Metabolic Determinants of Cardiac Bioactive Sphingolipids in Lipid Overload

描述（由申请人提供）：肥胖和代谢综合征是两种重叠的疾病，其中身体开始在非脂肪细胞（包括心脏）中储存多余的脂肪。这种异位脂质储存导致胰岛素抵抗和脂毒性，部分原因是这些脂质可以作为一类称为鞘脂的信号脂质合成的底物。最近，研究表明，使用不同种类的脂肪制成的鞘脂启动不同的细胞信号程序。因此，重要的是要了解哪些类型的脂肪优先用于鞘脂的合成，以及鞘脂水平和生产是如何调节不同的膳食脂肪。 神经酰胺是一种关键的鞘脂信号分子，与糖尿病心肌病和胰岛素抵抗的发生有关。神经酰胺含有两条酰基链，它们通过丝氨酸棕榈酰转移酶（SPT）和（二氢）神经酰胺合酶（CerS）依次添加。异构体特异性的底物利用模式允许合成结合不同酰基链长度的鞘脂，其可以具有不同的作用。拟议研究的目的是通过调节SPT组成和CerS表达和底物供应，确定脂质过载对组织特异性鞘脂谱的影响。 要做到这一点，我们将首先分析在体外替代底物利用的小鼠心脏SPT复合物，我们将分析亚基组成的酶在心脏的质谱。接下来，我们将量化CerS异构体对由这些替代底物制成的鞘脂的活性。之后，我们将确定外源脂肪酸如何管理SPTLC亚基和所有六个CerS亚型的表达。最后，我们将测量鞘脂谱，以确定膳食脂肪对鞘脂水平的影响。这一系列研究将揭示过量膳食脂肪组成与心脏鞘脂水平的关系，并将确定具有潜在生物相关性的脂质种类。 拟议的研究将支持一个强大的研究培训计划，包括课程，在科学会议上的演示，以及与其他科学家的结构化和非正式互动。课程将涉及特定的生物化学和生物医学主题以及统计方法，并将补充以前的拨款写作，科学道德以及动物和人类主题的利用。此外，学生将参加并参加期刊俱乐部和部门和计划赞助的研讨会系列。最后，学生将在东南地区脂质会议和国际查尔斯顿神经酰胺会议等会议上发表演讲或海报，并提交作品在同行评审期刊上发表。

DESCRIPTION (provided by applicant): Obesity and the metabolic syndrome are two overlapping conditions in which the body begins to store excess fat in non-adipose cells, including the heart. This ectopic lipid storage leads to insulin resistance and lipotoxicity, in part because these lipids can serve as substrate for synthesis of a class of signaling lipids known as sphingolipids. Recently, it has been shown that sphingolipids made using different kinds of fat initiate distinct cellular signaling programmes. It is thus important to understand what kinds of fat are preferentially used for sphingolipid synthesis, as well as how sphingolipid levels and production are regulated by different dietary fats. Ceramide, a key sphingolipid signaling molecule, has been implicated in the development of diabetic cardiomyopathy and insulin resistance. Ceramide contains two acyl chains, which are added sequentially by the enzymes serine palmitoyltransferase (SPT) and (dihydro)ceramide synthase (CerS). Isoform-specific patterns of substrate utilization allow synthesis of sphingolipids incorporating different acyl chain lengths, which can have distinct roles. The purpose of the proposed study is to determine the effect of lipid overload on tissue-specific sphingolipid profiles, via the regulation of SPT composition and CerS expression and substrate supply. To do this, we will first analyze in vitro alternative substrate utilization by the murine cardiac SPT complex, and we will analyze the subunit makeup of the enzyme in heart by mass spectrometry. Next, we will quantify the activity of the CerS isoforms toward sphingolipids made from these alternative substrates. Afterward, we will determine how exogenous fatty acids govern the expression of the SPTLC subunits and all six CerS isoforms. Finally, we will measure sphingolipid profiles to ascertain the impact of dietary fat on sphingolipid levels. This series of studies will reveal the relationship of excess dietary fat composition to sphingolipid levels in the heart and will identify lipid species with potential biological relevance. The proposed studies will support a robust research training plan incorporating coursework, presentations at scientific meetings, and both structured and informal interactions with other scientists. Classes will address specific biochemical and biomedical topics as well as statistical methods, and they will complement previous instruction on grant writing, scientific ethics, and utilization of animal and human subjects. Furthermore, the student will attend and participate in a journal club and both departmental and program-sponsored seminar series. Finally, the student will present talks or posters at meetings such as the Southeastern Regional Lipid Conference and the International Charleston Ceramide Conference and submit work for publication in peer-reviewed journals.