The mechanism of apoptosis induction by TGF-β stimuli and its clinical application

我们研究转化生长因子-β刺激诱导细胞凋亡的机制，为其在肿瘤治疗中的应用奠定基础。我们从单个肝癌细胞系中建立了几个对转化生长因子-β刺激部分敏感的可诱导和不可诱导的亚克隆。我们的初步研究表明，在可诱导凋亡的亚克隆中，肿瘤坏死因子家族成员的表达受到转化生长因子-β刺激的诱导。我们进一步研究了从转化生长因子-β刺激到细胞凋亡的信号通路。经转化生长因子-β刺激后，观察细胞DNA含量、肿瘤坏死因子家族的表达及半胱氨酸天冬氨酸氨基转移酶家族的激活情况。首先观察到一过性的G1/G2期停滞，然后依次发生了诱导肿瘤坏死因子家族表达、激活caspase-8和诱导细胞凋亡。我们研究了抗肿瘤坏死因子家族成员的中和抗体和caspase-8抑制剂Z-IETD-FMK对细胞凋亡的诱导作用，获得了约50%的细胞凋亡抑制率。NF-κB报告分析还表明，转化生长因子-κ刺激可激活NF-βB通路。我们的研究证实了从转化生长因子-β到细胞凋亡的信号通路是通过肿瘤坏死因子家族的诱导和caspase-8的激活来实现的。进一步的研究可能需要利用高密度的基因芯片来揭示诱导细胞凋亡的全部途径。NF-κB的激活可能与转化生长因子-β耐药有关，但其作用尚不明确。

We studied the mechanism of apoptosis induction by TGF-β stimuli for the purpose of applying in the cancer therapy. We established several apoptosis-inducible and -uninducible subclones derived from the single hepatoma cell line, which were partially sensitive to TGF-β stimuli. Our preliminary study using apoptosis-specific cDNA arrays showed that the expression of TNF family members were induced by TGF-β stimuli in the apoptosis-inducible subclones. We studied further the signaling pathways from TGF-β stimuli to apoptosis. After TGF-β stimuli, DNA contents, expression of TNF family, and activation of caspase family were sequentially studied. At first, the G1/G2 arrest was transiently observed, and then the induction of TNF family expression, caspase-8 activation, and apoptosis induction occurred sequentially. We studied the effects of neutralizing antibodies against TNF family members and caspase-8 inhibitor, Z-IETD-FMK against the apoptosis induction, and obtained about 50% inhibition of apoptosis. The NF- κB reporter analysis also showed the activation of NF-κB pathway by TGF-β stimuli. Our studies demonstrated the signaling pathway from TGF-β to apoptosis through TNF family induction and caspase-8 activation. Furhter studies using high-density cDNA array may be required to reveal the whole pathways of apoptosis induction. The activation of NF-κB might be associated with the TGF-β -resistance, but it remains to be determined.