CAJ | 학술논문

目的研究大鼠急性肺栓塞(APE)后肺组织中线粒体前凋亡蛋白(ARTS)的表达变化及其诱导的细胞凋亡.方法采用自体血栓栓塞颈总动脉制备大鼠APE模型.分别在APE前(对照)和APE后1、8、24和48 h开胸取肺脏.常规提取肺组织的总RNA和总蛋白,用半定量逆转录-聚合酶链反应(RT-PCR)方法检测ARTS mRNA表达水平,用蛋白质免疫印迹法(Western blotting)进一步验证ARTS以及ARTS诱导凋亡相关蛋白Bcl-2、Bcl-xL、XIAP和H2Ax的表达变化,用免疫组化法检测肺组织中ARTS在APE前后的表达变化及其组织分布情况,用原位末端缺刻标记法(TUNEL)检测肺组织内细胞凋亡情况.结果 APE后ARTS的mRNA及蛋白表达水平均升高,于1 h和48 h升高最明显(P<O.05或P<0.01).免疫组化显示ARTS在APE前肺组织内表达量很低,不易检测;APE后48 h ARTS表达明显升高,主要分布在支气管黏膜上皮细胞和肺泡上皮细胞.TUNEL染色发现,随着ARTS表达升高,肺组织出现明显的细胞凋亡现象,同时ARTS诱导凋亡系统中凋亡抑制蛋白Bcl-2、Bcl-xL和XIAP的表达下降,凋亡标记蛋白H2Ax表达升高(P<O.05或P<0.01).结论大鼠APE后肺组织内ARTS表达明显升高,ARTS介导的凋亡系统在APE后细胞凋亡中发挥了重要作用.
목적 연구대서급성폐전새(APE)후폐조직중선립체전조망단백(ARTS)적표체변화급기유도적세포조망.방법 채용자체혈전전새경총동맥제비대서APE모형.분별재APE전(대조)화APE후1、8、24화48 h개흉취폐장.상규제취폐조직적총RNA화총단백,용반정량역전록-취합매련반응(RT-PCR)방법검측ARTS mRNA표체수평,용단백질면역인적법(Western blotting)진일보험증ARTS이급ARTS유도조망상관단백Bcl-2、Bcl-xL、XIAP화H2Ax적표체변화,용면역조화법검측폐조직중ARTS재APE전후적표체변화급기조직분포정황,용원위말단결각표기법(TUNEL)검측폐조직내세포조망정황.결과 APE후ARTS적mRNA급단백표체수평균승고,우1 h화48 h승고최명현(P<O.05혹P<0.01).면역조화현시ARTS재APE전폐조직내표체량흔저,불역검측;APE후48 h ARTS표체명현승고,주요분포재지기관점막상피세포화폐포상피세포.TUNEL염색발현,수착ARTS표체승고,폐조직출현명현적세포조망현상,동시ARTS유도조망계통중조망억제단백Bcl-2、Bcl-xL화XIAP적표체하강,조망표기단백H2Ax표체승고(P<O.05혹P<0.01).결론 대서APE후폐조직내ARTS표체명현승고,ARTS개도적조망계통재APE후세포조망중발휘료중요작용.
Objective To study the expression changes in apoptosis related protein in transforming growth factors-β signaling pathway (ARTS) in the lung tissues in a rat acute pulmonary embolism (APE) model and its effects on cell apoptosis.Methods A rat APE model was reproduced.Samples of lung tissues were harvested at time points of 1,8,24 and 48 hours after APE.Healthy rats were used as control.The changes in mRNA level of ARTS were identified by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR),and the changes in its protein level,and also Bcl-2,Bcl-xL,XIAP and H2Ax proteins,which were related with ARTS-mediated cell apoptosis, were determined by Western blotting.Immunohistochemical method was employed to study the distribution and expression changes in ARTS in the lung tissue before and after APE.Apoptotic cells in lung tissue sections were identified by the terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling (TUNEL) method.Resuits At different time points,the mRNA levels and the protein levels of ARTS significantly increased in the lung tissues of rats with APE at 1 hour and 48 hours(P<0.05 or P<0.01).The immunohistochemical study showed that ARTS had low expression levels and could not be detected in the normal lung tissue.but il was up-regulated obviously at 48 hours after APE,mainly expressed in the bronchial epithelium and the lung alveolar epithelium.Apoptotic cells could be observed in the lung tissue by TUNEL after APE and at the same time when the lung tissue cells exhibited lower levels of the anti-apoptotic proteins Bcl-2.Bcl-xL,and XIAP,as compared with controls.Apoptosis level (as evaluated by H2Ax,apoptotic marker staining) in the lung tissue cells was obviously raised compared with controls(P<0.05 or P<0.01).Conclusion The expression of ARTS is up-regulated after APE,and ARTS-mediated apoptosis plays an important role in the cell apoptosis of lung tissue.