角蛋白23在胃癌中表达的临床意义和生物学作用

The clinical significance and biological effect of Keratin 23 expression in gastric cancer

目的分析角蛋白23(Keratin 23)在胃癌组织中表达的意义及生物学作用, 探讨其作用机制。方法收集2018年8月至2020年3月中山大学孙逸仙纪念医院69例胃癌患者术后胃癌组织和癌旁组织样本, 采用免疫组织化学法或蛋白质印迹法(Western blot)检测胃癌组织、癌旁组织及胃癌细胞中Keratin 23的表达水平, 通过χ^(2)检验结果分析Keratin 23表达水平与胃癌患者临床病理特征关系;利用Kaplan-Meier Plotter在线分析Keratin 23表达情况与胃癌患者预后关系;采用3-(4, 5-二甲基噻唑-2)-2, 5-二苯基四氮唑溴盐(MTT)法、流式细胞术、Transwell侵袭实验、Western blot分别检测敲除Keratin 23表达后胃癌细胞的增殖能力、凋亡情况、侵袭能力、p38丝裂素活化蛋白激酶(p38)和细胞外调节激酶(ERK)总蛋白及磷酸化表达水平。结果 Keratin 23在胃癌组织和胃癌细胞中高表达(癌组织3.99±1.28, 癌旁正常组织2.04±1.34, t=8.699, P<0.05), Keratin 23的表达与肿瘤的直径、分化程度及淋巴转移有显著关系(χ^(2)=1.806、0.255、0.569, P<0.05), 并且Keratin 23高表达的胃癌患者预后差[低表达33.7%(139/412), 高表达23.9%(111/463), P<0.05];当利用小干扰RNA(siRNA)敲低Keratin 23的表达后, 胃癌细胞增殖能力明显低于对照组(Western blot法:tBGC-823=3.154, tMGC-803=5.094, P<0.05;MTT实验:t24 h=2.381, t48 h=4.462, t72 h=8.618, P<0.05)、侵袭能力明显低于对照组(tBGC-823=4.778, tMGC-803=3.830, P<0.05)、细胞凋亡率明显高于对照组(tBGC-823=18.910, tMGC-803=19.570, P<0.05), 并可降低p38和ERK通路的磷酸化水平(tp-p38=4.171, tMGC-803=2.624, P<0.05)。结论 Keratin 23对胃癌具有促癌作用, 其可能通过p38和ERK相关通路介导胃癌的发展。

Objective To analyze the significance and biological effects of Keratin 23 expression level in gastric cancer tissues and the potential mechanism.Methods A total of 69 clinical samples of gastric cancer tissues and adjacent tissues from August 2018 to March 2020 were collected from Sun Yat-sen Memorial Hospital.The expression level of Keratin 23 in gastric cancer tissues,adjacent gastric tissues and gastric cancer cells were detected by immunohistochemistry or Western blotting.Chi-square test was used to analyze clinical correlations between the expression levels of Keratin 23 and the clinicopathological characteristics of gastric cancer patients.Kaplan-Meier Plotter was used to analyze the relationship between expression levels of Keratin 23 and the prognosis of gastric cancer patients.The methylthiazolyldiphenyl-tetrazolium bromide(MTT)method,flow cytometry,Transwell invasion experiment and Western blotting were used to detect the proliferation,apoptosis,invasion,protein expression level and its phosphorylation levels of p38 mitogen-activated protein kinase(p38)and extracellular signal-regulated kinases in Keratin 23-silenced gastric cancer cells.Results Keratin 23 was highly expressed in gastric cancer tissues and gastric cancer cells(gastric cancer:3.99±1.28,normal tissue:2.04±1.34,t=8.699,P<0.05).The expression of Keratin 23 was significantly correlated to tumor diameter,degree of differentiation and lymphatic metastasis(χ^(2)=1.806,0.255,0.569,P<0.05).Kaplan-Meier analysis indicated that gastric cancer patients with high expression of Keratin 23 had poor overallsurvival[poor:33.7%(139/412),high:23.9%(111/463),P<0.05].After knocking down Keratin 23 by small interfering RNA(siRNA),the proliferation ability was significantly inhibited(Western blotting:tBGC-823=3.154,tMGC-803=5.094,P<0.05;MTT:t24 h=2.381,t48 h=4.462,t72 h=8.618,P<0.05)and invasion ability was significantly inhibited(tBGC-823=4.778,tMGC-803=3.830,P<0.05)in gastric cancer cells.Consistently,the apoptosis rate was increased(tBGC-823=18.910,tMGC-803=19.570,P<0.05),and the phosphorylation level of p38 and extracellular signal-regulated kinases pathways decreased(tp-p38=4.171,tMGC-803=2.624,P<0.05).Conclusion Keratin 23 promoted the progression in gastric cancer,which mediated its biological function through p38 and extracellular signal-regulated kinases(ERK)-related pathways.Our study supported Keratin 23 may be an effective biomarker for prognosis in gastric cancer.