long non-coding RNA expression profiles in Paraquat-induced pulmonary fibrosis. Mus musculus

In this study, using the next generation transcriptome sequencing and microarray analysis, we profiled expression of long non-coding RNAs (lncRNAs) in a mouse model of paraquat-induced pulmonary fibrosis. We identified 513 up-regulated and 204 down-regulated lncRNAs (fold-change ≥5.0) when comparing fibrotic to normal lung tissues. Quantitative real-time PCR, Gene Ontology analysis, Pathway analysis and Bioinformatics analysis were performed for further research. Furthermore, we predicted and verified the target genes of two up-regulated lncRNAs, uc.77 and 2700086A05Rik, as Zeb2 and Hoxa3 respectively, both of which are important regulators of EMT. Overall design: BALB/c mice (7-8 weeks, male, 19.8 g±0.63 g) were provided by Experimental Animal Center of Nanjing Medical University, China. A total of 20 BALB/c mice were randomly divided into 2 groups (n=10 in each group): the normal control and pulmonary fibrosis model group. Mice in the model group were administered 10mg/kg paraquat (SIGMA Chemical Co., St. Louis, MO, USA) dissolved in sterile saline via intraperitoneal injection (once per day for 5 days). The normal control group were administered an equal volume of saline at the same time. Lung tissues were harvested on the 12st day following the first treatment. The motality of paraquat group was 70%. Three lung tissues from each group were pick up and mixed to prepare for the RNA extraction and the following microarray analysis.

本研究采用下一代转录组测序（next generation transcriptome sequencing）与微阵列分析技术，对百草枯诱导肺纤维化小鼠模型中的长链非编码RNA（long non-coding RNAs, lncRNAs）表达谱进行了分析。相较于正常肺组织，纤维化肺组织中共鉴定出513个上调、204个下调的lncRNAs（倍数变化≥5.0）。本研究通过实时定量PCR、基因本体（Gene Ontology, GO）分析、通路分析及生物信息学分析开展后续研究。进一步地，本研究预测并验证了两个上调lncRNAs——uc.77与2700086A05Rik——的靶基因分别为Zeb2与Hoxa3，二者均为上皮间质转化（epithelial-mesenchymal transition, EMT）的重要调控因子。
实验设计：中国南京医科大学实验动物中心提供20只7~8周龄雄性BALB/c小鼠，体质量为19.8 g±0.63 g。将小鼠随机分为两组（每组n=10）：正常对照组与肺纤维化模型组。模型组小鼠经腹腔注射溶于无菌生理盐水的10 mg/kg百草枯（美国密苏里州圣路易斯市西格玛化工有限公司产品），每日1次，连续给药5天；正常对照组同期注射等体积生理盐水。首次给药后第12天采集肺组织。百草枯模型组小鼠死亡率为70%。每组取3份肺组织混合后，用于RNA提取及后续微阵列分析。