Department of Clinical Biochemistry and Pharmacology, Faculty of Health Sciences, Ben-Gurion University of the Negev, Beer-Sheva, 84103, Israel.
The National Institute of Biotechnology in the Negev, Ben-Gurion University of the Negev, Beer-Sheva, 84103, Israel.
Mol Metab. 2017 May 6;6(7):725-736. doi: 10.1016/j.molmet.2017.05.003. eCollection 2017 Jul.
Obesity variably disrupts human health, but molecular-based patients' health-risk stratification is limited. Adipose tissue (AT) stresses may link obesity with metabolic dysfunction, but how they signal in humans remains poorly-characterized. We hypothesized that a transcriptional AT stress-signaling cascade involving E2F1 and ASK1 (MAP3K5) molecularly defines high-risk obese subtype.
ASK1 expression in human AT biopsies was determined by real-time PCR analysis, and chromatin immunoprecipitation (ChIP) adopted to AT explants was used to evaluate the binding of E2F1 to the promoter. Dual luciferase assay was used to measure promoter activity in HEK293 cells. Effects of E2F1 knockout/knockdown in adipocytes was assessed utilizing mouse-embryonal-fibroblasts (MEF)-derived adipocyte-like cells from WT and E2F1 mice and by siRNA, respectively. ASK1 depletion in adipocytes was studied in MEF-derived adipocyte-like cells from WT and adipose tissue-specific ASK1 knockout mice (ASK1-ATKO).
Human visceral-AT mRNA (N = 436) was associated with parameters of obesity-related cardio-metabolic morbidity. Adjustment for expression attenuated the association of with fasting glucose, insulin resistance, circulating IL-6, and lipids (triglycerides, HDL-cholesterol), even after adjusting for BMI. Chromatin-immunoprecipitation in human-AT explants revealed BMI-associated increased occupancy of the promoter by E2F1 (r = 0.847, p < 0.01). In adipocytes, siRNA-mediated E2F1-knockdown, and MEF-derived adipocytes of -knockout mice, demonstrated decreased ASK1 expression and signaling to JNK. Mutation/truncation of an E2F1 binding site in h promoter decreased E2F1-induced promoter activity, whereas E2F1-mediated sensitization of promoter to further activation by TNFα was inhibited by JNK-inhibitor. Finally, MEF-derived adipocytes from adipocyte-specific -knockout mice exhibited lower leptin and higher adiponectin expression and secretion, and resistance to the effects of TNFα.
AT E2F1 -ASK1 molecularly defines a metabolically-detrimental obese sub-phenotype. Functionally, it may negatively affect AT endocrine function, linking AT stress to whole-body metabolic dysfunction.
肥胖会对人体健康造成不同程度的影响，但目前基于分子的患者健康风险分层方法还很有限。脂肪组织（AT）的压力可能与代谢功能障碍有关，但它们在人体中的信号传递方式仍知之甚少。我们假设，涉及 E2F1 和 ASK1（MAP3K5）的转录 AT 应激信号级联反应从分子水平定义了高危肥胖亚型。
通过实时 PCR 分析确定人 AT 活检中的 ASK1 表达，并采用 AT 外植体进行染色质免疫沉淀（ChIP），以评估 E2F1 与 启动子的结合。使用双荧光素酶测定法测量 HEK293 细胞中的 启动子活性。利用 WT 和 E2F1 小鼠的鼠胚胎成纤维细胞（MEF）衍生的脂肪样细胞分别通过 E2F1 敲除/敲低和 siRNA 评估脂肪细胞中 E2F1 敲除/敲低的作用。在 WT 和脂肪组织特异性 ASK1 敲除（ASK1-ATKO）小鼠的 MEF 衍生的脂肪样细胞中研究了脂肪细胞中的 ASK1 耗竭。
在 436 名患者中，人内脏 AT mRNA（N）与肥胖相关的心脏代谢疾病的参数相关。调整 表达后，即使在调整 BMI 后，空腹血糖、胰岛素抵抗、循环 IL-6 和脂质（甘油三酯、高密度脂蛋白胆固醇）与 之间的关联也减弱了。在人 AT 外植体中进行的染色质免疫沉淀显示，BMI 相关的 E2F1 与 启动子的结合增加（r = 0.847，p < 0.01）。在脂肪细胞中，siRNA 介导的 E2F1 敲低和 -/-小鼠的 MEF 衍生的脂肪细胞中，发现 ASK1 表达和信号转导至 JNK 减少。h 启动子中 E2F1 结合位点的突变/截断降低了 E2F1 诱导的 启动子活性，而 JNK 抑制剂抑制了 E2F1 介导的 TNFα 对 启动子的进一步激活的敏感性。最后，脂肪细胞特异性 -/-敲除小鼠的 MEF 衍生的脂肪细胞表现出较低的瘦素和较高的脂联素表达和分泌，并且对 TNFα 的作用具有抗性。
AT E2F1-ASK1 从分子水平定义了一种代谢有害的肥胖亚型。从功能上讲，它可能会对 AT 内分泌功能产生负面影响，将 AT 应激与全身代谢功能障碍联系起来。
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