Objective

Obesity due to overnutrition causes adipose tissue dysfunction, which is a critical pathological step on the road to type 2 diabetes (T2D) and other metabolic disorders. In this study, we conducted an unbiased investigation into the fundamental molecular mechanisms by which adipocytes transition to an unhealthy state during obesity.

Methods

We used nuclear tagging and translating ribosome affinity purification (NuTRAP) reporter mice crossed with Adipoq-Cre mice to determine adipocyte-specific 1) transcriptional profiles (RNA-seq), 2) promoter and enhancer activity (H3K27ac ChIP-seq), 3) and PPARγ cistrome (ChIP-seq) profiles in mice fed chow or a high-fat diet (HFD) for 10 weeks. We also assessed the impact of the PPARγ agonist rosiglitazone (Rosi) on gene expression and cellular state of adipocytes from the HFD-fed mice. We integrated these data to determine the transcription factors underlying adipocyte responses to HFD and conducted functional studies using shRNA-mediated loss-of-function approaches in 3T3-L1 adipocytes.

Results

Adipocytes from the HFD-fed mice exhibited reduced expression of adipocyte markers and metabolic genes and enhanced expression of myofibroblast marker genes involved in cytoskeletal organization, accompanied by the formation of actin filament structures within the cell. PPARγ binding was globally reduced in adipocytes after HFD feeding, and Rosi restored the molecular and cellular phenotypes of adipocytes associated with HFD feeding. We identified the TGFβ1 effector protein SMAD to be enriched at HFD-induced promoters and enhancers and associated with myofibroblast signature genes. TGFβ1 treatment of mature 3T3-L1 adipocytes induced gene expression and cellular changes similar to those seen after HFD in vivo, and knockdown of Smad3 blunted the effects of TGFβ1.

Conclusions

Our data demonstrate that adipocytes fail to maintain cellular identity after HFD feeding, acquiring characteristics of a myofibroblast-like cell type through reduced PPARγ activity and elevated TGFβ-SMAD signaling. This cellular identity crisis may be a fundamental mechanism that drives functional decline of adipose tissues during obesity.

中文翻译：

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由于 PPARγ 活性降低和 TGFβ-SMAD 信号传导升高，脂肪细胞在肥胖期间无法维持细胞特性

客观的

营养过剩导致的肥胖会导致脂肪组织功能障碍，这是通往 2 型糖尿病 (T2D) 和其他代谢紊乱道路上的关键病理步骤。在这项研究中，我们对肥胖期间脂肪细胞转变为不健康状态的基本分子机制进行了公正的调查。

方法

我们使用核标记和翻译核糖体亲和纯化 (NuTRAP) 报告小鼠与 Adipoq-Cre 小鼠杂交来确定脂肪细胞特异性 1) 转录谱 (RNA-seq)，2) 启动子和增强子活性 (H3K27ac ChIP-seq)，3)喂食食物或高脂肪饮食 (HFD) 10 周的小鼠的PPAR γ cistrome (ChIP-seq) 谱。我们还评估了 PPAR γ激动剂罗格列酮 (Rosi) 对 HFD 喂养小鼠脂肪细胞基因表达和细胞状态的影响。我们整合了这些数据以确定脂肪细胞对 HFD 反应的转录因子，并在 3T3-L1 脂肪细胞中使用 shRNA 介导的功能丧失方法进行功能研究。

结果

来自 HFD 喂养的小鼠的脂肪细胞表现出脂肪细胞标志物和代谢基因的表达减少，而参与细胞骨架组织的肌成纤维细胞标志物基因的表达增加，伴随着细胞内肌动蛋白丝结构的形成。HFD 喂养后脂肪细胞中的PPAR γ结合全面降低，Rosi 恢复了与 HFD 喂养相关的脂肪细胞的分子和细胞表型。我们确定了TGF-β1效应蛋白SMAD在HFD诱导启动子和增强来丰富和肌成纤维细胞信号基因有关。成熟 3T3-L1 脂肪细胞的 TGFβ1 处理诱导的基因表达和细胞变化与体内HFD 后观察到的相似，并且 Smad3 的敲低减弱了 TGFβ1 的作用。

结论

我们的数据表明，脂肪细胞在 HFD 喂养后无法保持细胞特性，通过降低 PPAR γ活性和升高 TGFβ-SMAD 信号来获得肌成纤维细胞样细胞类型的特征。这种细胞身份危机可能是导致肥胖期间脂肪组织功能下降的基本机制。