盐酸吸入性急性肺损伤大鼠肺组织细胞凋亡及异丙酚对其的影响

A study of lung tissue cell apoptosis in rats with acute lung injury caused by hydrochloric acid aspiration and the impact of propofol on it

目的 观察盐酸吸入性急性肺损伤（ALI）大鼠肺组织细胞凋亡情况，以及异丙酚对肺组织细胞凋 亡的影响。 方法 采用气管内滴注盐酸复制ALI 大鼠模型。将40 只成年健康雄性SD 大鼠按简单随机化方法 分为假手术组（腹腔注射等量生理盐水）、模型组、异丙酚预处理48 h 和24 h 组及异丙酚治疗1 h 组（分别于盐 酸滴注前48 h、24 h 及盐酸滴注后1 h 腹腔注射异丙酚10 mg/kg） 5 组，每组8只。盐酸吸入后6 h 测定大鼠动 脉血氧分压（PaO2）、氧合指数（PaO2/FiO2）、支气管肺泡灌洗液（BALF）总蛋白含量、肺湿/ 干重（W/D）比值及肺 组织丙二醛（MDA）含量；光镜下观察肺组织病理学改变；用原位末端缺刻标记法（TUNEL）检测肺组织细胞凋 亡程度；免疫组化法检测肺组织天冬氨酸特异性半胱氨酸蛋白酶3（caspase-3）蛋白的表达及分布.结果 模 型组大鼠PaO2/FiO（2 mm Hg，1 mm Hg=0.133 kPa）较假手术组显著降低（211±31比379±28，P ＜0.01），BALF总 蛋白含量（mg/L ：250.82±98.26 比63.05±16.56）、肺W/D 比值（4.76±0.09 比4.11±0.07）、MDA 含量（nmol/g ： 3.1±0.8 比2.3±0.6）均较假手术组显著增加（均P ＜0.01）；模型组肺组织细胞凋亡指数（AI）较假手术组显著 增加〔（14.15±1.29）% 比（3.52±0.50）%，P ＜0.01〕，caspase-3 蛋白表达较假手术组显著增加〔（17.95±2.73）% 比（7.37±2.56）%，P ＜0.01〕。与模型组比较，异丙酚预处理24 h 组、异丙酚治疗1 h 组PaO2/FiO2 显著升高， BALF总蛋白含量、肺W/D 比值、MDA 含量、AI 及caspase-3 蛋白表达量均明显下降，以异丙酚治疗1 h 组改善 最为显著〔PaO2/FiO2 ：326±37，BALF总蛋白含量：132.23±47.41，肺W/D 比值：4.28±0.03，MDA：1.5±0.3，AI： （6.35±1.27）%，caspase-3：（ 11.35±1.13）%，均P ＜0.01〕，但与假手术组比较差异仍有统计学意义（均P ＜0.01）； 而异丙酚预处理48 h 组各指标与模型组比较差异无统计学意义（均P ＞0.05）.结论 肺组织细胞凋亡的增加 可能参与了大鼠盐酸吸入性ALI 的发病，而caspase-3 途径可能是肺组织细胞凋亡增加的机制之一；异丙酚可减 轻ALI 程度，减少AI；抑制凋亡可能是异丙酚减轻ALI 的机制之一.

Objective To observe the lung tissue cell apoptosis in rats with acute lung injury （ALI） caused by hydrochloric acid （HCl） aspiration and to investigate the impact of propofol on the lung tissue cell apoptosis. Methods The rat ALI model was reproduced by HCl intra-tracheal instillation. Forty adult healthy male Sprague-Dawley （SD）rats were randmnly divided into five groups ： sham operation group （intra-peritoneal injection of a same amount of normal saline rather than HCl）, model group, propofol pretreatment 48-hour group, propofol pretreatment 24-hour group, and propofol treatment 1 hour group （intra-peritoneal injection of 10 mg/kg propofol 48 hours, 24 hours before and 1 hour after intra-tracheal instillation of HCl respectively）,8 rats being in each group. Six hours after aspiration of HCI, arterial partial pressure of oxygen （PaO2） was measured and the oxygenation index （PaO2/FiO2 ） was calculated, total protein content in bronchoalveolar lavage fluid （BALF）, malondialdehyde （MDA） and wet / dry weight ratio （W/D） of lung tissue were calculated respectively ; the pathological changes of lung tissue were observed by light microscope, the level of apoptosis in lung tissue cells was detected by terminal deoxynucleotidyl transferase-mediated nick end labeling （TUNEL） , and the expression and distribution of caspase-3 protein in lung tissues was also determined by immunohistoehemistry methods. Results PaO2/FiO2 （ram Hg, 1 mm Hg= 0.133 kPa） in model group was significantly lower than that in sham group （211±31 vs. 379±28, P〈0.01 ）. The total protein content in BA LF （ mg/L ： 250.82± 98.26 vs. 63.05 ± 16.56）, W/D ratio （ 4.76± 0.09 vs. 4.11 ± 0.07 ） and MDA levels of lung tissues （nmol/g ： 3.1 ±0.8 vs. 2.3±0.6）were all increased significantly in model group compared to those in sham group （all P〈0.01 ）. Apoptosis index （AI）of lung tissue cell in model group was increased obviously compared to that in sham group [ （14.15± 1.29）% vs. （3.52±0.50）%, P〈0.011. The level of caspase-3 protein expression in model group was markedly higher than that in sham group [ （17.95±2.73） % vs. （7.37±2.56）%, P〈0.01 ]. Compared with model group, PaO2/FiO2 was increased significantly, total protein content in BALF, W/D ratio, MDA content, AI and the level of caspase-3 protein expression were all decreased obviously in propofol pretreatment 24-hour group and in propofol treatment 1 hour group, and the most significant changes of all above indexes happened in propofol treatment1 hour group [ PaO2/FiO2 ： 326±37, total protein content in BALF ： 132.23±47.41, W/D ratio ： 4.28±0.03, MDA ： 1.5±0.3, AI ： （6.35± 1.27 ）%, easpase-3 ：（ 11.35± 1.13）%, all P〈0.01 ], but there were still statistically significant differences in all above indexes between pretreatment 24 hours group and sham group as well as between propofol treatment 1 hour group and sham group （all P〈0.01 ）. Meanwhile, all above indexes in propofol pretreatment 48-hour group had no statistically significant differences compared with those in model group （all P〉0.05）. Conclusion The apoptotic increase of lung tissue cells through caspase-3 pathway may be one of the mechanisms involved in the pathogenesis of ALl induced by HCl inhalation, and the inhibition of the apoptotic increase of lung tissue cells by using propofol may be one of the mechanisms to decrease the degree of the ALI and AI.