CALCIUM IONOPHORE INCREASES AMYLOID-BETA PEPTIDE PRODUCTION BY CULTURED-CELLS
CALCIUM IONOPHORE INCREASES AMYLOID-BETA PEPTIDE PRODUCTION BY CULTURED-CELLS
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DOI：
10.1021/bi00181a016
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发表时间：
1994-04-19
期刊：
影响因子：
2.9
通讯作者：
SELKOE, DJ
中科院分区：
文献类型：
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作者：
QUERFURTH, HW;SELKOE, DJ
淀粉样蛋白β肽（A β）在正常代谢过程中释放到培养物中的各种细胞的培养基中。家族性阿尔茨海默病中β淀粉样前体蛋白（β APP）A β区域内或侧翼的几个错义突变的发现为β APP加工改变在该疾病发病机制中的作用提供了强有力的证据。调节分泌途径（导致β APP在A β结构域内裂解）和替代蛋白水解途径（产生完整A β）相对利用的细胞机制尚不清楚。据推测，许多神经退行性疾病，包括阿尔茨海默病，其特征在于钙代谢异常。我们研究了钙稳态失调对β APP cDNA转染的人肾293细胞中A β产生的影响。在代谢标记和脉冲追踪标记范例中，将从细胞条件培养基中免疫沉淀的β与免疫沉淀的全长和分泌形式的β APP进行比较。钙离子载体A23187持续增加A β的产生约3倍。这种效应依赖于完整细胞中细胞外钙的存在。咖啡因还增加了A β的产生，这可能是通过从细胞内储存中释放钙来实现的。A β的增加是cAMP非依赖性的，并且它不是由蛋白激酶C依赖性途径介导的，因为用佛波酯治疗降低了A β水平。还确定了离子载体对β APP成熟和磷酸化的影响。我们得出结论，细胞内钙离子水平的升高对β APP成熟和蛋白水解加工有重要影响，并大大增强了淀粉样蛋白A β肽的产生和释放。
Amyloid beta peptide (A beta) is released into the media of a variety of cells in culture during normal metabolism. The discovery of several missense mutations within or flanking the A beta region of the beta amyloid precursor protein (beta APP) in familial Alzheimer's disease provides strong evidence for a role of altered processing of beta APP in the pathogenesis of this disorder. The cellular mechanisms that regulate the relative utilization of the secretory pathway, which causes beta APP to be cleaved within the A beta domain, and the alternative proteolytic pathway, which produces intact A beta, are unknown. It is hypothesized that a number of neurodegenerative diseases, including Alzheimer's disease, are characterized by abnormal calcium metabolism. We investigated the effect of disordered calcium homeostasis on A beta production in human kidney 293 cells transfected with beta APP cDNA. A beta immunoprecipitated from the conditioned media of cells was compared to immunoprecipitated full-length and secreted forms of beta APP in both metabolic labeling and pulse-chase labeling paradigms. The calcium ionophore A23187 consistently increased the production of A beta approximately 3-fold. This effect was dependent on the presence of extracellular calcium in intact cells. Caffeine also increased A beta production, possibly through release of calcium from intracellular stores. The increase in A beta was cAMP-independent, and it was not mediated by a protein kinase C-dependent pathway, as treatment with phorbol esters decreased A beta levels. The effects of the ionophore on beta APP maturation and phosphorylation were also established. We conclude that elevation of intracellular calcium levels has an important effect on beta APP maturation and proteolytic processing and substantially enhances the production and release of the amyloidogenic A beta peptide.