Midkine protects hepatocellular carcinoma cells against TRAIL-mediated apoptosis through down-regulation of caspase-3 activity
Midkine protects hepatocellular carcinoma cells against TRAIL-mediated apoptosis through down-regulation of caspase-3 activity
复制标题
作者：
Ohuchida, T;Okamoto, K;Itoh, H
背景中期因子（midkine，MIC）是一种肝素结合生长因子，在肿瘤发生中起重要作用。然而，MK在肝细胞癌中的生物学机制至今仍不清楚。本研究的目的是探讨MK在人肝癌细胞系中的抗凋亡作用。采用人肝癌细胞株HepG 2研究MK的抗凋亡作用。采用2-（2-甲氧基-4-硝基苯基）-3-（4-硝基苯基）-5-（2，4-二磺基苯基）-2H-四唑鎓盐（WST-8）测定、半胱天冬酶-3活性测定、半胱天冬酶-8活性测定和流式细胞术分析检测肿瘤坏死因子相关凋亡诱导配体（TRAIL）/放线菌素D（ActD）诱导的细胞凋亡。TRAIL对HepG 2细胞中的细胞死亡具有强有力的、密切依赖的诱导作用，在500 ng/mL ActD的存在下，在100 ng/mL的TRAIL浓度下，HepG 2细胞的活细胞计数降低至对照计数的6.3%。流式细胞术用于证明由TRAIL/ActD诱导的凋亡实际上是细胞死亡的原因。根据WST-8试验，MK预处理导致HepG 2细胞中TRAIL/ActD介导的细胞凋亡受到抑制，尽管单独用MK处理HepG 2细胞时细胞活力并未增加。加入MK后，Caspase-3活性下调，但Caspase-8活性在MK存在和不存在时均升高。目前的研究结果表明，MK作为一种抗凋亡因子在HepG 2细胞通过下调caspase-3的活性。(C)2004年美国癌症协会。
BACKGROUND. It is believed that midkine (MIC), a heparin-binding growth factor, plays an important role in carcinogenesis. However, the biologic mechanism of MK in hepatocellular carcinoma has not been clarified to date. The objective of the current study was to investigate the antiapoptotic role of MK in a human hepatoma cell line.METHODS. The human hepatoma cell line HepG2 was used to study the antiapoptotic effect of MK. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)/actinomycin D (ActD)-induced apoptosis was detected using a 2-(2-methoxy-4-nitrophenyl) -3-(4-nitrophenyl)-5-(2,4-disulphophenyl)-2H-tetrazolium monosodium salt (WST-8) assay, a caspase-3 activity assay, a caspase-8 activity assay, and flow cytometric analysis.RESULTS. TRAIL had a potent, close-dependent inductive effect on cell death in HepG2 cells, for which viable cell counts decreased to 6.3% of the control count at a TRAIL concentration of 100 ng/mL in the presence of 500 ng/mL ActD. Flow cytometry was used to demonstrate that apoptosis induced by TRAIL/ActD was in fact the cause of cell death. According to the WST-8 assay, MK pretreatment resulted in the suppression of TRAIL/ActD-mediated apoptosis in HepG2 cells, although cell viability did not increase when HepG2 cells were treated with MK alone. Caspase-3 activity was down-regulated when MK was added, but caspase-8 activity was high in both the absence and presence of MK.CONCLUSIONS. The results of the current study indicate that MK acts as an antiapoptotic factor in HepG2 cells through the down-regulation of caspase-3 activity. (C) 2004 American Cancer Society.