The cerebral accumulation of beta-amyloid (Abeta) is a consistent feature of and likely contributor to the development of Alzheimer's disease. In addition to dysregulated production, increasing experimental evidence suggests reduced catabolism also plays an important role in Abeta accumulation. We have previously shown that neprilysin (NEP), the major protease which cleaves Abetain vivo, is modified by 4-hydroxy-nonenal (HNE) adducts in the brain of Alzheimer's disease patients. To determine if these changes affected Abeta, SH-SY5Y cells were treated with HNE or Abeta, and then NEP mRNA, protein levels, HNE adducted NEP, NEP activity and secreted Abeta levels were determined. Intracellular NEP developed HNE adducts after 24 h of HNE treatment as determined by immunoprecipitation, immunoblotting, and double immunofluorescence staining. HNE-modified NEP showed decreased catalytic activity, which was associated with elevations in Abeta1-40 in SH-SY5Y and H4 APP695wt cells. Incubation of cells with Abeta1-42 also induced HNE adduction of NEP. In an apparent compensatory response, Abeta-treated cells showed increased NEP mRNA and protein expression. Despite elevations in NEP protein, the activity was significantly lower compared with the NEP protein level. This study demonstrates that NEP can be inactivated by HNE-adduction, which is associated with, at least partly, reduced Abeta cleavage and enhanced Abeta accumulation.