α-fetoprotein causes apoptosis in tumor cells via a pathway independent of CD95, TNFR1 and TNFR2 through activation of caspase-3-like proteases
α-fetoprotein causes apoptosis in tumor cells via a pathway independent of CD95, TNFR1 and TNFR2 through activation of caspase-3-like proteases
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DOI：
10.1046/j.1432-1327.1999.00868.x
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发表时间：
1999-12-01
期刊：
影响因子：
--
通讯作者：
Sukhikh, G
中科院分区：
文献类型：
--
作者：
Dudich, E;Semenkova, L;Sukhikh, G
甲胎蛋白（AFP）是一种具有多种细胞生长调节、分化和免疫抑制活性的癌胚蛋白。以前的研究表明，在体外用1-10 μ M AFP处理肿瘤细胞，通过诱导剂量依赖性细胞毒性产生显著的肿瘤细胞生长抑制，但这些AFP功能的分子机制尚不清楚。在这里，我们表明，AEP的细胞毒性是密切相关的细胞凋亡，所示的细胞形态，核DNA片段化和caspase-3样活性，导致裂解的聚（ADP-核糖）聚合酶。CPP 32家族蛋白酶抑制剂显著抑制细胞凋亡，而一般的半胱天冬酶抑制剂没有抑制作用，显示出AFP介导的细胞死亡的一些增强。使用荧光caspase底物，我们发现caspase-3样蛋白酶在用15 μ M AFP处理Raji细胞后4小时就被激活，而caspase-1，caspase-8和caspase-9样活性在0.5-17小时的时间间隔内没有检测到。AFP处理的Raji细胞增加Bcl-2蛋白，表明AFP诱导的细胞凋亡不能用Bcl-2基因的下调来解释。这也表明AFP在Bcl-2敏感步骤的下游起作用。AFP显著降低可溶性和膜结合Fas配体的基础水平。AFP敏感的turner细胞（HepG 2，Raji）与中和性抗Fas，抗肿瘤坏死因子受体（TNFR）1或抗TNFR 2 mAb孵育不能阻止AFP诱导的凋亡，表明其不依赖于Fas依赖性和TNFR依赖性信号传导。此外，发现对TNF诱导的细胞凋亡（Raji）或Fas诱导的细胞凋亡（MCF-7）有抗性的细胞对AFP介导的细胞死亡敏感，相反，对Fas介导的细胞死亡敏感的细胞（Jurkat）对AFP完全有抗性。总的来说，我们的数据表明：（a）AFP诱导肿瘤细胞凋亡独立于Fas/Fas配体或TNFR/TNF信号通路，和（B）AFP介导的细胞死亡涉及激活效应caspase-3样蛋白酶，但独立于上游激活的起始caspase-1，caspase-8。和半胱天冬酶-9样蛋白酶。
alpha-Fetoprotein (AFP) is an oncoembryonal protein with multiple cell growth regulating, differentiating and immunosuppressive activities. Previous studies have shown that treatment of tumor cells in vitro with 1-10 mu M AFP produces significant suppression of tumor cell growth by inducing dose-dependent cytotoxicity, but the molecular mechanisms underlying these AFP functions are obscure. Here, we show that AEP cytotoxicity is closely related to apoptosis, as shown by cell morphology, nuclear DNA fragmentation and caspase-3-like activity resulting in cleavage of poly(ADP-ribose) polymerase. Apoptosis was significantly inhibited by a CPP32 family protease inhibitor whereas a general caspase inhibitor had no inhibitory effect, showing some enhancement of AFP-mediated cell death. Using fluorogenic caspase substrates, we found that caspase-3-like proteases were activated as early as 4 h after treatment of Raji cells with 15 mu M AFP,whereas caspase-1, caspase-8, and caspase-9-like activity was not detected during the time interval 0.5-17 h. AFP treatment of Raji cells increased Bcl-2 protein, showing that AFP-induced apoptosis is not explained by downregulation of the Bcl-2 gene. This also suggests that AFP operates downstream of the Bcl-2-sensitive step. AFP notably decreased basal levels of soluble and membrane-bound Fas ligand. Incubation of AFP-sensitive turner cells (HepG2, Raji) with neutralizing anti-Fas, anti-rumor necrosis factor receptor (TNFR)1 or anti-TNFR2 mAb did not prevent AFP-induced apoptosis, demonstrating its independence of Fas-dependent and TNFR-dependent signaling. In addition, it was found that cells resistant to TNF-induced (Raji) or Fas-induced (MCF-7) apoptosis are, nevertheless, sensitive to AFP-mediated cell death, in contrast, cells sensitive to Fas-mediated cell death (Jurkat) an completely resistant to AFP. Taken as a whole, our data demonstrate that: (a) AFP induces apoptosis in tumor cells independently of Fas/Fas ligand or TNFR/TNF signaling pathways, and (b) AFP-mediated cell death involves activation of the effector caspase-3-like proteases, but is independent of upstream activation of the initiator caspase-1, caspase-8. and caspase-9-like proteases.