Zhang Andrew Y, Yi Fan, Jin Si, Xia Min, Chen Qi-Zheng, Gulbins Erich, Li Pin-Lan
Department of Pharmacology and Toxicology, Medical College of Virginia Campus, Virginia Commonwealth University, Richmond, VA 23298, USA.
Antioxid Redox Signal. 2007 Jul;9(7):817-28. doi: 10.1089/ars.2007.1509.
This study examined the role of acid sphingomyelinase (ASM) and its redox amplification in mediating the formation of lipid raft (LR) redox signaling platforms in coronary arterial endothelial cells (CAECs). Using small interference RNA (siRNA) of ASM, Fas ligand (FasL)-induced increase in ASM activity, production of ceramide, and LR clustering in CAECs were blocked, and clustered Fas was also substantially reduced in detergent-resistant membrane fractions of CAECs. LR clustering, gp91(phox) aggregation, and p47(phox) translocation to the LR clusters induced by FasL were also blocked in ASM-siRNA transfected CAECs. Corresponding to this reduction of LR clustering with NAD(P)H oxidase subunits in ASM-siRNA transfected CAECs, superoxide (O(2)(-)) production was significantly decreased as measured by either ESR or fluorescent spectrometry. Interestingly, superoxide dismutase (SOD) not only scavenged (O(2)(-)), but also markedly attenuated LR clustering. Xanthine/xanthine oxidase, an exogenous (O(2)(-)) generating system, dramatically increased ASM activity and LR clustering in EC membrane and enhanced FasL-induced LR clustering, which were blocked by SOD. These results suggest that that ASM activates LR clustering to form redox signaling platforms, where (O(2)(-)) production enhances ASM activity, and thereby results in a forwarding amplification of LR and redox signaling. This ASM-mediated feedforwarding mechanism may be critical for an efficient transmembrane signaling through LRs.
本研究探讨了酸性鞘磷脂酶（ASM）及其氧化还原放大作用在介导冠状动脉内皮细胞（CAECs）中脂筏（LR）氧化还原信号平台形成中的作用。使用ASM的小干扰RNA（siRNA）可阻断Fas配体（FasL）诱导的CAECs中ASM活性增加、神经酰胺产生和LR聚集，并且CAECs耐去污剂膜组分中聚集的Fas也显著减少。在转染ASM-siRNA的CAECs中，FasL诱导的LR聚集、gp91(phox)聚集和p47(phox)向LR簇的转位也被阻断。与转染ASM-siRNA的CAECs中LR聚集以及NAD(P)H氧化酶亚基减少相对应，通过电子自旋共振（ESR）或荧光光谱法测定，超氧化物（O(2)(-)）的产生显著降低。有趣的是，超氧化物歧化酶（SOD）不仅清除了（O(2)(-)），还显著减弱了LR聚集。黄嘌呤/黄嘌呤氧化酶作为一种外源性（O(2)(-)）生成系统，可显著增加EC膜中的ASM活性和LR聚集，并增强FasL诱导的LR聚集，而这些均被SOD阻断。这些结果表明，ASM激活LR聚集以形成氧化还原信号平台，在该平台中（O(2)(-)）的产生增强了ASM活性，从而导致LR和氧化还原信号的正向放大。这种ASM介导的前馈机制可能对于通过LRs进行有效的跨膜信号传导至关重要。
