Desaturation Of Essential Fatty Acids Using Stable Isotope GC/MS

使用稳定同位素 GC/MS 进行必需脂肪酸的去饱和

• 批准号: 7591931
• 负责人: Norman Salem
• 金额: $ 38.78万
• 依托单位: NATIONAL INSTITUTE ON ALCOHOL ABUSE AND ALCOHOLISM
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7591931
• 关键词: Adipose tissue; Adult; Animal Feed; Animals; Arachidonic Acids; Area; Back; Biochemical; Birth; Blood; Blood Circulation; Brain; Condition; Deposition; Development; Diet; Docosahexaenoic Acids; Dose; Drug or chemical Tissue Distribution; Essential Fatty Acids; Extrahepatic; Fatty Acids; Female; Goals; Growth; Heart; High Density Lipoproteins; Human; Infant; Invasive; Kidney; Label; Life; Linoleic Acids; Lipoproteins; Liver; Low-Density Lipoproteins; Lung; Mammals; Metabolic; Metabolism; Methodology; Milk; Molecular; Mus; Myocardium; Nervous system structure; Oils; Oleic Acid; Oleic Acids; Omega-3 Fatty Acids; Oral; Organ; Pattern; Phospholipids; Plants; Plasma; Positron-Emission Tomography; Rate; Rattus; Relative (related person); Research; Research Project Grants; Rodent; Role; Smoke; Smoker; Source; Spleen; Stream; Suggestion; Techniques; Testis; Thalamic structure; Time; Tissues; Tracer; Triglycerides; Very low density lipoprotein; alpha-Linolenic Acid; base; day; fatty acid metabolism; feeding; gray matter; healthy volunteer; heart imaging; in vivo; latent nuclear antigen; low density lipoprotein inhibitor; male; non-smoker; novel; nutrition; problem drinker; pup; radiotracer; research study; stable isotope; uptake; white matter

Prior to the recent application of stable isotope based GC/MS methodology, little was known about in vivo essential fatty acid metabolism in animals or humans. Essential fatty acid metabolism was studies in human adults, both male and female, and those who smoked as well as non-smokers. This was a stable isotope study of in vivo metabolism of deuterated-LA and deuterated-LNA conversion after a single oral dose of these precursors. Our results indicated that female smokers had a two-fold increase in the percent of plasma dose and a higher fractional conversion rate for 22:5n-3 conversion to 22:6n-3 compared with non-smokers. Male smokers had elevated total plasma n-3 fatty acids, a more rapid turn over of D5-18:3n-3, a disappaerance rate of D5-20:5n-3 that was both delayed and slower, and a greater percentage of D5-20:5n-3 was directed into 22:5n-3 relative to non-smokers. Generally, smoking increased the bioavailablity of n-3 fatty acids from plasma, accelerated fractional conversion rates, and increased the percent formation for some long chain n-3 fatty acids. In rats, it was observed that addition of preformed DHA to the diet leads to a decreased accumulation of label from 18-C precursors into DHA and DPAn6 in several organs even though there was a significant increase in tissue DHA. Female rats accumulated more DHA and DPAn6 but less AA than males when fed a controlled diet containing 3 wt% alpha-linolenic acid. An n-3 fatty acid deficient diet led to a marked decline in labeling of liver 22:4n6 and 22:5n6 from the 18:2n6 precursor. A closely related research project concerns the origins of nervous system and other organ DHA. Possible sources are from dietary preformed DHA, from metabolism of the precursor, LNA, or from body stores of DHA. A novel technique has been developed that allows for the quantitative assessment of the amount of DHA accreted from LNA metabolism under various dietary conditions. For this study, it is necessary to control the diet from near birth up to a period where significant brain development has occurred. This has been accomplished thru the use of newly developed artifiicial rearing techniques using an artificial rat milk that was nearly devoid of n-3 fatty acids. The n-3 fatty acids are then added as deuterated-LNA and containing varying levels of DHA. In one major experiment, rat pups were fed diets with 0 or 2% DHA between days 8-29 of life. During this period, it could be calculated that 40% of the newly formed brain DHA in the animals fed D5-LNA as their only source of n-3 fatty acids were derived from preformed DHA and not from LNA metabolism. This was surprising as there was no DHA in the diet; thus, all preformed DHA deposited in the brain must have been derived from other organs via the blood stream. When DHA was added to the diet, there was a pronounced decrease in the rate of LNA metabolism to DHA, possibly due to a form of end-product inhibition, and 88% of brain DHA was derived from the preformed dietary DHA. The biochemical mechanisms underlying these metabolic effects of dietary DHA are being investigated. A decline in labeled DHA was also observed in liver, heart, muscle, kidney and testes but no such changes were observed in adipose tissues. There was also a higher level of brain DHA in the rats given preformed DHA indicating that metabolism could not provide an adequate source of brain DHA. Another finding of consequence for infants fed formulas without DHA was that several organs including the heart, lungs, kidney and spleen had a net loss of DHA content during a period of intense body growth when no preformed DHA was present in the diet. An attempt was made to determine what the underlying mechanisms for DHA transport into brain and other organs. Lipoproteins were purified and labeled with radiotracers and modified with a tracer levels of phospholipids acylated with DHA, AA or oleic acid (OA). The modified lipoproteins were intravenously injected in mice. The plasma and tissue distribution of the radiotracers were investigated as a function of time and the lipoproteins composition. We found that higher proportion of DHA in LDL results in an enhanced uptake of these lipoproteins by brain and heart. A similar enrichment of LDL in AA or OA did not result in any changes compared to control unaltered LDL. Tissue uptake of HDL did not depend on its fatty acid composition. We next compared the distribution in plasma pools and tissue uptake of 14C-DHA and 3H-(OA) intravenously injected in mice. We found that DHA is rapidly taken up by liver, selectively acylated into triglycerides and released back into the circulation in VLDL. Most of the DHA from VLDL and LDL appeared to be rapidly taken up by extrahepatic organs. This pattern seems to be unique for DHA, because no significant amount of non-essential oleic acid, traced in a similar way, was found in TG and VLDL fractions. In summary, these results point to the important role of VLDL and LDL in transport of DHA to extrahepatic tissues, and to the involvement of liver in the initial selectivity for DHA transport. A novel application of PET imaging for the study of C11-DHA incorporation into brain has been initiated. Brain and heart images from 17 healthy volunteers and 11 alcoholics have now been obtained. Extensive characterization of the fatty acid input function in plasma has been made in real time for the 11-C-DHA. Our findings thusfar are that the J(in) and K* values for male and female healthy volunteers are similar except for the K* values in the thalamus and the gray matter/white matter ratio. There is a suggestion from initial studies that alcoholics may have a lower incorporation of DHA in many areas of cortex than control subjects, but more subjects will be needed.

在最近使用稳定基于同位素的GC/MS方法论之前，关于动物或人类中的体内必需脂肪酸代谢知之甚少。 必需的脂肪酸代谢是对男性和女性的成年人以及吸烟和非吸烟者的研究。 这是对这些前体单一口服剂量后的氘化la和氘化LNA转化率的体内代谢的稳定同位素研究。 我们的结果表明，与非吸烟者相比，女性吸烟者的血浆剂量百分比增加了两倍，而22：5N-3转化率的分数转化率更高，为22：6n-3。 雄性吸烟者的总血浆N-3脂肪酸升高，更快地转移了D5-18：3N-3，不赞成率为D5-20：5N-3，既延迟又慢，而D5-20：5N-3的比例更高，将其定向到22：5n-3中，相对于非熟练者而言。 通常，吸烟增加了血浆中N-3脂肪酸的生物可利用性，分数转化率加速，并增加了一些长链N-3脂肪酸的形成百分比。 在大鼠中，观察到，在饮食中添加预先形成的DHA会导致在几个器官中从18-C前体中累积到DHA和DPAN6的标记降低，即使组织DHA显着增加。 雌性大鼠积累了更多的DHA和DPAN6，但与男性相比，喂食含有3 wt％α-亚麻酸的受控饮食时的AA。 N-3脂肪酸缺乏饮食导致肝脏的标记22：4n6和22：5n6的标记下降显着下降。 密切相关的研究项目涉及神经系统和其他器官DHA的起源。可能的来源来自饮食中预先形成的DHA，来自前体，LNA的代谢或DHA的体内储存。已经开发了一种新的技术，可以在各种饮食条件下对从LNA代谢中积累的DHA量进行定量评估。对于这项研究，有必要控制饮食从近出生到发生重大大脑发育的时期。通过使用新开发的人工饲养技术，使用几乎没有N-3脂肪酸的人造大鼠牛奶来实现这一目标。然后添加N-3脂肪酸作为氘代LNA并含有不同水平的DHA。在一个主要实验中，在生命的第8-29天之间，大鼠幼崽的饮食为0或2％DHA。在此期间，可以计算出，在饲喂D5-LNA的动物中，有40％的新形成的脑DHA是其唯一的N-3脂肪酸来源，它来自预先形成的DHA，而不是LNA代谢。这是令人惊讶的，因为饮食中没有DHA。因此，沉积在大脑中的所有预先形成的DHA必须通过血流源自其他器官。当将DHA添加到饮食中时，LNA代谢对DHA的速率明显降低，这可能是由于最终产物抑制的形式，而88％的脑DHA源自预先形成的饮食DHA。正在研究这些代谢性DHA代谢作用的生化机制。 在肝，心脏，肌肉，肾脏和睾丸中也观察到标记的DHA的下降，但在脂肪组织中未观察到这种变化。 鉴于预先形成的DHA，大鼠的大脑DHA水平也更高，表明新陈代谢无法提供足够的脑DHA来源。 在饮食中没有预先形成的DHA时，对未经DHA的配方喂养的婴儿的后果的另一个发现是，包括心脏，肺，肾脏和脾脏在内的多个器官净损失了DHA含量。 试图确定DHA运输到大脑和其他器官的基本机制。将脂蛋白纯化并用放射性示例标记，并用示踪剂水平的磷脂，用DHA，AA或油酸（OA）酰化。将改性的脂蛋白静脉注射在小鼠中。研究了放射性示例的血浆和组织分布，这是时间和脂蛋白组成的函数。我们发现，LDL中DHA的比例较高会导致大脑和心脏对这些脂蛋白的摄取增强。 与对照未改变的LDL相比，AA或OA中LDL的类似富集不会导致任何变化。 HDL的组织摄取不取决于其脂肪酸组成。 接下来，我们比较了在小鼠中注射14C-DHA和3H-（OA）的血浆池和组织摄取的分布。我们发现DHA被肝脏迅速吸收，有选择地酰化为甘油三酸酯，并释放回VLDL的循环。来自VLDL和LDL的大多数DHA似乎被肝外器官迅速占据。对于DHA，这种模式似乎是独一无二的，因为在TG和VLDL级分中没有发现大量的非必需油酸。 总之，这些结果表明，VLDL和LDL在DHA向肝外组织的运输中的重要作用，以及肝脏参与DHA转运的初始选择性。 启动了PET成像在研究C11-DHA掺入大脑中的新型应用。 现已获得来自17位健康志愿者和11名酗酒者的大脑和心脏图像。 对于11-C-DHA，已经实时对等离子体中的脂肪酸输入功能进行了广泛的表征。我们的发现因此，男性和女性健康志愿者的J（IN）和K*值除外，丘脑中的K*值和灰质/白质的比例相似。最初研究的建议是，酗酒者在许多皮质领域的掺入可能要比对照受试者较低，但需要更多的受试者。