A link between hyperbilirubinemia, oxidative stress and injury to neocortical synaptosomes
A link between hyperbilirubinemia, oxidative stress and injury to neocortical synaptosomes
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DOI：
10.1016/j.brainres.2004.07.063
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发表时间：
2004-11-05
期刊：
影响因子：
2.9
通讯作者：
Butterfield, DA
中科院分区：
文献类型：
--
作者：
Brito, MA;Brites, D;Butterfield, DA
非偶联胆红素的细胞毒性包括膜结构紊乱、兴奋毒性和细胞死亡。据报道，这些事件引发自由基的产生升高和钙稳态的损害，并导致细胞膜完整性的丧失。因此，本研究旨在探讨临床相关浓度的游离非偶联胆红素与突触体膜囊泡的相互作用是否与氧化应激、胞质钙积累和膜功能紊乱有关。从沙鼠皮质脑组织中制备突触体囊泡，并与纯化胆红素（小于或等于1mum）在37℃下孵育4小时。用二氯荧光素和BAPTA荧光探针分别测定细胞内活性氧（ROS）和钙的浓度。通过免疫细胞化学和膜联蛋白V结合的磷脂酰丝氨酸暴露评估膜蛋白和脂质氧化。还测定了还原性和氧化性谷胱甘肽（GSH和GSSG）的水平，以及Mg2+- atp酶氨基磷脂转位酶（翻转酶）和Na+，K+- atp酶的活性。我们的研究结果表明，胆红素诱导氧化应激，由于脂质上升(大于或等于10%，P
Cytotoxicity by unconjugated bilirubin involves disturbances of membrane structure, excitotoxicity and cell death. These events were reported to trigger elevated free radicals production and impairment of calcium homeostasis, and to result in loss of cell membrane integrity. Therefore, this study was designed to investigate whether interaction of clinically relevant concentrations of free unconjugated bilirubin with synaptosomal membrane vesicles could be linked to oxidative stress, cytosolic calcium accumulation and perturbation of membrane function. Synaptosomal vesicles were prepared from gerbil cortical brain tissue and incubated with purified bilirubin (less than or equal to1 muM), for 4 h at 37 degreesC. Intracellular concentrations of reactive oxygen species (ROS) and calcium were determined by dichlorofluorescin and BAPTA fluorescent probes, respectively. Membrane protein and lipid oxidation were evaluated by immunocytochemistry and phosphatidylserine exposure by annexin V binding. Levels of reduced and oxidized glutathione (GSH and GSSG, respectively), as well as activities of Mg2+-ATPase aminophospholipid translocase (flippase) and Na+,K+-ATPase, were also measured. Our results showed that bilirubin induced oxidative stress, due to a rise in lipid (greater than or equal to10%, P