长链非编码RNA人肺腺癌转移相关转录本1在早产儿高氧肺损伤中的表达及意义

Expression and significance of long non-coding RNA metastasis associated in lung denocarcinoma tran-script 1 in the process of hyperoxia-induced lung injury in preterm infants

目的 探讨长链非编码RNA（long noncoding RNA,lncRNA）人肺腺癌转移相关转录本1 （metastasis associated in lung denocarcinoma transcript 1,MALAT1）对早产儿高氧肺损伤的保护作用.方法 从基因表达数据库GEO（gene expression omnibus）下载芯片数据集GSE25286（Affymetrix Mouse Genome 4302.0 Array）,依据高氧暴露与否,比较14 d、29 d的小鼠正常肺组织和高氧肺损伤的肺组织MALAT1 mRNA的表达.从GEO中下载芯片数据集GSE43830（Human Exon 1.0 ST Arrays）,在WI 38细胞（肺成纤维细胞）中敲除MALAT1后和未敲除MALAT1的WI38细胞中,用反义RNA技术检测比较多种基因如细胞分裂周期蛋白6（cell division cycle 6,CDC6）、死亡效应结构域蛋白2（death effector domain containing 2,DEDD2）及细胞周期蛋白Cyclin B1（CCNB1）的表达变化.同时,收集早产儿外周血标本验证,2015年1月至2016年12月在我院NICU住院治疗的高氧肺损伤早产儿20例为试验组,无高氧肺损伤的20例早产儿为对照组,采集40例早产儿末梢外周血,抽提RNA,逆转录后进行Real time-PCR检测40例早产儿外周血MALAT1的表达,收集并分析40例早产儿的一般临床资料.结果（1）通过使用软件对芯片源文件进行预处理和差异表达基因筛选,从最终的差异表达基因中发现, lncRNA MALAT1在高氧肺损伤的小鼠肺组织中表达明显上调[差异倍数（fold change,FC） =2.33,P=0.001].（2）WI38细胞 MALAT1敲除后,MALAT1表达大幅下调（FC = -15.6,P =0.000）,CDC6 （FC= -2.37,P =0.001）及 CCNB1（FC = -2.16,P =0.002）均表达下调,DEDD2表达上调（FC =2.46,P=0.000）.（3）早产儿外周血标本验证结果表明,与对照组早产儿（0.2734 ± 0.0673）比较, MALAT1在高氧肺损伤早产儿的外周血中表达显著上调（0.3755 ± 0.0819,t =4.634,P =0.015）.结论 lncRNA MALAT1可能通过抑制细胞凋亡,对早产儿高氧肺损伤起保护作用,为临床防治早产儿高氧肺损伤提供新策略.

Objective To explore the protective effect of long non-coding RNA（lncRNA） metasta-sis associated in lung denocarcinoma transcript 1 （MALAT1） involved in hyperoxia-induced lung injury in preterm infants.Methods This study had downloaded chip data set GSE25286 （Mouse GEO Genome 430 2.0 Array） from gene expression database gene expression omnibus （GEO）,according to the state of hyperoxia exposure,the MALAT1 mRNA expression in rats normal lung tissues and hyperoxic lung tissues was compared at day 14th and 29th.In chip data set GSE43830（Human Exon 1.0 ST Arrays） from GEO,the expression of multi-ple genes[cell division cycle 6（CDC6）,death effector domain containing 2（DEDD2）,and Cyclin B1 （CCNB1）] in WI38 cells（lung fibroblasts） was compared before and after MALAT1 was knockout.At the same time,the peripheral blood samples of premature infants were collected to verify.Totally 40 premature infants were hospitalized in the department of neonatology in our hospital from Jan 2015 to Dec 2016,the pe-ripheral blood samples of 40 premature infants were collected.RNA was extracted and Real time-PCR was performed after reverse transcription,clinical data of these 40 cases were retrospectively analyzed. Results （1） By using Affymetrix Expression console and Affymetrix Transcriptome analysis console software source files of the chip of pretreatment and difference expression gene screening,the expression of lncRNA MALAT1 gene in lung tissues of hyperoxia lung injury mice significantly upregulated[fold change（FC） =2.33,P=0.001].（2） After MALAT1 in WI38 cell was knockout,MALAT1 expression was significantly reduced（FC= -15.6,P=0.000）,the expression of CDC6（FC= -2.37,P=0.001） and CCNB1（FC=-2.16,P=0.002） were down regulated,DEDD2 expression was up regulated（FC =2.46,P =0.000）. （3） The results of peripheral blood samples from preterm infants showed that the expression of MALAT1 was significantly increased in preterm infants with hyperoxia-induced lung injury（0.375 5 ± 0.081 9,t =4.634, P=0.015）,compared with normal preterm infants（0.273 4 ± 0.067 3）.Conclusion Through inhibiting cell apoptosis,lncRNA MALAT1 can protect preterm infants with hyperoxia-induced lung injury,it may provide a new strategy for prevention and treatment of hyperoxia-induced lung injury in premature infants.