Cohen Daniel, Papillon Joan, Aoudjit Lamine, Li Hongping, Cybulsky Andrey V, Takano Tomoko
Department of Medicine, McGill University, Montreal, Quebec, Canada.
Am J Physiol Renal Physiol. 2008 Mar;294(3):F469-79. doi: 10.1152/ajprenal.00372.2007. Epub 2008 Jan 2.
In experimental membranous nephropathy, complement C5b-9-induced glomerular epithelial cell (GEC) injury leads to morphological changes in GEC and proteinuria, in association with phospholipase A(2) (PLA(2)) activation. The present study addresses the role of calcium-independent PLA(2) (iPLA(2)) in GEC injury. iPLA(2)beta short and iPLA(2)gamma were expressed in cultured rat GEC and normal rat glomeruli. To determine whether iPLA(2) is involved in complement-mediated arachidonic acid (AA) release, GEC were stably transfected with iPLA(2)gamma or iPLA(2)beta cDNAs (GEC-iPLA(2)gamma; GEC-iPLA(2)beta). Compared with control cells (GEC-Neo), GEC-iPLA(2)gamma and GEC-iPLA(2)beta demonstrated greater expression of iPLA(2) proteins and activities. Complement-mediated release of [(3)H]AA was augmented significantly in GEC-iPLA(2)gamma compared with GEC-Neo, and the augmented [(3)H]AA release was inhibited by the iPLA(2)-directed inhibitor bromoenol lactone (BEL). For comparison, overexpression of iPLA(2)gamma also amplified [(3)H]AA release after incubation of GEC with H(2)O(2), or chemical anoxia followed by reexposure to glucose (in vitro ischemia-reperfusion injury). In parallel with release of [(3)H]AA, complement-mediated production of prostaglandin E(2) was amplified in GEC-iPLA(2)gamma. Complement-mediated cytotoxicity was attenuated significantly in GEC-iPLA(2)gamma compared with GEC-Neo, and the cytoprotective effect of iPLA(2)gamma was reversed by BEL, and in part by indomethacin. Overexpression of iPLA(2)beta did not amplify complement-dependent [(3)H]AA release, but nonetheless attenuated complement-mediated cytotoxicity. Thus iPLA(2)gamma may be involved in complement-mediated release of AA. Expression of iPLA(2)gamma or iPLA(2)beta induces cytoprotection against complement-dependent GEC injury. Modulation of iPLA(2) activity may prove to be a novel approach to reducing GEC injury.
在实验性膜性肾病中，补体C5b - 9诱导的肾小球上皮细胞（GEC）损伤会导致GEC形态改变和蛋白尿，这与磷脂酶A2（PLA2）激活有关。本研究探讨了钙非依赖性PLA2（iPLA2）在GEC损伤中的作用。iPLA2β短链和iPLA2γ在培养的大鼠GEC和正常大鼠肾小球中表达。为了确定iPLA2是否参与补体介导的花生四烯酸（AA）释放，用iPLA2γ或iPLA2β cDNA稳定转染GEC（GEC - iPLA2γ；GEC - iPLA2β）。与对照细胞（GEC - Neo）相比，GEC - iPLA2γ和GEC - iPLA2β表现出更高的iPLA2蛋白表达和活性。与GEC - Neo相比，GEC - iPLA2γ中补体介导的[³H]AA释放显著增加，并且增加的[³H]AA释放被iPLA2定向抑制剂溴苯醇内酯（BEL）抑制。为作比较，iPLA2γ的过表达在GEC与H₂O₂孵育后，或化学性缺氧后再暴露于葡萄糖（体外缺血 - 再灌注损伤）后也放大了[³H]AA释放。与[³H]AA释放平行，GEC - iPLA2γ中补体介导的前列腺素E₂产生增加。与GEC - Neo相比，GEC - iPLA2γ中补体介导的细胞毒性显著减弱，iPLA2γ的细胞保护作用被BEL逆转，部分被吲哚美辛逆转。iPLA2β的过表达未放大补体依赖性[³H]AA释放，但仍减弱了补体介导的细胞毒性。因此，iPLA2γ可能参与补体介导的AA释放。iPLA2γ或iPLA2β的表达诱导对补体依赖性GEC损伤的细胞保护作用。调节iPLA2活性可能被证明是减少GEC损伤的一种新方法。
