目的： 利用RNA干扰技术沉默人肝L-02细胞内可诱导的同型半胱氨酸内质网应激泛素样结构域1（homocysteine-inducible, endoplasmic reticulum stress-inducible, ubiquitin-like domain member 1, HerpUD1 ）基因表达，探讨其对人肝L-02细胞的辐射增敏作用。 方法： 根据 HerpUD1 基因序列设计shRNA片段，构建靶向 HerpUD1 基因的shRNA表达载体，稳定转染L-02细胞，采用real-time PCR和Western blotting检测转染后细胞中 HerpUD1 mRNA及蛋白的表达。以8 Gy X射线照射转染后细胞，Hoechst荧光染色观察细胞的凋亡情况。 结果： 成功构建靶向 HerpUD1 的真核表达干扰载体，并建立稳定转染shHerpUD1的L-02细胞株。在稳定转染的L-02细胞中， shHerpUD1-1420干扰片段干扰效果最好，与pGPU6-NC组（阴性对照组）相比，shHerpUD1-1420组 HerpUD1 mRNA表达显著下降\[（0.15±0.05） vs （1.00±0.08），P<0.05\]，其蛋白水平的表达也显著降低\[（0.19±0.01） vs （1.62±0.08），P<0.01\]。转染后细胞经8 Gy X射线照射后24 h，L-02-shHerpUD1-1420组细胞凋亡率显著高于L-02-NC组（阴性对照组）\[（10.23±3.37）%  vs （6.89±1.49）%，P<0.01\]，而L-02-NC组与L-02-Neo组相比则无显著差异（P>005）。 结论： HerpUD1 基因表达沉默显著增加X射线辐射引起的L-02细胞凋亡，具有辐射增敏作用。

Objective:To study the radiosensitization effect of homocysteine-inducible, endoplasmic reticulum stress-inducible, ubiquitin-like domain member 1 ( HerpUD1 ) on human liver L-02 cells through silencing the expression of HerpUD1 gene by RNA interference technology. Methods: Based on the HerpUD1 gene sequence, shRNA expression vector interfering HerpUD1 gene was constructed and L-02 cells were stably transfected. The expressions of HerpUD1 mRNA and protein in cells after transfection were detected by real-time PCR and Western blotting. The transfected cells were radiated by 8 Gy X ray and the cell apoptosis was analyzed by Hoechst fluorescence assay. Results: Eukaryotic expression interference vector targeting HerpUD1 gene was successfully constructed and L-02 cell line stably transfected with shHerpUD1 was established. The shHerpUD1-1420 interference fragment had the highest interference efficiency in stably transfected L-02 cells. Compared with pGPU6-NC (empty vector) group, the expression level of HerpUD1 mRNA in shHerpUD1-1420 group was significantly decreased (\[0.15±005\] vs \[1.00±0.08\], P<0.05), and the expression level of HerpUD1 protein was also significantly reduced (\[0.19±0.01\] vs \[1.62±0.08\], P<0.01). The apoptotic rate was higher in L-02-shHerpUD1-1420 group than that in L-02-NC group (negative control group) 24 h after 8 Gy X ray irradiation (\[1023±3.37\]% vs \[6.89±1.49\]%, P<0.05), while there was no significant difference between L-02-NC and L-02-Neo group. Conclusion:Silencing the expression of HerpUD1 gene significantly increases the X ray radiation-induced apoptosis of L-02 cells, therefore playing an role in radiosensitization.

国家自然科学基金资助项目（No. 30972460）；上海市自然科学基金资助项目（No. 09ZR1415400）