- Email: [email protected]
P4-168: Mitochondrial dysfunction and neurodegeneration in P301L tau transgenic AIF-deficient mice
T720
Poster Presentations P4:
quantitative PCR we evaluated the level of mtDNA damage at the 16s locus in these two brain regions from 10 LOAD subjects and 10 agematched controls. DNA was prepared using techniques that enrich the sample for mtDNA and reduce in vitro DNA damage. The proportion of nuclear DNA was quantified for each sample to determine the [mtDNA] since variation in mtDNA copy number has been reported. Results: Initial results suggest increased mtDNA damage in the temporal cortex, as compared to the cerebellum, for both AD and control subjects (p⬍0.004). In the temporal cortex we observed a significant difference in mtDNA damage between AD and control subjects (p⫽0.0011) but not in the cerebellum (p⫽0.2799), We also found a significant reduction in the proportion of nuclear to mitochondrial DNA in AD as compared to control samples from the temporal cortex (p⫽0.001). Conclusions: These data indicate that mtDNA damage may be more severe in tissue affected by LOAD pathology and suggest an increase in mtDNA copy number in the temporal cortex. Investigation of additional subjects and mitochondrial loci is underway to confirm these findings. P4-168
MITOCHONDRIAL DYSFUNCTION AND NEURODEGENERATION IN P301L TAU TRANSGENIC AIF-DEFICIENT MICE
Luka Kulic1, Marc Axel Wollmer1, Virginie Rhein2, Lucia Pagani3, Katrin Kuehnle1, Susann Cattepoel1, Jay Tracy1, Anne Eckert3, Roger M. Nitsch1, 1University of Zurich, Zurich, Switzerland; 2 University of Basel, Basel, Switzerland; 3University of Basel, Basel, Switzerland. Contact e-mail: [email protected] Background: Mitochondrial dysfunction and oxidative stress play an important role in the ageing process and in several age-related neurodegenerative diseases including Alzheimer’s disease (AD). AD and some AD-related conditions like frontotemporal dementia with parkinsonism linked to chromosome 17 (FTDP-17) belong to so-called tauopathies, neurodegenerative conditions characterized by the presence of intracellular accumulations of the hyperphosphorylated microtubule-associated protein tau. Methods: In this study we aimed to examine the in vivo effects of tau in a mouse model of impaired mitochondrial function and increased oxidative stress. Therefore, we crossbred P301L tau transgenic mice, an established model of human tauopathies, with mice bearing the Harlequin (Hq) mouse mutation which causes an 80-90% downregulation of the mitochondrial apoptosis-inducing factor (AIF). AIF deficiency in the Hq mutant mice has previously been associated with increased oxidative stress and a reduced activity of the complex I of the mitochondrial respiratory chain (MRC). Results: The Tau/Hq double mutant mice showed an impairment of the activity of multiple complexes of the MRC together with signs of exacerbated oxidative stress and a depletion of cerebral ATP levels. Moreover, they were affected by an increase of the tau pathology, aggravated motor deficits and neuronal degeneration in several brain regions with the most prominent changes in the dentate gyrus and cerebellum. Conclusions: The Tau/Hq double mutant mice may be an interesting mouse model for the study of tau-mediated neurotoxicity in a setting of impaired mitochondrial function, a characteristic of ageing and of several age-related neurodegenerative diseases. P4-169
MITOCHONDRIALLY TARGETED ANTIOXIDANT THERAPEUTICS IN ALZHEIMER’S DISEASE
P. Hemachandra Reddy, Maria Manczak, Wei Zhao, Oregon Health and Science University, Beaverton, OR, USA. Contact e-mail: [email protected] Background: Several lines of evvidence suggest that amyloid beta (Abeta) is associated with mitochondrial damage and synaptic dysfunction in AD. Our global gene expression studies have revealed an up-regulation of mitochondrial genes in AD transgenic mice (Tg2576 line) suggesting that mitochondrial metabolism is impaired by Abeta and that the up-regulation of mitochondrial genes may be compensatory response to Abeta (Reddy et
al 2004). We found Abeta monomers and oligomers in isolated mitochondria from the cortex of Tg2576 mice, and in isolated mitochondria from mouse neuroblastoma (N2a) cells expressing mutant APP (Manczak et al 2006). Our digitonin fractionation analysis of isolated mitochondria from Tg2576 mice and APP cells revealed Abeta in mitochondrial membranes and matrix, and that mitochondrial Abeta decreases cytochrome oxidase and increases H2O2 production and carbonyl proteins. These findings suggest that early mitochondrial targeted interventions may be effective in delaying AD progression. Methods: Recently, several mitochondrial targeted antioxidants (MTAs) including, MitoQ, MitoVitE and several small and peptide-based, antioxidants have been developed. These targeted antioxidants enter the mitochondrial membranes several hundred folds more than regular antioxidants, neutralize free radicals, inhibit mitochondrial permeability transition, and protect neurons. To determine whether MTAs reduces free radicals, oxidative damage and protect neurons from oxidative stress, we investigated the efficacies of MitoQ and SS31 in cell and AD mouse models. Results: Our preliminary studies of N2a cells grown in serum free media treated with MitoQ and SS31 have shown increased neurite outgrowth and increased expression of neuroprotective genes indicating that MitoQ and SS31 may have a neuroprotective role. We found decreased H2O2 production, cytochrome oxidase, and normal cellular ATP levels when N2a cells incubated with Abeta peptide 25-35 and pretreated with MitoQ and SS31 compared to N2a cells incubated with Abeta peptide alone. Further, we found neurite outgrowth is maintained in cells treated with MitoQ and SS31 and Abeta peptide. Conclusions: These findings suggest that MTAs may protect neurons from Abeta toxicity and oxidative damage. Our ongoing studies of Tg2576 mice treated with SS31 and MitoQ may provide new insights in understanding the role of MTAs in delaying AD progression and pathogenesis in Tg2576 mice. P4-170
PEROXIDATIVE NITROTYROSINE FORMATION IN ALZHEIMER’S DISEASE
George Perry1, Adam D. Cash2, Peggy L. Harris2, Sandra L. Siedlak2, Mark A. Smith2, Xiongwei Zhu2, 1University of Texas at San Antonio, San Antonio, TX, USA; 2Case Western Reserve University, Cleveland, OH, USA. Contact e-mail: [email protected] Background: Findings from numerous approaches in Alzheimer disease (AD) implicate oxidative stress as a major pathogenic pathway. We have focused our studies to elucidate the nature of the reactive species and sources of oxidative insult by considering cellular biomacromolecule modifications such as carbonyl and nitrotyrosine formation. These studies have strongly implicated metal catalyzed oxidation as critical to oxidative damage in AD. This led us to re-examine our prior work implicating non-metal catalyzed oxidation through peroxynitrite. Methods: In this study, we examine whether peroxynitrite or peroxidative nitration with nitrite and hydrogen peroxide is responsible for tyrosine nitration. Results: We found that peroxidative nitration yields nitrotyrosine in tissue with the characteristic cytoplasmic and nuclear distribution found in AD. Significantly, the process is dependent on redox active transition metals. In contrast, peroxynitrite produced lesswidespread damage without the anatomic signature of AD. Conclusions: This study suggests nitration of proteins in neurons in AD is in fact dependent on site directed metal-catalyzed processes, further evidence for the significant role of metals in the process of neurodegeneration. It is our conclusion that a reaction involving nitrite, hydrogen peroxide and the redox active metals shown to be elevated in AD, may play the primary roles of oxidative nitrotyrosine damage in AD. To truly target the in vivo mechanisms occurring in neurons in the brain, a complete picture of all nitrative events and consequences must be known. At the least, increasing awareness of the multiple possible pathways leading to the endpoints classified as nitrative damage will only increase the chances for therapeutic success.
Poster Presentations P4:
quantitative PCR we evaluated the level of mtDNA damage at the 16s locus in these two brain regions from 10 LOAD subjects and 10 agematched controls. DNA was prepared using techniques that enrich the sample for mtDNA and reduce in vitro DNA damage. The proportion of nuclear DNA was quantified for each sample to determine the [mtDNA] since variation in mtDNA copy number has been reported. Results: Initial results suggest increased mtDNA damage in the temporal cortex, as compared to the cerebellum, for both AD and control subjects (p⬍0.004). In the temporal cortex we observed a significant difference in mtDNA damage between AD and control subjects (p⫽0.0011) but not in the cerebellum (p⫽0.2799), We also found a significant reduction in the proportion of nuclear to mitochondrial DNA in AD as compared to control samples from the temporal cortex (p⫽0.001). Conclusions: These data indicate that mtDNA damage may be more severe in tissue affected by LOAD pathology and suggest an increase in mtDNA copy number in the temporal cortex. Investigation of additional subjects and mitochondrial loci is underway to confirm these findings. P4-168
MITOCHONDRIAL DYSFUNCTION AND NEURODEGENERATION IN P301L TAU TRANSGENIC AIF-DEFICIENT MICE
Luka Kulic1, Marc Axel Wollmer1, Virginie Rhein2, Lucia Pagani3, Katrin Kuehnle1, Susann Cattepoel1, Jay Tracy1, Anne Eckert3, Roger M. Nitsch1, 1University of Zurich, Zurich, Switzerland; 2 University of Basel, Basel, Switzerland; 3University of Basel, Basel, Switzerland. Contact e-mail: [email protected] Background: Mitochondrial dysfunction and oxidative stress play an important role in the ageing process and in several age-related neurodegenerative diseases including Alzheimer’s disease (AD). AD and some AD-related conditions like frontotemporal dementia with parkinsonism linked to chromosome 17 (FTDP-17) belong to so-called tauopathies, neurodegenerative conditions characterized by the presence of intracellular accumulations of the hyperphosphorylated microtubule-associated protein tau. Methods: In this study we aimed to examine the in vivo effects of tau in a mouse model of impaired mitochondrial function and increased oxidative stress. Therefore, we crossbred P301L tau transgenic mice, an established model of human tauopathies, with mice bearing the Harlequin (Hq) mouse mutation which causes an 80-90% downregulation of the mitochondrial apoptosis-inducing factor (AIF). AIF deficiency in the Hq mutant mice has previously been associated with increased oxidative stress and a reduced activity of the complex I of the mitochondrial respiratory chain (MRC). Results: The Tau/Hq double mutant mice showed an impairment of the activity of multiple complexes of the MRC together with signs of exacerbated oxidative stress and a depletion of cerebral ATP levels. Moreover, they were affected by an increase of the tau pathology, aggravated motor deficits and neuronal degeneration in several brain regions with the most prominent changes in the dentate gyrus and cerebellum. Conclusions: The Tau/Hq double mutant mice may be an interesting mouse model for the study of tau-mediated neurotoxicity in a setting of impaired mitochondrial function, a characteristic of ageing and of several age-related neurodegenerative diseases. P4-169
MITOCHONDRIALLY TARGETED ANTIOXIDANT THERAPEUTICS IN ALZHEIMER’S DISEASE
P. Hemachandra Reddy, Maria Manczak, Wei Zhao, Oregon Health and Science University, Beaverton, OR, USA. Contact e-mail: [email protected] Background: Several lines of evvidence suggest that amyloid beta (Abeta) is associated with mitochondrial damage and synaptic dysfunction in AD. Our global gene expression studies have revealed an up-regulation of mitochondrial genes in AD transgenic mice (Tg2576 line) suggesting that mitochondrial metabolism is impaired by Abeta and that the up-regulation of mitochondrial genes may be compensatory response to Abeta (Reddy et
al 2004). We found Abeta monomers and oligomers in isolated mitochondria from the cortex of Tg2576 mice, and in isolated mitochondria from mouse neuroblastoma (N2a) cells expressing mutant APP (Manczak et al 2006). Our digitonin fractionation analysis of isolated mitochondria from Tg2576 mice and APP cells revealed Abeta in mitochondrial membranes and matrix, and that mitochondrial Abeta decreases cytochrome oxidase and increases H2O2 production and carbonyl proteins. These findings suggest that early mitochondrial targeted interventions may be effective in delaying AD progression. Methods: Recently, several mitochondrial targeted antioxidants (MTAs) including, MitoQ, MitoVitE and several small and peptide-based, antioxidants have been developed. These targeted antioxidants enter the mitochondrial membranes several hundred folds more than regular antioxidants, neutralize free radicals, inhibit mitochondrial permeability transition, and protect neurons. To determine whether MTAs reduces free radicals, oxidative damage and protect neurons from oxidative stress, we investigated the efficacies of MitoQ and SS31 in cell and AD mouse models. Results: Our preliminary studies of N2a cells grown in serum free media treated with MitoQ and SS31 have shown increased neurite outgrowth and increased expression of neuroprotective genes indicating that MitoQ and SS31 may have a neuroprotective role. We found decreased H2O2 production, cytochrome oxidase, and normal cellular ATP levels when N2a cells incubated with Abeta peptide 25-35 and pretreated with MitoQ and SS31 compared to N2a cells incubated with Abeta peptide alone. Further, we found neurite outgrowth is maintained in cells treated with MitoQ and SS31 and Abeta peptide. Conclusions: These findings suggest that MTAs may protect neurons from Abeta toxicity and oxidative damage. Our ongoing studies of Tg2576 mice treated with SS31 and MitoQ may provide new insights in understanding the role of MTAs in delaying AD progression and pathogenesis in Tg2576 mice. P4-170
PEROXIDATIVE NITROTYROSINE FORMATION IN ALZHEIMER’S DISEASE
George Perry1, Adam D. Cash2, Peggy L. Harris2, Sandra L. Siedlak2, Mark A. Smith2, Xiongwei Zhu2, 1University of Texas at San Antonio, San Antonio, TX, USA; 2Case Western Reserve University, Cleveland, OH, USA. Contact e-mail: [email protected] Background: Findings from numerous approaches in Alzheimer disease (AD) implicate oxidative stress as a major pathogenic pathway. We have focused our studies to elucidate the nature of the reactive species and sources of oxidative insult by considering cellular biomacromolecule modifications such as carbonyl and nitrotyrosine formation. These studies have strongly implicated metal catalyzed oxidation as critical to oxidative damage in AD. This led us to re-examine our prior work implicating non-metal catalyzed oxidation through peroxynitrite. Methods: In this study, we examine whether peroxynitrite or peroxidative nitration with nitrite and hydrogen peroxide is responsible for tyrosine nitration. Results: We found that peroxidative nitration yields nitrotyrosine in tissue with the characteristic cytoplasmic and nuclear distribution found in AD. Significantly, the process is dependent on redox active transition metals. In contrast, peroxynitrite produced lesswidespread damage without the anatomic signature of AD. Conclusions: This study suggests nitration of proteins in neurons in AD is in fact dependent on site directed metal-catalyzed processes, further evidence for the significant role of metals in the process of neurodegeneration. It is our conclusion that a reaction involving nitrite, hydrogen peroxide and the redox active metals shown to be elevated in AD, may play the primary roles of oxidative nitrotyrosine damage in AD. To truly target the in vivo mechanisms occurring in neurons in the brain, a complete picture of all nitrative events and consequences must be known. At the least, increasing awareness of the multiple possible pathways leading to the endpoints classified as nitrative damage will only increase the chances for therapeutic success.