Adipocyte to neuron signaling in thermogenic programming of white adipose tissue

摘要 我们实验室的长期目标是了解和利用脂肪组织 对全身葡萄糖耐量和胰岛素敏感性进行强有力的控制。少量 小鼠棕色(BAT)或人类“米色”脂肪细胞移植到受体小鼠可改善血糖 动态平衡，强调了解脂肪褐变机制的重要性。基于 了解新生脂肪生成(DNL)途径的中间产物显示出有效的信号 功能(例如，乙酰辅酶A作为组蛋白乙酰化的底物，转录调节)和脂肪细胞 DNL受肥胖、禁食、寒冷暴露和运动的高度调节，我们假设脂肪细胞DNL是 新陈代谢的主要调节节点。我们的目标是通过干扰DNL途径来询问这一概念 通过选择性KO的DNL酶，三磷酸腺苷柠檬酸裂解酶(ACLY)和脂肪酸合成酶(FASN)。我们的 初步数据显示，FASN KO可上调脂肪细胞，从而支持这一方法 神经营养因子神经调节蛋白4(NRG4)促进SWAT交感神经元(SNS)扩张， 即使在热中性(30摄氏度)下也是如此。因此，DNL代谢物(乙酰辅酶A、丙二酰辅酶A)或DNL产物 (Palmitoyl CoA)似乎与控制脂肪SNS活性、脂肪能量消耗密切相关 和全身葡萄糖动态平衡。基于这些数据，该项目试图确定蜂窝 以及脂肪细胞向定位的SNS神经元发出信号并促进 SWAT中米色脂肪细胞的发育。目标1将确定iAdFASNKO中的米色脂肪细胞 小鼠是通过直接将成熟的白色脂肪细胞转化为米色脂肪细胞或通过旁分泌信号来获得的 诱导祖细胞向米色脂肪细胞分化。为了解决潜在的机制，目标2将 确定NRG4(和也可能上调的Negr1)是否介导FASN耗竭的影响 脂肪细胞在体内引起SNS的扩张。这一目标是基于令人兴奋的初步数据显示 这种脂肪细胞的条件培养液可以显著促进PC-12神经元的突起生长。 体外，NRG4沉默可抑制这种作用。我们开发的新技术将被用来删除脂肪细胞 NRG4和Negr1在植入受体小鼠前使用基于CRISPR的纳米颗粒和分析 它们对SNS神经支配的影响。最后，Aim 3测试脂肪细胞DNL中间代谢物乙酰 CoA/Malonyl CoA在iAdFASNKO小鼠中启动信号，导致NRG4表达和SNS扩张。这些 IAdFASNKO小鼠的脂肪细胞代谢物，以及它们对细胞蛋白的乙酰化和丙二酸化，将是 在双KO小鼠中，产生乙酰辅酶A的三磷酸腺苷柠檬酸裂解酶(ACLY)的KO逆转了这种作用。识别 调节脂肪细胞功能的DNL中间体将有助于确定其潜在的机制。 总之，这些实验具有很高的潜力来定义由DNL驱动的新信号通路 调节脂肪褐变的代谢物和2型糖尿病的新治疗策略。

Abstract The overarching long term goal of our laboratory is to understand and exploit how adipose tissues exert powerful control over whole body glucose tolerance and insulin sensitivity. Small amounts of mouse brown (BAT) or human “Beige” adipocytes transplanted into recipient mice can improve glucose homeostasis, highlighting the importance of understanding mechanisms of adipose browning. Based on the knowledge that intermediates of the de novo lipogenesis (DNL) pathway display potent signaling functions (e.g., Acetyl CoA as substrate for histone acetylation, transcriptional regulation) and that adipocyte DNL is highly regulated by obesity, fasting, cold exposure, and exercise, we hypothesize that adipocyte DNL is a major regulatory node in metabolism. We aim to interrogate this concept by perturbing this DNL pathway through selective KO of DNL enzymes ATP citrate lyase (ACLY) and fatty acid synthase (FASN). Our preliminary data encourage this approach by revealing that FASN KO upregulates adipocyte neurotrophic factor Neuregulin 4 (Nrg4) and enhances expansion of sWAT sympathetic neurons (SNS), even at thermo-neutrality (30C). Thus, DNL metabolites (Acetyl CoA, Malonyl CoA) or DNL product (Palmitoyl CoA) appear to be intimately linked to controlling adipose SNS activity, adipose energy expenditure and whole body glucose homeostasis. Based on these data, this project seeks to determine the cellular and molecular mechanisms whereby adipocytes can signal to localized SNS neurons and promote the development of Beige adipocytes in sWAT. Aim 1 will determine whether Beige adipocytes in iAdFASNKO mice are derived by direct “conversion” of mature white to beige adipocytes OR by paracrine signaling to induce differentiation of progenitor cells to Beige adipocytes. To address underlying mechanisms, Aim 2 will determine whether Nrg4 (and Negr1, which may also be upregulated) mediates the effect of FASN-depleted adipocytes to cause expansion of the SNS in vivo. This Aim is based on exciting preliminary data showing that conditioned media from such adipocytes cause marked neurite outgrowth in PC-12 neurons in vitro, which is inhibited by Nrg4 silencing. New technology we developed will be used to delete adipocyte Nrg4 and Negr1 using CRISPR-based nanoparticles prior to implantation into recipient mice and analysis of their effects on SNS innervation. Finally, Aim 3 tests whether adipocyte DNL intermediate metabolites Acetyl CoA/Malonyl CoA in iAdFASNKO mice initiate signaling to cause Nrg4 expression and SNS expansion. These adipocyte metabolites in iAdFASNKO mice, and their acetylation and malonylation of cellular proteins, will be reversed by KO of ATP citrate Lyase (ACLY), which generates the Acetyl CoA, in double KO mice. Identifying the DNL intermediates that modulate adipocyte function will enable defining their underlying mechanisms. Together, these experiments have high potential to define novel signaling pathways driven by DNL metabolites that regulate adipose browning and new therapeutic strategies for type 2 diabetes.