神经酰胺在辐射诱导的秀丽隐杆线虫生殖细胞旁效应中的机制研究

目的 利用秀丽隐杆线虫为研究对象，以单粒子束为辐射源，从活体水平探究神经酰胺在辐射旁效应中的作用。方法 以野生型N2和突变体L4期线虫的后食道球为辐射部位，分别给予2 000个质子的辐照剂量，检测和分析成虫的生殖腺凋亡细胞以及基因表达水平。结果 定点辐射N2的后食道球可显著诱导线虫旁区生殖腺细胞凋亡的增加（t=9.007，P<0.05），但在神经酰胺合酶基因突变体中未表现出这一现象（P>0.05）。实时定量PCR结果显示，辐射诱导N2和神经酰胺合酶基因突变体lagr-1（gk327）；hyl-1（ok976）线虫中凋亡核心通路基因egl-1和ced-13表达量的上升，但两品系线虫间的差异无统计学意义（P>0.05）；辐射可诱导N2和DNA损伤检验点突变体hus-1（op241）中hyl-1和lagr-1表达量的上升，且两品系间的差异无统计学意义（P>0.05）。非受体络氨酸激酶abl-1突变可显著增加辐射诱导的旁区细胞凋亡（t=13.241，P<0.05），而当神经酰胺合酶基因与abl-1同时突变时，凋亡增加的现象被抑制（t=13.462，P<0.05）。结论 神经酰胺参与线虫体内辐射引起的旁区凋亡过程，且可能与凋亡核心通路中的促凋亡蛋白egl-1和ced-13以及DNA损伤响应通路中的HUS-1协同发挥作用。抑凋亡基因abl-1与神经酰胺合酶在凋亡过程中表现出拮抗关系。

The role of ceramide in radiation-induced bystander effect on germ cells in Caenorhabditis elegans

Objective To investigate the role of ceramide in radiation induced bystander effect (RIBE) in vivo by irradiating Caenorhabditis elegans with proton microbeam. Methods The posterior pharynx of wide-type N2 and genetically mutated L4-staged worms was irradiated with 2 000 particles, then germ cell apoptosis and gene expression were analyzed. Results Point-fixed radiation to posterior pharynx of N2 worms significantly increased the number of apoptotic germ cells (t=9.007, P<0.05), but not in the worms with deletion mutants of ceramide synthase (CerS) genes (hyl-1 and lagr-1) (P>0.05). Real-time quantitative PCR assay indicated that in both N2 and lagr-1(gk327);hyl-1(ok976) worms, the genes of egl-1 and ced-13 in core apoptosis pathway were up-regulated after radiation to posterior pharynx, without statistical significance between these two strains (P>0.05). The gene expression levels of hyl-1 and lagr-1 were increased in both N2 and hus-1(op241) worms after radiation with no statistical significant difference between two strains (P>0.05). Furthermore, radiation-induced germ cell apoptosis was increased in abl-1(ok171) worms (t=13.241, P<0.05), rather than in lagr-1(gk327);hyl-1(ok976);abl-1(ok171) worms (t=13.462, P<0.05). Conclusions Ceramide is required for RIBE on germ cell apoptosis in C. elegans, and it might play function together with egl-1 and ced-13 in core apoptosis pathway and HUS-1 in DNA damage response pathway. But ceramide had antagonistic relationship with anti-apoptotic protein abl-1 in germ cell apoptosis.