利用稳定同位素和液相色谱-质谱联用技术，将人血小板作为监测代谢生物标志物的平台。
Human platelets as a platform to monitor metabolic biomarkers using stable isotopes and LC-MS.
作者信息
Basu Sankha S, Deutsch Eric C, Schmaier Alec A, Lynch David R, Blair Ian A
机构信息
Department of Pharmacology, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, PA 19104, USA.
出版信息
Bioanalysis. 2013 Dec;5(24):3009-21. doi: 10.4155/bio.13.269.
BACKGROUND
Intracellular metabolites such as CoA thioesters are modulated in a number of clinical settings. Their accurate measurement from surrogate tissues such as platelets may provide additional information to current serum and urinary biomarkers.
METHODS
Freshly isolated platelets from healthy volunteers were treated with rotenone, propionate or isotopically labeled metabolic tracers. Using a recently developed LC-MS-based methodology, absolute changes in short-chain acyl-CoA thioesters were monitored, as well as relative metabolic labeling using isotopomer distribution analysis.
RESULTS
Consistent with in vitro experiments, isolated platelets treated with rotenone showed decreased intracellular succinyl-CoA and increased β-hydroxybutyryl-CoA, while propionate treatment resulted in increased propionyl-CoA. In addition, isotopomers of the CoAs were readily detected in platelets treated with the [(13)C]- or [(13)C(15)N]-labeled metabolic precursors.
CONCLUSION
Here, we show that human platelets can provide a powerful ex vivo challenge platform with potential clinical diagnostic and biomarker discovery applications.
背景
细胞内代谢物如辅酶A硫酯在多种临床环境中受到调节。从血小板等替代组织中准确测量它们可能为当前的血清和尿液生物标志物提供额外信息。
方法
从健康志愿者新鲜分离的血小板用鱼藤酮、丙酸盐或同位素标记的代谢示踪剂处理。使用最近开发的基于液相色谱-质谱的方法，监测短链酰基辅酶A硫酯的绝对变化，以及使用同位素异构体分布分析的相对代谢标记。
结果
与体外实验一致，用鱼藤酮处理的分离血小板显示细胞内琥珀酰辅酶A减少，β-羟基丁酰辅酶A增加，而丙酸盐处理导致丙酰辅酶A增加。此外，在用[(13)C]-或[(13)C(15)N]-标记的代谢前体处理的血小板中很容易检测到辅酶A的同位素异构体。
结论
在此，我们表明人类血小板可以提供一个强大的体外挑战平台，具有潜在的临床诊断和生物标志物发现应用。
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