Lysophosphatidic acid (LPA) is a potent signaling lipid, and state-dependent alterations in LPA make it a promising diagnostic marker for various diseases. However, plasma LPA concentrations vary widely among reports, even under normal conditions. These variations can be attributed, at least in part, to the artificial metabolism of LPA after blood collection, thus complicating the use of plasma LPA as a clinical biomarker. Previous studies focused on suppressing LPA production by the LPA-producing enzyme autotaxin (ATX) but did not take the artificial LPA degradation into account. Here, we aimed to develop an optimized plasma preparation method that reflects the concentration of LPA in the circulating blood by finding conditions to suppress both the production and degradation of LPA after blood collection. The main features of the devised method were suppression of LPA production and degradation after blood collection by keeping whole blood samples at low temperature and followed by adding an ATX inhibitor to plasma samples. Using this devised method, the LPA level did not change for 30 minutes after blood collection, and mouse plasma LPA concentrations showed minimal variation across individual animals, as determined by LC-MS/MS. Additionally, human and mouse LPA levels were found to be much lower than those previously reported, ranging from 40 to 50 nM. Finally, the increased accuracy made it possible to detect circadian rhythms in the levels of certain LPA species in mouse plasma. These results demonstrate the usefulness of the devised plasma preparation method to determine accurate plasma LPA concentrations.

中文翻译：

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抑制采集后溶血磷脂酸 (LPA) 代谢可提高血浆 LPA 定量的精度。


溶血磷脂酸 (LPA) 是一种有效的信号脂质，LPA 的状态依赖性改变使其成为多种疾病的有前景的诊断标记物。然而，即使在正常条件下，各报告的血浆 LPA 浓度差异也很大。这些变化至少部分归因于血液采集后 LPA 的人工代谢，从而使血浆 LPA 作为临床生物标志物的使用变得复杂。之前的研究主要集中在通过产生 LPA 的酶自分泌运动因子 (ATX) 抑制 LPA 的产生，但没有考虑人工 LPA 降解。在这里，我们的目标是通过寻找在采血后抑制 LPA 产生和降解的条件，开发一种反映循环血液中 LPA 浓度的优化血浆制备方法。该方法的主要特点是通过将全血样本保持在低温下，然后在血浆样本中添加 ATX 抑制剂来抑制采血后 LPA 的产生和降解。使用这种设计的方法，LPA 水平在采血后 30 分钟内没有变化，并且根据 LC-MS/MS 测定，小鼠血浆 LPA 浓度在个体动物之间显示出最小的变化。此外，人类和小鼠的 LPA 水平比之前报道的要低得多，范围为 40 至 50 nM。最后，准确性的提高使得检测小鼠血浆中某些 LPA 物质水平的昼夜节律成为可能。这些结果证明了所设计的血浆制备方法对于确定准确的血浆 LPA 浓度的有用性。