背景与目的自噬相关基因(autophagy related genes,ARGs)可调控溶酶体的降解过程从而诱导细胞发生自噬,参与多种癌症的发生发展,肿瘤组织中ARGs的表达情况在预测患者生存方面具有很大的前景。本研究基于ARGs构建了肺腺癌(lung adenocarc...背景与目的自噬相关基因(autophagy related genes,ARGs)可调控溶酶体的降解过程从而诱导细胞发生自噬,参与多种癌症的发生发展,肿瘤组织中ARGs的表达情况在预测患者生存方面具有很大的前景。本研究基于ARGs构建了肺腺癌(lung adenocarcinoma,LUAD)预后风险评分模型。方法通过GeneCards数据库获得5,786个ARGs。从癌症基因组图谱(The Cancer Genome Atlas,TCGA)数据库收集了395个LUAD患者的基因表达谱及临床数据,提取所有ARGs的表达数据,利用R软件筛选差异表达的ARGs。对差异表达的ARGs进行生存分析,筛选有预后价值的ARGs并进行功能富集分析。利用套索(the least absolute shrinkage and selection operator,LASSO)回归和Cox回归模型构建ARGs的预后风险评分模型。绘制受试者工作特征曲线(receiver operating characteristic curve,ROC曲线)得到风险评分的最佳cut-off值,将患者分为高风险评分组和低风险评分组。计算ROC曲线下面积(area under curve,AUC)和绘制Kaplan-Meier生存曲线评估模型性能,并在外部数据集验证。最后利用单因素和多因素Cox回归分析评价模型是否具有独立预后价值,并分析其临床相关性。结果通过生存分析初步筛选了52个与预后相关的ARGs,以此为基础,利用LASSO回归和Cox回归分析构建了由5个ARGs(ADAM12、CAMP、DKK1、STRIP2和TFAP2A)组成的LUAD预后风险评分模型。该模型中,低风险评分组患者的生存时间明显优于高风险评分组(P<0.001),且在训练集(AUCmax=0.78)和两个外部验证集(AUCmax=0.88)中均展现出良好的预测性能。风险评分在单因素和多因素Cox回归分析中与LUAD患者预后显著相关(P<0.001),提示风险评分可作为LUAD潜在的独立预后因素。临床特征相关性分析表明高风险评分与高T分期、高肿瘤分期和发生不良预后密切相关。结论我们构建了一个由5个ARGs组成的LUAD风险评分模型,该模型可为预测LUAD患者预后提供参考,未来或可与恶性肿瘤(tumor node metastasis,TNM)分期联合应用于肺腺癌患者的预后预测。展开更多
Objective Fumonisin B1(FB1)is an important mycotoxin in nature worldwide.The biomechanical properties of cells are closely related to their structure and function,and the cytoskeleton is the structural and functional ...Objective Fumonisin B1(FB1)is an important mycotoxin in nature worldwide.The biomechanical properties of cells are closely related to their structure and function,and the cytoskeleton is the structural and functional basis of cells motility,and therefore,from a biomechanical point of view,the purpose of this study is to investigate the effects of FB1 on the biomechanical properties,migration capacity and cytoskeletal structure of human umbilical vein endothelial cells(HUVECs),which may lay an experimental foundation for further exploration of the toxicity mechanism of fumonisin.Methods HUVECs were cultured and treated with different concentrations of FB1.Then,CCK-8 kit was used to detect the effect of FB1 on the survival rate.The osmotic fragility of the cells was measured after treatment with different osmotic pressures for30 min.The cell membrane fluidity was measured by fluorescence polarization method.The cell electrophoretic mobility was measured by cell electrophoretic apparatus.The migration capacity of the cells was observed by scratch repair assay.The changes of reactive oxygen species and cytoskeletal structure were observed by confocal laser scanning microscopy.Finally,the mRNA and protein relative expression levels of cytoskeletal binding proteins were detected by real-time PCR,Western blotting and confocal laser scanning.Results The results of CCK-8 showed that FB1 could significantly inhibit the proliferation of HUVECs in a dose-and time-dependent manner.After treatment of HUVECs with FB1,the hypotonic resistance of the cell,cell surface charge,cell membrane fluidity and migration capacity were all weakened,while reactive oxygen species were significantly increased and the cytoskeletal structure was significantly reorganized.Furthermore,RTPCR results showed that the mRNA relative expression levels of cytoskeletal binding proteins,exception of actin,were down-regulated after treated with FB1.Besides,Western blotting and statistical analysis based on fluorescence intensity of laser confocal microscopy confirmed theses changes in protein level.Conclusions FB1 can significantly affect the biomechanical properties and motility of HUVECs,which may be directly correlated to the remodel of F-actin cytoskeleton,as well as the relative expression changes of cytoskeletal binding proteins.It is significant for further exploring the toxicity mechanism of fumonisin.展开更多
文摘背景与目的自噬相关基因(autophagy related genes,ARGs)可调控溶酶体的降解过程从而诱导细胞发生自噬,参与多种癌症的发生发展,肿瘤组织中ARGs的表达情况在预测患者生存方面具有很大的前景。本研究基于ARGs构建了肺腺癌(lung adenocarcinoma,LUAD)预后风险评分模型。方法通过GeneCards数据库获得5,786个ARGs。从癌症基因组图谱(The Cancer Genome Atlas,TCGA)数据库收集了395个LUAD患者的基因表达谱及临床数据,提取所有ARGs的表达数据,利用R软件筛选差异表达的ARGs。对差异表达的ARGs进行生存分析,筛选有预后价值的ARGs并进行功能富集分析。利用套索(the least absolute shrinkage and selection operator,LASSO)回归和Cox回归模型构建ARGs的预后风险评分模型。绘制受试者工作特征曲线(receiver operating characteristic curve,ROC曲线)得到风险评分的最佳cut-off值,将患者分为高风险评分组和低风险评分组。计算ROC曲线下面积(area under curve,AUC)和绘制Kaplan-Meier生存曲线评估模型性能,并在外部数据集验证。最后利用单因素和多因素Cox回归分析评价模型是否具有独立预后价值,并分析其临床相关性。结果通过生存分析初步筛选了52个与预后相关的ARGs,以此为基础,利用LASSO回归和Cox回归分析构建了由5个ARGs(ADAM12、CAMP、DKK1、STRIP2和TFAP2A)组成的LUAD预后风险评分模型。该模型中,低风险评分组患者的生存时间明显优于高风险评分组(P<0.001),且在训练集(AUCmax=0.78)和两个外部验证集(AUCmax=0.88)中均展现出良好的预测性能。风险评分在单因素和多因素Cox回归分析中与LUAD患者预后显著相关(P<0.001),提示风险评分可作为LUAD潜在的独立预后因素。临床特征相关性分析表明高风险评分与高T分期、高肿瘤分期和发生不良预后密切相关。结论我们构建了一个由5个ARGs组成的LUAD风险评分模型,该模型可为预测LUAD患者预后提供参考,未来或可与恶性肿瘤(tumor node metastasis,TNM)分期联合应用于肺腺癌患者的预后预测。
基金funded by the National Natural Science Foundation of China ( 31660258, 31771014,31860262,11762006)the Science and Technology Foundation of Guizhou Province ( 2019-2787,2018-1412, 2016-5676,2017-5718)+2 种基金the Science and Technology Innovative Talent Team of Guizhou Province ( 2015-4021)the 2011 Collaborative Innovation Program of Guizhou Province ( 2015-04 )the Cell and Gene Engineering Innovative Research Groups of Guizhou Province ( KY-2016-031)
文摘Objective Fumonisin B1(FB1)is an important mycotoxin in nature worldwide.The biomechanical properties of cells are closely related to their structure and function,and the cytoskeleton is the structural and functional basis of cells motility,and therefore,from a biomechanical point of view,the purpose of this study is to investigate the effects of FB1 on the biomechanical properties,migration capacity and cytoskeletal structure of human umbilical vein endothelial cells(HUVECs),which may lay an experimental foundation for further exploration of the toxicity mechanism of fumonisin.Methods HUVECs were cultured and treated with different concentrations of FB1.Then,CCK-8 kit was used to detect the effect of FB1 on the survival rate.The osmotic fragility of the cells was measured after treatment with different osmotic pressures for30 min.The cell membrane fluidity was measured by fluorescence polarization method.The cell electrophoretic mobility was measured by cell electrophoretic apparatus.The migration capacity of the cells was observed by scratch repair assay.The changes of reactive oxygen species and cytoskeletal structure were observed by confocal laser scanning microscopy.Finally,the mRNA and protein relative expression levels of cytoskeletal binding proteins were detected by real-time PCR,Western blotting and confocal laser scanning.Results The results of CCK-8 showed that FB1 could significantly inhibit the proliferation of HUVECs in a dose-and time-dependent manner.After treatment of HUVECs with FB1,the hypotonic resistance of the cell,cell surface charge,cell membrane fluidity and migration capacity were all weakened,while reactive oxygen species were significantly increased and the cytoskeletal structure was significantly reorganized.Furthermore,RTPCR results showed that the mRNA relative expression levels of cytoskeletal binding proteins,exception of actin,were down-regulated after treated with FB1.Besides,Western blotting and statistical analysis based on fluorescence intensity of laser confocal microscopy confirmed theses changes in protein level.Conclusions FB1 can significantly affect the biomechanical properties and motility of HUVECs,which may be directly correlated to the remodel of F-actin cytoskeleton,as well as the relative expression changes of cytoskeletal binding proteins.It is significant for further exploring the toxicity mechanism of fumonisin.
