PI3K/Akt信号通路在异丙酚抑制人肝癌细胞侵袭中的作用

Role of PI3K/Akt signaling pathway in propofol-induced invasion of human liver cancer cell line HepG2

目的评价磷脂酰肌醇3-激酶（PI3K）/蛋白质丝氨酸苏氨酸激酶（Akt）信号通路在异丙酚抑制人肝癌细胞侵袭中的作用。 方法将HepG2细胞用含10%胎牛血清的RPMI 1640培养基于37 ℃、5%CO2的培养箱中培养，当细胞稳定进入对数生长期时，采用随机数字表法分为6组（n＝18）：对照组（C组）；异丙酚组（P组）加入120 μg/ml异丙酚；PI3K/Akt信号通路激动剂IGF-1组（IGF组）加入10 nmol/L IGF-1；PI3K/Akt信号通路抑制剂LY294002组（LY组）加入10 μmol LY294002；IGF-1 ＋异丙酚组（IGF＋P组）先加入10 nmol/L IGF-1孵育24 h，再加入120 μg/ml异丙酚；LY294002 ＋异丙酚组（LY ＋ P组）先加入10 μmol LY294002孵育24 h，再加入120 μg/ml异丙酚。于孵育24 h时，采用Transwell法检测细胞的侵袭力，采用RT-PCR法检测PI3K和Akt的mRNA表达，采用Western blot法检测Akt、PI3K及磷酸化Akt（p-Akt）的表达。 结果与C组比较，P组、P＋IGF组、LY组和P＋LY组侵袭细胞计数减少，细胞PI3K及其mRNA表达下调，p-Akt/Akt比值下降，Akt mRNA表达下调，IGF组侵袭细胞计数增加，细胞PI3K及其mRNA表达上调，p-Akt/Akt比值升高，Akt mRNA表达上调（P〈0.05）；与P组比较，P＋IGF组侵袭细胞计数增多，细胞PI3K及其mRNA表达上调，p-Akt/Akt比值升高，Akt mRNA表达上调，P＋LY组侵袭细胞计数减少，细胞PI3K及其mRNA表达下调，p-Akt/Akt比值下降，Akt mRNA表达下调（P〈0.05）；与IGF组比较，P＋IGF组侵袭细胞数减少，细胞PI3K及其mRNA表达下调，p-Akt/Akt比值下降，Akt mRNA表达下调（P〈0.05）；与LY组比较，P＋LY组侵袭细胞数减少，细胞PI3K及其mRNA表达下调，p-Akt/Akt比值下降，Akt mRNA表达下调（P〈0.05）。 结论异丙酚抑制人肝癌HepG2细胞侵袭的机制与抑制PI3K/Akt信号通路的激活有关。

Objective To evaluate the role of phosphatidylinositol 3-kinase （PI3K）/serine-threo- nine kinase （Akt） signaling pathway in propofol-induced invasion of human liver cancer cell line HepG2. Methods HepG2 cells were cultured in RPMI 1640 medium supplemented with 10% fetal bovine serum ina 5% CO2 incubator at 37 ℃. HepG2 cells at the logarithmic growth phase were divided into 6 groups （n= 18 each） using a random number table： control group （group C）, propofol group （group P）, PI3K/Akt signaling pathway agonist IGF-1 group （group IGF）, PI3K/Akt signaling pathway inhibitor LY294002 group （group LY）, IGF-1 plus propofol group （group IGF＋P） and LY29g002 plus propofol group （group LY ＋ P）. Propofol 120 μg/ml was added in group P. IGF-1 10 nmol/L was added in group IGF. LY294002 10 μmol was added in group LY. In group IGF＋P, 10 nmol/L IGF-1 was added, cells were in- cubated for 24 h, and then 120 μg/ml propofol was added. In group LY ＋ P, 10 μmol LY294002 was add- ed, cells were incubated for 24 h, and then 120 μg/ml propofol was added. The invasion of cells was measured by Transwell invasion assay at 24 h of incubation. The expression of PI3K and Akt mRNA in cells was determined by real-time polymerase chain reaction. The expression of Akt, PI3K and phosphorylated Akt （p-Akt） was detected by using Western blot. Results Compared with group C, the invasive cell count was significantly reduced, the expression of PI3K protein and mRNA was down-regulated, p-Akt/Akt ratio was decreased, and the expression of Akt mRNA was down-regulated in P, P＋IGF, LY and P＋EY groups, and the invasive cell count was significantly increased, the expression of PI3K protein and mRNA was up-regulated, p-Akt/Akt ratio was increased, and the expression of Akt mRNA was up-regulated in group IGF （P〈0. 05）. Compared with group P, the invasive cell count was significantly increased, the expression of PI3K protein and mRNA was up-regulated, p-Akt/Akt ratio was increased, and the expres- sion of Akt mRNA was up-regulated in group P＋IGF, and the invasive cell count was significantly reduced, the expression of PI3K protein and mRNA was down-regulated, p-Akt/Akt ratio was decreased, and the ex- pression of Akt mRNA was down-regulated in group P＋LY （P〈0.05）. The invasive cell count was signifi- cantly reduced, the expression of PI3K protein and mRNA was down-regulated, p-Akt/Akt ratio was de- creased, and the expression of Akt mRNA was down-regulated in group P＋IGF as compared with group IGF （P〈0.05） and in group P＋LY as compared with group LY （P〈0. 05）. Conclusion The mechanism by which propofol inhibits invasion of HepG2 cells is related to inhibiting activation of PI3K/Akt signaling path- ways.