Parfitt Geraint J, Brown Donald J, Jester James V
The Gavin Herbert Eye Institute, School of Medicine, University of California, Irvine, CA.
Mol Vis. 2016 May 24;22:518-27. eCollection 2016.
Dry eye disease is a common condition associated with age-related meibomian gland dysfunction (ARMGD). We have previously shown that ARMGD occurs in old mice, similar to that observed in human patients with MGD. To begin to understand the mechanism underlying ARMGD, we generated transcriptome profiles of eyelids excised from young and old mice of both sexes.
Male and female C57BL/6 mice were euthanized at ages of 3 months or 2 years and their lower eyelids removed, the conjunctival epithelium scrapped off, and the tarsal plate, containing the meibomian glands, dissected from the overlying muscle and lid epidermis. RNA was isolated, enriched, and transcribed into cDNA and processed to generate four non-stranded libraries with distinct bar codes on each adaptor. The libraries were then sequenced and mapped to the mm10 reference genome, and expression results were gathered as reads per length of transcript in kilobases per million mapped reads (RPKM) values. Differential gene expression analyses were performed using CyberT.
Approximately 55 million reads were generated from each library. Expression data indicated that about 15,000 genes were expressed in these tissues. Of the genes that showed more than twofold significant differences in either young or old tissue, 698 were identified as differentially expressed. According to the Gene Ontology (GO) analysis, the cellular, developmental, and metabolic processes were found to be highly represented with Wnt function noted to be altered in the aging mouse.
The RNA sequencing data identified several signaling pathways, including fibroblast growth factor (FGF) and Wnt that were altered in the meibomian glands of aging mice.
干眼症是一种与年龄相关性睑板腺功能障碍（ARMGD）相关的常见病症。我们之前已经表明，ARMGD在老年小鼠中出现，类似于在患有睑板腺功能障碍的人类患者中观察到的情况。为了开始了解ARMGD的潜在机制，我们生成了从年轻和老年雌雄小鼠切除的眼睑的转录组图谱。
将3个月或2岁的雄性和雌性C57BL/6小鼠安乐死，切除其下眼睑，刮除结膜上皮，并从覆盖的肌肉和眼睑表皮中解剖出含有睑板腺的睑板。分离RNA，进行富集，转录成cDNA，并进行处理以生成四个非链文库，每个接头带有不同的条形码。然后对文库进行测序并映射到mm10参考基因组，并将表达结果收集为每百万映射读数中每千碱基转录本长度的读数（RPKM）值。使用CyberT进行差异基因表达分析。
每个文库产生了约5500万个读数。表达数据表明，这些组织中约有15000个基因表达。在年轻或老年组织中显示出超过两倍显著差异的基因中，有698个被鉴定为差异表达基因。根据基因本体论（GO）分析，发现细胞、发育和代谢过程高度富集，并且注意到衰老小鼠中Wnt功能发生改变。
RNA测序数据确定了几种信号通路，包括成纤维细胞生长因子（FGF）和Wnt，它们在衰老小鼠的睑板腺中发生了改变。
