Rodrigo A. Quintanilla

Peroxisome Proliferator-activated Receptor γ Up-regulates the Bcl-2 Anti-apoptotic Protein in Neurons and Induces Mitochondrial Stabilization and Protection against Oxidative Stress and Apoptosis

Peroxisome proliferator-activated receptor γ (PPARγ) has been proposed as a therapeutic target for neurodegenerative diseases because of its anti-inflammatory action in glial cells. However, PPARγ agonists preventβ-amyloid (Aβ)-induced neurodegeneration in hippocampal neurons, and PPARγ is activated by the nerve growth factor (NGF) survival pathway, suggesting a neuroprotective anti-inflammatory independent action. Here we show that the PPARγ agonist rosiglitazone (RGZ) protects hippocampal and dorsal root ganglion neurons against Aβ-induced mitochondrial damage and NGF deprivation-induced apoptosis, respectively, and promotes PC12 cell survival. In neurons and in PC12 cells RGZ protective effects are associated with increased expression of the Bcl-2 anti-apoptotic protein. NGF-differentiated PC12 neuronal cells constitutively overexpressing PPARγ are resistant to Aβ-induced apoptosis and morphological changes and show functionally intact mitochondria and no increase in reactive oxygen species when challenged with up to 50 μm H2O2. Conversely, cells expressing a dominant negative mutant of PPARγ show increased Aβ-induced apoptosis and disruption of neuronal-like morphology and are highly sensitive to oxidative stress-induced impairment of mitochondrial function. Cells overexpressing PPARγ present a 4- to 5-fold increase in Bcl-2 protein content, whereas in dominant negative PPARγ-expressing cells, Bcl-2 is barely detected. Bcl-2 knockdown by small interfering RNA in cells overexpressing PPARγ results in increased sensitivity to Aβ and oxidative stress, further suggesting that Bcl-2 up-regulation mediates PPARγ protective effects. PPARγ prosurvival action is independent of the signal-regulated MAPK or the Akt prosurvival pathways. Altogether, these data suggest that PPARγ supports survival in neurons in part through a mechanism involving increased expression of Bcl-2.

The Permeability Transition Pore Controls Cardiac Mitochondrial Maturation and Myocyte Differentiation

Although mature myocytes rely on mitochondria as the primary source of energy, the role of mitochondria in the developing heart is not well known. Here, we find that closure of the mitochondrial permeability transition pore (mPTP) drives maturation of mitochondrial structure and function and myocyte differentiation. Cardiomyocytes at embryonic day (E) 9.5, when compared to E13.5, displayed fragmented mitochondria with few cristae, a less-polarized mitochondrial membrane potential, higher reactive oxygen species (ROS) levels, and an open mPTP. Pharmacologic and genetic closing of the mPTP yielded maturation of mitochondrial structure and function, lowered ROS, and increased myocyte differentiation (measured by counting Z bands). Furthermore, myocyte differentiation was inhibited and enhanced with oxidant and antioxidant treatment, respectively, suggesting that redox-signaling pathways lie downstream of mitochondria to regulate cardiac myocyte differentiation.

Peroxisomal Proliferation Protects from β-Amyloid Neurodegeneration

Alzheimer disease is a neurodegenerative process that leads to severe cognitive impairment as a consequence of selective death of neuronal populations. The molecular pathogenesis of Alzheimer disease involves the participation of the β-amyloid peptide (Aβ) and oxidative stress. We report here that peroxisomal proliferation attenuated Aβ-dependent toxicity in hippocampal neurons. Pretreatment with Wy-14.463 (Wy), a peroxisome proliferator, prevent the neuronal cell death and neuritic network loss induced by the Aβ peptide. Moreover, the hippocampal neurons treated with this compound, showed an increase in the number of peroxisomes, with a concomitant increase in catalase activity. Additionally, we evaluate the Wy protective effect on β-catenin levels, production of intracellular reactive oxygen species, cytoplasmic calcium uptake, and mitochondrial potential in hippocampal neurons exposed to H2 O2 and Aβ peptide. Results show that the peroxisomal proliferation prevents β-catenin degradation, reactive oxygen species production, cytoplasmic calcium increase, and changes in mitochondrial viability. Our data suggest, for the first time, a direct link between peroxisomal proliferation and neuroprotection from Aβ-induced degenerative changes.

Mutant Huntingtin Expression Induces Mitochondrial Calcium Handling Defects in Clonal Striatal Cells FUNCTIONAL CONSEQUENCES

Huntington disease (HD) is caused by a pathological elongation of CAG repeats in the huntingtin protein gene and is characterized by atrophy and neuronal loss primarily in the striatum. Mitochondrial dysfunction and impaired Ca2+ homeostasis in HD have been suggested previously. Here, we elucidate the effects of Ca2+ on mitochondria from the wild type (STHdhQ7/Q7) and mutant (STHdhQ111/Q111) huntingtin-expressing cells of striatal origin. When treated with increasing Ca2+ concentrations, mitochondria from mutant huntingtin-expressing cells showed enhanced sensitivity to Ca2+, as they were more sensitive to Ca2+-induced decreases in state 3 respiration and ΔΨm, than mitochondria from wild type cells. Further, mutant huntingtin-expressing cells had a reduced mitochondrial Ca2+ uptake capacity in comparison with wild type cells. Decreases in state 3 respiration were associated with increased mitochondrial membrane permeability. The ΔΨm defect was attenuated in the presence of ADP and the decreases in Ca2+ uptake capacity were abolished in the presence of Permeability Transition Pore (PTP) inhibitors. These findings clearly indicate that mutant huntingtin-expressing cells have mitochondrial Ca2+ handling defects that result in respiratory deficits and that the increased sensitivity to Ca2+ induced mitochondrial permeabilization maybe a contributing mechanism to the mitochondrial dysfunction in HD.

Caspase-cleaved Tau Expression Induces Mitochondrial Dysfunction in Immortalized Cortical Neurons IMPLICATIONS FOR THE PATHOGENESIS OF ALZHEIMER DISEASE

In Alzheimer disease (AD) mitochondrial abnormalities occur early in the pathogenic process and likely play a significant role in disease progression. Tau is a microtubule-associated protein that is abnormally processed in AD, and a connection between tau pathology and mitochondrial impairment has been proposed. However, few studies have examined the relationship between pathological forms of tau and mitochondrial dysfunction. We recently demonstrated that inducible expression of tau truncated at Asp-421 to mimic caspase cleavage (T4C3) was toxic to immortalized cortical neurons compared with a full-length tau isoform (T4). In this study we investigated the effects of T4C3 on mitochondrial function. Expression of T4C3 induced mitochondrial fragmentation and elevated oxidative stress levels in comparison with T4-expressing cells. Thapsigargin treatment of T4 or T4C3 cells, which causes an increase in intracellular calcium levels, resulted in a significant decrease in mitochondrial potential and loss of mitochondrial membrane integrity in T4C3 cells when compared with cells expressing T4. The mitochondrial fragmentation and mitochondrial membrane damage were ameliorated in T4C3 cells by pretreatment with cyclosporine A or FK506, implicating the calcium-dependent phosphatase calcineurin in these pathogenic events. Increased calcineurin activity has been reported in AD brain, and thus, inhibition of this phosphatase may provide a therapeutic target for the treatment of AD.

Role of mitochondrial dysfunction in the pathogenesis of Huntington's disease.

Huntingtons disease (HD) is an autosomal dominant neurodegenerative disorder that is caused by a pathological expansion of CAG repeats within the gene encoding for a 350 kD protein called huntingtin. This polyglutamine expansion within huntingtin is the causative factor in the pathogenesis of HD, however the underlying mechanisms have not been fully elucidated. Nonetheless, it is becoming increasingly clear that alterations in mitochondrial function play key roles in the pathogenic processes in HD. The net result of these events is compromised energy metabolism and increased oxidative damage, which eventually contribute to neuronal dysfunction and death. Mitochondria from striatal cells of a genetically accurate model of HD take up less calcium and at a slower rate than mitochondria from striatal cells derived from normal mice. Further, respiration in mitochondria from these mutant huntingtin-expressing cells is inhibited at significantly lower calcium concentrations compared to mitochondria from wild-type cells. Considering these and other findings this review explores the evidence suggesting that mutant huntingtin, directly or indirectly impairs mitochondrial function, which compromises cytosolic and mitochondrial calcium homeostasis, and contributes to neuronal dysfunction and death in HD.

Rosiglitazone Treatment Prevents Mitochondrial Dysfunction in Mutant Huntingtin-expressing Cells: POSSIBLE ROLE OF PEROXISOME PROLIFERATOR-ACTIVATED RECEPTOR-γ (PPARγ) IN THE PATHOGENESIS OF HUNTINGTON DISEASE*

Peroxisome proliferator-activated receptor-γ (PPARγ) is a member of the PPAR family of transcription factors. Synthetic PPARγ agonists are used as oral anti-hyperglycemic drugs for the treatment of non-insulin-dependent diabetes. However, emerging evidence indicates that PPARγ activators can also prevent or attenuate neurodegeneration. Given these previous findings, the focus of this report is on the potential neuroprotective role of PPARγ activation in preventing the loss of mitochondrial function in Huntington disease (HD). For these studies we used striatal cells that express wild-type (STHdhQ7/Q7) or mutant (STHdhQ111/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. PPARγ 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 PPARγ, as all protective effects were prevented by the PPARγ antagonist GW9662. Additionally, the PPARγ signaling pathway was significantly impaired in the mutant striatal cells with decreases in PPARγ expression and reduced PPARγ transcriptional activity. Treatment with rosiglitazone increased mitochondrial mass levels, suggesting a role for the PPARγ pathway in mitochondrial function in striatal cells. Altogether, this evidence indicates that PPARγ 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.

Truncated tau and Aβ cooperatively impair mitochondria in primary neurons

Mitochondrial dysfunction is likely a significant contributing factor to Alzheimer disease pathogenesis, and both amyloid peptide (Aβ) and pathological forms of tau may contribute to this impairment. Cleavage of tau at Asp421 occurs early in Alzheimer disease, and Asp421-cleaved tau likely negatively impacts neuronal function. Previously we showed that expression of Asp421-cleaved tau in a neuronal cell model resulted in mitochondrial impairment. To extend these findings we expressed either full length tau or Asp421-cleaved tau (truncated tau) in primary cortical neurons and measured different aspects of mitochondrial function with or without the addition of sublethal concentrations of Aβ. The expression of truncated tau alone induced significant mitochondrial fragmentation in neurons. When truncated tau expression was combined with Aβ at sublethal concentrations, increases in the stationary mitochondrial population and the levels of oxidative stress in cortical neurons were observed. Truncated tau expression also enhanced Aβ-induced mitochondrial potential loss in primary neurons. These new findings show that Asp421-cleaved tau and Aβ cooperate to impair mitochondria, which likely contributes to the neuronal dysfunction in Alzheimer disease.

Mitochondrial permeability transition pore induces mitochondria injury in Huntington disease

BackgroundMitochondrial impairment has been implicated in the pathogenesis of Huntington’s disease (HD). However, how mutant huntingtin impairs mitochondrial function and thus contributes to HD has not been fully elucidated. In this study, we used striatal cells expressing wild type (STHdhQ7/Q7) or mutant (STHdhQ111/Q111) huntingtin protein, and cortical neurons expressing the exon 1 of the huntingtin protein with physiological or pathological polyglutamine domains, to examine the interrelationship among specific mitochondrial functions.ResultsDepolarization induced by KCl resulted in similar changes in calcium levels without compromising mitochondrial function, both in wild type and mutant cells. However, treatment of mutant cells with thapsigargin (a SERCA antagonist that raises cytosolic calcium levels), resulted in a pronounced decrease in mitochondrial calcium uptake, increased production of reactive oxygen species (ROS), mitochondrial depolarization and fragmentation, and cell viability loss. The mitochondrial dysfunction in mutant cells was also observed in cortical neurons expressing exon 1 of the huntingtin protein with 104 Gln residues (Q104-GFP) when they were exposed to calcium stress. In addition, calcium overload induced opening of the mitochondrial permeability transition pore (mPTP) in mutant striatal cells. The mitochondrial impairment observed in mutant cells and cortical neurons expressing Q104-GFP was prevented by pre-treatment with cyclosporine A (CsA) but not by FK506 (an inhibitor of calcineurin), indicating a potential role for mPTP opening in the mitochondrial dysfunction induced by calcium stress in mutant huntingtin cells.ConclusionsExpression of mutant huntingtin alters mitochondrial and cell viability through mPTP opening in striatal cells and cortical neurons.

Understanding Risk Factors for Alzheimer's Disease: Interplay of Neuroinflammation, Connexin-based Communication and Oxidative Stress

Alzheimers disease (AD) is an age-related neurodegenerative disease characterized by dementia and the presence of amyloid plaques and anomalous tau aggregates. Although pathophysiological mechanisms are still unclear, neuroinflammation and glial cell dysfunction have been identified as conspicuous components of AD. Glial cell dysfunction is associated with dysregulated production of inflammation mediators and generation of both reactive oxygen species (ROS) and reactive nitrogen species (RNS), which affect synapses and induce neuronal damage. Importantly, both increased neuroinflammation and ROS/RNS production by glia dysregulate communication mediated by connexin-based channels in brain cells, which could further affect oxidative balance and neuronal viability. Recent evidence suggests that connexin-based channels could be involved in AD pathogenesis. Here we discuss how aging affects neuroinflammation, oxidative stress, and connexin-based channels and the potential relevance of these changes for AD. Understanding how they cooperate as pathogenic mechanisms of AD is promising for the discovery of new therapeutic strategies against neurodegenerative disorders.