沙眼衣原体小鼠肺炎株呼吸道感染诱导肺组织中性粒细胞浸润及活化

Infiltration and activation of neutrophils in lung tissues during Chlamydia muridarum respiratory tract infection

目的：探讨小鼠沙眼衣原体肺炎感染中中性粒细胞（ polymorphonuclear neutrophils， PMN）的募集、活化及诱导其趋化的免疫学机制。方法 C57BL/6（C57）小鼠鼻腔吸入3×10^3 IFU（in-clusion-forming units）沙眼衣原体小鼠肺炎株（Chlamydia muridarum，Cm）建立小鼠沙眼衣原体肺炎模型。取感染后不同时间点小鼠肺组织进行组织病理学染色；通过检测肺组织内髓过氧化物酶（ myelo-peroxidase，MPO）的含量，检测 PMN 在肺组织聚集；分离小鼠肺组织单个核细胞，Giemsaˊs 染色计数肺组织炎症细胞种类；流式细胞术检测 CD11b^＋ Gr1^＋ PMN 细胞百分率及 CD11b^＋ PMN 活性水平；应用RT-PCR 技术检测 Cm 感染后肺组织内与 PMN 募集相关的趋化性细胞因子（MIP-2、LIX、KC、MCP-1）的表达，探讨小鼠 Cm 呼吸道感染过程中 PMN 趋化机制。结果一定剂量的 Cm 呼吸道吸入诱导C57小鼠衣原体肺炎，与对照组（感染第0天）比较，肺组织内有大量炎性细胞浸润，主要包括 PMN、单核细胞、淋巴细胞，其中 PMN 在肺内的聚集早于单核细胞；Cm 感染诱导小鼠肺组织 MPO 含量升高，在感染第7天达到高峰，第14天虽然有所下降，但仍明显高于对照组；流式结果显示，Cm感染后，CD11b^＋Gr1^＋细胞百分比、PMN 总数均显著高于对照组，同时 PMN CD11b 分子的表达在感染后也显著升高，说明 Cm 感染能诱导小鼠肺组织内 PMN 大量聚集并活化。RT-PCR 结果显示，Cm 感染后小鼠肺组织内与 PMN 募集相关的趋化性细胞因子（MIP-2、LIX、KC、MCP-1）的含量明显升高，提示Cm感染上调 PMN 相关趋化性细胞因子的表达，从而吸引大量 PMN 在肺部聚集参与炎症反应。结论一定剂量的 Cm 可以诱导感染小鼠肺组织内 PMN 相关趋化性细胞因子的分泌，从而使 PMN 在肺组织内大量浸润并活化，在抵抗衣原体感染中发挥重要作用。

Objective To investigate the possible mechanism of aggregation and activation of neu-trophils（polymorphonuclear neutrophils,PMN）in mice with chlamydial pneumonitis. Methods C57BL/ 6 mice were inoculated intranasally with 3×10^3 inclusionforming units（IFU）of Chlamydia muridarum（Cm） to induce the murine model of chlamydial pneumonitis. Samples of lung tissues collected at different time points after infection were stained by hematoxylin and eosin for histopathological assessment of inflammation. The levels of myeloperoxidase（MPO）were detected for the evaluation of PMN aggregation. The mononu-clear cells were isolated from lung tissues. The inflammatory cells were counted with Giemsaˊs staining. CD11b^＋Gr1^＋ cell population and CD11b expression in lung mononuclear cells were analyzed by flow cytome-try. The expression of chemokines（MIP-2,LIX,KC and MCP-1）in lung tissues at mRNA level was meas-ured by RT-PCR. Results Chlamydial pneumonitis was induced in mice by intranasal inoculation of 3×10^3 IFU of Cm. Compared with the mice from control group,large amounts of inflammatory cells including PMN, monocytes and lymphocytes were induced in lung tissues of mice with Cm infection. PMN responded earlier than monocytes to the infection. The levels of MPO were significantly increased in mice with Cm infection and reached the highest level on the 7th day after infection. A decline in MPO levels was observed on the 14th day but the levels were still higher than those on day 0. The percentages and total numbers of CD11b^＋Gr1^＋ cells were significantly increased after Cm infection. Moreover,an increased expression of PMN CD11b was also detected by flow cytometry. The expression of chemokines（MIP-2,LIX,KC and MCP-1）was in-creased in lung tissues of mice after Cm infection. The results of the study indicated that Cm infection induced the expression of PMN chemoattractants,resulting in the recruitment of PMN. Conclusion The infiltration and activation of PMN in lung tissues of mice were induced by Cm infection through increasing the expression of chemokines. PMN played an important role in immune responses against Cm infection.