Mäkelä Kari A, Karhu Toni, Jurado Acosta Alicia, Vakkuri Olavi, Leppäluoto Juhani, Herzig Karl-Heinz
Research Unit of Biomedicine, Physiology, University of Oulu, Oulu, Finland.
Biocenter Oulu, University of Oulu, Oulu, Finland.
Front Endocrinol (Lausanne). 2018 Dec 5;9:710. doi: 10.3389/fendo.2018.00710. eCollection 2018.
Orexin-A (OXA) has been originally isolated from a precursor peptide prepro-orexin from the lateral hypothalamus. The orexin system has been attributed to important functions in sleep, arousal and regulation of energy homeostasis. In addition to its high levels in cerebrospinal fluid, OXA is present in blood. However, reported peptide concentrations in plasma vary significantly depending on the method used. Therefore, a specific and sensitive OXA radioimmunoassay (RIA) with solid phase extraction method was developed to determine whether plasma OXA concentrations is affected by acute feeding and/or wake and sleep in young healthy males. Blood samples were collected for 24 h from nine healthy males (aged 20-24 years; BMI 20.7-26.5) every 2 h starting at 11 a.m. Food was served at 12 p.m, 5:30 p.m, 8 p.m and 8 a.m and the sleep time was between 10 p.m and 7 a.m. Plasma samples were analyzed in addition for cortisol and melatonin levels. Blood pressure was monitored through the experimental period. OXA antibody was raised in rabbits. OXA antiserum had only minor cross-reactivity with prepro-orexin precursor (<0.001%), amino-terminal peptide (<0.001%), carboxy-terminal peptide (0.001%), and orexin-B (0.3%) with high sensitivity (0.15 pg/tube). Plasma OXA levels varied between 0.5 and 16 pg/ml in seven subjects and were undetectable (below 0.5 pg/ml) in two subjects. The OXA concentrations did not correlate to feeding nor wake/sleep, whereas cortisol, melatonin and mean arterial blood pressure presented a clear circadian rhythm in each subject. In conclusion, OXA is present in blood in low amounts and its levels do not follow autonomic nor neuroendocrine circadian rhythms. Thereby, studies examining regulatory mechanisms and influences of OXA from blood samples should interpret results very cautiously.
食欲素A（OXA）最初是从下丘脑外侧的前体肽前食欲素中分离出来的。食欲素系统在睡眠、觉醒和能量稳态调节中具有重要作用。除了在脑脊液中含量较高外，OXA也存在于血液中。然而，报道的血浆中该肽的浓度因所用方法的不同而有显著差异。因此，开发了一种采用固相萃取法的特异性和灵敏性OXA放射免疫分析法（RIA），以确定年轻健康男性的血浆OXA浓度是否受急性进食和/或清醒与睡眠的影响。从9名健康男性（年龄20 - 24岁；体重指数20.7 - 26.5）中，从上午11点开始，每2小时采集一次血样，共采集24小时。中午12点、下午5:30、晚上8点和早上8点提供食物，睡眠时间为晚上10点至早上7点。此外，还分析了血浆样本中的皮质醇和褪黑素水平。在整个实验期间监测血压。用兔子制备OXA抗体。OXA抗血清与前食欲素前体（<0.001%）、氨基末端肽（<0.001%）、羧基末端肽（0.001%）和食欲素B（0.3%）的交叉反应性很小，灵敏度高（0.15 pg/管）。7名受试者的血浆OXA水平在0.5至16 pg/ml之间，2名受试者检测不到（低于0.5 pg/ml）。OXA浓度与进食、清醒/睡眠均无相关性，而每个受试者的皮质醇、褪黑素和平均动脉血压呈现明显的昼夜节律。总之，OXA在血液中的含量较低，其水平不遵循自主神经或神经内分泌昼夜节律。因此，从血样中研究OXA的调节机制及其影响的研究在解释结果时应非常谨慎。
