The effect of tacrine and leupeptin on the secretion of the beta-amyloid precursor protein in HeLa cells.
The senile plaque in Alzheimer's disease (AD) consists mainly of the amyloid beta-peptide (A beta) derived from a larger beta-amyloid precursor protein (betaAPP). The majority of betaAPP is processed by either a secretory or lysosomal/endosomal pathway. Soluble derivatives of betaAPP (sAPP) and A beta generated by the proteolytic processing of full-length betaAPP are normally secreted into the conditioned medium of cultured cells. Tacrine, a centrally active potent cholinesterase inhibitor that has been shown to improve cognitive functions in some patients with AD, inhibits the secretion of sAPP. Here we have investigated whether leupeptin, a lysosomal protease inhibitor, could influence this effect of tacrine. We analyzed levels of betaAPP derivatives in cultured HeLa cells by immunoblotting cell lysates and conditioned media using the monoclonal antibody 22C11. Levels of sAPP normally present in conditioned media were severely reduced by treating cells with tacrine. The treatment of cells with tacrine resulted in a small decrease in the intracellular levels of betaAPP. The effect of treating the cells with tacrine did not depend upon the growing state of the cells as a similar effect was observed when the drug was added either during initial plating of the cells or after the attachment of the cells. The effect of tacrine was not affected by preincubating the cells with low serum in the culture medium. The treatment of cells with tacrine plus leupeptin reduced the secretion of sAPP in the medium to the same degree as did the treatment with tacrine alone, suggesting that the tacrine-mediated inhibition of sAPP release may not involve leupeptin-sensitive proteolytic pathways. The results suggest that the inhibitory effect of tacrine on sAPP secretion is not due to the proteolytic cleavage of the holoprotein in the medium.
Lahiri
D K
DK
Institute of Psychiatric Research, Department of Psychiatry, Indiana University School of Medicine, Indianapolis 46202, USA.
Farlow
M R
MR
Sambamurti
K
K
Nall
C
C
D000544:Alzheimer Disease; D016564:Amyloid beta-Protein Precursor / Q000378:metabolism*; D002217:Carbachol / Q000494:pharmacology; D002800:Cholinesterase Inhibitors / Q000494:pharmacology*; D017077:Culture Media, Conditioned; D006367:HeLa Cells / Q000187:drug effects* / Q000378:metabolism*; D006801:Humans; D015151:Immunoblotting; D007976:Leupeptins / Q000494:pharmacology*; D010277:Parasympathomimetics / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology; D013619:Tacrine / Q000494:pharmacology*
D016564:Amyloid beta-Protein Precursor; D002800:Cholinesterase Inhibitors; D017077:Culture Media, Conditioned; D007976:Leupeptins; D010277:Parasympathomimetics; D011480:Protease Inhibitors; D013619:Tacrine; D002217:Carbachol; C032854:leupeptin