糖皮质激素对晶状体上皮细胞生物学功能调控作用的生物信息学分析

Regulatory effect of glucocorticoid on the biological function of lens epithelial cells: a bioinformatics analysis

目的利用生物信息学方法分析糖皮质激素对晶状体上皮细胞(LECs)生物学功能的影响,并预测相关的微小RNA(miRNA)。方法下载GSE3040数据集,设置1 μmol/L地塞米松处理的人LECs细胞株HLE-B3细胞为实验组,1 μmol/L二甲基亚砜(DMSO)处理的HLE-B3细胞为对照组,利用GEO2R工具分析2个组间的差异表达基因。利用Metascape网站进行差异基因功能富集分析,通过EdU细胞增生实验检测2个组间细胞增生差异。通过STRING网站和cytoscape软件构建蛋白互作网络图,cytohubba app计算出hub基因,采用实时荧光定量PCR法检测2个组间hub基因的表达差异。利用mirCode数据库预测与hub基因相关的miRNA。结果实验组与对照组间分析出341个差异基因,其中上调基因为300个,下调基因为41个。下调基因中差异最显著的5个基因是SLC12A1、MED13L、ALDH5A1、SLC15A3和WWC1基因;上调基因中差异最显著的5个基因是SCNN1A、ANKRD36、FKBP5、PYY和ADH1B基因。列出了富集量排名前20的生物学功能关键词,结果显示富集量最大的是对HLE-B3细胞增生的负向调节。实验组细胞增生率为(8.09±0.20)%,低于对照组的(39.63±0.80)%,差异有统计学意义(t=38.43,P<0.01)。前10位hub基因分别是SST、CXCL8、GRM1、GNRH1、CXCL5、PPBP、CX3CR1、PYY、EDNRA和GRK5,实时荧光定量PCR结果显示2个组间SST、CXCL8、GRM1、PYY、EDNRA和GRK5 mRNA相对表达量比较差异均有统计学意义(均P<0.05)。与hub基因相关性强的前6位miRNA分别是miR-15abc、miR-214、miR-23abc、miR-129-5p、miR-132和miR-24。结论 1 μmol/L地塞米松即可对HLE-B3细胞的增生产生负性调控作用。SST、CXCL8、GRM1、PYY、EDNRA和GRK5基因可能是糖皮质激素的作用靶点,miR-15abc、miR-214、miR-23abc、miR-129-5p、miR-132和miR-24最可能与hub基因相关。

Objective To analyze the effect of glucocorticoid on the biological function of lens epithelial cells(LECs)by bioinformatics and predict related microRNA(miRNA).Methods GSE3040 database was downloaded and the human LECs line(HLE-B3)cells in the experimental group were treated with 1μmol/L dexamethasone,and HLE-B3 cells in the control group were treated with 1μmol/L dimethyl sulfoxide(DMSO).GEO2R was used to analyze the differentially expressed genes between the two groups.Metascape website was employed to analyze the functional enrichment of differentially expressed genes,and EdU cell proliferation assay was performed to detect the difference in cell proliferation between the two groups.STRING website and cytoscape software were used to construct protein-protein interaction network.Hub genes were calculated by cytohubba app,and quantitative real-time PCR was performed to detect the expression levels of hub genes between the two groups.MirCode website was used to predict the related miRNAs.Results A total of 341 differentially expressed genes were detected between the experimental group and the control group,among which there were 300 up-regulated genes and 41 down-regulated genes.SLC12A1,MED13L,ALDH5A1,SLC15A3 and WWC1 were the top five down-regulated genes and SCNN1A,ANKRD36,FKBP5,PYY and ADH1B were the top five up-regulated genes.The top 20 terms of functional enrichment were listed,and the negative regulation of HLE-B3 cells proliferation showed the most enrichment.Cell proliferation rate in the experimental group was(8.09±0.20)%,which was significantly lower than(39.63±0.80)%in the control group(t=38.43,P<0.01).The top ten hub genes were SST,CXCL8,GRM1,GNRH1,CXCL5,PPBP,CX3CR1,PYY,EDNRA and GRK5,and quantitative real time PCR confirmed that the expression levels of SST,CXCL8,GRM1,PYY,EDNRA and GRK5 mRNA were statistically different(all at P<0.05).The top six miRNAs which might be associated with hub genes were miR-15abc,miR-214,miR-23abc,miR-129-5p,miR-132 and miR-24.Conclusions The 1μmol/L glucocorticoid can negatively regulate the proliferation of HLE-B3 cells.SST,CXCL8,GRM1,PYY,EDNRA and GRK5 may be hub genes and miR-15abc,miR-214,miR-23abc,miR-129-5p,miR-132,miR-24 are most likely to relate to them.