Investigation of radiosensitizers with radiation-induced apoptosis

电离辐射激活半胱氨酸天冬氨酸酶，诱导不同类型细胞的凋亡。然而，电离辐射如何激活半胱氨酸天冬氨酸酶并随后诱导细胞死亡仍有待明确阐明。在这里，我们提供了电离辐射激活caspase-8和Bid并通过两条不同的途径破坏淋巴细胞白血病细胞线粒体膜电位(ΔΨm)的证据。辐射诱导的半胱氨酸天冬氨酸氨基转移酶-8、-3和BID的裂解以及随后的DNA断裂被四肽z-IETD-FMK显著阻断，但不能抑制ΔΨm的断裂，这表明ΔΨm的丢失不依赖于半胱氨酸酶的激活。此外，单独抑制caspase的激活不足以防止辐射诱导的细胞死亡。相反，12-O-十四酰佛波醇-3-乙酸酯通过阻断Caspase-8/Bid的激活和ΔΨm的丢失而显著抑制辐射诱导的细胞死亡。TPA介导的抗凋亡活性可被MEK抑制剂PD98059取消，但不能被PKC抑制剂19-27假底物所消除，后者反而增强了抗凋亡活性。这些数据有力地表明，至少有两条不同的途径促进辐射诱导的细胞凋亡，即Caspase激活和Caspase非依赖性ΔΨm反应，后者是辐射诱导的细胞凋亡中的一个关键过程。这两条通路都受到MEK/ERK信号的负调控，表明其激活可能是辐射抵抗的一种机制。这些研究表明，利用辐射诱导的细胞凋亡的新途径作为新的放射增敏剂来调节辐射敏感性。

Ionizing radiation activates caspases and induces apoptosis in different cell types. However, how ionizing radiation activates caspases and subsequently induces cell death remains to be definitively elucidated. Here we provide evidence that ionizing radiation activates caspase-8 and Bid and disrupts mitochondrial membrane potential (ΔΨm) by two distinct pathways in lymphocytic leukemia cells. Radiation-induced cleavage of caspase-8, -3 and Bid, and subsequent DNA fragmentation were substantially blocked by pre-treatment with the tetrapeptide z-IETD-FMK, however it could not suppress the breakdown of ΔΨm, indicating that the ΔΨm loss occurred independently of the caspase activation. Moreover, suppression of caspase activation alone was insufficient to prevent the radiation-induced cell death. In contrast, 12-O-tetradecanoylphorbol-3-acetate (TPA) substantially inhibited the radiation-induced cell death through blockage of both caspase-8/Bid activation and the ΔΨm loss. TPA-mediated anti-apoptotic activity was abrogated by a MEK inhibitor PD98059, but not by a PKC inhibitor 19-27 pseudosubstrate, which rather augmented the anti-apoptotic activity. These data strongly suggest that there are at least two distinct pathways to facilitate radiation-induced apoptosis, i.e., caspase activation and caspase-independent ΔΨm reakdown, the latter being a crucial process in the apoptosis. These two pathways are negatively regulated by the MEK/ERK-mediated signal, signifying that its activation is a possible mechanism for radioresistance.These studies demonstrate that the new approach using radiation-induced apoptosis modulates the radiosensitivity as new radiosensitizers.