Metabolic State Determines Sensitivity to Cellular Stress in Huntington Disease: Normalization by Activation of PPARγ

Impairments in mitochondria and transcription are important factors in the pathogenesis of Huntington disease (HD), a neurodegenerative disease caused by a polyglutamine expansion in the huntingtin protein. This study investigated the effect of different metabolic states and peroxisome proliferator-activated receptor γ (PPARγ) activation on sensitivity to cellular stressors such as H2O2 or thapsigargin in HD. Striatal precursor cells expressing wild type (STHdhQ7) or mutant huntingtin (STHdhQ111) were prepared in different metabolic conditions (glucose vs. pyruvate). Due to the fact that STHdhQ111 cells exhibit mitochondrial deficits, we expected that in the pyruvate condition, where ATP is generated primarily by the mitochondria, there would be greater differences in cell death between the two cell types compared to the glucose condition. Intriguingly, it was the glucose condition that gave rise to greater differences in cell death. In the glucose condition, thapsigargin treatment resulted in a more rapid loss of mitochondrial membrane potential (ΔΨm), a greater activation of caspases (3, 8, and 9), and a significant increase in superoxide/reactive oxygen species (ROS) in STHdhQ111 compared to STHdhQ7, while both cell types showed similar kinetics of ΔΨm-loss and similar levels of superoxide/ROS in the pyruvate condition. This suggests that bioenergetic deficiencies are not the primary contributor to the enhanced sensitivity of STHdhQ111 cells to stressors compared to the STHdhQ7 cells. PPARγ activation significantly attenuated thapsigargin-induced cell death, concomitant with an inhibition of caspase activation, a delay in ΔΨm loss, and a reduction of superoxide/ROS generation in STHdhQ111 cells. Expression of mutant huntingtin in primary neurons induced superoxide/ROS, an effect that was significantly reduced by constitutively active PPARγ. These results provide significant insight into the bioenergetic disturbances in HD with PPARγ being a potential therapeutic target for HD

Cigarette smoke extracts and cadmium induce COX-2 expression through γ-secretase-mediated p38 MAPK activation in C6 astroglia cells.

Exposure to cigarette smoke has been implicated in the progression of cerebrovascular and neurological disorders like stroke through inflammation and blood-brain barrier disruption. In this study, we investigated the signaling cascade activated by cigarette smoke extracts (CSE) and cadmium (Cd) resulting in the COX-2 induction in C6 rat astroglia cells. CSE or Cd induced Notch1 cleavage and activated p38 MAPK and CREB signaling pathways in C6 astroglia cells. Knockdown of nicastrin using siRNA or γ-secretase inhibitors, DAPT and L-685,486, reduced Notch1 cleavage and phosphorylation of p38 MAPK and CREB, while phosphorylation of ERK and JNK remained unaffected. Additionally, the blockage of γ-secretase activity did not show any effect on the phosphorylation of AKT, another upstream activator of CREB, indicating that γ-secretase-mediated CREB activation occurs via p38 MAPK. γ-secretase inhibitor also inhibited the CSE and Cd-mediated increase in the expression of COX-2. Furthermore, recombinant overexpression of Notch1 intracellular domain resulted in an increase in the expression of COX-2. Notch signaling induced by CSE and Cd induced apoptosis in C6 cells. Our results demonstrate that CSE exposure activated the p38 MAPK and CREB-mediated induction in COX-2 expression in astrocytes via γ-secretase-mediated Notch1 signaling. Our data provides novel insights into the potential mechanism of pro-inflammatory response activated by exposure to cigarette smoke

Altered expression of Notch1 in Alzheimer's disease.

Notch signaling is an evolutionarily conserved pathway that regulates cell-cell interactions through binding of Notch family receptors to their cognate ligands. Notch signaling has an essential role in vascular development and angiogenesis. Recent studies have reported that Notch may be implicated in Alzheimer's disease (AD) pathophysiology. We measured the levels of soluble Notch1 (sNotch1) in the plasma samples from 72 dementia patients (average age 75.1 y), 89 subjects with amnestic mild cognitive impairment (MCI) (average age 73.72 y), and 150 cognitively normal controls (average age 72.34 y). Plasma levels of sNotch1 were 25.27% lower in dementia patients as compared to healthy control subjects. However, the level of Notch1 protein was significantly increased in human brain microvascular endothelial cells (HBMECs) after amyloid-beta treatment. Also, Notch1 mRNA level was significantly increased in HBMECs and iPSC-derived neuronal cells from AD patient compared to normal control. These results indicate that altered expression of Notch1 might be associated with the risk of Alzheimer's disease

Metabolic conditions differentiate ΔΨm-loss in response to TG.

<p><i>A</i>, In the glucose condition STHdh^Q111 cells undergo ΔΨm-loss in response to TG at a significantly faster rate than STHdh^Q7 cells. In the pyruvate condition, ΔΨm of both cell types shows similar kinetics in response to TG. Arrows indicate the point at which ΔΨm begins to drop below the baseline. <i>n</i> = 4. <i>B</i>, RSG slightly but significantly delays TG-induced ΔΨm-loss in glucose and pyruvate conditions. GW9662 abrogates the delayed ΔΨm-loss by RSG treatment. <i>n</i> = 5–9. Data shown are mean ± SE. * <i>P</i><0.05.</p

Mutant huntingtin expression sensitizes striatal cells to stressors in the glucose condition.

<p><i>A</i>, H₂O₂ treatment in the glucose condition results in significantly greater cell death in STHdh^Q111 than STHdh^Q7 cells, while both cell types show similar cell death responses to H₂O₂ in pyruvate condition. <i>n</i> = 4. <i>B</i>, RSG treatment does not protect striatal cells from H₂O₂ toxicity. <i>n</i> = 3–4. <i>C</i>, TG treatment in the glucose condition results in much greater cell death in STHdh^Q111 than STHdh^Q7 cells, while both cell types show similar cell death responses to TG in the pyruvate condition. <i>n</i> = 4–6. <i>D</i>, RSG significantly attenuates TG-induced cell death in the glucose and pyruvate conditions. <i>n</i> = 3–4. RP, rolipram. Data shown are mean ± SE. * <i>P</i><0.05, ** <i>P</i><0.01.</p

Mutant huntingtin results in repressed transcriptional activities, and reduced PPARγ activity is independent of the protein level.

<p>B3 and E4 STHdh^Q7 cells and 1A and 6L STHdh^Q111 cells were transiently transfected with PPRE, CRE, or PGC-1α promoter luciferase reporter plasmids. The basal activities of PPRE (<i>A</i>), CRE (<i>B</i>), and PGC-1α promoter (<i>C</i>) reporters were dramatically reduced in both 1A and 6L STHdh^Q111 cells. <i>n</i> = 3–4 Data shown are mean ± SE. <i>D</i>, PPARγ protein levels were variable in B3 and E4 STHdh^Q7 cells and 1A and 6L STHdh^Q111 cells, while the original STHdh^Q111 cells exhibited lower levels of PPARγ than the original STHdh^Q7 cells. Sixty micrograms of protein was run in each lane.</p