CAJ | 학술논문

为研究外源性血管紧张素Ⅱ(angiotensin Ⅱ,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(175～200 g)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20 h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15 min静脉注入BODIPY-白蛋白).TUNEL测定显示,灌注ANG 6 h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所敛的DNA片断增加可被同时灌注ZVAD-frnk或Iosartan阻断.灌注ANG后免疫标定caspase 3 阳性细胞数量显著增多(P<0.01),ZVAD-fmk或losartan 同样显著减少caspase 3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.
위연구외원성혈관긴장소Ⅱ(angiotensin Ⅱ,ANG)재급성폐손상화폐포상피세포조망중적작용,경기관분별급웅성Wistar대서(175～200 g)관주ANG、ANG가caspase억제제ZVAD-fmk、ANG가ANG수체1조단제losartan화부관주린산염완충용액(PBS).6혹20 h후재체관세동물폐장,측정관세액중혈홍단백(hemoglobin,Hb)화형광물질(BODIPY)표정적백단백함량(재관세전15 min정맥주입BODIPY-백단백).TUNEL측정현시,관주ANG 6 h후,지기관화폐포상피세포내표정적DNA편단현저증가(P<0.05);ANG소렴적DNA편단증가가피동시관주ZVAD-frnk혹Iosartan조단.관주ANG후면역표정caspase 3 양성세포수량현저증다(P<0.01),ZVAD-fmk혹losartan 동양현저감소caspase 3양성세포적수량.관주ANG현저증가폐포관세액중형광표정적백단백(P<0.01)화Hb적함량(P<0.05);ZVAD-fmk혹losartan역현저억제형광백단백화Hb함량적변화.결과표명,폐포상피세포재체폭로우외원성ANG족이인기ANG수체1개도적상피세포조망화폐포병장손상.
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 pmol/L),ANG plus the caspase inhibitor ZVAD.fmk(60 μmol/L),ANG plus the ANG receptor AT1 antagonist Iosartan(LOS,100 μmol/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 deoxvnucleotidyl 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 afterinstillationofANG(P<0.01);the salnedoses 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.