LIPID-DIABET

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。2型糖尿病（T2DM）、肥胖、血脂异常、高血压和大血管疾病都与胰岛素作用受损（胰岛素抵抗）有关。骨骼肌是体内最大的胰岛素反应器官，因此已被确定为胰岛素刺激葡萄糖摄取和处置的主要部位。研究证实，它是胰岛素抵抗的主要位点。脂质过剩导致肥胖、血浆游离脂肪酸、甘油三酯升高，甚至过量的膳食脂肪摄入都与胰岛素抵抗有关。有趣的是，从公共健康的角度来看，脂肪的质量和数量会影响胰岛素抵抗的程度：饱和脂肪、单不饱和脂肪、多不饱和脂肪（尤其是omega - 3脂肪酸油）。事实上，在横断面研究中，肌膜磷脂的饱和脂肪酸酰基链的较大程度与胰岛素抵抗的程度显著相关。动物和人类的研究都证实，血浆游离脂肪酸形式的脂质过剩可急性导致胰岛素抵抗状态，与肥胖或T2DM3观察到的状态没有区别，这表明脂质底物过剩是诱导胰岛素抵抗的原因。甘油三酯可能储存在肌肉中，已知该储存库与血浆室提供的FFA底物处于稳态平衡状态。在大多数情况下，肌肉细胞优先氧化脂质，并使用细胞内池来满足其基础氧化需求的60%。最近有人提出，如果细胞内甘油三酯池增加，也可能在胰岛素抵抗的发生或持续中起病因学作用。许多动物和人体活检研究证实了这种关联，但这种关联相对较弱且不一致。PI认为，迄今为止的结果已经被所使用的活检技术的问题所混淆：即使是1%的脂肪污染也可以预测导致甘油三酯分析的100-200%的误差。为了解决这个问题，他最近验证了使用磁共振质子光谱测量骨骼肌细胞内甘油三酯。高分辨率体积定位质子光谱使Stein博士和他的同事能够在不受脂肪污染的情况下，对这一重要的代谢变量进行定量。Stein博士已经证明了细胞内甘油三酯储存与胰岛素抵抗程度之间存在很强的反比关系。令人印象深刻的是，在多变量回归分析后，这种关系仍然很强，并且比体脂百分比、BMI、年龄和区域肥胖等经典变量更能预测胰岛素敏感性。目前还不清楚过量的脂质是如何引起胰岛素抵抗的。经典地，FFA的增加及其氧化的影响导致NADH/NAD和乙酰CoA/CoA比率的增加，这反过来导致糖酵解途径的反馈，葡萄糖6-磷酸的增加，从而抑制葡萄糖磷酸化，限制葡萄糖进入细胞（Randle效应）。对人类受试者的研究往往与这一教条相矛盾，表明FFA最初可能直接抑制葡萄糖转运，导致葡萄糖6-磷酸水平下降，随后产生直接抑制糖原合成的特异性作用。因此，在通过输注脂质乳剂提高血浆FFA的人体研究中，葡萄糖氧化抑制迅速发生（60分钟），随后葡萄糖转运/磷酸化缺陷在约2小时出现，糖原合成酶在约3-4小时进一步抑制。有研究表明，游离脂肪酸通过脂质衍生介质如长链酰基辅酶a、二酰基甘油（DAG）或磷脂的局部积累而对胰岛素作用产生有害影响。另一个原因可能是葡萄糖通量转向己糖胺途径，这也可能具有独立的受体后效应来对抗胰岛素的作用。这些可能包括胰岛素应答基因转录的改变，抑制糖原合成酶等酶活性，以及改变质膜激活/葡萄糖转运蛋白（Glut 4）囊泡运输。在稳态条件下，细胞内甘油三酯池被认为与这些脂质衍生的部分处于平衡状态。在动态环境下，这个池与胰岛素敏感性的关系尚不清楚。目前尚不清楚在急性高脂血症的情况下，细胞内甘油三酯池是否与胰岛素抵抗的发生有关。另一个相关的问题是，是否所有类型的脂肪在诱导胰岛素抵抗的能力上都是一样的。人体脂质输注研究使用了一种富含不饱和脂肪酸（18:2和18:3）的大豆油乳剂，其平衡大部分为18:1（油酸）。到目前为止，还没有研究检验各种脂肪酸对诱导胰岛素抵抗的时间过程的急性影响，以及胰岛素反应途径的特定方面是否受到不同的影响。事实上，持续数周的中期饮食研究无法证明在啮齿类动物的高脂肪喂养模型中观察到的胰岛素抵抗的相同诱导。那么哪个更重要呢？是细胞内储存池中甘油三酯的总量，还是饱和与不饱和甘油三酯的相对数量？通过比较质子光谱或13C光谱的峰高，无需侵入性活检即可计算脂肪酸的饱和/不饱和程度。以前利用核磁共振光谱研究脂质乳诱导的人体葡萄糖摄取和储存缺陷的时间过程的研究已经在2.1 - 4.7T进行。本研究将在GCRC MR核心实验室使用4.0T全身磁铁进行，该磁铁对监测糖原具有高灵敏度。目前还没有高场（3.0T）下的细胞内脂质代谢研究，Stein博士的初步研究都是在1.5T下进行的。这不仅可以提高脂质定量的灵敏度，而且增加的光谱分辨率有望允许特定的脂肪种类定量，即细胞内甘油三酯饱和程度。1.具体目标与不饱和脂肪相比，饱和膳食脂肪中血浆游离脂肪酸的增加是否会更快、更大程度地降低全身和肌肉葡萄糖的处置率。2. 脂肪诱导的葡萄糖处理抑制的时间过程是否与总和特定饱和长链细胞内甘油三酯的积累平行或先于。3. 与不饱和脂肪相比，饱和膳食脂肪中血浆游离脂肪酸的增加是否会改变总脂肪酸氧化率

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Type 2 diabetes mellitus (T2DM), obesity, dyslipidemia, hypertension and macrovascular disease have all been linked to impaired insulin action (insulin resistance). Skeletal muscle is the largest insulin responsive organ in the body and as such has been has been identified as the primary site of insulin stimulated glucose uptake and disposal. Studies have confirmed that it is the major locus of insulin resistance. Excess availability of lipid giving rise to obesity, to elevated plasma FFA, triglycerides1 and even excess dietary fat intake have been correlated with insulin resistance. Interestingly, from the public health perspective, the quality as well as the quantity of fat influences the degree of insulin resistance: saturated fats monounsaturates polyunsaturates (especially omega 3 fatty acid oils). In fact, a greater degree of saturated fatty acid acyl chains of muscle membrane phospholipids correlates significantly with the degree of insulin resistance in cross sectional studies. Studies both in animals and humans have confirmed that excess availability of lipid in the form of plasma FFA can acutely bring on a state of insulin resistance indistinguishable from that observed with obesity or T2DM3, suggesting that excess availability of lipid substrate is causal in the induction of insulin resistance. Triglycerides may be stored in muscle, and it is known that this pool is in homeostatic equilibrium with the supply of FFA substrate from the plasma compartment. Muscle cells preferentially oxidize lipid under most conditions and use the intracellular pool for up to 60% of their basal oxidative requirements. It has recently been proposed that the intramyocellular pool of triglycerides, if increased, could also play an etiologic role in the onset or perpetuation of insulin resistance. A number of animal and human biopsy studies have confirmed such an association, however the association has been relatively weak and inconsistent. The PI believes that the results to date have been confounded by problems with the biopsy techniques used: contamination by as little as 1% fat can be predicted to cause a 100-200% error in the triglyceride analysis. In order to resolve this issue, he has recently validated the use of magnetic resonance proton spectroscopy for the measurement of intracellular triglycerides in skeletal muscle. High resolution volume localized proton spectroscopy has allowed Dr. Stein and colleagues to specifically quantify this important metabolic variable in isolation from contaminating adipose fat. Dr. Stein has demonstrated a very strong inverse relationship between intramyocellular triglyceride stores and the degree of insulin resistance. Impressively, after multivariate regression analysis, the relationship remained strong and a better predictor of insulin sensitivity than classical variables such as percent body fat, BMI, age, and regional adiposity. It is not yet known how excess lipid causes insulin resistance. Classically, the effect of an increase in FFA and its oxidation results in an increase in the NADH/NAD and acetyl CoA/CoA ratio which in turn causes a feedback along the glycolytic pathway, an increase in glucose 6-phosphate such that glucose phosphorylation is inhibited limiting glucose entry into the cell (the Randle effect). Studies in human subjects have tended to contradict this dogma, suggesting that FFA may directly inhibit glucose transport initially resulting in decreased glucose 6- phosphate levels, followed by specific effects to inhibit glycogen synthesis directly. Thus in human studies where plasma FFA have been raised with infusion of a lipid emulsion, inhibition in glucose oxidation occurred rapidly (by 60 minutes) and subsequent defects on glucose transport/phosphorylation appeared at ~2 hours and further inhibition of glycogen synthase at ~3-4 hours. It has been suggested that FFA cause their deleterious effects on insulin action via local accumulation of lipid derived mediators such as long chain acyl-CoA's, diacyl glycerol (DAG) or phospholipids. Another cause may be diversion of glucose flux into the hexosamine pathway which may also have independent post receptors effects to antagonize insulin action. These may include alteration in transcription of insulin responsive genes, inhibiting enzyme activity such as glycogen synthase, and altering plasma membrane activation/Glucose transporter (Glut 4) vesicle trafficking. The intramyocellular pool of triglycerides is thought to be in equilibrium with these lipid derived moieties under steady state conditions. How this pool is related to insulin sensitivity in a dynamic setting is unknown. At present, it is currently unknown whether under conditions of acutely imposed hyperlipacidemia, whether the intramyocellular pool of triglyceride tracks with the onset of insulin resistance. Another and related question is whether all types are fat are equal in their ability to induce insulin resistance. The lipid infusion studies in humans used a soybean oil emulsion which is highly enriched in unsaturated fatty acids (18:2 and 18:3), with the balance being mostly 18:1 (oleic) acid. No studies to date have examined the acute effects of various fatty acids on the time course of inducing insulin resistance and whether specific aspects of the insulin response pathway are affected differentially. In fact medium term dietary studies of several weeks duration have been unable to demonstrate the same induction of insulin resistance as seen in high fat feeding models in rodents. Thus what is more important? Is it the total amount of triglyceride within the intramyocellular storage pool, or is it the relative amounts of saturated vs unsaturated triglycerides? By comparing peak heights, either from the proton spectrum or from the 13C spectrum, the degree of saturation/unsaturation of afatty acids can be calculated without invasive biopsy Previous studies utilizing NMR spectroscopy to study the time course of lipid emulsion induced defects in glucose uptake and storage in human subjects have been performed at 2.1 - 4.7T. The present study will be performed at the GCRC MR Core Laboratory using a 4.0T whole body magnet which has high sensitivity for monitoring glycogen. No studies have been performed on intramyocellular lipid metabolism at high field (3.0T), and Dr. Stein's preliminary studies were all performed at 1.5T. This will not only allow increased sensitivity for quantification of lipids, but increased spectral resolution promises to allow specific quantification of fatty species, i.e. the degree of intracellular triglyceride saturation. SPECIFIC AIMS 1. Whether increases in plasma FFA from saturated dietary fats will decrease whole body and muscle glucose disposal rates faster and to a greater degree compared to unsaturated fat. 2. Whether the time course of fat induced inhibition of glucose disposal is paralleled or preceded by the accumulation of total and specifically saturated long chain intramyocellular triglycerides. 3. Whether increases in plasma FFA from saturated dietary fats will alter total fatty acid oxidation rates compared to unsaturated fat