沉默GRAMD1A及抑制STAT5信号通路对肝癌细胞Huh7放射敏感性的影响

目的 探讨沉默GRAMD1A及抑制STAT5信号对肝癌细胞Huh7放射敏感性的影响，旨在为肝癌临床联合治疗提供新思路。方法 慢病素感染构建沉默GRAMD1A的Huh7细胞株，采用qPCR和Western blot进行验证，qPCR和荧光素酶报告实验检测沉默GRAMD1A后对Huh7细胞中STAT5及其下游基因表达的影响；以克隆形成率和细胞凋亡为指标检测沉默GRA株。结果 构建后的Huh7细胞经2 Gy照射后，沉默GRAMD1A联合照射组细胞克隆形成能力较阴性对照联合照射组显著降低，差异有统计学意义（t=8.494，P<0.05）；沉默GRAMD1A联合照射组细胞凋亡较阴性对照联合照射组显著增加，差异有统计学意义（t=3.560，P<0.05）。沉默GRAMD1A后Huh7细胞放射敏感性明显增加，且细胞中STAT5及其下游基因表达显著降低。SH-4-54抑制剂联合照射组较二甲基亚砜联合照射组细胞克隆存活能力显著降低，差异有统计学意义（t=8.660，P<0.05），SH-4-54抑制STAT5通路后，Huh7细胞放射敏感性显著增加。结论 沉默GRAMD1A可能通过STAT5信号通路增强肝癌细胞Huh7的放射敏感性，表明GRAMD1A在肝癌发生发展中起重要作用，将可能为肝癌靶向治疗及联合治疗提供新靶点。

Effect of silencing GRAMD1A and inhibiting STAT5 signaling pathway on radiosensitivity of hepatocellular carcinoma cell line Huh7

Objective To investigate the effect of silencing GRAMD1A and inhibiting STAT5 signaling pathway on the radiosensitivity of Huh7 cells in order to provide new ideas for the clinical combined therapy of hepatocellular carcinoma.Methods The Huh7 cells silencing GRAMD1A was constructed by infecting lentivirus and verified by qPCR and Western blot. QPCR and luciferase reporter assays were used to detect the effect of silencing GRAMD1A on the expression of STAT5 and its downstream genes. Colony formation and apoptosis were detected to evaluate the effects of silencing GRAMD1A and STAT5 inhibitor SH-4-54 on cell radiosensitivity.Results After 2 Gy exposure of the constructed Huh7 cells, the colony formation ability of the silencing GRAMD1A combined irradiation group was significantly lower than that of the negative control combined irradiation group, and the difference was statistically significant (t=8.494, P<0.05).Silence apoptosis in the GRAMD1A combined irradiation group was significantly increased compared with the negative control combined irradiation group (t=3.560, P<0.05).After silencing GRAMD1A, the radiosensitivity of Huh7 cells was significantly increased, and the expression of STAT5 and its downstream genes was significantly reduced in cells.The survival rate of the SH-4-54 inhibitor combined irradiation group was significantly lower than that of the dimethyl sulfoxide combined irradiation group, and the difference was statistically significant (t=8.660, P<0.05). SH-4-54 inhibited STAT5 after passage, the radiosensitivity of Huh7 cells was significantly increased.Conclusions Silencing GRAMD1A could significantly enhance the radiosensitivity of Huh7 cells via STAT5 signaling pathway, indicating that GRAMD1A plays an important role in the development and progression of HCC. This finding may provide a new target for HCC therapy.