摘要
目的探讨内毒素（LPS）激活X盒结合蛋白1（XBP1）信号转导通路对移植肝缺血再灌注损伤的影响及其机制。方法以雄性SD大鼠为供、受者，分为冷缺血转染组、活体转染组和对照组，以两袖套法建立肝移植模型。冷缺血转染组大鼠于冷缺血期经门静脉灌注转染携带XBP1短发夹干扰RNA的质粒（pSIXBP1）；活体转染组大鼠在门静脉袖套吻合完成后经门静脉分支注入pSIXBPI；对照组不予任何处理。分别于移植肝门静脉恢复再灌注后60和180min处死大鼠，光镜观察肝组织病理损害程度，逆转录聚合酶链反应及蛋白免疫印记法测定肝组织XBP1mRNA和XBP的表达水平；酶连免疫吸附试验检测肝组织核因子xB（NF-~B）的活性及血清肿瘤坏死因子a（TNF-n）的含量。结果再灌注后180min，冷缺血转染组病理损害程度、XBP1mRNA含量和XBP1含量低于活体转染组和对照组（P〈0．05），该组TNF-a的表达水平为（584．94±15．97）pg／ml，低于活体转染组和对照组（P〈0．05）。而再灌注后180min时，3组NF-KB活性的差异无统计学意义（P〉0．05）。结论XBP1通路与NF-kB通路在大鼠肝脏缺血再灌注损伤中对TNF-a基因的调控作用是两个相对独立的环节，XBP1短发夹RNA干扰技术能有效减轻肝移植时的缺血再灌注损伤。
Objective To explore the regulation mechanism of X box binding protein 1 （XBPI） signal transduction pathway for TNF-a and effective approach in ischemia/reperfusion （I/R） injury of liver transplantation for short hairpin RNA （shRNA） interference used to gene therapy in liver graft. Methods Male Sprague-Dawley rats were divided into three groups： the cold ischemia transfection group （CIT）, the in vivo transfection group （IVT） and the control group. Experiments of ortbotopic liver transplantation were performed by two cuff method. The rats in CIT were perfused with XBP1- shRNA plasmid （pSIXBP1） during cold ischemia phase, those in IVT received the equivalent volume （2 ml） of pSIIRAK 4 after portal vein inoculation, and those in the control group were not subjected to any treatment. Rats were killed at 60 or 180 rain after restoring reperfusion of hepatic portal vein. Histopathological damage degree of graft liver was observed by light microscope. The expression levels of XBP1 gene and protein were detected by RT-PCR and Western blotting. The activities of NF- KB and the serum TNF-a level were detected by ELISA. Results All the indexes including the degree of histopathological damage, the expression levels of XBP1 mRNA and protein and the TNF-a level were significantly decreased in CIT as compared with IVT and control group （P〈0. 05）. However, there was no significant difference in NF-kB activity among the three groups （P〉0. 05）. Conclusion The role of XBP1 pathway in TNF-a gene regulation and that of NF-kB pathway in rat liver I/R injury are two relatively independent aspects, and the depression of XBP1 expression with XBP1 shRNA through portal vein perfusion during cold ischemia phase could effectively alleviate graft hepatic I/R injury.
出处
《中华器官移植杂志》
CAS
CSCD
北大核心
2011年第2期69-72,共4页
Chinese Journal of Organ Transplantation
基金
基金项目：国家自然科学基金（30801126）
关键词
肝移植
再灌注损伤
X盒结合蛋白1
Liver transplantation
Reperfusion injury
X-box binding protein 1
