Manson Anne, Winter Tanja, Aukema Harold M
Food and Human Nutritional Sciences, University of Manitoba, Winnipeg MB, Canada; Canadian Centre for Agri-Food Research in Health and Medicine (CCARM), Winnipeg MB, Canada.
Food and Human Nutritional Sciences, University of Manitoba, Winnipeg MB, Canada; Canadian Centre for Agri-Food Research in Health and Medicine (CCARM), Winnipeg MB, Canada.
Prostaglandins Leukot Essent Fatty Acids. 2023 Apr;191:102555. doi: 10.1016/j.plefa.2023.102555. Epub 2023 Feb 28.
Phospholipase A (PLA) enzymes cleave cell membrane phospholipids and release polyunsaturated fatty acids (PUFA), which can be converted into oxylipins. However, little is known about PLA preference for PUFA, and even less is known about how this further impacts oxylipin formation. Therefore, we investigated the role of different PLA groups in PUFA release and oxylipin formation in rat hearts. Sprague-Dawley rat heart homogenates were incubated without or with varespladib (VAR), methyl arachidonyl fluorophosphonate (MAFP) or EDTA. Free PUFA and oxylipins were determined by HPLC-MS/MS, and isoform expressions by RT-qPCR. Inhibition of sPLA IIA and/or V by VAR reduced the release of ARA and DHA, but only DHA oxylipins were inhibited. MAFP reduced the release of ARA, DHA, ALA, and EPA, and the formation of ARA, LA, DGLA, DHA, ALA, and EPA oxylipins. Interestingly, cyclooxygenase and 12-lipoxygenase oxylipins were not inhibited. mRNA expression levels of sPLA and iPLA isoforms were highest whereas levels of cPLA were low, consistent with activity. In conclusion, sPLA enzymes lead to the formation of DHA oxylipins, while iPLA is likely responsible for the formation of most other oxylipins in healthy rat hearts. Oxylipin formation cannot be implied from PUFA release, thus, both should be evaluated in PLA activity studies.
磷脂酶A（PLA）可裂解细胞膜磷脂并释放多不饱和脂肪酸（PUFA），后者可转化为氧化脂质。然而，关于PLA对PUFA的偏好知之甚少，而关于这如何进一步影响氧化脂质的形成则了解更少。因此，我们研究了不同PLA组在大鼠心脏中PUFA释放和氧化脂质形成中的作用。将斯普拉格-道利大鼠心脏匀浆在无或有伐瑞普拉德（VAR）、甲基花生四烯酰氟磷酸酯（MAFP）或乙二胺四乙酸（EDTA）的情况下孵育。通过高效液相色谱-串联质谱法测定游离PUFA和氧化脂质，并通过逆转录定量聚合酶链反应测定同工型表达。VAR对分泌型磷脂酶A IIA和/或V的抑制降低了花生四烯酸（ARA）和二十二碳六烯酸（DHA）的释放，但仅DHA氧化脂质受到抑制。MAFP降低了ARA、DHA、α-亚麻酸（ALA）和二十碳五烯酸（EPA）的释放，以及ARA、亚油酸（LA）、二高γ-亚麻酸（DGLA）、DHA、ALA和EPA氧化脂质的形成。有趣的是，环氧化酶和12-脂氧合酶氧化脂质未受抑制。分泌型磷脂酶A和内膜磷脂酶同工型的mRNA表达水平最高，而胞质磷脂酶的水平较低，这与活性一致。总之，分泌型磷脂酶导致DHA氧化脂质的形成，而内膜磷脂酶可能是健康大鼠心脏中大多数其他氧化脂质形成的原因。氧化脂质的形成不能从PUFA释放推断得出，因此，在PLA活性研究中应同时评估两者。
