Role of ACSL5 in Intestinal and Liver Triacylglycerol Metabolism

描述(申请人提供)：肥胖症的流行与有害的代谢并发症有关。对我们理解这些代谢并发症至关重要的是描述调节脂肪酸代谢命运的机制。当脂肪酸进入细胞时，它们在酰基-辅酶A合成酶(ACSL)的作用下转化为酰基-COA。新出现的数据表明，五种ACSL亚型中的每一种都将酰基-COA导向特定的代谢命运。在这项赠款申请中，我们建议研究ACSL5在调节肠道和肝脏三酰甘油代谢中的作用。ACSL5在肠道和肝脏中都有高表达，但我们对其在这些组织中的功能了解有限。为了解决ACSL5在细胞特异性和系统代谢中的作用，我们产生了两种不同的ACSL5基因敲除小鼠：一种是基因在所有组织中结构性中断(空)，另一种是条件性基因敲除(Acsl5loxP/loxP)，使我们能够扰乱ACSL5在特定组织中的表达。在这项应用的第一个特定目的中，我们建议用肠道特异性消融ACSL5来建立小鼠，以确定：1)肠道ACSL5在肠细胞脂肪酸运输中的作用，2)肠道的影响 ACSL5对全身代谢和信号的影响，以及肠道微生物区系的改变3)ACSL5肠道细胞消融调节能量代谢和体内脂肪积累的机制基础。为了进行这些研究，我们将我们的条件性ACSL5基因敲除小鼠系(Acsl5loxP/loxP)与由Villin启动子驱动的表达Cre的小鼠配对，以特异性地去除肠道中ACSL5的表达(Acsl5int-/-)。在第二个特定目标中，我们将确定ACSL5在调节肝脂和全身能量代谢以及胰岛素抵抗方面的作用。肥胖和高能量饮食导致肝脏ACSL5表达增加。我们假设ACSL5是由SREBP转录因子在肝脏中上调的，SREBP转录因子被认为是诱导肝脏脂肪变性的原因，因为它催化了脂肪生成的限速酶mtGPAT1所利用的酰基-COAs的形成。在这一目标中，我们将产生ACSL5在肝细胞中特异性表达被消融的小鼠(Acsl5L-/-)。我们将用高脂肪、高蔗糖饮食喂养我们的Acsl5L-/-小鼠，以确定肝脏特异性切除ACSL5如何调节肝脏脂肪代谢，包括TAG积累和胰岛素-葡萄糖动态平衡，并确定ACSL5的丢失是否会将脂肪酸直接转化为其他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ACSL5表达在介导这两种蛋白的造脂作用中的作用。最后，我们将确定ACSL5消融的能力 肥胖/肥胖小鼠的脂肪变性。我们期望这些研究将使我们对脂肪酸代谢的了解取得重大进展，从而为肥胖、肝脏脂肪变性和相关疾病的治疗提供新的治疗途径。

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.