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目的 :探讨Janus激酶(Janus kinase,JAK)-信号转导及转录激活因子(signal transducer and activator of transcription,STAT)信号转导通路阻断剂AG490对肝癌SMMC-7721细胞及移植瘤生长和凋亡的影响,及可能的作用机制。方法 :用人肝癌SMMC-7721细胞株建立肝细胞癌裸鼠移植瘤模型,待肿瘤生长至约150 mm3时随机分为对照组、AG490单药组、顺铂(cisplatin,DDP)单药组和AG490联合DDP组,用AG490单药或联合DDP处理各组小鼠,绘制各组小鼠的肿瘤生长曲线并计算抑瘤率,FCM法检测细胞凋亡率,免疫组织化学法和蛋白质印迹法检测移植瘤中磷酸化STAT3(phospho-STAT3,p-STAT3)及caspase-3蛋白表达的情况。结果 :与对照组相比,DDP组、AG490组和AG490联合DDP组在腹腔注射药物第10和13天时,移植瘤的体积均明显较小(P值均<0.05);AG490联合DDP组的抑瘤率为60.24%,明显高于DDP组的51.73%和AG490组的34.58%(P值均<0.05)。FCM检测结果显示,AG490组、DDP组和AG490联合DDP组移植瘤组织中肿瘤细胞的凋亡率分别为(22.4±0.8)%、(40.1±1.2)%和(43.2±1.5)%,与对照组的(11.5±0.4)%相比,差异均具有统计学意义(P值均<0.05)。免疫组织化学法检测结果显示,对照组、AG490组、DDP组和AG490联合DDP组移植瘤组织中p-STAT3蛋白的阳性表达率依次为(95.4±1.8)%、(65.8±2.4)%、(37.4±2.1)%和(20.3±1.2)%,caspase-3蛋白的阳性表达率依次为(10.7±1.1)%、(26.4±1.6)%(、53.9±3.2)%和(82.5±2.8)%。蛋白质印迹法检测结果显示,与对照组相比,AG490组、DDP组和AG490联合DDP组中p-STAT3蛋白的表达水平均明显降低(P值均<0.05),而caspase-3蛋白的表达水平均明显升高(P值均<0.05)。结论 :AG490P联合DDP可进一步提高DDP对肿瘤生长的抑制作用,阻断JAK-STAT通路中p-STAT3蛋白的表达水平,而上调凋亡相关蛋白caspase-3的表达可能是其作用机制之一。
AIM: To investigate the effect of Janus kinase (JAK) - signal transducer and activator of transcription (STAT) signal transduction pathway inhibitor AG490 on the growth and apoptosis of hepatoma SMMC-7721 cells and xenografts The impact, and possible mechanism of action. Methods: Hepatocellular carcinoma xenograft model was established with human hepatocellular carcinoma SMMC-7721 cell line. When the tumor grew to about 150 mm3, it was randomly divided into control group, AG490 single group, cisplatin (DDP) monotherapy group and AG490 In combination with DDP group, mice in each group were treated with AG490 alone or in combination with DDP, the tumor growth curves of the mice in each group were plotted and the tumor inhibition rate was calculated. FCM was used to detect the apoptosis rate, immunohistochemistry and Western blotting The expression of phospho-STAT3 (p-STAT3) and caspase-3 protein in tumor. Results: Compared with the control group, the volume of tumor xenografts in DDP group, AG490 group and AG490 combined with DDP group were significantly smaller (P <0.05) after intraperitoneal injection of drugs on the 10th and 13th days. Compared with the DDP group, The tumor incidence was 60.24%, significantly higher than 51.73% in DDP group and 34.58% in AG490 group (all P <0.05). The results of FCM showed that the apoptotic rate of tumor cells in AG490 group, DDP group and AG490 combined with DDP group was (22.4 ± 0.8)%, (40.1 ± 1.2)% and (43.2 ± 1.5)%, respectively, Group (11.5 ± 0.4)%, the difference was statistically significant (P <0.05). The results of immunohistochemistry showed that the positive rates of p-STAT3 protein in control group, AG490 group, DDP group and AG490 combined with DDP group were (95.4 ± 1.8)%, (65.8 ± 2.4)% and The positive rates of caspase-3 protein were (10.7 ± 1.1)%, (26.4 ± 1.6)% (53.9 ± 3.2)% and (82.5 ± 2.8)%, respectively . The results of Western blotting showed that compared with the control group, the expression of p-STAT3 protein in AG490 group, DDP group and AG490 combined with DDP group were significantly decreased (P <0.05), while the expression of caspase-3 protein Were significantly higher (P <0.05). Conclusion: AG490P combined with DDP can further increase the inhibitory effect of DDP on tumor growth and block the expression of p-STAT3 protein in JAK-STAT pathway. Upregulation of the expression of caspase-3 may be one of its mechanisms.