Li Qiaozhuan, Li Ming, Wang Ya-Lan, Fauzee Nilufer Jasmine Selimah, Yang Yi, Pan Juan, Yang Lian, Lazar Alexander
Department of Pathology, Molecular Medicine and Cancer Research Center, Chongqing Medical University, Chongqing, China.
Cell Physiol Biochem. 2012;29(3-4):361-72. doi: 10.1159/000338491. Epub 2012 Apr 3.
To investigate the role and mechanism of PARG inhibition of metastatic behavior in colonic carcinoma cells.
We examined the effects of PARG protein knockdown by RNA interference on invasion, migration and matrix adhesion of colon carcinoma cell lines in vitro and using a murine in vivo model of liver metastasis. Metastasis related genes were detected using mRNA and protein levels. Moreover, LY294002, an Akt phosphorylation inhibitor, was used to determine whether the suppression of metastatic behavior of colon carcinoma cells was mediated by Akt phosphorylation that was confirmed by EMSA. Pyrrolidine dithiocarbamate (PDTC) was used as a selective NFκ-B inhibitor to clarify the relationship between PARG, PARP and NF-κB.
PARG protein was undetectable following specific shRNA transfection; mRNA and protein levels of PARP were significantly decreased. PARG-shRNA cells showed high levels of phosphorylated Akt with decreased expression of NF-κB (both total & nuclear), MMP2 and MMP9. However, no additional changes were noted following inhibition of PI3K/Akt pathway by LY294002 in the PARG-shRNA cells; these cells displayed reduced number of liver metastases when characterized in the murine in vivo model.
PARG knockdown, concomitant with inhibition of PARP, suppressed the metastatic potency of colon carcinoma cells by activation of PI3K/Akt signaling pathway.
研究聚（ADP-核糖）糖苷酶（PARG）抑制结肠癌细胞转移行为的作用及机制。
我们通过RNA干扰检测PARG蛋白敲低对结肠癌细胞系体外侵袭、迁移和基质黏附的影响，并使用小鼠肝转移体内模型进行研究。采用mRNA和蛋白水平检测转移相关基因。此外，使用Akt磷酸化抑制剂LY294002来确定结肠癌细胞转移行为的抑制是否由Akt磷酸化介导，这一点通过电泳迁移率变动分析（EMSA）得以证实。使用吡咯烷二硫代氨基甲酸盐（PDTC）作为选择性核因子κB（NFκ-B）抑制剂来阐明PARG、聚（ADP-核糖）聚合酶（PARP）与NF-κB之间的关系。
特异性短发夹RNA（shRNA）转染后未检测到PARG蛋白；PARP的mRNA和蛋白水平显著降低。PARG-shRNA细胞显示出高水平的磷酸化Akt，同时NF-κB（总蛋白和核蛋白）、基质金属蛋白酶2（MMP2）和基质金属蛋白酶9（MMP9）的表达降低。然而，在PARG-shRNA细胞中用LY294002抑制PI3K/Akt途径后未观察到其他变化；在小鼠体内模型中，这些细胞的肝转移数量减少。
PARG敲低与PARP抑制同时发生，通过激活PI3K/Akt信号通路抑制结肠癌细胞的转移潜能。
