Development and application of targeted lipidomics of oxygenated polyunsaturated fatty acids in different oxidative damage study models

Abstract

For the past one decade or so, there have been growing interest in the research of lipids. In addition to being the fuel source and essential component of the cell plasma membrane, lipids have been increasingly reported to play key roles in cellular activities. Lipidome profiling by mass spectrometry in a biological system has undeniably advanced lipid research. However, the lack of pure internal and external lipid standards has greatly compromised the accuracy and confidence of targeted lipdomic analysis. This thesis explores the use of in-house synthesised chemical standards of a few novel lipid mediators derived from docosahexaenoic acid (DHA), namely 4(RS)-4-F4t-NeuroP, 10-F4t-NeuroP and 4(RS)-ST-Δ5-8-NeuroF, and from and adrenic acid (AdA), namely 7(RS)-7-F2t-dihomo-IsoP, 17(RS)-17-F2t-dihomo-IsoP, 7(RS)-ST-Δ8-11-dihomo-IsoF and 17(RS)-10-epi-SC-Δ15-11-dihomo-IsoF. Together with other commercially available lipid molecules, they were characterised and identified in biological samples using a newly established method, which is high in selectivity, linearity, recovery, signal-to-noise ratio, and reproducibility. Subsequently, the abundance of these lipid mediators developed in vivo were profiled, and investigated for their significance in three settings. First study demonstrated that the exposure of perfluorooctanesulfonic acid (PFOS), an endocrine disruptor, to pregnant mice led to an exhaustive induction of oxidative stress in foetuses, particularly in liver and kidney tissues, but were far less severe in the dams, which were partially alleviated by the maternal catalase and superoxide dismutase antioxidant enzymes. In contrast, production of enzymatically derived lipid mediators was restricted in the maternal-foetal study. PFOS-exposed foetuses, therefore, have a far more debilitating effect than the dams, and posed potential damaging effects to their early stages of development. Second study demonstrated that supplementation of dietary fish oil to chronic low-grade inflamed rodents (induced by carbon tetrachloride; CCl4) did not result in the alleviation of liver scarring and lipid accumulations. Lipidomic analysis revealed that dietary supplementation of fish oil, whether in the presence of CCl4 or not, showed similar lipid profiles, implying that both fish oil and CCl4 treatments shared a common destination but possibly via different routes that is yet to be confirmed. Thirdly, while non-enzymatic mediator from DHA, 4(RS)-4-F4t-NeuroP has shown to be anti-arrhythmia, cardiac protective and inhibit breast cancer cells, its effects in the brain where it is theoretically more abundant than other parts of the body was evaluated. It was found that high concentration of peripheral 4(RS)-4-F4t- NeuroP could lead to its accumulation in organs like liver and brain, probably via the formation of 4(RS)-4-F4t-NeuroP-albumin complex. Hepatic and neural monohydroxy derivatives of DHA (HDoHEs) were consistently elevated after one day of 4(RS)-4- F4t-NeuroP exposure, which warrants further investigation on its gene regulatory pathways. Taken together, this thesis has exhibited the importance of targeted lipidomic analysis on non-enzymatically derived lipid mediators to shed further mechanistic insights into how they might take on the role as a bioactive messenger.