Role of phospholipase A2 in the cytotoxic effects of oxalate in cultured renal epithelial cells
Role of phospholipase A2 in the cytotoxic effects of oxalate in cultured renal epithelial cells
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DOI：
10.1046/j.1523-1755.1999.00683.x
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发表时间：
1999-10-01
影响因子：
19.6
通讯作者：
Honeyman, TW
中科院分区：
文献类型：
--
作者：
Kohjimoto, Y;Kennington, L;Honeyman, TW
背景。草酸盐是肾结石的常见成分，对肾上皮细胞具有细胞毒性。尽管草酸盐诱导的细胞死亡的确切机制尚不清楚，但对包括肾上皮细胞在内的各种细胞类型的研究表明磷脂酶 A(2) (PLA(2)) 是细胞损伤的重要介质。因此，这些研究检验了 PLA(2) 在草酸盐细胞毒性作用中的作用。方法。测量预标记 Madin-Darby 犬肾 (MDCK) 细胞释放的 [H-3]-花生四烯酸 (AA) 或 [H-3]油酸 (OA)，作为 PLA(2) 活性的指标。通过排除乙锭同型二聚体-1来评估细胞活力。结果。草酸盐暴露（不含175至550μM）增加了MDCK细胞中[H-3]-AA的释放，但对[H-3]-OA的释放没有影响。草酸盐诱导的 [H-3]-AA 释放被胞质 PLA(2) (cPLA(2)) 的选择性抑制剂花生四烯基三氟甲基酮 (AACOCF(3)) 消除，但不受分泌性 PLA(2) 和钙非依赖性 PLA(2) 的选择性抑制剂的影响。 [H-3]-AA 的释放可以在暴露于草酸盐后 15 分钟内得到证实，这比观察到的细胞活力变化要早得多。此外，AACOCF(3) 显着降低 MDCK 细胞中的草酸盐毒性。结论。草酸通过涉及 cPLA(2) 的过程增加 MDCK 细胞的 AA 释放。此外，根据使用该异构体的选择性抑制剂获得的证据，该酶的活性似乎至少部分地导致了草酸盐的细胞毒性作用。草酸盐可以触发已知的脂质信号通路的发现可能为肾结石发病机制的初始事件提供新的见解。
Background. Oxalate, a common constituent of kidney stones, is cytotoxic for renal epithelial cells. Although the exact mechanism of oxalate-induced cell death remains unclear, studies in various cell types, including renal epithelial cells, have implicated phospholipase A(2) (PLA(2)) as a prominent mediator of cellular injury. Thus, these studies examined the role of PLA(2) in the cytotoxic effects of oxalate.Methods. The release of [H-3]-arachidonic acid (AA) or [H-3]oleic acid (OA) from prelabeled Madin-Darby canine kidney (MDCK) cells was measured as an index for PLA(2) activity. The cell viability was assessed by the exclusion of ethidium homodimer-l.Results. Oxalate exposure (175 to 550 mu M free) increased the release of [H-3]-AA in MDCK cells but had no effect on the release of [H-3]-OA. Oxalate-induced [H-3]-AA release was abolished by arachidonyl trifluoromethyl ketone (AACOCF(3)), a selective inhibitor of cytosolic PLA(2) (cPLA(2)), but was not affected by selective inhibitors of secretory PLA(2) and calcium-independent PLA(2). The [H-3]-AA release could be demonstrated within 15 minutes after exposure to oxalate, which is considerably earlier than the observed changes in cell viability. Furthermore, AACOCF(3) significantly reduced oxalate toxicity in MDCK cells.Conclusions. Oxalate increases AA release from MDCK cells by a process involving cPLA(2). In addition, based on the evidence obtained using a selective inhibitor of this isoform, it would appear that the activity of this enzyme is responsible, at least in part, for the cytotoxic effects of oxalate. The finding that oxalate can trigger a known lipid-signaling pathway may provide new insight into the initial events in the pathogenesis of nephrolithiasis.