MST4表达对MHCC97H肝癌细胞细胞因子、ERK蛋白、p-ERK蛋白表达的影响及其意义
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摘要
目的:探讨MHCC97H肝癌细胞中丝氨酸/苏氨酸蛋白激酶4/(MST4)的表达与细胞因子、ERK蛋白、p-ERK蛋白表达的关系及其意义。方法:将MHCC97H肝癌细胞培养至对数生长期后,接种于96孔板上培养,分为空白组、高表达组(MST4转染高表达)、低表达组(MST4转染siRNA),采用Western-blot法检测各组共培养24 h后的MST4蛋白表达水平,采用ELISA法检测各组培养24 h后上清液中白细胞介素-2(IL-6)、白细胞介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)、趋化因子-2(CCL2),采用Western-blot法检测各组的细胞外信号调节激酶(ERK)、磷酸化细胞外信号调节激酶(p-ERK)蛋白的表达,采用MTT实验检测3组肿瘤细胞的增殖能力,采用Transwell实验检测各组细胞的侵袭迁移能力。结果:高表达组的MST4蛋白表达显著高于空白组和低表达组(P<0.05);空白组和低表达组的MST4蛋白表达水平差异无统计学意义(P>0.05);高表达组的上清液中IL-6、IL-1β、TNF-α、CCL2水平均显著高于空白组和低表达组(P<0.05);空白组和低表达组的上清液中IL-6、IL-1β、TNF-α、CCL2水平差异无统计学意义(P>0.05);空白组、高表达组和低表达组的ERK蛋白表达差异无统计学意义(P>0.05);高表达组的pERK蛋白显著高于空白组和低表达组(P<0.05);空白组和低表达组的p-ERK蛋白差异无统计学意义(P>0.05);高表达组的MTT实验吸光度值、Transwell实验MHCC97H肝癌细胞迁移数目高于空白组和低表达组(P<0.05);空白组和低表达组的MTT实验吸光度值、Transwell实验MHCC97H肝癌细胞迁移数目差异无统计学意义(P>0.05)。结论:MHCC97H肝癌细胞中MST4高表达,将会激活p-ERK蛋白表达,从而提高细胞因子的表达,提高MHCC97H肝癌细胞的增殖、侵袭能力。
Objective: To investigate the relationship between MST4 expression and cytokines, ERK protein and p-ERK protein expression in MHCC97 H hepatoma cells and its significance. Methods:After the MHCC97 H hepatoma cells were cultured at the logarithmic growth stage,they were inoculated on 96 orifice plates and divided into blank group-high expression group(MST4 transfection high expression) and low expression group(MST4 transfection siRNA). The ELISA method was used to detect interleukin-2(IL-6),interleukin-1 beta(IL-1 beta) and tumor bad in the supernatant of each group after 24 h Death factor-alpha(TNF-alpha) and chemokine-2(CCL2) were used to detect the expression of extracellular signal regulated kinase(ERK) and phosphorylated extracellular signal regulated kinase(pERK) protein in each group by Western-blot method. The increment ability of tumor cellsof three groups was detected by MTT test and the invasion of the cells was detected by Transwell test. Migration ability. Results:The expression of MST4 protein in the high expression group was significantly higher than that in the blank group and the low expression group(P<0.05). There was no significant difference in the expression of MST4 protein between the blank group and the low expression group(P>0.05).The levels of IL-6,IL-1 beta,TNF-α,and CCL2 in the supernatant of high expression group were significantly higher than those in the blank group and low expression group(P<0.05),and the differences in IL-6,IL-1 beta,TNF-α and CCL2 levels of the supernatant of the blank group and low expression group were not significant(P>0.05),but the negative expressions of the ERK protein in the blank group,the high expression group and the low expression group were poor. The p-ERK protein in the high expression group was significantly higher than that in the blank group and the low expression group(P<0.05),and the difference in the p-ERK protein of the blank group and the low expression group was not statistically signifi cant(P>0.05); the MTT experimental absorbance value of the high expression group and the number of migration of the liver cancer cells in the Transwell experiment MHCC97 H were higher than those of the empty group(P>0.05). In the white group and the low expression group(P<0.05),there was no significant difference between the MTT experimental absorbance value of the blank group and the low expression group and the number of MHCC97 H hepatoma cell migration in the Transwell experiment(P>0.05). Conclusion:The high expression of MST4 in MHCC97 H hepatoma cells mayactivate the expression of p-ERK protein,enhance the expression of cytokines,and in crease the value added and invasion of MHCC97 H hepatoma cells.
Objective: To investigate the relationship between MST4 expression and cytokines, ERK protein and p-ERK protein expression in MHCC97 H hepatoma cells and its significance. Methods:After the MHCC97 H hepatoma cells were cultured at the logarithmic growth stage,they were inoculated on 96 orifice plates and divided into blank group-high expression group(MST4 transfection high expression) and low expression group(MST4 transfection siRNA). The ELISA method was used to detect interleukin-2(IL-6),interleukin-1 beta(IL-1 beta) and tumor bad in the supernatant of each group after 24 h Death factor-alpha(TNF-alpha) and chemokine-2(CCL2) were used to detect the expression of extracellular signal regulated kinase(ERK) and phosphorylated extracellular signal regulated kinase(pERK) protein in each group by Western-blot method. The increment ability of tumor cellsof three groups was detected by MTT test and the invasion of the cells was detected by Transwell test. Migration ability. Results:The expression of MST4 protein in the high expression group was significantly higher than that in the blank group and the low expression group(P<0.05). There was no significant difference in the expression of MST4 protein between the blank group and the low expression group(P>0.05).The levels of IL-6,IL-1 beta,TNF-α,and CCL2 in the supernatant of high expression group were significantly higher than those in the blank group and low expression group(P<0.05),and the differences in IL-6,IL-1 beta,TNF-α and CCL2 levels of the supernatant of the blank group and low expression group were not significant(P>0.05),but the negative expressions of the ERK protein in the blank group,the high expression group and the low expression group were poor. The p-ERK protein in the high expression group was significantly higher than that in the blank group and the low expression group(P<0.05),and the difference in the p-ERK protein of the blank group and the low expression group was not statistically signifi cant(P>0.05); the MTT experimental absorbance value of the high expression group and the number of migration of the liver cancer cells in the Transwell experiment MHCC97 H were higher than those of the empty group(P>0.05). In the white group and the low expression group(P<0.05),there was no significant difference between the MTT experimental absorbance value of the blank group and the low expression group and the number of MHCC97 H hepatoma cell migration in the Transwell experiment(P>0.05). Conclusion:The high expression of MST4 in MHCC97 H hepatoma cells mayactivate the expression of p-ERK protein,enhance the expression of cytokines,and in crease the value added and invasion of MHCC97 H hepatoma cells.
引文
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[12]陈翔,陆宝华,袁杰,等.老年结直肠癌血清可溶性细胞间黏附分子-1及炎性因子表达的临床意义[J].重庆医学,2016,45(16):2202
[13]Chen X,Xu H,Wu N,et al.Interaction between granulin A and enolase 1 attenuates the migration and invasion of human hepatoma cells:[J].Oncotarget,2017,8(18):30305
[14]张莉莉,陈伟,梁国强.北虫草素对体外小鼠肝癌H22细胞增殖及肝癌鼠血清中IL-2/TNF-α水平的影响[J].吉林中医药,2015,35(2):184
[15]Feng X J,Pan Q,Wang S M,et al.MAP4K4 promotes epithelialmesenchymal transition and metastasis in hepatocellular carcinoma[J].Tumor Biol,2016,37(8):11457
[2]张志杨,朱磊,贾平,等.柯里拉京对肝癌细胞增殖、迁移和侵袭的影响[J].中国实验方剂学杂志,2017,22(21):153
[3]Gong J,Shen S,Yang Y,et al.Inhibition of FASN suppresses migration,invasion and growth in hepatoma carcinoma cells by deregulating the HIF-1α/IGFBP1 pathway[J].International J Oncol,2017,50(3):88
[4]张思,刘元林,李雪,等.顺铂通过抑制转移抑制基因1-细胞外信号调节激酶及丝氨酸/苏氨酸激酶激活抑制He La细胞增殖[J].中国药理学与毒理学杂志,2016,30(4):350
[5]梁彦玲,崔付超,姬广森.原发性肝癌患者行肝癌介入化疗栓塞术的可行性及安全性研究[J].癌症进展,2018,16(7):858
[6]吴爱兵,黎明春,麦宗炯,等.CK2α通过PI3K/Akt/GSK-3β信号通路调控肺腺癌A549细胞的侵袭及迁移[J].中国肺癌杂志,2017,20(4):233
[7]Sun J,Luo Q,Liu L,et al.Biomechanical profile of cancer stem-like cells derived from MHCC97H cell lines[J].J Biomechanics,2016,49(1):45
[8]李天然,黄晓斌,黄楚恒,等.骨髓间充质干细胞对经转化生长因子β1、骨桥蛋白基因沉默肝癌细胞MHCC97-H的影响[J].中国组织工程研究,2017,38(5):687
[9]Lin W,Zhong M,Liang S,et al.Emodin inhibits migration and invasion of MHCC-97H human hepatocellular carcinoma cells[J].Exper Therap Med,2016,12(5):3369
[10]李聪,邢秀亚,张巍,等.肝癌中星形细胞上调基因1作用于microRNA-885-5p/基质金属蛋白酶9信号通路促进肝癌转移的机制研究[J].中华消化外科杂志,2016,15(2):161
[11]Cu i L,Gao B O,Cao Z,et al.Downregulation of B7-H4 in the MHCC97-H hepatocellular carcinoma cell line by arsenic trioxide[J].Mol Med Rep,2016,13(3):2032
[12]陈翔,陆宝华,袁杰,等.老年结直肠癌血清可溶性细胞间黏附分子-1及炎性因子表达的临床意义[J].重庆医学,2016,45(16):2202
[13]Chen X,Xu H,Wu N,et al.Interaction between granulin A and enolase 1 attenuates the migration and invasion of human hepatoma cells:[J].Oncotarget,2017,8(18):30305
[14]张莉莉,陈伟,梁国强.北虫草素对体外小鼠肝癌H22细胞增殖及肝癌鼠血清中IL-2/TNF-α水平的影响[J].吉林中医药,2015,35(2):184
[15]Feng X J,Pan Q,Wang S M,et al.MAP4K4 promotes epithelialmesenchymal transition and metastasis in hepatocellular carcinoma[J].Tumor Biol,2016,37(8):11457