靶向小干扰RNA对小鼠肺缺血/再灌注损伤时细胞凋亡及CCAAT/增强子结合蛋白同源蛋白的作用

doi:10.3760/cma.j.issn.1001-9030.2013.08.018

科研成果详情

题名	靶向小干扰RNA对小鼠肺缺血/再灌注损伤时细胞凋亡及CCAAT/增强子结合蛋白同源蛋白的作用
其他题名	Effects of targeted small interfering RNA on pneumocyte apoptosis and CCAAT/enhancer-binding protein homologous protein in pulmonary ischemia/reperfusion injury in mice
作者	周俊辉 1,2,3; 王良荣1; 郝卯林2; 应磊2; 孙勤 4; 王万铁2
发表日期	2013-08-08
发表期刊	中华实验外科杂志影响因子和分区
语种	中文
原始文献类型	学术期刊
关键词	CCAAT/增强子结合蛋白同源蛋白肺缺血再灌注损伤小干扰RNA 脱噬作用
其他关键词	C/EBP homologous protein ; Lung ischemia ; Reperfusion injure ; Small interfering RNA ; Apoptosis
摘要	目的观察靶向小干扰RNA(siRNA)对小鼠肺缺血/再灌注损伤(PIRI)时细胞凋亡及CCAAT/增强子结合蛋白同源蛋白(CHOP)的影响。方法荧光标记法观察靶向siRNA在小鼠体内的分布并评估转染效率。C57BL/6J小鼠50只,随机分为5组(n=10):假手术组(Sham组)、缺血/再灌注组(I/R组)、I/R+载体组(I/R+Vehicle组)、I/R+阴性对照组(I/R+Control-siRNA组)和I/R+CHOP-siRNA组(L/R+CHOP-siRNA组)。取左肺,检测肺湿/干重比(W/D)、总肺水含量(TLW)和肺泡损伤定量评估(IQA),光镜和电镜观察肺组织结构改变,逆转录-聚合酶链反应(RT-PCR)、Western blot检测CHOP、葡萄糖调节蛋白78(GRF78)mRNA和蛋白表达水平,原位末端转移酶标记(TUNEL)法检测细胞凋亡指数(AI)。结果靶向siRNA通过滴鼻法可有效分布于肺脏中。与Sham组比较,I/R组中CHOP、GRP78 mRNA和蛋白表达水平(0.80±0.03、0.80±0.02;0.80±0.04、0.84±0.02)均升高(P<0.05),W/D(5.24±0.49)、TLW(4.24±0.49)、IQA[(42.80±4.21)%]和AI[(34.50±1.51)%]均明显增加(P<0.01);肺组织形态结构发生明显损伤;与I/R+Vehicle组和I/R+Control-siRNA组比较,I/R+CHOP-siRNA组中GRP78 mRNA和蛋白表达水平(0.84±0.04、0.85±0.04)无明显变化(P>0.05),而CHOP mRNA和蛋白表达水平(0.40±0.03、0.44±0.04)均降低(P<0.05),W/D(2.54±0.24)、TLW(1.54±0.24)、IQA[(16.71±2.33)%]和AI[(11.50±1.58)%]亦均降低(P<0.01);肺组织形态结构损伤减轻。结论靶向siRNA对I/R损伤肺具有良好的保护作用,其机制可能与其对抗过度的未折叠蛋白反应(UPR)中CHOP介导的细胞凋亡有关。
其他摘要	Objective To explore the effects of targeted small interfering RNA (siRNA) on pneumocyte apoptosis and CCAAT/enhancer-binding protein homologous protein (CHOP) in pulmonary ischemia/reperfusion injury (PIRI) in mice. Methods The location of targeted siRNA and transfection efficiency in mice were determined by using fluorescence labeling method. Fifty mice were randomly allocated into five groups (n = 10): sham operation group (Sham group), ischemia/reperfusion group (I/R group), I/R + vehicle group (I/R + Vehicle group), I/R + negative control group (I/R + Control-siRNA group), and I/R + CHOP-siRNA group (I/R + CHOP-siRNA group). Left lung tissue was excised. Wet lung weight to dry lung weight (W/D), total lung water content (TLW) and index of quantitative evaluation for alveolar damage (IQA) were tested. Under light microscope and electron microscope, changes in lung tissues were observed. The expression levels of CHOP and glucose regulated protein (GRP78) was detected by using reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blotting. Apoptosis index (AI) of lung tissue was measured by using TdT-mediated dUTP nick end labeling (TUNEL) method. Results Targeted siRNA could be effectively distributed in the lung by intranasal delivery. Compared to Sham group, the expression levels of CHOP and GRP78 mRNA and protein (0. 80 ±0. 03,0. 80 ± 0. 02; 0.80 ±0.04,0.84 ±0.02) were all significantly increased (P < 0.05) in I/R group, and W/D (5.24 ±0.49),TLW (4. 24 ±0.49), IQA [(42. 80 ±4. 21)%] and AI [(34. 50 ±1.51)%] were all very notably increased (P <0.01). The structural injury in lung tissue was notably observed in I/R group. Compared to I/R + Vehicle group and I/R + Control-siRNA group, there was no significant difference (P > 0. 05) in the expression levels of GRP78 mRNA and protein (0. 84 ± 0. 04; 0. 85 ± 0. 04) in I/R + CHOP-siRNA group, but the expression levels of CHOP mRNA and protein (0. 40 ±0. 03,0. 44 ±0. 04) were reduced (P<0.05), W/D (2.54±0.24), TLW (1.54 ±0.24),IQA [(16.71 ±2.33)%] and AI [(11.50 ± 1.58)% ] were all decreased (P <0.01). The structural injury in lung tissue was alleviated in I/R + CHOP-siRNA group. Conclusion Targeted siRNA has great effect on lung protection against I/R injury, which may be related to inhibition of pneumocyte apoptosis mediated by CHOP in excessive unfolded protein response (UPR).
资助项目	浙江省公益技术应用研究项目（2011C37092）;浙江省医药卫生科技计划项目（2010KYA132）;浙江省中医药科技计划项目（2010ZB088）;浙江省中医药重点研究计划项目（013Z2011）;浙江省中医药重点研究学科建设计划项目（2012-XK-A28）
ISSN	1001-9030
卷号	30 期号:08 页码:1608-1611+1783
DOI	10.3760/cma.j.issn.1001-9030.2013.08.018
页数	5
收录类别	CNKI ; 万方 ; 北大核心 ; ISTIC ; CSCD ; PKU
学科领域	医药、卫生 ; 外科学
URL	查看原文
CSCD记录号	CSCD:4926069
引用统计
文献类型	期刊论文
条目标识符	https://kms.wmu.edu.cn/handle/3ETUA0LF/53353
专题	附属第一医院其他_附属慈溪医院（慈溪市人民医院）
作者单位	1.温州医科大学附属第一医院麻醉科; 2.温州医科大学缺血/再灌注损伤研究室; 3.郑州市人民医院麻醉科; 4.温州医科大学附属慈溪医院ICU
第一作者单位	附属第一医院; 麻醉科; 温州医科大学
第一作者的第一单位	附属第一医院
推荐引用方式 GB/T 7714	周俊辉,王良荣,郝卯林,等. 靶向小干扰RNA对小鼠肺缺血/再灌注损伤时细胞凋亡及CCAAT/增强子结合蛋白同源蛋白的作用[J]. 中华实验外科杂志,2013,30(08):1608-1611+1783.
APA	周俊辉, 王良荣, 郝卯林, 应磊, 孙勤, & 王万铁. (2013). 靶向小干扰RNA对小鼠肺缺血/再灌注损伤时细胞凋亡及CCAAT/增强子结合蛋白同源蛋白的作用. 中华实验外科杂志, 30(08), 1608-1611+1783.
MLA	周俊辉,et al."靶向小干扰RNA对小鼠肺缺血/再灌注损伤时细胞凋亡及CCAAT/增强子结合蛋白同源蛋白的作用".中华实验外科杂志 30.08(2013):1608-1611+1783.