Green tea component epigallocatechin-3-gallate decreases expression of osteopontin via a decrease in mRNA half-life in cell lines of metastatic hepatocellular carcinoma.
Green tea component epigallocatechin-3-gallate decreases expression of osteopontin via a decrease in mRNA half-life in cell lines of metastatic hepatocellular carcinoma.
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DOI：
10.1016/j.surg.2015.06.011
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发表时间：
2015-10
期刊：
影响因子：
3.8
通讯作者：
Mi, Zhiyong
中科院分区：
文献类型：
--
作者：
Zapf, Matthew A. C.;Kothari, Anal N.;Weber, Cynthia E.;Arffa, Matthew L.;Wai, Phillip Y.;Driver, Joseph;Gupta, Gopal N.;Kuo, Paul C.;Mi, Zhiyong
骨桥蛋白(OPN)介导肝细胞癌的转移和侵袭。绿茶中发现的表没食子儿茶素没食子酸酯(EGCG)在体外可以抑制肝癌肿瘤的生长。我们试图研究EGCG在转移性肝细胞癌细胞系中调节骨桥蛋白的作用。实验肝癌细胞系包括高水平表达OPN的HepG2和MHCC-97H肝癌细胞，以及低水平表达OPN的Hep3B细胞。采用酶联免疫吸附试验和逆转录聚合酶链式反应检测骨桥蛋白的含量。EGCG0.02~20μg/m L处理细胞。划痕试验检测细胞迁移。用染色质免疫印迹法检测EGCG处理后OPN启动子与RNA PolII的结合。用含有人OPN启动子不同缺失片段的荧光素酶报告质粒研究OPN的转录调控。用放线菌素D测定骨桥蛋白的半衰期。2μg/mLEGCG和20μg/mLEGCG作用于人肝癌细胞株97H和HepG2细胞后，骨桥蛋白的分泌水平分别下降约6倍和90倍(P均<0.01)。当EGCG0.2~20μg/ml时，OPN mRNA表达降低(P均<0.01)。3-(4，5-dimethylthiazolyl-2)-2，5-diphenyltetrazolium)比色分析表明，OPN表达的差异不是由于肝癌细胞系的生存能力所致。启动子分析和染色质免疫沉淀分析表明，EGCG对OPN的转录调控没有影响。转录后，EGCG将OPN mRNA的半衰期从16.8小时(95%可信区间9.0-125.1)降低到2.5小时(95%可信区间2.1-3.2)(P<；.001)。EGCG处理后24小时和48小时细胞迁移率分别下降(8.0±2.4%比21.2±10.8%，P<0.01)和(13.2±3.6%比53.5±19.8%，P<；0.01)。我们提供的证据表明，EGCG通过缩短OPN mRNA的半衰期来降低MHCC-97H细胞中OPN mRNA和OPN蛋白的分泌水平。EGCG对肝癌患者的可译性是有希望的，因为EGCG是一种廉价、容易获得的化学物质，具有广泛的安全性历史。
Osteopontin (OPN) mediates metastasis and invasion of hepatocellular carcinoma (HCC). Epigallocatechin-3-gallate (EGCG), found in green tea, suppresses HCC tumor growth in vitro. We sought to investigate the role of EGCG in modulating OPN in cell lines of metastatic HCC. Experimental HCC cell lines included HepG2 and MHCC-97H HCC cells, which express high levels of OPN, and the Hep3B cells, which express lesser levels of OPN. Cells were treated with EGCG (0.02–20 μg/mL) before measurement of OPN with enzyme-linked immunosorbent assay and reverse transcriptase-polymerase chain reaction. Scratch assay measured cell migration. Binding of the OPN promoter to RNA pol II was evaluated by the use of Chromatin-IP assay after EGCG treatment. Transcriptional regulation of OPN was investigated with luciferase reporter plasmids containing various deletion fragments of the human OPN promoter. Measurement of the half-life of OPN mRNA was conducted using actinomycin D. Treatment of MHCC-97H and HepG2 cells with 2 μg/mL and 20 μg/mL EGCG caused a ~6-fold and ~90-fold decrease in secreted protein levels of OPN (All P < .001). OPN mRNA was decreased with EGCG concentrations of 0.2–20 μg/ml (All P < .001). The 3-(4, 5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide (ie, MTT) assay revealed that differences in OPN expression were not due to viability of the HCC cell lines. Promoter assay and chromatin immunoprecipitation analysis revealed no effect of EGCG on the transcriptional regulation of OPN. Posttranscriptionally, EGCG decreased the half-life of OPN mRNA from 16.8 hours (95% confidence interval 9.0–125.1) to 2.5 hours (95% confidence interval 2.1–3.2) (P < .001). Migration was decreased in EGCG treated cells at 24 hours (8.0 ± 2.4% vs 21.2 ± 10.8%, P < .01) and at 48 hours (13.2 ± 3.6% vs 53.5 ± 19.8%, P < .001). We provide evidence that EGCG decreases OPN mRNA and secreted OPN protein levels by decreasing the half-life of OPN mRNA in MHCC-97H cells. The translatability of EGCG for patients with HCC is promising, because EGCG is an inexpensive, easily accessible chemical with an extensive history of safety.
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影响因子：
4.6
作者：
Dube, Admire;Nicolazzo, Joseph A.;Larson, Ian
通讯作者：
Larson, Ian
影响因子：
5.7
作者：
Shirakami, Yohei;Shimizu, Masahito;Moriwaki, Hisataka
通讯作者：
Moriwaki, Hisataka
影响因子：
3.8
作者：
Bhattacharya SD;Garrison J;Guo H;Mi Z;Markovic J;Kim VM;Kuo PC
通讯作者：
Kuo PC
影响因子：
5.6
作者：
Tsang, Wing Pui;Kwok, Tim Tak
通讯作者：
Kwok, Tim Tak
影响因子：
4.8
作者：
Umeda, Daisuke;Yano, Satomi;Tachibana, Hirofumi
通讯作者：
Tachibana, Hirofumi