Hyperphosphorylation and accumulation of neurofilament proteins in Alzheimer disease brain and in okadaic acid-treated SY5Y cells

We investigated the role of neurofilament (NF) proteins in Alzheimer diseas e (AD) neurofibrillary degeneration. The levels and degree of phosphorylati on of NF proteins in AD neocortex were determined by Western blots develope d with a panel of phosphorylation-dependent NF antibodies. Levels of all th ree NF subunits and the degree of phosphorylation of NF-H and NF-M were sig nificantly increased in AD as compared to Huntington disease brains used as control tissue. The increase in the levels of NF-H and NF-M was 1.7- and 1 .5-fold (P < 0.01) as determined by monoclonal antibody SM133, and was 1.6- fold (P < 0.01) in NF-L using antibody NR4. The phosphorylation of NF-H and NF-M in AD was increased respectively at the SM131 epitope by 1.6- and 1.9 -fold (P <less than> 0.05) and at the SM133 epitope by 2.7- and 1.3-fold (P < 0.01 and P < 0.05). Essentially similar effects were observed in SY5Y hu man neuroblastoma cells when treated with okadaic acid, an inhibitor of pro tein phosphatase (PP)-2A and -1. This is the first biochemical evidence whi ch unambiguously demonstrates the hyperphosphorylation and the accumulation of NF subunits in AD brain, and shows that the inhibition of PP-2A/PP-1 ac tivities can lead to the hyperphosphorylation of NF-H and NF-M subunits. (C ) 2001 Published by Elsevier Science B.V. on behalf of the Federation of Eu ropean Biochemical Societies.