Burton Gregory R, Nagarajan Radhakrishnan, Peterson Charlotte A, McGehee Robert E
University of Arkansas for Medical Sciences, Little Rock, AR 72205, USA.
Gene. 2004 Mar 31;329:167-85. doi: 10.1016/j.gene.2003.12.012.
During cellular differentiation and development, it is recognized that many complex molecular mechanisms as well as precise patterns of differentially expressed genes occur in directing precursor cells toward a given lineage. Using microarray-based technology, we examined gene expression across the course of 3T3-L1 adipocyte differentiation. Total cellular RNA was isolated at times 0, 2, 8, 16, 24, 48, and 96 h following treatment with either standard hormonal inducers of differentiation; insulin, dexamethasone, isobutylmethylxanthine (IDX), or IDX plus trichostatin A (TsA), a histone deacetylase inhibitor and potent adipogenic inhibitor. cRNA was synthesized from cellular RNA and hybridized to high density Affymetrix MG_U74Av2 microarray gene chips containing 12,488 cDNA/Expressed Sequence Tags (ESTs) probe sets. From the IDX-only treated cells, all probe sets that were either unchanged or differentially expressed less than 2-fold throughout differentiation with respect to time 0 preadipocytes were excluded from further analyses. This selection resulted in a net of 1686 transcripts, 859 were increased in expression, and 827 were decreased in expression at least 2-fold across differentiation. To focus in on genes that were more specific to differentiation, the same analysis was performed on IDX plus TsA-treated non-differentiating cells and all probe sets from the IDX-only group that exhibited similar expression profiles in the non-differentiating TsA-treated group were excluded leaving a total of 1016 transcripts that were regulated only under differentiating conditions. Six hundred and thirty-six of these transcripts were elevated at least 2-fold and 380 exhibited a decrease in expression relative to time 0 preadipocytes. This group of genes was further analyzed using hierarchical clustering and self-organizing maps and resulted in the identification of numerous genes not previously known to be regulated during adipocyte differentiation. Many of these genes may well represent novel adipogenic mediators and markers of adipogenesis.
在细胞分化和发育过程中，人们认识到，在引导前体细胞走向特定谱系时会出现许多复杂的分子机制以及差异表达基因的精确模式。我们使用基于微阵列的技术，研究了3T3-L1脂肪细胞分化过程中的基因表达情况。在用标准激素分化诱导剂（胰岛素、地塞米松、异丁基甲基黄嘌呤（IDX）或IDX加曲古抑菌素A（TsA），一种组蛋白脱乙酰酶抑制剂和强效脂肪生成抑制剂）处理后的0、2、8、16、24、48和96小时，分离总细胞RNA。从细胞RNA合成cRNA，并与包含12,488个cDNA/表达序列标签（EST）探针组的高密度Affymetrix MG_U74Av2微阵列基因芯片杂交。对于仅用IDX处理的细胞，所有在整个分化过程中相对于0时前脂肪细胞不变或差异表达小于2倍的探针组都被排除在进一步分析之外。这种筛选产生了1686个转录本的净值，其中859个表达增加，827个在整个分化过程中表达至少降低2倍。为了专注于更特异于分化的基因，对用IDX加TsA处理的未分化细胞进行了相同的分析，并排除了仅用IDX处理组中在未分化的TsA处理组中表现出相似表达谱的所有探针组，留下总共1016个仅在分化条件下受调控的转录本。相对于0时前脂肪细胞，这些转录本中有636个升高至少2倍，380个表达降低。使用层次聚类和自组织图对这组基因进行了进一步分析，结果鉴定出许多以前不知道在脂肪细胞分化过程中受调控的基因。其中许多基因很可能代表新的脂肪生成介质和成脂标记物。
