Activation of the pluripotency factor OCT4 in smooth muscle cells is atheroprotective. doi: 10.1038/nm.4109. Mus musculus

The multiple claims about reactivation of the embryonic stem cell (ESC) pluripotency factor OCT4 in somatic cells are highly controversial due to the fact that there is no direct evidence that OCT4 has a functional role in cells other than ESCs. Herein we demonstrate that smooth muscle cell (SMC)-specific knockout of Oct4 within atherosclerotic mice resulted in increased lesion size and multiple changes consistent with decreased plaque stability. SMC-lineage tracing studies showed that lesions from SMC-specific conditional Oct4 KO mice had a reduced number of SMCs likely due to impaired SMC migration. RNA-seq analysis of lesion specimens showed that loss of Oct4 in SMCs was associated with marked activation of genes associated with inflammation and suppression of genes associated with cell migration, a number of which were shown to be activated in cultured SMCs by the combination of hypoxia and oxidized phospholipids in an OCT4-dependent manner. Activation of Oct4 within SMCs was associated with hydroxymethylation of the Oct4 promoter and was HIF1α- and KLF4-dependent. Results provide the first genetic evidence that OCT4 plays a functional role in somatic cells and highlight the importance of further investigation of possible OCT4 functions in somatic cells. Overall design: In vivo: mRNA profiles of 18 week fed Western diet wild type (WT) and Oct4-/- mice were generated by deep sequencing, four animals per group, using Illumina HiSeq 2000. In vitro: a smooth muscle cell wild type (WT) and Oct4-/- (KO) primary aortic cell line was generated and used. mRNA profiles were generated by deep sequencing, in triplicates, using Illumina HiSeq 2000, for the following groups: WT-normoxia-vehicle; WT-normoxia-POVPC; KO-normoxia-vehicle; KO-normoxia-POVP; WT-hypoxia-vehicle; WT-hypoxia-POVPC; KO-hypoxia-vehicle; and KO-hypoxia-POVPC.

此前关于在体细胞中重激活胚胎干细胞（embryonic stem cell, ESC）多能性因子OCT4的多项研究主张存在高度争议，原因是尚无直接证据表明OCT4在胚胎干细胞以外的细胞中具备功能性作用。本文证实，在动脉粥样硬化小鼠体内进行平滑肌细胞（smooth muscle cell, SMC）特异性的Oct4基因敲除，会导致病变体积增大，并出现多种与斑块稳定性降低相符的特征性变化。平滑肌细胞谱系示踪实验显示，平滑肌细胞特异性条件性Oct4敲除小鼠的动脉粥样硬化病变中，平滑肌细胞数量减少，这可能是由于平滑肌细胞迁移能力受损所致。对病变标本的RNA测序（RNA-seq）分析显示，平滑肌细胞中Oct4的缺失与炎症相关基因的显著激活以及细胞迁移相关基因的抑制密切相关；其中多个基因已被证实，在缺氧联合氧化磷脂处理的培养平滑肌细胞中，会以OCT4依赖的方式被激活。平滑肌细胞中Oct4的激活与Oct4启动子的羟甲基化修饰相关，且该过程依赖于HIF1α和KLF4。本研究结果首次提供了遗传学证据，证实OCT4在体细胞中发挥功能性作用，并凸显了进一步探究OCT4在体细胞中潜在功能的重要性。实验设计概述：体内实验：通过Illumina HiSeq 2000平台进行深度测序，获取喂食西方饮食18周的野生型（wild type, WT）及Oct4基因敲除（Oct4-/-）小鼠的mRNA表达谱，每组包含4只小鼠。体外实验：构建并使用了野生型（WT）及Oct4基因敲除（KO）原代主动脉平滑肌细胞系。通过Illumina HiSeq 2000平台进行深度测序，获取以下各组的mRNA表达谱，每组设置3次生物学重复：WT-常氧-溶剂对照组；WT-常氧-POVPC处理组；KO-常氧-溶剂对照组；KO-常氧-POVP处理组；WT-缺氧-溶剂对照组；WT-缺氧-POVPC处理组；KO-缺氧-溶剂对照组；KO-缺氧-POVPC处理组。