with the PAO-JP2 group: During the early stages of lung infection(3 days after infection), the IFN-gamma level in lung homogenates wassignificantly higher and the percentage of PMNs, the concentration of C response protein(CRP) were significantly lower. During the middle stages of the infection(7 days after infection), the percentage of the PMNs and the concentration of CRP in peripheral blood, the levels of IFN-γ and IL-4 in lung homogenates of PAO1-infected rats were significantly higher. In the later stages of infection(14 to 28 days after infection), levies of EFN-γ in lung homogenates, IgG2a in sera were lower, while levels of IL-4 in lung homogenates, IgG, IgG1 in sera were higher persist...