海水淹溺型急性肺损伤兔外周血中性粒细胞凋亡的动态变化

Dynamic changes in apoptosis of polymorphonuclear neutrophil in rabbit with acute lung injury induced by seawater drowning

目的探讨海水淹溺型急性肺损伤(SWD-ALI)时外周血中性粒细胞(PMN)凋亡的演变规律以及细胞因子对其可能的影响。方法36只新西兰兔用随机数字表法均分为对照组(作为0h基线值)和海水灌注后1、3、6、12、24h组(分别于灌注后1、3、6、12、24h处死)。观察各组血象和血气分析指标的变化,计算肺湿/干重比(W/D)和肺微血管通透指数;流式细胞术检测PMN凋亡情况,ELISA法检测血清TNF-α、IL-1β、IL-10水平;对肺组织进行病理学观察,并计算病理评分(LPS)。结果W/D值在3h达高峰,肺微血管通透指数以6h最高,炎症细胞浸润以6～12h最显著。LPS在1h已显著高于对照组(P<0.01),6h达最高。PMN凋亡率于灌注后1h一过性升高,3h显著抑制,6h最低,以后逐渐接近对照水平。PMN计数与凋亡率呈负相关(P<0.05)。各海水灌注组血清TNF-α、IL-1β、IL-10均显著高于对照组(P<0.05或P<0.01),与LPS呈正相关(P<0.05或P<0.01),但与PMN凋亡率无相关性(P>0.05)。结论PMN凋亡在SWD-ALI炎症反应早期受到显著抑制,是最主要的炎症效应细胞。TNF-α、IL-1β、IL-10在SWD-ALI中发挥重要作用,但对PMN凋亡抑制无显著影响。

Objective To explore the dynamic changes in apoptosis of polymorphonuclear neutrophil （PMN） during the course of seawater drowning-induced acute lung injury （SWD-ALI）, and the possible roles of cytokines in the process. Methods Thirty-six NZ rabbits were randomly divided into 6 groups by using a random number table., control group （0h, as baseline value） and 5 seawater drowning groups （S1h, S3h, S6h, S12h and S24h）. Blood gas analysis and hemogram were observed at different time points. The ratio of wet to dry lung weight （W/D） and lung permeability index （LPI） were calculated. The changes in PMN apoptosis were analyzed by flow cytometer. The concentrations of tumor necrosis factor alpha （TNF-α）, interleukin- （IL-1） and IL-10 in serum were detected by ELISA. Meanwhile, the pathological changes in lung tissue were examined with hematoxylin-eosin （HE） staining, and lung pathologic scores （LPS） were evaluated. Results The W/D ratio reached its peak value at 3h, while LPI at 6h. The histopathologic findings indicated that inflammatory cell infiltration was most massive at 6-12h. LPS increased at 1h, significantly higher than that of control group （P〈0. 01）, while reached its peak level at 6h. The apoptotic rate of PMNs rose transiently at lh after seawater drowning, then decreased significantly at 3h, reaching its lowest value at 6h, and then returned slowly to baseline value from 12h to 24h. There was a negative correlation between the apoptotie rate and cell count of PMN （P〈0. 05）. The concentrations of TNF-α, IL-1 and IL-10 were significantly higher in the seawater drowning groups than i control group （P〈0. 05 or P〈0.01）. The levels of serum cytokines was positively correlated with LPS （P〈0. 05 or P〈0. 01）, but it had no correlation with the apoptotic rate of PMN （P〉0. 05）. Conclusions The apoptosis of PMN, the main inflammatory effector cells, is significantly inhibited in the early stage of SWD-ALI. The serum cytokines, TNF-α, IL-1 and IL-10, play important roles in the pathogenesis of SWD-ALI, but have no evident effects on the inhibition of PMN apoptosis.