小细胞肺癌肌球蛋白重链9表达下调参与细胞机械感知功能缺失  

Downregulation of myosin heavy chain 9 impairs the mechano-sensing function in small cell lung cancer

作　　者：李明君[1] 杨宁宁[2] 罗明 董卫萍 王文刚[5] Li Mingjun;Yang Ningning;Luo Ming;Dong Weiping;Wang Wengang(Department of Oncology,the First Affiliated Hospital of Zhengzhou University,Zhengzhou 450052,China;Department of Emergency,the First Affiliated Hospital of Zhengzhou University,Zhengzhou 450052,China;Department of Orthopedics,Zhongnan Hospital of Wuhuan University,Wuhan 430062,China;Department of Breast Surgery,the First Affiliated Hospital of Zhengzhou University,Zhengzhou 450052,China;Department of Orthopedics,the First Affiliated Hospital of Zhengzhou University,Zhengzhou 450052,China)

机构地区：[1]郑州大学第一附属医院肿瘤科,郑州450052 [2]郑州大学第一附属医院急诊科,郑州450052 [3]武汉大学中南医院骨科,武汉430062 [4]郑州大学第一附属医院乳腺外科,郑州450052 [5]郑州大学第一附属医院骨科,郑州450052

出　　处：《中华实验外科杂志》2024年第6期1241-1244,共4页Chinese Journal of Experimental Surgery

基　　金：河南省高等学校重点科研项目计划(22A320008、24A320066);河南省科技攻关项目(242102310075);吴阶平医学基金会临床科研专项(320.6750.2023-1-24)。

摘　　要：目的探究小细胞肺癌的机械感知功能及其关键蛋白肌球蛋白重链9(MYH9)在细胞失巢凋亡中的作用。方法采用3 kPa和12 kPa基质刚度水凝胶,比较小细胞肺癌细胞的Paxillin面积和细胞极化。组织和细胞系水平从11个重要机械感知蛋白中筛选关键蛋白MYH9。通过慢病毒敲减和过表达MYH9表达水平,明确MYH9对细胞机械感知功能的作用,采用流式细胞膜联蛋白V(Annexin V)/碘化丙锭(PI)双染法测定MYH9对细胞失巢凋亡能力的影响。两组连续变量采用独立样本的t检验,3组组间比较采用One-Way ANOVA分析。结果人支气管上皮细胞的软基质组黏着斑面积[(0.79±0.17)μm^(2)比(1.68±0.30)μm^(2),t=8.26,P<0.01]和细胞形态比值(1.30±0.22比1.81±0.26,t=4.65,P<0.01)低于硬基质组。小细胞肺癌细胞的软基质组黏着斑面积[(0.48±0.21)μm^(2)比(0.53±0.22)μm^(2),t=0.51,P>0.05]和细胞形态比值(1.26±0.20比1.36±0.22,t=1.06,P>0.05)较硬基质组差异无统计学意义。筛选基因表达综合数据库(GEO)中小细胞肺癌组织的转录组测序数据,小细胞肺癌细胞组MYH9蛋白表达(0.12±0.02比0.30±0.02,t=11.44,P<0.01)低于对照组。过表达小细胞肺癌细胞中MYH9后,软基质组黏着斑面积[(0.69±0.21)μm^(2)比(1.12±0.43)μm^(2),t=2.85,P<0.05]和细胞形态比值(1.37±0.18比1.70±0.34,t=2.72,P<0.05)低于硬基质组,漂浮培养下的早期凋亡细胞比例[(12.81±1.92)%比(4.81±1.30)%,t=7.69,P<0.01]高于硬基质组。结论小细胞肺癌细胞的机械感知功能缺失,其关键功能蛋白MYH9显著下调。过表达MYH9在恢复小细胞肺癌细胞机械感知功能的同时,削弱其抗脱落凋亡能力。Objective To investigate the mechano-sensing function of small cell lung cancer(SCLC)cells and the role of the key protein myosin heavy chain 9(MYH9)in cell anoikis.Methods Using 3 kPa and 12 kPa matrix stiffness hydrogels,the Paxillin area and cell polarization of SCLC cells were compared.At both tissue and cellular levels,the key protein MYH9 was identified from 11 important mechanosensitive proteins.By manipulating MYH9 expression levels through lentiviral knockdown and overexpression,the role of MYH9 in cell mechanosensing function was elucidated.Furthermore,its effect on cell anoikis ability was further determined using flow cytometry Annexin V/propidium iodide(PI)double staining.The two groups of continuous variables were subjected to independent samples t-test,while the comparison among three groups was analyzed using One-Way ANOVA.Results Compared to stiff substrates,the adhesion plaque area[(0.79±0.17)vs.(1.68±0.30)μm^(2),t=8.26,P<0.01]and cell aspect ratio(1.30±0.22 vs.1.81±0.26,t=4.65,P<0.01)of human bronchial epithelial cells on soft substrates were significantly decreased,while the adhesion plaque area[(0.48±0.21)vs.(0.53±0.22)μm^(2),t=0.51,P>0.05]and cell aspect ratio(1.26±0.20 vs.1.36±0.22,t=1.06,P>0.05)of SCLC cells showed no significant changes.Analysis of transcriptome sequencing data of SCLC tissues from the gene expression omnibus(GEO)database revealed significant downregulation of MYH9 protein expression in SCLC cells(0.12±0.02 vs.0.30±0.02,t=11.44,P<0.01).Lentivirus-mediated overexpression of MYH9 in SCLC cells led to a significant decrease in adhesion plaque area[(0.69±0.21)vs.(1.12±0.43)μm^(2),t=2.85,P<0.05]and cell aspect ratio(1.37±0.18 vs.1.70±0.34,t=2.72,P<0.05)on soft substrates compared to stiff substrates,and an increased proportion of early apoptotic cells in floating culture[(12.81±1.92)%vs.(4.81±1.30)%,t=7.69,P<0.01].Conclusion Impairment of mechano-sensing function in SCLC cells is associated with significant downregulation of the key protein MYH9.Overexpressi

关 键 词：小细胞肺癌 机械感知 脱落凋亡 

分 类 号：R734.2[医药卫生—肿瘤]