p75^NTR蛋白体外通过TrkA^NGFR／p75^NTR异源二聚体信号途径促进L02细胞增殖  

P75^NTR protein promotes L02 cells proliferation in vitro via TrkANGFR/P75^NTR heterodimer signal pathway

作　　者：李俊峰[1] 陈莲香[2] 吕霞[2] 舒建昌[2] 

机构地区：[1]暨南大学附属第一医院消化科,广州510630 [2]暨南大学附属第四医院消化科

出　　处：《中华消化杂志》2012年第4期256-261,共6页Chinese Journal of Digestion

基　　金：基金项目：广东省自然科学基金管理委员会科研项目（9151051501000083）

摘　　要：目的观察神经生长因子（NGF）及受体（TrkA^NGFR、P75^NTR）在肝细胞中的表达，探讨外源性P75^NTR蛋白对肝细胞的生物学作用。方法体外培养L02肝细胞，免疫细胞化学和荧光定量PCR法分别检测NGF、TrkA^NGFR、P75^NTR在L02细胞中的表达。XTT法检测外源性P75^NTR、NGF、NGF＋P75^NTR、抗TrkA^NGFR、抗P75^NTR对L02细胞增殖的作用。流式细胞术（膜联蛋白V／碘化丙啶）检测外源性P75^NTR对L02细胞凋亡的作用。流式细胞术（碘化丙啶）检测外源性P75^NTR对L02细胞周期的影响。结果P75^NTR促进L02细胞增殖，呈剂量依赖性。NGF和NGF＋P75^NTR组吸光度值（A）值分别为0．4916±0．0565和0．5839±0．0733，与阴性对照组比较差异有统计学意义（0．3601±0．0310，P〈0．05）；抗TrkA^NGFR A值为0．2689±0．0229，与阴性对照组比较差异有统计学意义（P=0．003）；抗P75^NTR A值为0．3524±0．0312，与阴性对照组比较差异无统计学意义（P=1．000）。外源性P75^NTR对L02细胞有抗凋亡趋势，加入100ng／mlP75^NTR抗凋亡作用最强，表达量为3．70±0．26，但与对照组比较差异无统计学意义（4．10±0．62，P=1．000）。P75^NTR作用于L02细胞周期的S期，呈剂量依赖性，呈倒置的U形曲线，浓度为100ng／ml时作用最强（25．60±0．40），与对照组比较差异有统计学意义（20．10±1．00，P=0．000）。外源性NGF、P75^NTR、NGF＋P75^NTR上调了L02细胞NGFmRNA、TrkA^NGFR mRNA、P75^NTR mRNA的基因表达，与对照组比较差异有统计学意义（P〈0．05）；抗TrkA^NGFR、抗P75^NTR对NGFmRNA、TrkA^NGFR mRNA、P75^NTR mRNA的基因表达无明显影响，与对照组比较差异无统计学意义（P〉0．05）。结论L02细胞表达NGF及受体TrkA^NGFR、P75^NTR，合适剂量的外源性P75^NTR可能通过TrkA^NGFR／P75^NTR异源二聚体信号途径促进L02细胞增殖。Objective To observe the expression of nerve growth factor（NGF） and its receptors （TrkA^NGFR and P75^NTR） in hepatocytes and to explore the biological effects of exogenous P75^NTR protein on hepatocytes. Methods L02 hepatocytes were cultured in vitro. The expression of NGF, TrkA^NGFR and P75NTR in L02 cells were assessed by immunocytochemistry and fluorescent quantitation polymerase chain reaction （PCR）. The effects on L02 cell proliferation by exogenous P75^NTR, NGF, NGF＋ P75^NTR , anti-TrkA^NGFR and anti-P75^NTR were detected by XTT assay. The effect of exogenous P75^NTR on L02 cell apoptosis was measured by flow cytometry （Annexin V/PI） and the effect of exogenous P75^NTR on L02 cell cycle was determined by flow cytometry （PI）. Results L02 cell proliferation was promoted by P75NTR and in dose-dependent manner. The A value of NGF group and NGF-kP75^NTR group was 0.4916±0. 0565 and 0. 5839±0. 0733, respectively., and there was statistical significance compared with control group （0. 3601± 0. 0310, P〈0.05）. The A value of anti-TrkA^NGFR group was 0. 2689±0.0229, and there was statistical significance compared with control group （P=0. 003）. The A value of anti-P75^NTR was 0. 3524 ± 0. 0312, and there was no statistical significance compared with control group （P = 1. 000）. Exogenous P75^NTR had anti-apoptosis effect on L02 cells, the antiapoptosis effect was strongest when 100 ng/ml P75^NTR was added and the expression quantity was 3.70 ±0.26. However there was no statistical significant compared with control group （4.10 ± 0.62, P= 1. 000）. P75^NTR affected the cell cycle S phase of L02 cells and in dose-dependent manner, which was inverted U-shaped curve. The effect was strongest when the concentration was 100 ng/ml （25.60± 0.40） and there was statistical significance compared with control group （20.10 ± 1.00, P= 0, 000）, Exogenous NGF, P75^NTR and NGF q-P75^NTR up regulated the gene expression of NGFmRNA, TrkA^NGFR mRNA and P75^NTR mRNA in L02 cells and

关 键 词：神经组织蛋白质类 受体 神经生长因子类 肝 细胞分裂 细胞凋亡 细胞 培养的 基因表达 

分 类 号：R363[医药卫生—病理学]