Expression profiling of developmental and regenerating liver in mice

Normal adult liver is uniquely capable of renewal and repair after injury. Whether this response represents simple hyperplasia of various liver elements or requires recapitulation of the genetic program of the developing liver is not known. To study these possibilities, we examined transcriptional programs of adult liver after partial hepatectomy and contrasted these with developing embryonic liver. Principal component analysis demonstrated that the time series of gene expression during liver regeneration does not segregate according to developmental transcription patterns. Gene ontology analysis revealed that liver restoration after hepatectomy and liver development differ dramatically with regard to transcription factors and chromatin structure modification. In contrast, the tissues are similar with regard to proliferationassociated genes. Consistent with these findings, realtime polymerase chain reaction showed transcription factors known to be important in liver development are not induced during liver regeneration. These three lines of evidence suggest that at a transcriptional level, restoration of liver mass after injury is best described as hepatocyte hyperplasia and not true regeneration. We speculate this novel pattern of gene expression may underlie the unique capacity of the liver to repair itself after injury. Keywords: time course In order to elucidate the molecular similarities between regenerating and developing liver, we performed high-density microarray analysis using Affymetrix MG 430 2.0 chips for targets at 0, 1, 2, 6, 12, 18, 24, 30, 48, and 72 hours after partial hepatectomy and at 10.5, 11.5, 12.5, 13.5, 14.5, and 16.5 days post conception (dpc). Each experimental time point is represented by two separate samples, each consisting of at least 3 pooled tissues from different animals. For example, 6 hepatectomies were performed for the 1 hour post-hepatectomy time point. Time 0 is used as control.

正常成年肝脏在损伤后拥有独特的再生修复能力。目前学界尚未明确，该修复反应究竟是肝脏各类组分的单纯增生，还是需要重现胚胎发育阶段肝脏的遗传程序。为探究这两种潜在机制，我们分析了部分肝切除术（partial hepatectomy）后成年小鼠肝脏的转录调控程序，并将其与胚胎发育阶段的肝脏转录组进行对比。

主成分分析（principal component analysis, PCA）结果显示，肝脏再生过程中的基因表达时间序列并未与发育转录模式聚类分离。基因本体论（gene ontology, GO）分析表明，肝切除术后的肝脏修复与肝脏发育在转录因子及染色质结构修饰通路中存在显著差异；与之相对，二者在增殖相关基因的表达谱上具有高度相似性。

与上述发现一致，实时聚合酶链式反应（real-time polymerase chain reaction）实验证实，在肝脏再生过程中，此前被证实对肝脏发育至关重要的转录因子并未被诱导激活。基于这三项独立证据，我们认为从转录组层面来看，损伤后肝脏体积的恢复更倾向于肝细胞增生（hepatocyte hyperplasia），而非真正的器官再生。我们推测，这种独特的基因表达模式或许是肝脏具备损伤后自我修复独特能力的分子基础。

关键词：时间进程（time course）

为阐明再生肝脏与发育肝脏之间的分子相似性，我们采用Affymetrix MG 430 2.0芯片开展高密度微阵列分析，检测样本的时间点涵盖：部分肝切除术后0、1、2、6、12、18、24、30、48及72小时，以及受孕后10.5、11.5、12.5、13.5、14.5及16.5天（days post conception, dpc）。每个实验时间点设置两份独立生物学重复样本，每份样本均由至少3只不同个体的肝脏组织混合制备。例如，针对肝切除术后1小时的时间点，我们共完成6例手术取材。以术后0小时的样本作为实验对照。