Adipocyte nuclei captured from VAT and SAT.
Adipocyte nuclei captured from VAT and SAT.
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DOI：
10.1186/s40608-016-0112-6
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发表时间：
2016
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影响因子：
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通讯作者：
Meagher RB
中科院分区：
文献类型：
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作者：
Ambati S;Yu P;McKinney EC;Kandasamy MK;Hartzell D;Baile CA;Meagher RB
肥胖相关的合并症被认为是由许多组织和细胞类型中的表观基因组重编程引起的，特别是内脏和皮下脂肪组织中的成熟脂肪细胞、VAT和SAT。VAT和SAT中成熟脂肪细胞的细胞类型特异性染色质重塑知之甚少，部分原因是难以从脂肪组织中分离和操作大型脆性成熟脂肪细胞。我们使用脂联素（ADIPOQ）启动子（ADNp）构建了MA-INTACT（成熟脂肪细胞-特定细胞类型中标记的核分离）小鼠，以报告蛋白标记成熟脂肪细胞核的表面。SUN 1 mRFP 1Flag报告基因由核跨膜蛋白SUN 1的片段、荧光蛋白mRFP 1和Flag表位标签的三个拷贝组成。成熟的脂肪细胞核从MA-INTACT小鼠的VAT和SAT（分别为MVA和MSA核）被快速有效地免疫捕获。MVA和MSA细胞核中脂肪细胞特异性转录物ADIPOQ、PPARg 2、EDNRB和LEP的水平比未捕获的细胞核高1,000至10,000倍，而后者表达的白细胞和内皮细胞标志物IKZF 1、RETN、SERPINF 1、SERPINE 1、ILF 3和TNFA的水平更高。MVA和MSA细胞核差异表达了与脂肪形成或肥胖相关的健康风险相关的几个因子，包括CEBPA，KLF 2，RETN，SERPINE 1和TNFA。不同的细胞核群体显著差异表达的转录本编码染色质重塑蛋白，调节DNA胞嘧啶甲基化和羟甲基化（TdR，DNMT，TDG，GADD 45）和核小体组蛋白修饰（ARID 1A，KAT 2B，KDM 4A，PRMT 1，PRMT 5，PAXIP 1）。值得注意的是，MSA和MVA核表达200至1000倍高水平的产热标记转录PRDM 16和UCP 1。MA-INTACT小鼠能够以简单的方式对来自VAT和SAT的高度纯化的成熟脂肪细胞核进行细胞类型特异性分析，并提高了收集的脂肪细胞数据的统计学显著性。孤立的增值税和SAT脂肪细胞核表达不同的模式的转录编码染色质重塑因子和蛋白质相关的糖尿病，心血管疾病和产热。MA-INTACT小鼠是测试热量摄入、膳食营养素、运动和药物对表观基因组诱导的肥胖健康风险的影响的有用模型。本文的在线版本（doi：10.1186/s40608-016-0112-6）包含补充材料，可供授权用户使用。
Obesity-related comorbidities are thought to result from the reprogramming of the epigenome in numerous tissues and cell types, and in particular, mature adipocytes within visceral and subcutaneous adipose tissue, VAT and SAT. The cell-type specific chromatin remodeling of mature adipocytes within VAT and SAT is poorly understood, in part, because of the difficulties of isolating and manipulating large fragile mature adipocyte cells from adipose tissues. We constructed MA-INTACT (Mature Adipocyte-Isolation of Nuclei TAgged in specific Cell Types) mice using the adiponectin (ADIPOQ) promoter (ADNp) to tag the surface of mature adipocyte nuclei with a reporter protein. The SUN1mRFP1Flag reporter is comprised of a fragment of the nuclear transmembrane protein SUN1, the fluorescent protein mRFP1, and three copies of the Flag epitope tag. Mature adipocyte nuclei were rapidly and efficiently immuno-captured from VAT and SAT (MVA and MSA nuclei, respectively), of MA-INTACT mice. MVA and MSA nuclei contained 1,000 to 10,000-fold higher levels of adipocyte-specific transcripts, ADIPOQ, PPARg2, EDNRB, and LEP, relative to uncaptured nuclei, while the latter expressed higher levels of leukocyte and endothelial cell markers IKZF1, RETN, SERPINF1, SERPINE1, ILF3, and TNFA. MVA and MSA nuclei differentially expressed several factors linked to adipogenesis or obesity-related health risks including CEBPA, KLF2, RETN, SERPINE1, and TNFA. The various nuclear populations dramatically differentially expressed transcripts encoding chromatin remodeler proteins regulating DNA cytosine methylation and hydroxymethylation (TETs, DNMTs, TDG, GADD45s) and nucleosomal histone modification (ARID1A, KAT2B, KDM4A, PRMT1, PRMT5, PAXIP1). Remarkably, MSA and MVA nuclei expressed 200 to 1000-fold higher levels of thermogenic marker transcripts PRDM16 and UCP1. The MA-INTACT mouse enables a simple way to perform cell-type specific analysis of highly purified mature adipocyte nuclei from VAT and SAT and increases the statistical significance of data collected on adipocytes. Isolated VAT and SAT adipocyte nuclei expressed distinct patterns of transcripts encoding chromatin remodeling factors and proteins relevant to diabetes, cardiovascular disease, and thermogenesis. The MA-INTACT mouse is an useful model to test the impact of caloric intake, dietary nutrients, exercise, and pharmaceuticals on the epigenome-induced health risks of obesity. The online version of this article (doi:10.1186/s40608-016-0112-6) contains supplementary material, which is available to authorized users.