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为研究外源性血管紧张素Ⅱ(angiotensinⅡ,ANG)在急性肺损伤和肺泡上皮细胞凋亡中的作用,经气管分别给雄性Wistar大鼠(175～200g)灌注ANG、ANG加caspase抑制剂ZVAD-fmk、ANG加ANG受体1阻断剂losartan和仅灌注磷酸盐缓冲溶液(PBS)。6或20h后在体灌洗动物肺脏,测定灌洗液中血红蛋白(hemoglobin,Hb)和荧光物质(BODIPY)标定的白蛋白含量(在灌洗前15min静脉注入BODIPY-白蛋白)。TUNEL测定显示,灌注ANG6h后,支气管和肺泡上皮细胞内标定的DNA片断显著增加(P<0.05);ANG所致的DNA片断增加可被同时灌注ZVAD-fmk或losartan阻断。灌注ANG后免疫标定caspase3阳性细胞数量显著增多(P<0.01),ZVAD-fmk或losartan同样显著减少caspase3阳性细胞的数量。灌注ANG显著增加肺泡灌洗液中荧光标定的白蛋白(P<0.01)和Hb的含量(P<0.05);ZVAD-fmk或losartan亦显著抑制荧光白蛋白和Hb含量的变化。结果表明,肺泡上皮细胞在体暴露于外源性ANG足以引起ANG受体1介导的上皮细胞凋亡和肺泡屏障损伤。
To investigate the effect of exogenous angiotensin Ⅱ (Angiotensin Ⅱ) on acute lung injury and alveolar epithelial cell apoptosis, male Wistar rats (175 ~ 200g) were infused with ANG and ANG with caspase inhibitor ZVAD -fmk, ANG plus ANG receptor 1 blocker losartan and only perfusion with phosphate buffered saline (PBS). After 6 or 20 h, the albumin contents of the lavage fluid in the lavage were measured and the albumin content of the hemoglobin (Hb) and fluorescent substance (BODIPY) in the lavage was measured (intravenous injection of BODIPY-albumin 15 min before lavage). TUNEL assay showed that the DNA fragments in bronchial and alveolar epithelial cells were significantly increased after ANG6h perfusion (P <0.05). The increase of ANG-induced DNA fragments could be blocked by simultaneous infusion of ZVAD-fmk or losartan. After immunization with ANG, the number of immuno-labeled caspase3 positive cells was significantly increased (P <0.01). The number of caspase 3 positive cells was also significantly decreased by ZVAD-fmk or losartan. Angiotensin Ⅱ (Angiotensin II) increased the levels of albumin (P <0.01) and Hb (P <0.05) in bronchoalveolar lavage fluid, while ZVAD-fmk or losartan also significantly inhibited the changes of fluorescence albumin and Hb. The results showed that exposure of alveolar epithelial cells to exogenous ANG was sufficient to induce ANG receptor 1-mediated epithelial cell apoptosis and alveolar barrier injury.