烟雾吸入性损伤大鼠肺组织丝裂原活化蛋白激酶通路的变化

目的 了解烟雾吸入性损伤大鼠肺组织丝裂原活化蛋白激酶(MAPK)通路及炎性细胞因子含量的变化,探讨其损伤机制. 方法建立密闭舱内烟雾吸入性损伤模型,将30只SD大鼠分为烟雾吸入性损伤后1、6、24、72 h及7 d组,另设正常对照组(6只).取各组大鼠肺组织行病理学观察,检测肺组织匀浆液中肿瘤坏死因子α(TNF-α)、巨噬细胞炎性蛋白2(MIP-2)和白细胞介素1β(IL-1β)含量,用蛋白质印迹法检测肺组织p38MAPK、c-Jun氨基末端激酶(JNK)、细胞外信号调节激酶1/2(ERK1/2)及各酶磷酸化水平.收集大鼠支气管肺泡灌洗液(BALF),检测TNF-α、MIP-2、IL-1β含量并行粒细胞分类、计数. 结果烟雾吸入使大鼠产生急性肺损伤样病理改变.伤后1 h组大鼠肺组织及BALF中TNF-α和IL-1β含量均高于正常对照组(P<0.01).各组肺组织MIP-2水平与正常对照组接近(P>0.05),伤后1 h组BALF中MIP-2水平高于正常对照组(P<0.01).各致伤组p38MAPK、ERK1/2、JNK水平接近正常对照组,但此3种酶的磷酸化水平在伤后不同时相组有高表达.伤后1 h组大鼠BALF粒细胞总数为(0.36±0.08)×106个/L,较正常对照组(0.61±0.09)×106个/L明显减少(P<0.05),伤后7 d组粒细胞总数[(1.71±0.67)×106个/L]却高于正常对照组(P<0.05).伤后6 h～7 d组中性粒细胞数多于正常对照组(P<0.05).伤后1 h组巨噬细胞数少于正常对照组(P<0.05),但6 h～7 d组逐渐增多.各组大鼠淋巴细胞数量接近(P>0.05).结论 密闭舱室内非金属材料燃烧释放的毒性气体能诱导肺组织产生明显的炎性反应,激活细胞MAPK通路中重要激酶的表达,这可能是毒性混合气体导致肺损伤的重要机制之一.

Study on activation of mitogen-activated protein kinase pathway in lung tissue of rats with smoke inhalation injury

Objective To investigate the activation of mitogen-activated protein kinase(MAPK) path- way and the expression of inflammatory cytokine in lung tissue of rats with smoke inhalation injury, and to explore the injury mechanism. Methods The model of smoke inhalation injury in airtight cabin was estab- lished. Thirty-six SD rats were randomly divided into normal control group( n = 6,NC group) and inhalation injury group (n = 30, Ⅱ group). The rats in Ⅱ group were observed at 1,6,24,72 post injury hour(PIH) and 7 post injury day(PID). The pathological changes in lung tissue were observed by optical microscope. The contents of TNF-α, MIP-2, IL-1β in lung tissue homogenate were examined. The level of p38MAPK, JNK, ERK1/2 and their phosphorylation in lung tissue were measured by Western blotting. The contents of TNF-α, MIP-2,and IL-1β and granulocytes counts in BALF( bronchoalveolar lavage fluid) were detected by ELISA . Results Pathological change of smoke inhalation was similar to acute lung injury . The contents of TNF-α and 1L-1β of lung tissue and BALF in Ⅱ group at 1 PIH were higher than those in NC group( P < 0. 01 ). The levels of MIP-2 of lung tissue in Ⅱ group at each time point were similar to that in NC group( P > 0.05) ,while that of BALF in Ⅱ group at 1 PIH were obviously higher than that in NC group( P <0.01 ). Compared with those in NC group,the levels of p38MAPK, JNK, ERK1/2 in Ⅱ group at each time point were similar, and their phosphorylation levels increased with different degree. Compared with that in NC group[ (0.61±0.09) × 106 cells/L] , the total number of grauulocytes in Ⅱ group was significantly de- creased at 1PIH[(0.36±0.08)×106 cells/L],increased on7PID [ (1.71±0.67)×106 cells/L, P < 0. 05]. The number of neutrophils in Ⅱ group during 6PIH～7 PID were more than that in NC group. The number of lymphocyte in Ⅱ group at 1 PIH less than that in N C group( P < 0. 05 ) ,which increased gradual- ly during 6 PIH ～ 7PID. There were no obvious difference in number of lymphocyte between two groups ( P > 0.05). Conclusion The obvious inflammatory response in lung tissue can be induced by the poisonous gas released from the combustion of nonmetal materials in airtight cabin, and activate the expression of kina- ses in MAPK pathway, which may be one of mechanisms for lung injury induced by poisonous gas.