Department of Pathology and Laboratory Medicine, and Waisman Center, University of Wisconsin School of Medicine and Public Health, Madison, WI 53706, USA.
J Alzheimers Dis. 2010;19(1):179-89. doi: 10.3233/JAD-2010-1226.
As one of the dominant amyloid-beta peptide (Abeta) proteases, neprilysin (NEP) plays a crucial role in maintaining a physiologic balance between Abeta production and catabolism. We have previously shown that NEP is modified by 4-hydroxynonenal (HNE) adducts, resulting in decreased activity in the brain of AD patients and cultured cells. In order to determine whether antioxidants can rescue NEP, SH-SY5Y cells were treated with HNE or Abeta, together with N-acetylcysteine for 24 hours, prior to analysis of NEP protein levels, activity, and oxidative modifications. Intracellular NEP developed HNE adducts after 24 hours of HNE or Abeta treatment as determined by immunoprecipitation, immunoblotting, and double immunofluorescence staining. N-acetylcysteine at 10 to 100 microM alleviated HNE adduction after HNE or Abeta treatment. In keeping with previous reports, HNE-modified NEP showed decreased catalytic activity. The present study demonstrates that antioxidants can be used to spare NEP from oxidative modification, suggesting a potential mechanism underlying the neuroprotective effects of antioxidants in aging or Alzheimer's disease.
作为主要的淀粉样β肽（Abeta）蛋白酶之一， Neprilysin（NEP）在维持 Abeta 产生和代谢之间的生理平衡方面起着至关重要的作用。我们之前已经表明，NEP 被 4-羟基壬烯醛（HNE）加合物修饰，导致 AD 患者和培养细胞中的活性降低。为了确定抗氧化剂是否可以挽救 NEP，将 SH-SY5Y 细胞用 HNE 或 Abeta 处理 24 小时，然后分析 NEP 蛋白水平、活性和氧化修饰。通过免疫沉淀、免疫印迹和双免疫荧光染色确定，NEP 在 24 小时的 HNE 或 Abeta 处理后形成了 HNE 加合物。用 10 至 100μM 的 N-乙酰半胱氨酸可减轻 HNE 加合物的形成。与以前的报告一致，HNE 修饰的 NEP 显示出催化活性降低。本研究表明，抗氧化剂可用于防止 NEP 发生氧化修饰，提示抗氧化剂在衰老或阿尔茨海默病中的神经保护作用的潜在机制。
