Rosiglitazone Treatment Prevents Mitochondrial Dysfunction in Mutant Huntingtin-expressing Cells

Abstract

Peroxisome proliferator-activated receptor-␥ (PPAR␥) is a member of the PPAR family of transcription factors. Synthetic PPAR␥ agonists are used as oral anti-hyperglycemic drugs for the treatment of non-insulin-dependent diabetes. However, emerging evidence indicates that PPAR␥ activators can also prevent or attenuate neurodegeneration. Given these previous findings, the focus of this report is on the potential neuroprotective role of PPAR␥ activation in preventing the loss of mitochondrial function in Huntington disease (HD). For these studies we used striatal cells that express wild-type (STHdh Q7/Q7 ) or mutant (STHdh Q111/Q111 ) huntingtin protein at physiological levels. Treatment of mutant cells with thapsigargin resulted in a significant decrease in mitochondrial calcium uptake, an increase in reactive oxygen species production, and a significant decrease in mitochondrial membrane potential. PPAR␥ activation by rosiglitazone prevented the mitochondrial dysfunction and oxidative stress that occurred when mutant striatal cells were challenged with pathological increases in calcium. The beneficial effects of rosiglitazone were likely mediated by activation of PPAR␥, as all protective effects were prevented by the PPAR␥ antagonist GW9662. Additionally, the PPAR␥ signaling pathway was significantly impaired in the mutant striatal cells with decreases in PPAR␥ expression and reduced PPAR␥ transcriptional activity. Treatment with rosiglitazone increased mitochondrial mass levels, suggesting a role for the PPAR␥ pathway in mitochondrial function in striatal cells. Altogether, this evidence indicates that PPAR␥ activation by rosiglitazone attenuates mitochondrial dysfunction in mutant huntingtin-expressing striatal cells, and this could be an important therapeutic avenue to ameliorate the mitochondrial dysfunction that occurs in HD.

Caspase-cleaved Tau Expression Induces Mitochondrial Dysfunction in Immortalized Cortical Neurons

“…The fluorescence changes determined by Fluo-3 represent the cytoplasmic calcium changes (20), and Rhod-2 fluorescence indicates calcium changes in the mitochondria (21)(22)(23). To estimate Rhod-2 fluorescence pattern in live mitochondria, we used MitoTracker Green TM (MTG) to mark the mitochondria (22,23). Cells were washed 3 times and left in KRH-glucose buffer for 10 min until cell fluorescence equilibrated.…”

“…Cells were washed 3 times and left in KRH-glucose buffer for 10 min until cell fluorescence equilibrated. Fluorescence was imaged with a confocal laser scanning microscope (Leica TCS SP2) using a 40ϫ water immersion lens, as described previously (22,24). Images were acquired using a 488-nm argon laser to excite Fluo-3 fluorescence and a 563-nm He-Ne laser to excite Rhod-2 fluorescence.…”

“…The fluorescence intensity variation was recorded from 10 -20 cells on average per experiment. Estimation of fluorescence intensities were presented as the pseudoratio (⌬F/F o ), which was calculated using the formula ⌬F/F o ϭ (F Ϫ F base )/(F base Ϫ B), where F is the measured fluorescence intensity of the indicator, F base is the fluorescence intensity before the stimulation, and B is the background signal determined from the average of areas adjacent to the cells (22).…”

“…Images were quantified using Image-Pro Plus 6 software. Results in intensity units were expressed as the average of fluorescence signal (F) minus background fluorescence (F o ) in every image (22).…”

“…Mitochondrial Membrane Potential Determination in Live Cells-Mitochondrial membrane potential was determined using TMRM (22). Before thapsigargin treatment, the cells were loaded for 30 min with TMRM (100 nM) in KRH-glucose buffer containing 0.02% pluronic acid, then washed and allowed to equilibrate for 20 min.…”