Role of ACSL5 in Intestinal and Liver Triacylglycerol Metabolism

描述（由申请人提供）：肥胖症的流行与有害的代谢并发症有关。我们理解这些代谢并发症的关键是描绘调节脂肪酸代谢命运的机制。当脂肪酸进入细胞时，它们通过酰基辅酶A合成酶（ACSL）的作用转化为酰基辅酶A。新出现的数据表明，五个ACSL亚型中的每一个都将酰基辅酶A导向特定的代谢命运。在本研究中，我们拟研究ACSL 5在调节肠道和肝脏三酰甘油代谢中的作用。ACSL5在肠道和肝脏中高度表达，但我们对其在这些组织中的功能的了解有限。为了解决ACSL 5在细胞特异性和全身代谢中的作用，我们已经产生了两种不同的ACSL 5敲除小鼠系：一种是在所有组织中基因已被组成性破坏（无效），另一种是允许我们破坏特定组织中ACSL 5表达的条件性敲除（Acsl5loxP/loxP）。在本申请的第一个具体目的中，我们提出产生具有ACSL 5的精氨酸特异性消融的小鼠，以确定：1）肠ACSL 5对肠上皮细胞脂肪酸运输的作用，和2）肠ACSL 5对肠上皮细胞脂肪酸运输的影响。 ACSL 5对全身代谢和信号传导的影响，以及肠道微生物群的改变3）ACSL 5的肠上皮细胞消融调节能量代谢和体脂积累的机制基础。为了进行这些研究，我们将我们的条件性ACSL 5敲除小鼠系（Acsl5loxP/loxP）与表达由绒毛蛋白启动子驱动的Cre的小鼠交配，以特异性消除肠中的ACSL 5表达（Acsl5int-/-）。在第二个具体目标中，我们将确定ACSL 5在调节肝脏脂质和全身能量代谢以及胰岛素抵抗中的作用。肥胖和能量密集饮食导致肝脏中ACSL 5表达增加。我们假设ACSL 5在肝脏中通过SREBP转录因子上调，已知SREBP转录因子诱导肝脂肪变性，因为它催化酰基辅酶A的形成，酰基辅酶A被脂肪生成中的限速酶mtGPAT 1利用。为此，我们将产生ACSL 5表达在肝细胞内特异性消除的小鼠（Acsl5L-/-）。我们将用高脂肪、高蔗糖饮食喂养我们的Acsl5L-/-小鼠系，以确定ACSL 5的肝脏特异性消融如何调节肝脏脂质代谢，包括TAG积累和胰岛素-葡萄糖稳态，并确定ACSL 5的损失是否将脂肪酸引导至其他ACSL亚型和氧化。我们将在我们新开发的Acsl5L-/-小鼠系中过表达SREBP-1c和mtGPAT，以确定肝脏ACSL 5表达在介导这两种蛋白质的脂肪生成作用中的作用。最后，我们将确定ACSL 5消融缓解 ob/ob小鼠脂肪变性。我们预计这些研究将导致我们对脂肪酸代谢的理解取得重大进展，从而为治疗肥胖症，肝脂肪变性和相关疾病提供新的治疗途径。

DESCRIPTION (provided by applicant): The epidemic of obesity is associated with detrimental metabolic complications. Critical to our understanding of these metabolic complications is delineating the mechanisms that regulate the metabolic fate of fatty acids. When fatty acids enter cells they are converted to acyl-CoAs by the actions of acyl-CoA synthetases (ACSL). Emerging data suggests that each of the five ACSL isoforms directs acyl-CoAs to specific metabolic fates. In this grant application we propose to investigate the role of ACSL5 in regulating intestinal and liver triacylglycerol metabolism. ACSL5 is highly expressed in both the intestine and liver, yet our understanding of its functions in these tissues is limited. T address the role of ACSL5 in cell-specific and systemic metabolism we have generated two different lines of ACSL5 knockout mice: one in in which the gene has been constitutively disrupted in all tissues (null) and a second conditional knockout (Acsl5loxP/loxP) that allows us to disrupt ACSL5 expression in specific tissues. In the first specific aim of this application, we propose to generate mice with an intestine-specific ablation of ACSL5 in order to determine: 1) the role of intestinal ACSL5 on enterocyte fatty acid trafficking and, 2) the effects of intestinal ACSL5 on whole body metabolism and signaling, and alterations in gut microbiota 3) the mechanistic basis by which enterocyte ablation of ACSL5 modulates energy metabolism and body fat accumulation. To perform these studies, we will mate our line of conditional ACSL5 knockout mice (Acsl5loxP/loxP) to mice expressing Cre driven by a villin promoter to specifically ablate ACSL5 expression in the intestine (Acsl5int-/-). In a second specific aim, we will determine the role of ACSL5 in regulating hepatic lipid and whole-body energy metabolism and insulin resistance. Obesity and energy dense diets result in increased ACSL5 expression in livers. We hypothesize ACSL5, is upregulated in the liver by SREBP transcription factors, known to induce hepatic steatosis because it catalyzes the formation of acyl-CoAs that are utilized by mtGPAT1 the rate limiting enzyme in lipogenesis. In this aim we will generate mice in which ACSL5 expression is ablated specifically within hepatocytes (Acsl5L-/-). We will feed our line of Acsl5L-/- mice a high fat, high sucrose diet to determine how liver specific ablation of ACSL5 regulates hepatic lipid metabolism, including TAG accumulation and insulin-glucose homeostasis, and determine whether loss of ACSL5 direct fatty acids to other ACSL isoforms and oxidation. We will overexpress SREBP-1c and mtGPAT in our newly developed line of Acsl5L-/- mice to determine the role of hepatic ACSL5 expression in mediating the lipogenic actions of these two proteins. Lastly, we will determine the ability of ACSL5 ablation to alleviate steatosis in ob/ob mice. We expect these studies will result in major advancements into our understanding of fatty acid metabolism and, thereby, provide novel therapeutic avenues for the treatment of obesity, hepatic steatosis and related diseases.