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经气管灌注血管紧张素Ⅱ导致凋亡性急性肺损伤

庄甲举, 李小鹏, Uhal B D, Yian K R

白求恩军医学院生理教研室.河北,石家庄 050081；美国密歇根州立大学生理学系.密歇根州东兰辛市 48824

摘要

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({sl 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({sl 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({sl P}<0.01)和Hb的含量({sl 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。

关键词： 凋亡; 肺泡上皮细胞; 急性肺损伤; Ⅱ型肺泡细胞

Apoptosis--dependent acute pulmonary injury after intratracheal instillation of angiotensin Ⅱ

Zhuang Jiaju, Li Xiaopeng, Uhal B D, Yian K R

Department of Physiology,Bethune Military Medical College.Shijiazhuang 050081,Hebei；USA

Abstract

To test the hypothesis that exogenous purified angiotensin Ⅱ(ANG) might cause apoptosis of alveolar epithelial cells(AECs) and acute lung injury,male Wistar rats were intratracheally instilled with purified ANG(10 #mu#mol/L),ANG plus the caspase inhibitor ZVAD-fmk(60 #mu#mol/L),ANG plus the ANG receptor AT1 antagonist losartan(LOS,100 #mu#mol/L) or sterile phosphate-buffered saline(PBS) vehicle alone.Six or 20 h later,the lungs were lavaged in situ for determination of bronchoalveolar lavage(BAL) fluid content of hemoglobin(Hb) and fluorescent(BODIPY)-albumin,a bolus of which was injected intravenously 15 min prior to BAL.Terminal deoxynucleotidyl transferase-mediated nick-end labeling(TUNEL) revealed that instillation of ANG,but not PBS alone,increased labeling of fragmented DNA in bronchiolar epithelial cells and in AECs(P<0.05) at 6 h post-ANG.Increased TUNEL was abrogated by concurrent instillation of ZVAD-fmk or LOS.Significant increased numbers of caspase-positive cells were observed by anti-caspase 3 immunolabeling after instillation of ANG(P<0.01);the same doses of LOS or ZVAD-fmk that blocked TUNEL also blocked the activation of caspase 3(P<0.01).Intratracheal instillation of ANG also remarkably increased BAL BODIPY-albumin(P< 0.01) and Hb(P<0.05),both of which were eliminated by ZVAD-fmk or LOS.These data indicate that exposure of AECs to ANG in vivo is sufficient to induce apoptosis and alveolar epithelial barrier injury mediated by ANG receptor AT1.

Key words: Apoptosis;alveolar epithelial cell;acute pulmonary injury;type Ⅱ pneumocyte

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引用本文：

庄甲举, 李小鹏, Uhal B D, Yian K R. 经气管灌注血管紧张素Ⅱ导致凋亡性急性肺损伤[J]. 生理学报 2008; 60 (6): 715-722.

Zhuang Jiaju, Li Xiaopeng, Uhal B D, Yian K R. Apoptosis--dependent acute pulmonary injury after intratracheal instillation of angiotensin Ⅱ. Acta Physiol Sin 2008; 60 (6): 715-722 (in Chinese with English abstract).