Winkler Jeremy W, Libreros Stephania, De La Rosa Xavier, Sansbury Brian E, Norris Paul C, Chiang Nan, Fichtner David, Keyes Gregory S, Wourms Nicholas, Spite Matthew, Serhan Charles N
Center for Experimental Therapeutics and Reperfusion Injury, Department of Anesthesiology, Perioperative and Pain Medicine, Building for Transformative Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, MA, USA.
Cayman Chemical, Ann Arbor, MI, USA.
J Leukoc Biol. 2018 Jan 29. doi: 10.1002/JLB.3MI0617-254R.
Local production and downstream metabolism of specialized proresolving lipid mediators (SPMs) are pivotal in regulating their biological actions during resolution of inflammation. Resolvin D4 (RvD4: 4S,5R,17S-trihydroxydocosa-6E,8E,10Z,13Z,15E,19Z hexaenoic acid) is one of the more recently elucidated SPMs with complete stereochemistry biosynthesized from docosahexaenoic acid . Here, we report a new multimilligram commercial synthesis that afforded enough material for matching, validation, and further evaluation of RvD4 functions. Using LC-MS-MS profiling, RvD4 was identified at bioactive amounts in human (1 pg/mL) and mouse bone marrow (12 pg/femur and tibia). In mouse bone marrow, ischemia increased the formation of RvD4 > 37-fold (455 pg/femur and tibia). Two separate mouse ischemic injury models were used, where RvD4 reduced second organ reperfusion lung injury > 50%, demonstrating organ protection. Structure-function relationships of RvD4 demonstrated > 40% increase in neutrophil and monocyte phagocytic function in human whole blood in comparison with 2 separate trans-containing double bond isomers that were inactive. These 2 isomers were prepared by organic synthesis: 4S,5R,17S-trihydroxydocosa-6E,8E,10E,13Z,15E,19Z-hexaenoic acid (10-trans-RvD4), a natural isomer, and 4S,5R,17S-trihydroxydocosa-6E,8E,10E,13E,15E,19Z-hexaenoic acid (10,13-trans-RvD4), a rogue isomer. Compared to leukotriene B , D-series resolvins (RvD1, RvD2, RvD3, RvD4, or RvD5) did not stimulate human neutrophil chemotaxis monitored via real-time microfluidics chambers. A novel 17-oxo-containing-RvD4 product of eicosanoid oxidoreductase was identified with human bone marrow cells. Comparison of 17-oxo-RvD4 to RvD4 demonstrated that with human leukocytes 17-oxo-RvD4 was inactive. Together, these provide commercial-scale synthesis that permitted a second independent validation of RvD4 complete stereochemical structure as well as evidence for RvD4 regulation in tissues and its stereoselective phagocyte responses.
在炎症消退过程中，特殊促消退脂质介质（SPM）的局部产生和下游代谢对于调节其生物学作用至关重要。消退素D4（RvD4：4S，5R，17S-三羟基二十二碳-6E，8E，10Z，13Z，15E，19Z-六烯酸）是最近阐明的一种SPM，具有由二十二碳六烯酸生物合成的完整立体化学结构。在此，我们报告了一种新的多毫克级商业合成方法，该方法提供了足够的材料用于RvD4功能的匹配、验证和进一步评估。使用液相色谱-串联质谱分析，在人（1 pg/mL）和小鼠骨髓（12 pg/股骨和胫骨）中鉴定出具有生物活性量的RvD4。在小鼠骨髓中，缺血使RvD4的形成增加了37倍以上（455 pg/股骨和胫骨）。使用了两种不同的小鼠缺血性损伤模型，其中RvD4使第二器官再灌注肺损伤降低了50%以上，证明了器官保护作用。RvD4的结构-功能关系表明，与两种无活性的单独反式含双键异构体相比，人全血中的中性粒细胞和单核细胞吞噬功能增加了40%以上。这两种异构体通过有机合成制备：天然异构体4S，5R，17S-三羟基二十二碳-6E，8E，10E，13Z，15E，19Z-六烯酸（10-反式-RvD4）和有害异构体4S，5R，17S-三羟基二十二碳-6E,8E,10E,13E,15E,19Z-六烯酸（10,13-反式-RvD4）。与白三烯B相比,D系列消退素（RvD1、RvD2、RvD3、RvD4或RvD5）不会刺激通过实时微流控室监测的人中性粒细胞趋化性。在人骨髓细胞中鉴定出一种新型的类花生酸氧化还原酶含17-氧代的RvD4产物。将17-氧代-RvD4与RvD4进行比较表明,对于人白细胞，17-氧代-RvD4无活性。总之，这些提供了商业规模的合成方法，允许对RvD4的完整立体化学结构进行第二次独立验证，以及RvD4在组织中的调节及其立体选择性吞噬细胞反应的证据。
