Sphingolipid-mediated skeletal muscle pathology in response to free fatty acids.

描述（由申请人提供）： 由肥胖引起的血浆游离脂肪酸（FFA）升高是代谢综合征的标志，并导致2型糖尿病。数据表明，升高的血浆FFA有助于与2型糖尿病相关的骨骼肌病理，包括炎症、萎缩和肌病。棕榈酸（PAL），最丰富的血浆FFA，作为鞘脂合成的底物，数据表明，血浆FFA的升高，如发生在肥胖症和糖尿病，增加肌肉鞘脂。由于鞘脂包含一组不同的生物活性脂质介质，并影响细胞程序，包括自噬，凋亡，炎症和增殖，我们假设PAL介导的骨骼肌毒性部分通过鞘脂合成。此外，我们建议，不同的鞘脂的生物学功能，可以确定使用特定的药理学抑制途径引起这些代谢产物。为了验证这一假设，我们将1）确定PAL对骨骼肌模型系统中鞘脂的影响，并确定是否可以使用药理学试剂减弱特定生物活性鞘脂（包括神经酰胺和1-磷酸鞘氨醇）。然后，我们将2）确定PAL及其鞘脂代谢产物神经酰胺和1-磷酸鞘氨醇诱导的基因表达变化，然后鉴定这些代谢产物激活的信号通路。最后，我们建议3）使用饮食诱导的肥胖小鼠模型，该模型显示血浆脂肪酸的时间依赖性升高，以确定这些动物骨骼肌中的鞘脂含量，并确定它们是否反映了细胞培养模型中的鞘脂谱、鞘脂靶点的激活以及炎症标志物和细胞因子的调节。然后，我们将在给予鞘脂合成抑制剂myriocin的小鼠以及鞘氨醇激酶缺陷小鼠（SK 1-/-靶向基因缺失小鼠）中应用相同的策略。这些研究将为骨骼肌中鞘脂功能提供新的见解，并将允许评估在血浆FFA升高的情况下治疗靶向鞘脂生物合成以保持骨骼肌健康的可行性。血浆FFA升高会导致肌病、肌肉萎缩、肌肉炎症和2型糖尿病。退伍军人患2型糖尿病的比率高于普通人群。因此，这些研究与退伍军人管理局的使命高度相关。 公共卫生相关性： 近几十年来，肥胖和代谢性疾病（包括代谢综合征、2型糖尿病、动脉粥样硬化、心血管疾病）的发病率急剧上升。这些健康问题困扰着大部分人口，2型糖尿病在退伍军人中的发病率高于美国普通人群。代谢性疾病的一个关键特征是血浆游离脂肪酸（FFA）的增加。这种过度供应增加了脂肪酸沉积到外周组织，包括骨骼肌。正常情况下，骨骼肌是血浆葡萄糖摄取和利用的主要功能，因此，该组织的健康在糖尿病中起着重要作用。此外，在表现出血浆FFA升高的个体中，观察到许多肌肉病理，包括纤维化、消耗、炎症和肌病。该提案阐述了FFA介导骨骼肌中这些病理过程的机制。具体来说，我们假设血浆FFA的升高导致鞘脂的不适当合成，鞘脂是参与许多细胞和生物体过程的关键脂质介质。我们建议使用细胞培养和动物模型来确定特定鞘脂信号分子在异常基因调控和随后的骨骼肌病理学（包括炎症和肌肉萎缩）中的作用。 该提案直接解决了退伍军人的主要健康问题，即2型糖尿病。在一般人群中，糖尿病的发病率约为6%（http：//www.va.gov/pressrel/diabtsfs.htm）。然而，正如在退伍军人事务部题为“糖尿病护理进展”的情况说明书中所发现的那样，“糖尿病对退伍军人事务部（VA）特别重要，因为VA患者的患病率-六分之一，或16%-远远高于一般人群。. . VA是为糖尿病患者提供护理的最大综合医疗保健系统。“因此，几乎没有什么争论说糖尿病及其后遗症是VA的主要关注点。此外，这种疾病与老龄化有关，随着人口平均年龄的增加，DMII将成为一个更大的问题。我们的建议旨在确定肥胖、代谢综合征和糖尿病中骨骼肌健康损失的机制。我们的具体目标的完成将提供新的见解，这些条件降低骨骼肌健康和功能的机制。

DESCRIPTION (provided by applicant): Elevated plasma free fatty acids (FFAs) resulting from obesity is a hallmark of the metabolic syndrome and contributes to type 2 diabetes. Data indicate that elevated plasma FFAs contribute to skeletal muscle pathologies associated with type 2 diabetes including inflammation, atrophy, and myopathy. Palmitate (PAL), the most abundant plasma FFA, serves as substrate for sphingolipid synthesis, and data indicate that elevation of plasma FFA as occurs in obesity and diabetes, increases muscle sphingolipids. Since sphingolipids comprise a diverse group of bioactive lipid mediators and impact cell programs including autophagy, apoptosis, inflammation, and proliferation, we hypothesize that PAL-mediated toxicity in skeletal muscle results in part via sphingolipid synthesis. Moreover, we propose that biological functions of distinct sphingolipids can be determined using specific pharmacological inhibition of pathways giving rise to these metabolites. To test the hypothesis we will 1) determine the impact of PAL on sphingolipids in a skeletal muscle model system and determine whether specific bioactive sphingolipids including ceramides and sphingosine-1-phosphate can be attenuated using pharmacological agents. We will then 2) determine gene expression changes induced by PAL and its sphingolipid metabolites, ceramide and sphingosine-1-phosphate, followed by identification of signaling pathways activated by these metabolites. Finally we propose 3) to use the diet-induced obesity mouse model, which demonstrates a time-dependent elevation of plasma fatty acids, to determine sphingolipid content in skeletal muscle of these animals, and determine whether they mirror the cell culture model in terms of sphingolipid profiles, activation of sphingolipid targets, and regulation of inflammatory markers and cytokines. We will then apply the same strategy in mice administered with the sphingolipid synthesis inhibitor myriocin, as well as in mice deficient for sphingosine kinase (SK1 -/- targeted gene deletion mice). These studies will provide novel insights into sphingolipid functions in skeletal muscle and will allow assessment of the feasibility of therapeutically targeting sphingolipid biosynthesis to preserve skeletal muscle health in the presence of elevated plasma FFA. Elevated plasma FFA contributes to myopathies, muscle wasting, muscle inflammation, and type 2 diabetes. Veterans suffer from type 2 diabetes at a higher rate than the general population. Therefore these studies are highly relevant to the VA mission. PUBLIC HEALTH RELEVANCE: Project Narrative Recent decades have witnessed a dramatic increase in obesity and metabolic disease (including metabolic syndrome, type 2 diabetes, atherosclerosis, cardiovascular disease). These health problems plague large sections of the population, and type 2 diabetes occurs at a higher rate in veterans than in the general United States population. A key feature of metabolic disease is the increase in plasma free fatty acids (FFA). This oversupply increases fatty acid deposition into peripheral tissues, including skeletal muscle. Normally, skeletal muscle serves a major function of plasma glucose uptake and utilization, and therefore, the health of this tissue plays a major role in diabetes. Moreover, in individuals who exhibit elevated plasma FFA, many muscle pathologies are observed, including fibrosis, wasting, inflammation, and myopathy. This proposal addresses the mechanism(s) by which FFA mediate these pathological processes in skeletal muscle. Specifically, we hypothesize that elevation of plasma FFA causes inappropriate synthesis of sphingolipids, key lipid mediators involved in numerous cell and organism processes. We propose to use cell culture and animal models to determine roles for specific sphingolipid signaling molecules in aberrant gene regulation and subsequent skeletal muscle pathology including inflammation and muscle wasting. This proposal directly addresses a primary health concern for veterans, namely, type 2 diabetes. In the general population, the incidence of diabetes is around 6%, (http://www.va.gov/pressrel/diabtsfs.htm). However, as found on the Department of Veterans Affairs Fact Sheet entitled "Advances in Diabetes Care", "Diabetes has particular importance for the Department of Veterans Affairs (VA) because the prevalence among VA patients -- one in six, or 16 percent -- is substantially higher than in the general population. . . VA is the largest integrated health care system to provide care to persons with diabetes." Therefore, there is little argument that diabetes and its sequelae represent a major concern for the VA. Furthermore, this disease is associated with aging, and as the average age of the population is increasing, DMII will become an even greater concern. Our proposal seeks to identify mechanisms of loss of skeletal muscle health in obesity, the metabolic syndrome, and diabetes. Completion of our specific aims will provide novel insights into the mechanisms by which these conditions decrease skeletal muscle health and function.