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Activation of phosphatidylcholine signalling during oxidative stress in synaptic endings.

机译:突触末端氧化应激过程中磷脂酰胆碱信号的激活。

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The purpose of the present study was to investigate the involvement of phosphatidylcholine (PC) signalling in synaptic endings incubated under oxidative stress conditions. Synaptosomes purified from adult rats (4 months old) cerebral cortex were exposed to oxidative insult (FeSO(4), 50microM) or vehicle, and diacylglycerol (DAG) generation and free fatty acid (FFA) release were subsequently evaluated using exogenous [(14)C]PC as substrate. DAG formation increased after 5, 30, and 60min of Fe(2+)-exposure with respect to the control conditions. The contribution of PC-specific phospholipase C (PC-PLC) and phospholipase D (PLD) pathways to DAG generation was evaluated using ethanol in the enzyme assays. Phosphatidylethanol (PEth) production was measured as a marker of PLD activity. In the presence of ethanol (2%) iron significantly stimulated DAG and PEth production at all times assayed. FFA release from PC, however, was inhibited after 5 and 60min of iron exposure. Similar results were observed in aged animals (28 months old) when compared with adult animals. DAG generation from PC was also evaluated in the presence of the tyrosine kinase inhibitors genistein and herbimycin A. Inhibition of tyrosine kinase activity did not modify the stimulatory effect exerted by iron on PC-PLC and PLD activities. Moreover, the presence of LY294002 (a specific PI3K inhibitor) did not alter DAG production. Our results demonstrate that oxidative stress induced by free iron stimulates the generation of the lipid messenger DAG from PC in synaptic endings in adult and aged rats.
机译:本研究的目的是研究磷脂酰胆碱(PC)信号在氧化应激条件下孵育的突触末端中的参与。从成年大鼠(4个月大)大脑皮层纯化的突触小体暴露于氧化性损伤(FeSO(4),50microM)或媒介物,随后使用外源性评估二酰基甘油(DAG)的产生和游离脂肪酸(FFA)的释放[[14 )C] PC作为底物。相对于对照条件,Fe(2+)暴露5、30和60分钟后,DAG的形成增加。在酶测定中使用乙醇评估了PC特异性磷脂酶C(PC-PLC)和磷脂酶D(PLD)途径对DAG产生的贡献。测量磷脂酰乙醇(PEth)的产生作为PLD活性的标志。在存在乙醇(2%)的情况下,铁在任何时候均能显着刺激DAG和PEth的产生。但是,铁暴露5分钟和60分钟后,从PC中释放的FFA受到抑制。与成年动物相比,在老年动物(28个月大)中观察到了相似的结果。在酪氨酸激酶抑制剂染料木黄酮和除草素A存在下,还评估了PC产生的DAG。抑制酪氨酸激酶活性不会改变铁对PC-PLC和PLD活性的刺激作用。此外,LY294002(一种特定的PI3K抑制剂)的存在不会改变DAG的产生。我们的结果表明,游离铁诱导的氧化应激刺激成年和成年大鼠突触末端PC中脂质信使DAG的生成。

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