Application of Next Generation Sequencing (RNA-seq and miRNA-seq) to study the molecular signature of Aluminium hydroxide adjuvant in ovine encephalon. [RNA-Seq]

We report the application of RNA sequencing technology for high-throughput profiling of the aluminium hydroxide adjuvant mechanism of action in an in vivo experiment on sheep treated with commercial vaccines, with the adjuvant alone or with PBS (control group). We mapped about 74.1 million sequence reads per sample to the sheep genome (build Oar3.1) and after filtering 16,369 genes were expressed, of which 14,387 were annotated genes in Ensembl and 1,982 were candidate lncRNAs. A total of 13, 63 and 76 differentialy expressed genes were identified in the Vaccine vs. Control, Adjuvant vs. Control and Adjuvant vs. Vaccine comparisons, respectively. Previously reported genes related to neuronal development, brain transport and neurotransmission, brain injury and neurodegenerative diseases associated with aluminum were found differentially expressed, namely VCAM1, TRPM4, GDF10 and NTN1. Overall design: Encephalon Total RNA profiles of twelve age-matched Rasa sheep after inoculation of nine different commercial vaccines based of aluminium hydroxide (the vaccine group composed of 4 animals), with the aluminium adjuvant alone in an equivalent dose (the adjuvant group composed of 4 animals) or with PBS (the control group composed of 4 animals) were generated by deep sequencing on an Illumina HiSeq2000 sequencer.

本研究报道了一项体内实验的相关结果：利用高通量RNA测序技术，对接种商业疫苗、单独使用氢氧化铝佐剂或磷酸盐缓冲液（Phosphate Buffered Saline, PBS）对照组的绵羊进行检测，以解析氢氧化铝佐剂的作用机制。本研究将每个样本约7410万条测序读段比对至绵羊参考基因组（版本Oar3.1），经数据过滤后共检出16369个表达基因，其中14387个为Ensembl数据库注释基因，1982个为候选长链非编码RNA（long non-coding RNA, lncRNA）。在疫苗组vs对照组、佐剂组vs对照组及佐剂组vs疫苗组的三组比较中，分别鉴定得到13、63和76个差异表达基因。研究发现，此前已报道的与铝相关的神经元发育、脑物质转运、神经传递、脑损伤及神经退行性疾病相关的基因呈现差异表达，具体包括VCAM1、TRPM4、GDF10及NTN1。实验设计：本研究纳入12只年龄匹配的拉萨绵羊（Rasa sheep），将其分为三组（每组4只）：疫苗组接种9种基于氢氧化铝的商业疫苗，佐剂组接种等效剂量的单独氢氧化铝佐剂，对照组接种PBS。采集各组绵羊的脑组织总RNA，通过Illumina HiSeq2000测序仪进行深度测序，以获取全脑组织的转录组表达谱。