Effect of UVC radiation on mouse fibroblasts deficient for FAS-associated protein with death domain
Effect of UVC radiation on mouse fibroblasts deficient for FAS-associated protein with death domain
复制标题
DOI：
10.1080/09553002.2016.1186298
复制
发表时间：
2016-08-01
影响因子：
2.6
通讯作者：
Marijanovic, Inga
中科院分区：
文献类型：
--
作者：
Begovic, Lidija;Antunovic, Maja;Marijanovic, Inga
目的：紫外线（UV）辐射诱导的细胞凋亡使我们能够研究DNA损伤的机制，并研究细胞如何避免DNA损伤的后果。具有广泛DNA损伤的细胞激活细胞凋亡的外在和内在途径。外源性途径与FAS相关蛋白死亡结构域（FADD），一个衔接蛋白分子介导细胞凋亡信号所必需的通过cell.Materials和方法：野生型和FADD缺陷小鼠胚胎成纤维细胞的活力和凋亡进行了研究暴露于三种剂量（50，75和300 J/m2）的UVC辐射后1，3，24和48小时。形态学变化进行了观察，使用DNA结合染料（Hoechst和碘化丙啶），而生化变化进行了监测，使用免疫检测的聚（ADP-核糖）聚合酶（PARP）蛋白裂解和caspase-3活性assay.Results：结果表明，野生型和FADD缺陷细胞之间的细胞死亡反应的差异依赖于剂量和孵育时间暴露于UVC辐射后。FADD缺陷细胞对UVC辐射更敏感。尽管FADD缺陷细胞缺乏凋亡外源性途径的衔接蛋白，但较高剂量的UVC触发了它们的凋亡反应，而野生型细胞主要由于坏死而死亡。辐射后24 h，两种细胞系之间的caspase 3活性和PARP裂解的不同模式被观察到，证实FADD缺陷cells.Conclusions：野生型细胞可以通过线粒体和受体介导的途径来执行凋亡，而FADD缺陷细胞只能激活内源性途径。两种细胞系之间的UVC辐射反应存在差异，表明FADD在细胞死亡方式选择中的作用。
Purpose: Ultraviolet (UV) radiation-induced apoptosis enabled us to study the mechanism of DNA damage and to investigate how cells avoid consequences of damaged DNA. Cells with extensive DNA damage activate extrinsic and intrinsic pathways of apoptosis. The extrinsic pathway is coupled to a FAS-associated protein with death domain (FADD), an adaptor protein molecule necessary for mediating apoptotic signals through the cell.Materials and methods: Viability and apoptosis of wild-type and FADD-deficient mouse embryonic fibroblasts were investigated 1, 3, 24 and 48h after exposure to three doses (50, 75 and 300 J/m(2)) of UVC radiation. Morphological changes were observed using DNA binding dyes (Hoechst and propidium iodide) while biochemical changes were monitored using immunodetection of the poly (ADP-ribose) polymerase (PARP) protein cleavage and caspase-3 activity assay.Results: Results showed that the difference in cell death response between wild-type and FADD-deficient cells depended on dose and incubation time after exposure to UVC radiation. FADD-deficient cells are more sensitive to UVC radiation. Even though FADD-deficient cells lack an adapter protein of apoptotic extrinsic pathway, higher doses of UVC triggered their apoptotic response, while wild-type cells die mainly due to necrosis. A different pattern of caspase 3 activity and PARP cleavage was observed 24h after radiation between two cell lines confirming higher apoptotic response in FADD-deficient cells.Conclusions: Wild-type cells can execute apoptosis via both, the mitochondrial and the receptor-mediated pathway whereas FADD-deficient cells can only activate the intrinsic pathway. There is a difference in UVC radiation response between two cell lines indicating the role of FADD in the selection of cell death modality.