Tyurin V A, Balasubramanian K, Winnica D, Tyurina Y Y, Vikulina A S, He R R, Kapralov A A, Macphee C H, Kagan V E
1] Center for Free Radical and Antioxidant Health, University of Pittsburgh, Pittsburgh, PA, USA [2] Department of Environmental and Occupational Health, University of Pittsburgh, Pittsburgh, PA, USA.
1] Center for Free Radical and Antioxidant Health, University of Pittsburgh, Pittsburgh, PA, USA [2] Department of Environmental and Occupational Health, University of Pittsburgh, Pittsburgh, PA, USA [3] Department of Biophysics, MV Lomonosov Moscow State University, Moscow, Russia.
Cell Death Differ. 2014 May;21(5):825-35. doi: 10.1038/cdd.2014.1. Epub 2014 Jan 24.
Diversified anionic phospholipids, phosphatidylserines (PS), externalized to the surface of apoptotic cells are universal phagocytic signals. However, the role of major PS metabolites, such as peroxidized species of PS (PSox) and lyso-PS, in the clearance of apoptotic cells has not been rigorously evaluated. Here, we demonstrate that H2O2 was equally effective in inducing apoptosis and externalization of PS in naive HL60 cells and in cells enriched with oxidizable polyunsaturated species of PS (supplemented with linoleic acid (LA)). Despite this, the uptake of LA-supplemented cells by RAW264.7 and THP-1 macrophages was more than an order of magnitude more effective than that of naive cells. A similar stimulation of phagocytosis was observed with LA-enriched HL60 cells and Jurkat cells triggered to apoptosis with staurosporine. This was due to the presence of PSox on the surface of apoptotic LA-supplemented cells (but not of naive cells). This enhanced phagocytosis was dependent on activation of the intrinsic apoptotic pathway, as no stimulation of phagocytosis occurred in LA-enriched cells challenged with Fas antibody. Incubation of apoptotic cells with lipoprotein-associated phospholipase A2 (Lp-PLA2), a secreted enzyme with high specificity towards PSox, hydrolyzed peroxidized PS species in LA-supplemented cells resulting in the suppression of phagocytosis to the levels observed for naive cells. This suppression of phagocytosis by Lp-PLA2 was blocked by a selective inhibitor of Lp-PLA2, SB-435495. Screening of possible receptor candidates revealed the ability of several PS receptors and bridging proteins to recognize both PS and PSox, albeit with diverse selectivity. We conclude that PSox is an effective phagocytic 'eat-me' signal that participates in the engulfment of cells undergoing intrinsic apoptosis.
多样化的阴离子磷脂，即磷脂酰丝氨酸（PS），外化到凋亡细胞表面是普遍的吞噬信号。然而，主要的PS代谢产物，如PS的过氧化物种（PSox）和溶血磷脂酰丝氨酸，在凋亡细胞清除中的作用尚未得到严格评估。在此，我们证明H2O2在诱导幼稚HL60细胞以及富含可氧化多不饱和PS物种（补充亚油酸（LA））的细胞中凋亡和PS外化方面同样有效。尽管如此，RAW264.7和THP-1巨噬细胞对补充LA的细胞的摄取比幼稚细胞的摄取效率高出一个数量级以上。在用星形孢菌素触发凋亡的富含LA的HL60细胞和Jurkat细胞中观察到类似的吞噬刺激。这是由于凋亡的补充LA的细胞（而非幼稚细胞）表面存在PSox。这种增强的吞噬作用依赖于内源性凋亡途径的激活，因为在用Fas抗体攻击的富含LA的细胞中未发生吞噬刺激。用脂蛋白相关磷脂酶A2（Lp-PLA2）孵育凋亡细胞，Lp-PLA2是一种对PSox具有高特异性的分泌酶，可水解补充LA的细胞中的过氧化PS物种，导致吞噬作用抑制至幼稚细胞观察到的水平。Lp-PLA2对吞噬作用的这种抑制被Lp-PLA2的选择性抑制剂SB-435495阻断。对可能的受体候选物的筛选揭示了几种PS受体和桥接蛋白识别PS和PSox的能力，尽管具有不同的选择性。我们得出结论，PSox是一种有效的吞噬“吃我”信号，参与内源性凋亡细胞的吞噬。
