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P4‐165: Role of chaperone‐mediated autophagy in Huntington's disease
Author(s) -
Qi Lin
Publication year - 2011
Publication title -
alzheimer's and dementia
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 6.713
H-Index - 118
eISSN - 1552-5279
pISSN - 1552-5260
DOI - 10.1016/j.jalz.2011.05.2187
Subject(s) - huntingtin , autophagy , polyglutamine tract , microbiology and biotechnology , lysosome , batten disease , biology , western blot , transfection , chaperone (clinical) , chemistry , mutant , cell culture , biochemistry , gene , genetics , apoptosis , enzyme , medicine , pathology
cases of AD. Evidences suggest that aggregation of a-syn is an important intermediate in the development of mitochondrial dysfunction and neurodegeneration in PD. Dimebon was originally developed as an antihistamine drug. This drug was later found to have cognition-enhancing effects in rodents and neuroprotective and pro-neurogenic functions, possibly through a mitochondrial-related mechanism, and is currently in Phase 3 clinical trials for the treatment of AD and Huntington’s disease (HD). Methods: SH-SY5Y cells were treated with hydrogen peroxide (300 uM), the calcium ionophore 4-Br-A23187 (2 uM) or the mitochondrial toxin rotenone (150 uM) in the presence ofvehicleorDimebon for 12and24hours and cell viabilitymeasured by lactate dehydrogenase (LDH) release. In addition,we used a Tet-off system to induce the expression ofwild type a-syn inSH-SY5Ycells (VekrellisKet al. J. of Neurochemistry 2009; 109: 1348) to study the effect ofDimebon on a-syn mRNA and protein expression and cytotoxicity. Cells were treated with retinoic acid (10 uM) (RA) and vehicle orDimebon in the absence of doxycycline. Cultures were maintained with RA and drugs treatments for 14 days. Cell viabilitywas byLDHrelease andMTS tetrazolium reduction assays. Expression of a-syn protein was by western blotting and by immunocytochemistry and levels of a-syn mRNA was by quantitative PCR. Levels of a-syn mRNA were normalized to beta-tubulin mRNA.Results: Dimebon at concentrations of between 1 and 100 nMwere associatedwith a statistically significant reduction in LDH release in SH-SY5Y cells treated with hydrogen peroxide (1520% reduction), 4-Br-A23187 (15-20%) and rotenone (55-60%). Elimination of doxycycline from the culture media induces a-syn expression by the Tet/off SH-SY5Y cells and increases LDH release and reduces MTS activity. Dimebon treatment (0.1 to 100 nM) was associated with a statistically significant improvement in cell viability asmeasuredby theLDHandMTSassays.Dimebon (10 nM) treatment for 14 days decreases total levels of a-syn protein,measured bywestern blot analysis and immunocytochemistry. Bywestern blotting itwas found that levels ofTritonX-100 insoluble a-syn aggregates aswells soluble monomers were reduced. No significant differences were found in the levels of a-synmRNAmeasured by quantitative PCR.Conclusions: These results indicate that in cell cultures Dimebon enhances the viability of neuroblastoma cells under different cytotoxic conditions including oxidative stress (hydrogen peroxide), high intracellular calcium (4-Br-A23187), mitochondrial poison (rotenone) and high a-syn protein levels. Dimebon reduces a-syn protein but not mRNA suggesting an effect on protein turnover rather than synthesis. These studies suggest that Dimebon has mitochondrial actions that could play a role in modulating cell processes involving protein turnover which in the setting of neurodegenerative proteinopathies may be beneficial.