胚胎干细胞分化为表皮样细胞的基因表达谱不同

胚胎干细胞分化为表皮样细胞的基因表达谱

不同

张仁礼 刘彩霞 孟锦绣 闻安民 张丽丽 韩冬

【摘要】 【目的】用全基因表达谱芯片技术挑选小鼠胚胎干细胞(ESC)定向分化为表皮样细胞(ELC)的不同表达的基因并对其进行分析,以进一步说明ESC分化为ELC的分子机制?【方式】 利用人羊膜成立体外诱导体系,将小鼠ESC诱导定向分化为ELC,做3次生物学重复,别离取未分化的ESC和诱导分化的ELC提取总RNA进行扩增?Cy3标记,与NimbleGen 135k 小鼠全基因表达谱芯片(含44170个基因)杂交,荧光扫描,挑选出不同表达基因,并进行GO分析和pathway分析,用实时定量PCR对结果进行验证?【结果】 芯片挑选结果显示,分化后的ELC与ESC间的不同表达的基因多达4856个(Cut off:2)?基因本体(GO)分析显示与生物学进程相关的不同表达基因最多的是细胞进程,1931个;与亚细胞组分(CC)相关的差基因中最多的是细胞和细胞组分相关基因,2391个;与分子功能相关的不同基因中最多是的结合功能(binding)基因,1869个?Pathway分析显示,与DNA复制通路相关的不同基因有23个;与蛋白酶体通路相关的24个;剪接体通路相关有47个;细胞周期相关的有44个?【结论】 ESC体外诱导定向分化为ELC进程中,基因表达谱发生了庞大的转变,提示细胞分化是一个众多基因参与的复杂的生物学进程,这些基因在分化进程所起的作用和作用机制尚需大量的实验研究说明?

【关键词】 全基因组表达芯片; 胚胎干细胞; 表皮样细胞; GO分析; Pathway分析

Abstract: 【Objective】 To screen and analyze different expressed genes between mouse embryonic stem cells (ESC) and their differentiated epidermal-like cells (ELC) with the whole genomic expression profile chip, and explore the molecular mechanisms of differentiation further.【Methods】 Mouse ESC was induced differentiation into ELC in vitro by human amnion repeated for three times. Total RNA of ESC and ELC was extracted, and then amplified by RT-PCR. The product cDNA was labeled with Cy3 and hybridized on NimbleGen 135k mouse whole gene chip (containing 44, 170 genes). Fluorescence scanning, Gene Ontology (GO) and pathway analysis were used to screen the different expressed genes. The analytic result was validated by Real-time Quantitative PCR. 【Results】 A total of 4,856 genes showed differentially expressed between ESC and ELC (Cut off: 2). GO analysis showed that the differential expressed genes between ESC and ELC mainly involved cellular process (CP) genes of 1931, Cell/Cell part genes of 2391,and binding function genes of 1869, which belong to biological process (BP), cellular

component (CC), and molecular function (MF), respectively. The pathway analysis showed that the differential expressed genes mainly involved genes related to DNA replication, proteasome pathway, spliceosome pathway, and cell cycle with the number of 23, 24, 47, and 44, respectively. 【Conclusion】 The change of expression profile between ESC and ELC indicated that large variety of genes were involved in this complex cellular differentiation process, and how these gene worked and co worked to regulate the differentiation procedure need further investigations.

Key words: whole genome expression chip; ES cells; epidermal-like cells; GO analysis; pathway analysis

胰岛素抗击不仅是2型糖尿病的发病基础,更是贯穿多种代谢相关疾病的一起病理生理基础?胰岛素抗击加速了2型糖尿病的发生和进展,也增加了患心血管系统疾病的危险性[1]?因此,如何防治胰岛素抗击十分重要?最近的调查说明,白藜芦醇(resveratrol,RSV)对高脂诱导的胰岛素抗击综合征[2]具有爱惜作用,但分子机制仍不清楚?骨骼肌是机体要紧的胰岛素靶器官,胰岛素介导的葡萄糖摄取,75%以上由骨骼肌细胞负责[3]?咱们以往的研究说明,雌激素可通过上调骨骼肌细胞微囊蛋白-3(caveolin-3,CAV-3)的表达,增进葡萄糖摄取[4]?