博莱霉素诱导人肺泡上皮细胞过程中核转录因子-κB对核因子2相关因子2信号通路的调控机制  

Mechanism of nuclear factor-κB signaling pathway regulated nuclear factor erythroid 2-related factor 2 in human alveolar epithelial cells by bleomycin-induced

作　　者：赵印敏[1] 汤中文[2] 郑卉[3] 粟波[1] 唐亮[1] 陈昶[3] 

机构地区：[1]同济大学附属上海市肺科医院中心实验室,上海200433 [2]武汉市医疗救治中心 [3]同济大学附属上海市肺科医院胸外科,上海200433

出　　处：《中华实验外科杂志》2014年第8期1650-1653,共4页Chinese Journal of Experimental Surgery

基　　金：上海市科委资助项目（10JC1413500）

摘　　要：目的 观察人肺泡上皮细胞HPAEpiC中核转录因子-κB （NF-κB）基因沉默前后核因子2相关因子2（Nrf2）及其相关表达产物含量的变化,探讨博莱霉素诱导的过程中NF-κB对Nrf2信号通路的调控机制.方法 应用慢病毒技术沉默人肺泡上皮细胞HPAEpiC中的NF-κB基因,采用荧光定量聚合酶链反应（FQ-PCR）技术检测博莱霉素诱导前后HPAEpiC细胞中的Nrf2、胞浆蛋白伴侣分子（Keap1）、Cullin3蛋白（Cul3）、Rbx1、蛋白激酶C（PKC）、磷酸肌醇3激酶（PI3K）、细胞外信号调节激酶（ERK）、NF-κB、κB抑制蛋白（IκB）的mRNA相对表达量,采用流式细胞术膜联蛋白V（Annexin V）-红色荧光蛋白（RFP）单染法检测各实验组的细胞凋亡率,采用Western blot检测NF-κB、Nrf2及其相关蛋白的表达量.结果 慢病毒感染HPAEpiC细胞后NF-κB基因表达下降90.6％,博莱霉素诱导后NF-κB沉默的人肺泡上皮细胞中Nrf2基因表达量上升142.6％,并与NF-κB、κB抑制蛋白（IκB）、Keap1、Cul3、Rbx1表达呈负相关,PI3K表达增加11.6倍而PKC、ERK无明显变化,其凋亡率与对照组比较增高.结论 在博莱霉素诱导的过程中,HPAEpiC细胞的NF-κB表达量增加可以抑制Nrf2的功能.在NF-κB被沉默的情况下,Nrf2通过PI3K途径并同时降低Keap1-Cu13-RBx1复合物表达实现活化,可能通过启动其下游的抗氧化反应,减少博莱霉素导致的氧自由基对细胞的损伤.Objective Observe nuclear factor （NF）-erythroid 2-related factor 2 （Nrf2） and its related gene expression changes after silencing of NF-κB in human alveolar epithelial cell （HPAEpiC） and to explore the mechanism of NF-κB signaling pathway regulated Nrf2 expression by bleomycin-induced.Methods Silencing of NF-κB using lentiviral technology in human alveolar epithelial cell （HPAEpiC）.Using real-time fluorescent quantitative polymerase chain reaction （FQ-PCR） detect mRNA expression of Nrf2,Keap1,Cul3,Rbx1,protein kinase C （PKC）,phosphatidylinositol 3 kinase （PI3K）,extracellular signalregulated kinase （ERK）,NF-κB and inhibitory κB （IκB） in human alveolar epithelial cells by Bleomycininduced.Cell apoptosis were determined by Annexin V-RFP single staining.NF-κB,Nrf2 and the associatedprotein expression were determined by Western blotting.Results Lentiviral infected HPAEpiC cells NF-κBgene expression decreased 90.6％,HPAEpiC cells which NF-κB silenced the gene expression of Nrf2 in-creased 142.6％ after Bleomycin-induced,it negatively correlated with NF-κB,IκB,Keap1,Cul3,Rbx1expression.PI3K expression increased 11.6-fold and PKC,ERK no significant change.The apoptosis rateincreased compared with the control group.Conclusion In process of bleomycin-induced,NF-κB in humanalveolar epithelial cell （HPAEpiC） can inhibit the function of Nrf2.NF-κB in the case of silence,Nrf2 viathe PI3K pathway while reducing Keap1-Cul3-RBx1 complex expression achieve activation,it may start itsdownstream oxidation reaction and reduce that bleomycin oxygen radicals cause damage to cells.

关 键 词：肺纤维化 核转录因子-ΚB 核因子2相关因子2 信号转导 

分 类 号：R363[医药卫生—病理学]