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目的探讨铜绿假单胞菌(Pseudomonas aeruginosa,PA)群体感应系统(quorum sensing system,QS)对CD4+CD25+Treg细胞的影响及作用机制。方法60只雄性SD大鼠,随机分为铜绿假单胞菌野生菌株PAO1组,变异菌株PAO-JP2组(lasI、rhlI基因双变异)和空白对照组。将PA包被的硅胶管置入大鼠一侧主支气管建立慢性肺部感染模型,对照组置入无菌硅胶管。28 d后,FACs测外周血CD4+CD25+Treg细胞数量,ELISA测血清IL-10、TGF-β,RT-PCR测脾Foxp3mRNA表达水平,肺组织行HE染色。结果CD4+CD25+Treg/CD4+T淋巴细胞的百分比:PAO1组(19.79±6.45)%,PAO-JP2组(11.03±3.92)%,对照组(5.15±0.47)%,PAO1组的百分比明显高于PAO-JP2组和对照组(P<0.05)。IL-10:PAO1组(231.52±54.48)pg/mL,PAO-JP2组(60.10±29.64)pg/mL,对照组(35.43±23.56)pg/mL,PAO1组表达水平高于JP2、对照组(P<0.05)。TGF-β:PAO1组、JP2组、对照组水平分别为(121.05±7.98)pg/mL、(63.11±5.73)pg/mL、(36.02±8.94)pg/mL,PAO1组与PAO-JP2、对照组比较均有统计学意义。Foxp3mRNA相对含量:PAO1组(0.80±0.044),PAO-JP2组(0.41±0.044),对照组(0.25±0.054),PAO1组较其他两组含量显著升高(P<0.05)。HE染色可见,PAO1组肺组织大量淋巴细胞聚集、纤维增生明显,脓肿形成;PAO-JP2组炎症细胞浸润叫PAO1组明显减轻,对照组未见明显异常。结论PA群体感应系统通过提高IL-10、TGF-β和Foxp3mRNA的表达水平,上调CD4+CD25+Treg细胞的数量及功能,在慢性肺部感染中发挥重要作用。
Objective To investigate the effect and mechanism of quorum sensing system (QS) on CD4 + CD25 + Treg cells by Pseudomonas aeruginosa (PA). Methods Sixty male Sprague-Dawley rats were randomly divided into three groups: PAO1 group, PAO-JP2 group (double mutation of lasI and rhlI gene) and blank control group. The PA-coated silicone tube was inserted into the main bronchus of the rat to establish a chronic lung infection model, and the control group was placed in a sterile silicone tube. After 28 days, the number of CD4 + CD25 + Treg cells in peripheral blood was measured by FACs. The expression of Foxp3 mRNA in the spleen was detected by ELISA, serum IL-10, TGF-β and RT-PCR. Results The percentages of CD4 + CD25 + Treg / CD4 + T lymphocytes were significantly higher in PAO1 group (19.79 ± 6.45)%, PAO-JP2 group (11.03 ± 3.92)% and control group (5.15 ± 0.47)% In PAO-JP2 group and control group (P <0.05). The levels of IL-10 in the PAO1 group (231.52 ± 54.48) pg / mL, the PAO-JP2 group (60.10 ± 29.64) pg / mL and the control group (35.43 ± 23.56) pg / mL in the PAO1 group were higher than those in the control group P <0.05). The levels of TGF-β in PAO1 group, JP2 group and control group were (121.05 ± 7.98) pg / mL, 63.11 ± 5.73 pg / mL and 36.02 ± 8.94 pg / mL, Group comparison was statistically significant. The relative content of Foxp3mRNA in PAO1 group (0.80 ± 0.044), PAO-JP2 group (0.41 ± 0.044) and control group (0.25 ± 0.054), PAO1 group was significantly higher than the other two groups (P <0.05). HE staining showed that a large number of lymphocytes in PAO1 group were aggregated, fibrogenesis was obvious and abscess was formed. PAO-JP2 group inflammatory cell infiltration was significantly reduced in PAO1 group and no obvious abnormality in control group. Conclusion PA population sensing system plays an important role in chronic pulmonary infection by increasing the expression of IL-10, TGF-β and Foxp3 mRNA and up-regulating the number and function of CD4 + CD25 + Treg cells.