Nicergoline stimulates protein kinase C mediated alpha-secretase processing of the amyloid precursor protein in cultured human neuroblastoma SH-SY5Y cells

We investigated the ability of the antidementia agents, nicergoline, anirac etam and hydergine to stimulate PKC mediated alpha-secretase amyloid precur sor protein (APP) processing in cultured human neuroblastoma SH-SY5Y cells. Western immunoblotting of cell conditioned media using the Mabs 22C11 and 6E10 revealed the presence of 2 bands with molecular mass of 90 and 120 kDa , corresponding to possible alternatively glycosylated forms of secreted AP P (APPs). Short-term (30 min and 2 h) treatment of cells with nicergoline g ave an increased intensity of both bands, compared to non-treated cells, Ma ximal nicergoline effects, of the order of 150-200% over basal APPs release , were seen at concentrations between 1 and 10 mu M. Under the same conditi on, 1 mu M PdBu, used as a positive control, gave 500-1000% increases of ba sal APPs release. In contrast, aniracetam and hydergine, did not show any e ffect on APPs secretion. 2 h treatment with nicergoline had no effect on ce llular full-length APP levels, as determined by inmunoblotting of cell extr acts with 22C11 and CT15 antibodies. Immunoblotting with PKC isoform specif ic antibodies of soluble and membrane fractions prepared from 2 h treated c ells, showed that nicergoline (50 mu M) and PdBu (1 mu M) both induced tran slocation of PKC alpha, gamma and epsilon, but not PKC beta. The involvemen t of PKC in mediating nicergoline stimulated APPs release was also studied using specific inhibitors. 1 mu M calphostin C, a broad range PKC inhibitor , significantly reduced both PdBu (1 mu M) and nicergoline (10 mu M) induce d APPs release, In contrast, Go6976 (1 mu M), a selective PKC alpha and bet a 1 inhibitor, as well as the cAMP-dependent protein kinase inhibitor, H89 (1 mu M) were without effect. These results indicate that nicergoline can m odulate alpha-secretase APP processing by a PKC dependent mechanism that is likely to involve the gamma and epsilon isoforms of this enzyme. (C) 1999 Elsevier Science Ltd. All rights reserved.