Flow cytometry-based method to analyze the change in Tau phosphorylation in a hGSK-3beta and hTau over-expressing EcR-293 cell line.
Neurofibrillary tangles are composed of insoluble aggregates of microtubule-associated protein Tau. In the pathology of Alzheimer's disease (AD), accumulation of hyperphosphorylated Tau results in formation of paired helical filaments. One of the main candidate to hyperphosphorylate Tau in AD is glycogen synthase kinase 3beta (GSK-3beta). Here we introduce a non-neuronal cell line, stably co-expressing human Tau and GSK-3beta proteins, where the effect of potential kinase inhibitors on Tau phosphorylation can be monitored. The aim of our study was to establish a new flow-cytometry-based method to quantitatively analyze the changing of Tau phosphorylation, which is a suitable alternative to the well-accepted but non-quantitative Western blot technique. Our results demonstrate that the flow cytometry-based method is a convenient tool to analyze the effect of GSK-3beta inhibitors on Tau phosphorylation. This new approach provides appropriate throughput for screening purposes in preclinical research for characterization of GSK-3beta inhibitors, as potential drug candidate to cure Alzheimer's disease.
Kurko
Dalma
D
Pharmacological and Drug Safety Research, Gedeon Richter Ltd., Budapest, Hungary.
Boros
András
A
Dezso
Péter
P
Urbányi
Zoltán
Z
Sárvári
Miklós
M
Nagy
József
J
Szombathelyi
Zsolt
Z
Szendrei
Györgyi I
GI
D000544:Alzheimer Disease / Q000188:drug therapy / Q000201:enzymology / Q000235:genetics; D002460:Cell Line; D004353:Drug Evaluation, Preclinical / Q000379:methods; D004791:Enzyme Inhibitors / Q000494:pharmacology* / Q000627:therapeutic use; D005434:Flow Cytometry / Q000379:methods*; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D016874:Neurofibrillary Tangles / Q000187:drug effects / Q000201:enzymology; D010766:Phosphorylation; D012333:RNA, Messenger / Q000187:drug effects / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects / Q000502:physiology; D016875:tau Proteins / Q000235:genetics / Q000378:metabolism*
D004791:Enzyme Inhibitors; D012333:RNA, Messenger; D016875:tau Proteins; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; D038362:Glycogen Synthase Kinase 3