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血红素加氧酶-1对高氧急性肺损伤大鼠Ⅱ型肺泡上皮细胞凋亡的影响

Effect of heme oxygenase-1 on the apoptosis of type Ⅱ alveolar epithelial cells in rats with hyperoxia-induced acute lung injury

摘要:

目的 探讨血红素加氧酶-1(HO-1)对高氧急性肺损伤(HALI)大鼠Ⅱ型肺泡上皮细胞(AECⅡ)凋亡的影响.方法 按随机数字表法将24只健康雄性SD大鼠分为对照组(空气中饲养)、HALI模型组(高氧箱中饲养,氧浓度90%以上,温度25~27℃,湿度50%~70%,CO2浓度<0.5%)、HO-1干预组(高氧饲养前给予0.2 μmol/L 的HO-1)、HO-1抑制剂组〔高氧饲养前给予20 μmol/L的锌原卟啉Ⅸ(ZnPP)〕,每组6只.高氧处理48 h后处死大鼠,取左上肺组织行苏木素-伊红(HE)染色,光镜下观察肺组织病理学改变;取左下肺测量湿/干重(W/D)比值;取右肺组织提取AECⅡ,用流式细胞仪检测细胞凋亡率,用蛋白质免疫印迹试验(Western Blot)检测凋亡相关蛋白Bcl-2、天冬氨酸特异性半胱氨酸蛋白酶3(caspase-3)的表达.结果 ① 光镜下观察可见:对照组肺组织结构清晰;HALI模型组及HO-1抑制剂组肺组织结构紊乱,出现肺泡壁断裂破坏融合成肺大泡,肺泡间隔明显水肿、增宽,大量炎性细胞浸润,肺泡腔内出现水肿液及炎性细胞;HO-1干预组肺组织病理学改变较HALI模型组明显减轻.② 与对照组比较,HALI模型组肺W/D比值、AECⅡ凋亡率、Bcl-2蛋白表达明显增加,caspase-3蛋白表达明显降低〔肺W/D比值:4.61±0.41比3.68±0.45,AECⅡ凋亡率:(42.44±0.93)%比(24.74±0.64)%,Bcl-2蛋白表达(IA值):0.72±0.18比0.41±0.12,caspase-3蛋白表达(IA值):1.32±0.32比1.81±0.69,均P<0.05〕.与HALI模型组比较,HO-1干预组肺W/D比值、AECⅡ凋亡率、Bcl-2蛋白表达明显降低,caspase-3蛋白表达明显增加〔肺W/D比值:3.82±0.28比4.61±0.41,AECⅡ凋亡率:(26.67±1.58)%比(42.44±0.93)%,Bcl-2蛋白表达(IA值):0.39±0.08比0.72±0.18,caspase-3蛋白表达(IA值):1.78±0.46比1.32±0.32,均P<0.05〕.HO-1抑制剂组各指标与HALI模型组比较差异均无统计学意义.结论HO-1可降低HALI大鼠AECⅡ凋亡率,其调节机制可能与凋亡相关蛋白Bcl-2及caspase-3表达有关.

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abstracts:

Objective To investigate the effect of heme oxygenase-1 (HO-1) on the apoptosis of type Ⅱalveolar epithelial cells (AEC-Ⅱ) in rats with hyperoxia-induced acute lung injury (HALI). Methods Twenty-four healthy male Sprague-Dawley (SD) rats were randomly divided into 4 groups (n = 6): control group, HALI group, HO-1 group, and HO-1 inhibition group. The control group was fed in the room air; the HALI group was fed in the hyperoxia box (the oxygen concentration was more than 90%, the temperature was kept at 25-27 ℃, the humidity was maintained at 50%-70%, and the CO2concentration was less than 0.5%); the HO-1 group was fed in the hyperoxia box after HO-1 (0.2 μmol/L) treatment; and the HO-1 inhibition group was fed in the hyperoxia box after treatment with zinc protoporphyrin Ⅸ (20 μmol/L). After 48 hours of hyperoxia treatment, rats were sacrificed, left upper lung tissue was stained with hematoxylin-eosin (HE) and the pathological changes of lung tissue were observed under light microscope. The ratio of wet/dry weight (W/D) was measured in the lower left lung. AECⅡ was extracted from the right lung tissue, the apoptosis rate was detected by flow cytometry, and the expressions of apoptosis-related proteins Bcl-2 and caspase-3 were detected by Western Blot. Results ①It was shown by light microscopy that the lung tissue structure of the control group was clear. In HALI group and HO-1 inhibitor group, the lung tissue structure was disordered, alveolar wall was broken and fused into pulmonary alveoli, alveolar septum was obviously swollen and widened, a large number of inflammatory cells infiltrated, and edema fluid and inflammatory cells appeared in alveolar cavity. The pathological changes of lung tissue in HO-1 group were significantly less than those in HALI group. ② Compared with the control group, the lung W/D ratio, the apoptosis rate of AECⅡand the expression of Bcl-2 protein in the HALI group and the HO-1 inhibitor group were significantly increased, and the expression of caspase-3 was significantly decreased [lung W/D ratio: 4.61±0.41 vs. 3.68±0.45, apoptosis rate of AECⅡ: (42.44±0.93) % vs. (24.74±0.64) %, Bcl-2 (integral absorbance): 0.72±0.18 vs. 0.41±0.12, caspase-3 (integral absorbance): 1.32±0.32 vs. 1.81±0.69, all P < 0.05]. Compared with the HALI group, the lung W/D ratio, the apoptosis rate of AECⅡ, the expression of Bcl-2 protein in HO-1 group were significantly decreased, and the expression of caspase-3 was significantly increased [lung W/D ratio: 3.82±0.28 vs. 4.61±0.41, apoptosis rate of AECⅡ: (26.67±1.58) % vs. (42.44±0.93) %, Bcl-2 (integral absorbance): 0.39±0.08 vs. 0.72±0.18, caspase-3 (integral absorbance): 1.78±0.46 vs. 1.32±0.32, all P < 0.05]. There was no significant difference between HO-1 inhibitor group and HALI group. Conclusions HO-1 can reduce the apoptosis rate of AECⅡin rats with HALI, which may be related to the expressions of apoptosis related proteins Bcl-2 and caspase-3.

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作者: 刘国跃 [1] 陈涛 [1] 陈博文 [1]
期刊: 《中华危重病急救医学》2018年30卷10期 1001-1005页 MEDLINEISTICPKUCSCD
栏目名称: 基础研究
DOI: 10.3760/cma.j.issn.2095-4352.2018.10.020
发布时间: 2018-12-05
基金项目:
贵州省科技计划项目(黔科合LH字[2017]7115) Guizhou Province Science and Technology Program Project of China
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