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• 11. [Article] Influence of aging and behavioral experience on expression of GluN1 splice variants of the NMDA receptor in prefrontal cortex of mice brain

  As the aging population continues to grow the world over, age related complications become more and more apparent among the elderly population. One such complication is age associated memory impairment, ...

  Citation × Citation Citation Abstract Copy Title: Influence of aging and behavioral experience on expression of GluN1 splice variants of the NMDA receptor in prefrontal cortex of mice brain Author: Das, Siba Ranjan As the aging population continues to grow the world over, age related complications become more and more apparent among the elderly population. One such complication is age associated memory impairment, which makes the elderly more dependent on caregivers early on. NMDA receptors in the brain are important for memory formation, consolidation and retrieval. Expression of NMDA receptors declines with age, which is associated with declines in memory observed during aging. Age-related changes in the protein and mRNA expression of some of the splice forms of the GluN1 (GluN1, NR1) subunit of the NMDA receptor have been seen in mice and rats. The present study was designed to determine whether individual splice forms of the GluN1 subunit of the NMDA receptor within prefrontal / frontal cortical regions contribute to memory deficits during aging and whether experience in learning tasks can influence the expression of the splice forms. mRNA expression of 4 splice forms GluN1[subscript X11], GluN1[subscript X10], GluN1[subscript 0XX] and GluN1[subscript 1XX] (GluN1-1, GluN1-3, GluN1-a and GluN1-b, respectively) and mRNA for all known splice forms (GluN1-pan) were examined by in situ hybridization. mRNA for the C-terminal splice forms, GluN1[subscript X11] (GluN1-1; +C1 and +C2 cassettes) and GluN1[subscript X10] (GluN1-3; +C1 and +C2'), showed significant declines during aging in several brain regions even though overall GluN1-pan mRNA expression was not significantly affected by aging. This work provides evidence that these splice forms are more influenced by aging than the subunit as a whole. There was an increase in the expression of GluN1[subscript 0XX] (GluN1-a; -N1 cassette) splice form in the behaviorally-experienced old mice relative to the younger groups. Old mice with the highest levels of mRNA expression for the GluN1[subscript 0XX] (GluN1-a) splice form in orbital cortex showed the best performances in spatial working and reference memory tasks, but the poorest performances in a cued, associative learning task. These results suggest that the GluN1[subscript 0XX] subunit splice variant may be important for spatial memory performance in the old animals. Protein expression of GluN1 subunits containing C-terminal cassettes C2 or C2' were observed to decline with increasing age, regardless of experience. In middle-age animals, higher expressions of the GluN1 subunit and C2' cassette proteins were associated with good reference memory on initial search. In the aged animals, higher protein expression of GluN1 subunits containing C1 cassettes and the whole population of GluN1 subunits were found to be associated with better performance in the final phase of probe trials but this appeared to be due to perseveration or delays in applying an accurate search. These results provide support for the theory that there is heterogeneity in the effect of aging on the expression of the GluN1 subunits containing different splice cassettes. It also suggests that the GluN1 subunit might be most important for good reference memory during middle age. The next study was undertaken to determine if the GluN1[subscript 0XX] splice form is required for good performance in reference memory tasks in young mice. Mice were injected with 5µl of either siRNA specific to GluN1[subscript 0XX], control siRNA or vehicle alone into ventro-lateral orbital regions of both sides of the brain using a stereotaxic apparatus. A fourth group of mice did not receive any injections. Five days post-injection, mice were tested for their performance in a spatial reference memory task for 4 days using the Morris water maze. There was a 10 -19% reduction in GluN1[subscript 0XX] splice variant expression for mice after siRNA treatment in ventro-lateral orbital regions of the brain. Decline in performance in the first half of reference memory were observed in the mice receiving siRNA specific for GluN1[subscript 0XX] splice form, as compared to the mice injected with control siRNA and/or vehicle. These results suggest an important role of the GluN1[subscript 0XX] splice variant in orbital regions for spatial reference memory acquisition and/or consolidation in the early stages of memory training. These results suggest that there is a complex interaction between GluN1 splice form expression and performance of memory tasks during aging. Future studies designed to differentiate between involvement of splice variants in particular stages of memory formation would be helpful. Title: Influence of aging and behavioral experience on expression of GluN1 splice variants of the NMDA receptor in prefrontal cortex of mice brain Author: Das, Siba Ranjan As the aging population continues to grow the world over, age related complications become more and more apparent among the elderly population. One such complication is age associated memory impairment, which makes the elderly more dependent on caregivers early on. NMDA receptors in the brain are important for memory formation, consolidation and retrieval. Expression of NMDA receptors declines with age, which is associated with declines in memory observed during aging. Age-related changes in the protein and mRNA expression of some of the splice forms of the GluN1 (GluN1, NR1) subunit of the NMDA receptor have been seen in mice and rats. The present study was designed to determine whether individual splice forms of the GluN1 subunit of the NMDA receptor within prefrontal / frontal cortical regions contribute to memory deficits during aging and whether experience in learning tasks can influence the expression of the splice forms. mRNA expression of 4 splice forms GluN1[subscript X11], GluN1[subscript X10], GluN1[subscript 0XX] and GluN1[subscript 1XX] (GluN1-1, GluN1-3, GluN1-a and GluN1-b, respectively) and mRNA for all known splice forms (GluN1-pan) were examined by in situ hybridization. mRNA for the C-terminal splice forms, GluN1[subscript X11] (GluN1-1; +C1 and +C2 cassettes) and GluN1[subscript X10] (GluN1-3; +C1 and +C2'), showed significant declines during aging in several brain regions even though overall GluN1-pan mRNA expression was not significantly affected by aging. This work provides evidence that these splice forms are more influenced by aging than the subunit as a whole. There was an increase in the expression of GluN1[subscript 0XX] (GluN1-a; -N1 cassette) splice form in the behaviorally-experienced old mice relative to the younger groups. Old mice with the highest levels of mRNA expression for the GluN1[subscript 0XX] (GluN1-a) splice form in orbital cortex showed the best performances in spatial working and reference memory tasks, but the poorest performances in a cued, associative learning task. These results suggest that the GluN1[subscript 0XX] subunit splice variant may be important for spatial memory performance in the old animals. Protein expression of GluN1 subunits containing C-terminal cassettes C2 or C2' were observed to decline with increasing age, regardless of experience. In middle-age animals, higher expressions of the GluN1 subunit and C2' cassette proteins were associated with good reference memory on initial search. In the aged animals, higher protein expression of GluN1 subunits containing C1 cassettes and the whole population of GluN1 subunits were found to be associated with better performance in the final phase of probe trials but this appeared to be due to perseveration or delays in applying an accurate search. These results provide support for the theory that there is heterogeneity in the effect of aging on the expression of the GluN1 subunits containing different splice cassettes. It also suggests that the GluN1 subunit might be most important for good reference memory during middle age. The next study was undertaken to determine if the GluN1[subscript 0XX] splice form is required for good performance in reference memory tasks in young mice. Mice were injected with 5µl of either siRNA specific to GluN1[subscript 0XX], control siRNA or vehicle alone into ventro-lateral orbital regions of both sides of the brain using a stereotaxic apparatus. A fourth group of mice did not receive any injections. Five days post-injection, mice were tested for their performance in a spatial reference memory task for 4 days using the Morris water maze. There was a 10 -19% reduction in GluN1[subscript 0XX] splice variant expression for mice after siRNA treatment in ventro-lateral orbital regions of the brain. Decline in performance in the first half of reference memory were observed in the mice receiving siRNA specific for GluN1[subscript 0XX] splice form, as compared to the mice injected with control siRNA and/or vehicle. These results suggest an important role of the GluN1[subscript 0XX] splice variant in orbital regions for spatial reference memory acquisition and/or consolidation in the early stages of memory training. These results suggest that there is a complex interaction between GluN1 splice form expression and performance of memory tasks during aging. Future studies designed to differentiate between involvement of splice variants in particular stages of memory formation would be helpful. Close

• 12. [Article] A role for ceramide accumulation in age-related cardiac mitochondrial dysfunction

  Ceramides are a group of lipids in the sphingolipid family that are potent cell signaling molecules. Elevations in ceramide levels above the norm generally lead to apoptosis. Evidence from in vitro studies ...

  Citation × Citation Citation Abstract Copy Title: A role for ceramide accumulation in age-related cardiac mitochondrial dysfunction Author: Monette, Jeffrey S. Ceramides are a group of lipids in the sphingolipid family that are potent cell signaling molecules. Elevations in ceramide levels above the norm generally lead to apoptosis. Evidence from in vitro studies suggests that accumulation of ceramides within mitochondria leads to dysfunction of the mitochondria. This includes inhibition of the electron transport chain and release of reactive oxygen species (ROS). Large elevations of mitochondrial ceramide may also lead to death of the cell as they promote membrane permeability transition, which allows the release of cytochrome c and other apoptogenic factors. With age, cardiac mitochondria show a similar dysfunctional phenotype to that found in conditions of acute ceramide elevation. We therefore hypothesize that ceramides may be accumulating in cardiac mitochondria from aged animals. To elucidate the characteristics of the sphingolipidome, we developed both mitochondrial isolation techniques and tandem mass spectrometry assays to specifically and sensitively monitor mitochondrial sphingolipids. Using these techniques, it was found that mitochondria contain six distinct ceramide species with highly saturated lipid moieties. Using young (3-6 months old; which corresponds to a post-adolescent human) and old (24 to 28 months old; which corresponds to an elderly human) Fischer 344 rat hearts, we found that aging leads to a significant increase in total mitochondrial ceramides (32%, p < 0.03), with C₁₆-, C₁₈-, and C₂₄:₁-ceramides showing the largest percent increases (72.3%, 73.4%, and 77.7%, respectively, p < 0.05). Furthermore, the age-associated elevation in ceramide levels correlated to a 28% decrease in the activity of complex IV of the electron transport chain (p < 0.05), which could be replicated in vitro by inducing a ceramide accumulation in mitochondria isolated from young animals. Mitochondria do not contain enzymes for de novo ceramide biosynthesis, rather, these organelles have sphingomyelinases, a family of enzymes that cleave the phosphorylcholine headgroup from nascent pools of sphingomyelin. Specifically, mitochondria contain the magnesium-requiring isoform of sphingomyelinase with a neutral pH optima (nSMase). Recent work has shown that nSMase activity is inversely regulated by glutathione status. Because cardiac mitochondrial glutathione (mGSH) declines by up to 60% with age, we hypothesized that the loss in mGSH leads to an increase in ceramides through the upregulation of nSMase. To determine whether loss of mGSH plays a role in the regulation of mitochondrial nSMase activity, mGSH levels were depleted by treating freshly isolated hepatocytes with 3-hydroxy-4-pentenoate (3HP). It was found that 3HP rapidly depleted mGSH in a concentration-dependent manner (EC₅₀ = 232 μM, p < 0.05). Moreover, this depletion led to an increase in nSMase activity (24 ± 3% at 250 μM 3HP, p < 0.05), and an increase in total ceramide levels (27%, p < 0.05). These findings suggest that mGSH status plays a critical role in the maintenance of ceramide levels within mitochondria. Furthermore, because nSMase activity is regulated by mGSH levels, we hypothesized that any agent promoting an increase in mGSH would reverse the ceramide accumulation seen in cardiac mitochondria from aged animals. Lipoic acid (LA) is a naturally occurring dithiol compound used for many years as an anti-inflammatory agent. LA-supplementation has been shown to increase cellular and mGSH by increasing cellular cysteine levels in the aging heart. In order to determine whether LA reverses the age-associated ceramidosis in cardiac mitochondria, young and old F344 rats were pair-fed LA [0.2% (w/w) in the diet] against controls for two weeks and cardiac mitochondria were subsequently isolated and analyzed. It was found that LA-treatment reversed the age-associated decline in mGSH levels [decreased 43% with age (p < 0.05)], and reduced nSMase activity [increased 103% with age (p < 0.05)]. Ceramide levels were reduced [elevated 32% with age (p < 0.03)] so that they were no longer different from young controls and complex IV activity restored t youthful levels [declined 28% with age (p < 0.05)]. In conclusion, this dissertation provides evidence to support a new mechanism that explains, at least in part, the progression of mitochondrial dysfunction in the aging heart, and may also contribute to understanding the age-related loss of cardiomyocytes. It also provides mechanistic insights into the overall health benefits of LA supplementation and supports its use as a safe, natural, and "age-essential" micronutrient. Title: A role for ceramide accumulation in age-related cardiac mitochondrial dysfunction Author: Monette, Jeffrey S. Ceramides are a group of lipids in the sphingolipid family that are potent cell signaling molecules. Elevations in ceramide levels above the norm generally lead to apoptosis. Evidence from in vitro studies suggests that accumulation of ceramides within mitochondria leads to dysfunction of the mitochondria. This includes inhibition of the electron transport chain and release of reactive oxygen species (ROS). Large elevations of mitochondrial ceramide may also lead to death of the cell as they promote membrane permeability transition, which allows the release of cytochrome c and other apoptogenic factors. With age, cardiac mitochondria show a similar dysfunctional phenotype to that found in conditions of acute ceramide elevation. We therefore hypothesize that ceramides may be accumulating in cardiac mitochondria from aged animals. To elucidate the characteristics of the sphingolipidome, we developed both mitochondrial isolation techniques and tandem mass spectrometry assays to specifically and sensitively monitor mitochondrial sphingolipids. Using these techniques, it was found that mitochondria contain six distinct ceramide species with highly saturated lipid moieties. Using young (3-6 months old; which corresponds to a post-adolescent human) and old (24 to 28 months old; which corresponds to an elderly human) Fischer 344 rat hearts, we found that aging leads to a significant increase in total mitochondrial ceramides (32%, p < 0.03), with C₁₆-, C₁₈-, and C₂₄:₁-ceramides showing the largest percent increases (72.3%, 73.4%, and 77.7%, respectively, p < 0.05). Furthermore, the age-associated elevation in ceramide levels correlated to a 28% decrease in the activity of complex IV of the electron transport chain (p < 0.05), which could be replicated in vitro by inducing a ceramide accumulation in mitochondria isolated from young animals. Mitochondria do not contain enzymes for de novo ceramide biosynthesis, rather, these organelles have sphingomyelinases, a family of enzymes that cleave the phosphorylcholine headgroup from nascent pools of sphingomyelin. Specifically, mitochondria contain the magnesium-requiring isoform of sphingomyelinase with a neutral pH optima (nSMase). Recent work has shown that nSMase activity is inversely regulated by glutathione status. Because cardiac mitochondrial glutathione (mGSH) declines by up to 60% with age, we hypothesized that the loss in mGSH leads to an increase in ceramides through the upregulation of nSMase. To determine whether loss of mGSH plays a role in the regulation of mitochondrial nSMase activity, mGSH levels were depleted by treating freshly isolated hepatocytes with 3-hydroxy-4-pentenoate (3HP). It was found that 3HP rapidly depleted mGSH in a concentration-dependent manner (EC₅₀ = 232 μM, p < 0.05). Moreover, this depletion led to an increase in nSMase activity (24 ± 3% at 250 μM 3HP, p < 0.05), and an increase in total ceramide levels (27%, p < 0.05). These findings suggest that mGSH status plays a critical role in the maintenance of ceramide levels within mitochondria. Furthermore, because nSMase activity is regulated by mGSH levels, we hypothesized that any agent promoting an increase in mGSH would reverse the ceramide accumulation seen in cardiac mitochondria from aged animals. Lipoic acid (LA) is a naturally occurring dithiol compound used for many years as an anti-inflammatory agent. LA-supplementation has been shown to increase cellular and mGSH by increasing cellular cysteine levels in the aging heart. In order to determine whether LA reverses the age-associated ceramidosis in cardiac mitochondria, young and old F344 rats were pair-fed LA [0.2% (w/w) in the diet] against controls for two weeks and cardiac mitochondria were subsequently isolated and analyzed. It was found that LA-treatment reversed the age-associated decline in mGSH levels [decreased 43% with age (p < 0.05)], and reduced nSMase activity [increased 103% with age (p < 0.05)]. Ceramide levels were reduced [elevated 32% with age (p < 0.03)] so that they were no longer different from young controls and complex IV activity restored t youthful levels [declined 28% with age (p < 0.05)]. In conclusion, this dissertation provides evidence to support a new mechanism that explains, at least in part, the progression of mitochondrial dysfunction in the aging heart, and may also contribute to understanding the age-related loss of cardiomyocytes. It also provides mechanistic insights into the overall health benefits of LA supplementation and supports its use as a safe, natural, and "age-essential" micronutrient. Close

• 13. [Article] Perceptions of residential grab bars among community dwelling seniors

  Despite the perceived importance of grab bars to facilitate aging-in-place and healthy aging, many community-dwelling older adults do not have them installed. The aim of this study was to investigate predictors ...

  Citation × Citation Citation Abstract Copy Title: Perceptions of residential grab bars among community dwelling seniors Author: Thrall, Patti L. Despite the perceived importance of grab bars to facilitate aging-in-place and healthy aging, many community-dwelling older adults do not have them installed. The aim of this study was to investigate predictors of grab bar installation among well-educated community dwelling seniors. Data was collected quantitatively through an electronic survey of Oregon residents 50 years of age and older. The research analysis was completed using logistic regression with SPSS and qualitative analysis for the open questions. Title: Perceptions of residential grab bars among community dwelling seniors Author: Thrall, Patti L. Despite the perceived importance of grab bars to facilitate aging-in-place and healthy aging, many community-dwelling older adults do not have them installed. The aim of this study was to investigate predictors of grab bar installation among well-educated community dwelling seniors. Data was collected quantitatively through an electronic survey of Oregon residents 50 years of age and older. The research analysis was completed using logistic regression with SPSS and qualitative analysis for the open questions. Close

• 14. [Article] Disengagement of older people in an urban setting

  This dissertation reevaluates the controversial Disengagement Theory of aging introduced by Elaine Cumming and William E. Henry in their book, Growing Old (1961), with particular emphasis on aging in the ...

  Citation × Citation Citation Abstract Copy Title: Disengagement of older people in an urban setting Author: Somers, Melvina Year: 1977 This dissertation reevaluates the controversial Disengagement Theory of aging introduced by Elaine Cumming and William E. Henry in their book, Growing Old (1961), with particular emphasis on aging in the city. Disengagement theory embodies the antithesis to the traditional activity theory of aging based on a work ethic. Disengagement theory holds that normal aging is an inevitable mutual withdrawal or disengagement between the aging person and others in the social system, that it is universal, and that, by impl ication,it is a mutually satisfying condition since it results in a new equilibrium between society and the aging individual. The theory proposes that withdrawal happens automatically in the aging person because of ego changes which are "programmed" into the human organism. Activity theory, on the contrary, claims that maintaining physical, mental, or social activity is a prerequisite to successful aging. It implies that a meaningful social role is necessary to selfesteem, and that, allowing for biological impairments, the psychological needs of older people are not much different from those of middle-aged persons. Since the United States population aged 65 and over has a net annual increase of over 300,000 people, the questions as to whether they tend to be engaged or disengaged, and whether they should be integrated in or separated from the rest of society, become very important. According to the 1970 Census, over sixty percent of persons sixty-five and over live in metropolitan areas. Therefore their housing, transportation, medical care, and other needs have to be considered in urban policies. In planning services for older people it is imperative to know what kind of services are needed, and hence engagement or disengagement becomes a crucial question. It is the assumption in this research that the city and its older population have an investment in one another, and that the city is interested in the welfare of its older citizens. Theoretically, the study was guided by the principles of symbolic interactionism. Several methods were used in the approach to the research problem because it is believed that what is known as a "triangulated" perspective can come closest to revealing the various aspects of empirical reality. Specifically, the methods employed were the social survey, the depth interview, participant observation, unobtrusive observation, and the case study and life history technique. This research has used especially Weber's method of Verstehen or empathic unders tandi rig. Three hypotheses guided the research: Hypothesis One: Disengagement is not an intrinsic or inevitable phenomenon. If it occurs, it can be traced to various causes other than aging per se, such as ill health, personal misfortunes (such as widowhood), or social pressures (such as forced retirement). Hypthesis Two: Older people do not enjoy a disengaged state. There is continuity in personality and life style, and if older people choose nonengagement, it is a continuation of former habits. Hypothesis Three: A livable urban environment is a determinant in the life satisfaction of the aged. These hypotheses were tested empirically, using both qualitative and quantitative data; and our hypotheses have been confirmed by prevalence and recurrence of expected patterns in the behavior of our samples. The use of the inductive method has given consistent results in that no instances of intrinsic disengagement were discovered. We have been able to show that other causal factors were responsible for relative nonengagement if it occurred. The dissertation also reports many suggestions which were received on how the urban environment could be improved and how the city could help older people lead better lives. Title: Disengagement of older people in an urban setting Author: Somers, Melvina Year: 1977 This dissertation reevaluates the controversial Disengagement Theory of aging introduced by Elaine Cumming and William E. Henry in their book, Growing Old (1961), with particular emphasis on aging in the city. Disengagement theory embodies the antithesis to the traditional activity theory of aging based on a work ethic. Disengagement theory holds that normal aging is an inevitable mutual withdrawal or disengagement between the aging person and others in the social system, that it is universal, and that, by impl ication,it is a mutually satisfying condition since it results in a new equilibrium between society and the aging individual. The theory proposes that withdrawal happens automatically in the aging person because of ego changes which are "programmed" into the human organism. Activity theory, on the contrary, claims that maintaining physical, mental, or social activity is a prerequisite to successful aging. It implies that a meaningful social role is necessary to selfesteem, and that, allowing for biological impairments, the psychological needs of older people are not much different from those of middle-aged persons. Since the United States population aged 65 and over has a net annual increase of over 300,000 people, the questions as to whether they tend to be engaged or disengaged, and whether they should be integrated in or separated from the rest of society, become very important. According to the 1970 Census, over sixty percent of persons sixty-five and over live in metropolitan areas. Therefore their housing, transportation, medical care, and other needs have to be considered in urban policies. In planning services for older people it is imperative to know what kind of services are needed, and hence engagement or disengagement becomes a crucial question. It is the assumption in this research that the city and its older population have an investment in one another, and that the city is interested in the welfare of its older citizens. Theoretically, the study was guided by the principles of symbolic interactionism. Several methods were used in the approach to the research problem because it is believed that what is known as a "triangulated" perspective can come closest to revealing the various aspects of empirical reality. Specifically, the methods employed were the social survey, the depth interview, participant observation, unobtrusive observation, and the case study and life history technique. This research has used especially Weber's method of Verstehen or empathic unders tandi rig. Three hypotheses guided the research: Hypothesis One: Disengagement is not an intrinsic or inevitable phenomenon. If it occurs, it can be traced to various causes other than aging per se, such as ill health, personal misfortunes (such as widowhood), or social pressures (such as forced retirement). Hypthesis Two: Older people do not enjoy a disengaged state. There is continuity in personality and life style, and if older people choose nonengagement, it is a continuation of former habits. Hypothesis Three: A livable urban environment is a determinant in the life satisfaction of the aged. These hypotheses were tested empirically, using both qualitative and quantitative data; and our hypotheses have been confirmed by prevalence and recurrence of expected patterns in the behavior of our samples. The use of the inductive method has given consistent results in that no instances of intrinsic disengagement were discovered. We have been able to show that other causal factors were responsible for relative nonengagement if it occurred. The dissertation also reports many suggestions which were received on how the urban environment could be improved and how the city could help older people lead better lives. Close

• 15. [Article] Effects of Aging on the Molecular Circadian Oscillations in Drosophila

  This is the author's peer-reviewed final manuscript, as accepted by the publisher. The published article is copyrighted by Informa Healthcare USA, Inc. and can be found at: http://informahealthcare.com...

  Citation × Citation Citation Abstract Copy Title: Effects of Aging on the Molecular Circadian Oscillations in Drosophila Author: Guzik, Elzbieta M., Giebultowicz, Jadwiga M., Rakshit, Kuntol, Krishnan, Natraj, Pyza, Elzbieta This is the author's peer-reviewed final manuscript, as accepted by the publisher. The published article is copyrighted by Informa Healthcare USA, Inc. and can be found at: http://informahealthcare.com/cbi. Title: Effects of Aging on the Molecular Circadian Oscillations in Drosophila Author: Guzik, Elzbieta M., Giebultowicz, Jadwiga M., Rakshit, Kuntol, Krishnan, Natraj, Pyza, Elzbieta This is the author's peer-reviewed final manuscript, as accepted by the publisher. The published article is copyrighted by Informa Healthcare USA, Inc. and can be found at: http://informahealthcare.com/cbi. Close

• 16. [Article] A Comparative Analysis of Rodent Species Shows Oxidative Stress Resilience and HSP70 Expression Levels Correlate with Species Longevity

  Researchers of healthy aging have determined trends between a species’ body mass and its maximal life span (MLSP). Actual MLSPs are then compared to predicted MLSPs and reported as a longevity quotient ...

  Citation × Citation Citation Abstract Copy Title: A Comparative Analysis of Rodent Species Shows Oxidative Stress Resilience and HSP70 Expression Levels Correlate with Species Longevity Author: Sarker, Minhazur R. Researchers of healthy aging have determined trends between a species’ body mass and its maximal life span (MLSP). Actual MLSPs are then compared to predicted MLSPs and reported as a longevity quotient (LQ). Long-lived mammals, such as humans and naked mole rats (NMR), have LQs ranging from 5.0 to 10.0, meaning they live 5 to 10 times longer than expected. This LQ similarity leads researchers to believe the NMR may be an ideal model organism for human aging research. Among the proposed theories of aging, the oxidative stress theory, which states that cells have a chronic presence of oxidative stress that increases with age, is one of the most widely accepted. Our work delves deeper into this theory as it pertains to the NMR. We observed that NMR fibroblasts (NMRFs) have a much greater resilience to menadione, an oxidative stressor, than mouse fibroblasts (MFs), which, under the oxidative stress theory of aging, may explain why NMR have delayed onsets of aging phenotypes. We then observed that compared to MFs and rat hepatocytes, NMRFs have a greater basal expression of heat-­shock protein 70 (HSP70), a primary role player in homeostatic responses to oxidative stress. This greater HSP70 expression may grant the NMRFs a stronger initial defense allowing them to maintain viability at greater levels of oxidative stress. Finally, our work investigated the use of geranylgeranyl acetone (GGA) as an HSP70 inducer to slow the insults from oxidative stress. We were not able to show GGA as an efficient HSP70 inducer. Ultimately, our work has discovered some key aspects of the naked mole rat, however, further investigation is needed to determine if naked mole rats may serve as the ideal model species for human healthy aging research. Title: A Comparative Analysis of Rodent Species Shows Oxidative Stress Resilience and HSP70 Expression Levels Correlate with Species Longevity Author: Sarker, Minhazur R. Researchers of healthy aging have determined trends between a species’ body mass and its maximal life span (MLSP). Actual MLSPs are then compared to predicted MLSPs and reported as a longevity quotient (LQ). Long-lived mammals, such as humans and naked mole rats (NMR), have LQs ranging from 5.0 to 10.0, meaning they live 5 to 10 times longer than expected. This LQ similarity leads researchers to believe the NMR may be an ideal model organism for human aging research. Among the proposed theories of aging, the oxidative stress theory, which states that cells have a chronic presence of oxidative stress that increases with age, is one of the most widely accepted. Our work delves deeper into this theory as it pertains to the NMR. We observed that NMR fibroblasts (NMRFs) have a much greater resilience to menadione, an oxidative stressor, than mouse fibroblasts (MFs), which, under the oxidative stress theory of aging, may explain why NMR have delayed onsets of aging phenotypes. We then observed that compared to MFs and rat hepatocytes, NMRFs have a greater basal expression of heat-­shock protein 70 (HSP70), a primary role player in homeostatic responses to oxidative stress. This greater HSP70 expression may grant the NMRFs a stronger initial defense allowing them to maintain viability at greater levels of oxidative stress. Finally, our work investigated the use of geranylgeranyl acetone (GGA) as an HSP70 inducer to slow the insults from oxidative stress. We were not able to show GGA as an efficient HSP70 inducer. Ultimately, our work has discovered some key aspects of the naked mole rat, however, further investigation is needed to determine if naked mole rats may serve as the ideal model species for human healthy aging research. Close

• 17. [Article] Age-related loss of Nrf2, a novel mechanism for the potential attenuation of xenobiotic detoxification capacity

  Nuclear Factor, Erythroid Derived 2, Like 2 (NFE2L2 or Nrf2) is the primary transcription factor in cellular defense against oxidative and xenobiotic stresses in higher eukaryotes. This basic leucine zipper ...

  Citation × Citation Citation Abstract Copy Title: Age-related loss of Nrf2, a novel mechanism for the potential attenuation of xenobiotic detoxification capacity Author: Smith, Eric Jonathan Nuclear Factor, Erythroid Derived 2, Like 2 (NFE2L2 or Nrf2) is the primary transcription factor in cellular defense against oxidative and xenobiotic stresses in higher eukaryotes. This basic leucine zipper transcription factor regulates over 200 antioxidant, detoxification, and lipid metabolizing genes by binding to the Antioxidant Response Element (ARE; a cis-acting element) as a transcription enhancer. Our group, and others, have identified that the nuclear steady state levels of this highly inducible transcription factor declines with age which coincides with decreased expression of many ARE regulated genes. To elucidate the mechanism(s) involved in lowered Nrf2 levels we used hepatocytes isolated from young and old rats as a relevant cellular model that maintains the aging phenotype with respect to Nrf2 levels. Our results show that steady state Nrf2 levels decline by ~40% with age (N=4, p<0.05), which led us to investigate the multifaceted regulation of Nrf2 homeostasis. Specifically, analysis of Nrf2 mRNA levels, and the rate of Nrf2 protein turnover and translation were investigated. Surprisingly, Keap1-mediated degradation of Nrf2, its primary negative regulator, did not account for the age-related Nrf2 decline, nor did mRNA levels change with age. Rather, Nrf2 protein synthesis declines 5.3-fold with age (N=3, p<0.05). Furthermore, we identify that rno-miR-146a, a key inflammation regulating miRNA, both inhibits Nrf2 translation and increases significantly with age. Taken together, our data suggest that the age-related loss of Nrf2 stems from the loss of its translation and may be mediated through miR regulation. Our lab previously demonstrated in rats, that the age-related loss of Nrf2 consequentially attenuates synthesis and steady state levels of liver glutathione (GSH) synthesis and manifests as a diminished level of GSH. This antioxidant is the most abundant non-protein thiol in most cells and is pivotal for the detoxification of many xenobiotics. The rate-limiting enzyme in GSH synthesis, γ-glutamyl-cysteine ligase (Gcl), is composed of a catalytic (Gclc), and a modulatory (Gclm) subunit. In mice and humans, it is established that Nrf2, through the ARE, regulates Gclc expression. However, this had not been established in rats. From a bioinformatics analysis of the 5’ upstream sequence of the rat Gclc gene, we identified 3 putative AREs (ARE1, ARE3, and ARE4), and one cis-element containing the core but not the flanking nucleotides of the ARE (ARE2). Luciferase reporter plasmids containing each individual ARE were transfected into H4IIE rat hepatoma cells to test for ARE activity. Only one, designated “ARE4”, produced appreciable luciferase activity and was dependent on Nrf2 expression, suggesting that this ARE is a “true” enhancer element. Chromatin immunoprecipitation (ChIP) and qPCR identified that Nrf2 binds ARE4 but not ARE1-3. Further ChIP analysis identified that known partner transcription factors of Nrf2 (small maf, c-Jun and c-Fos) were also bound to ARE4. Taken together, our data demonstrates that Nrf2 regulates, in part, the expression of the rat Gclc gene through ARE4. Having identified the Gclc-AREs in the rat we sought to describe the molecular consequence of the age-related decrement of Nrf2 in hepatocytes isolated from young and old rats. ChIPs and reporter assays (ARE1-4) were used to investigate known ARE-binding transcription factors and the ARE mediated enhancement of Gclc transcription. We identified a decline in Nrf2 occupancy of ARE4 (40%) and a loss of transcriptional activity (70%; P < 0.05), consistent with the loss in Gclc levels. In addition, hepatic chromatin from old rat samples demonstrated ARE4 enrichment of the transcriptional repressor Bach1, and a loss of the histone acetyltransferase CREB-Binding Protein (CBP). These results demonstrate that the active Gclc transcriptional complex is remodeled into a repressive motif in the liver of old rats. Furthermore, ChIPs identified that Nrf2 binds ARE2 in liver lysate isolated from old rats but not young. In agreement with this finding, results from luciferase reporter assays suggest that ARE2 facilitates an Nrf2-dependent enhancement of Gclc expression in the old. Thus, a promoter switching mechanism may occur with age. In summary, we have identified key mechanisms of how the age-related decrement in Nrf2 protein levels is controlled and also gathered preliminary evidence that a pro-inflammatory state may contribute the loss of Nrf2 synthesis via increased miR levels. Finally, we have characterized the consequences of the loss of Nrf2 to GSH synthesis as an example of age-related impairment of cellular stress response. Title: Age-related loss of Nrf2, a novel mechanism for the potential attenuation of xenobiotic detoxification capacity Author: Smith, Eric Jonathan Nuclear Factor, Erythroid Derived 2, Like 2 (NFE2L2 or Nrf2) is the primary transcription factor in cellular defense against oxidative and xenobiotic stresses in higher eukaryotes. This basic leucine zipper transcription factor regulates over 200 antioxidant, detoxification, and lipid metabolizing genes by binding to the Antioxidant Response Element (ARE; a cis-acting element) as a transcription enhancer. Our group, and others, have identified that the nuclear steady state levels of this highly inducible transcription factor declines with age which coincides with decreased expression of many ARE regulated genes. To elucidate the mechanism(s) involved in lowered Nrf2 levels we used hepatocytes isolated from young and old rats as a relevant cellular model that maintains the aging phenotype with respect to Nrf2 levels. Our results show that steady state Nrf2 levels decline by ~40% with age (N=4, p<0.05), which led us to investigate the multifaceted regulation of Nrf2 homeostasis. Specifically, analysis of Nrf2 mRNA levels, and the rate of Nrf2 protein turnover and translation were investigated. Surprisingly, Keap1-mediated degradation of Nrf2, its primary negative regulator, did not account for the age-related Nrf2 decline, nor did mRNA levels change with age. Rather, Nrf2 protein synthesis declines 5.3-fold with age (N=3, p<0.05). Furthermore, we identify that rno-miR-146a, a key inflammation regulating miRNA, both inhibits Nrf2 translation and increases significantly with age. Taken together, our data suggest that the age-related loss of Nrf2 stems from the loss of its translation and may be mediated through miR regulation. Our lab previously demonstrated in rats, that the age-related loss of Nrf2 consequentially attenuates synthesis and steady state levels of liver glutathione (GSH) synthesis and manifests as a diminished level of GSH. This antioxidant is the most abundant non-protein thiol in most cells and is pivotal for the detoxification of many xenobiotics. The rate-limiting enzyme in GSH synthesis, γ-glutamyl-cysteine ligase (Gcl), is composed of a catalytic (Gclc), and a modulatory (Gclm) subunit. In mice and humans, it is established that Nrf2, through the ARE, regulates Gclc expression. However, this had not been established in rats. From a bioinformatics analysis of the 5’ upstream sequence of the rat Gclc gene, we identified 3 putative AREs (ARE1, ARE3, and ARE4), and one cis-element containing the core but not the flanking nucleotides of the ARE (ARE2). Luciferase reporter plasmids containing each individual ARE were transfected into H4IIE rat hepatoma cells to test for ARE activity. Only one, designated “ARE4”, produced appreciable luciferase activity and was dependent on Nrf2 expression, suggesting that this ARE is a “true” enhancer element. Chromatin immunoprecipitation (ChIP) and qPCR identified that Nrf2 binds ARE4 but not ARE1-3. Further ChIP analysis identified that known partner transcription factors of Nrf2 (small maf, c-Jun and c-Fos) were also bound to ARE4. Taken together, our data demonstrates that Nrf2 regulates, in part, the expression of the rat Gclc gene through ARE4. Having identified the Gclc-AREs in the rat we sought to describe the molecular consequence of the age-related decrement of Nrf2 in hepatocytes isolated from young and old rats. ChIPs and reporter assays (ARE1-4) were used to investigate known ARE-binding transcription factors and the ARE mediated enhancement of Gclc transcription. We identified a decline in Nrf2 occupancy of ARE4 (40%) and a loss of transcriptional activity (70%; P < 0.05), consistent with the loss in Gclc levels. In addition, hepatic chromatin from old rat samples demonstrated ARE4 enrichment of the transcriptional repressor Bach1, and a loss of the histone acetyltransferase CREB-Binding Protein (CBP). These results demonstrate that the active Gclc transcriptional complex is remodeled into a repressive motif in the liver of old rats. Furthermore, ChIPs identified that Nrf2 binds ARE2 in liver lysate isolated from old rats but not young. In agreement with this finding, results from luciferase reporter assays suggest that ARE2 facilitates an Nrf2-dependent enhancement of Gclc expression in the old. Thus, a promoter switching mechanism may occur with age. In summary, we have identified key mechanisms of how the age-related decrement in Nrf2 protein levels is controlled and also gathered preliminary evidence that a pro-inflammatory state may contribute the loss of Nrf2 synthesis via increased miR levels. Finally, we have characterized the consequences of the loss of Nrf2 to GSH synthesis as an example of age-related impairment of cellular stress response. Close

• 18. [Article] Changes in gene expression related to the decline of the aging immune system

  In this study, the gene expression levels between 6 month old mice and 26 month old mice were compared. The purpose of this research was to identify genes that are differentially regulated due to aging. ...

  Citation × Citation Citation Abstract Copy Title: Changes in gene expression related to the decline of the aging immune system Author: Gombart, Adrian F., Tran, Jenny K. In this study, the gene expression levels between 6 month old mice and 26 month old mice were compared. The purpose of this research was to identify genes that are differentially regulated due to aging. The results showed 148 significantly differentiated genes in the analysis of a total of 44,170 genes with 16 genes found to be related to immunity and inflammation. Title: Changes in gene expression related to the decline of the aging immune system Author: Gombart, Adrian F., Tran, Jenny K. In this study, the gene expression levels between 6 month old mice and 26 month old mice were compared. The purpose of this research was to identify genes that are differentially regulated due to aging. The results showed 148 significantly differentiated genes in the analysis of a total of 44,170 genes with 16 genes found to be related to immunity and inflammation. Close

• 19. [Article] Building a State for All Ages: Tapping the Potential of Older Oregonians

  This report explores how the aging population—including native Oregonians and senior adults moving to the state—represent an asset to Oregon. Aging in America has often been viewed from a perspective of ...

  Citation × Citation Citation Abstract Copy Title: Building a State for All Ages: Tapping the Potential of Older Oregonians Author: Percy, Stephen, Ramaley, Judith A., Tapogna, John, Neal, Margaret B., Bloom, Jay, Reece, Alexandra Year: 2016 This report explores how the aging population—including native Oregonians and senior adults moving to the state—represent an asset to Oregon. Aging in America has often been viewed from a perspective of deficit and decline. We wholeheartedly challenge this perspective. This report identifies how older adults can play vital roles in the workforce, entrepreneurship and innovation, and supporting community revitalization through volunteering of time and talent. Title: Building a State for All Ages: Tapping the Potential of Older Oregonians Author: Percy, Stephen, Ramaley, Judith A., Tapogna, John, Neal, Margaret B., Bloom, Jay, Reece, Alexandra Year: 2016 This report explores how the aging population—including native Oregonians and senior adults moving to the state—represent an asset to Oregon. Aging in America has often been viewed from a perspective of deficit and decline. We wholeheartedly challenge this perspective. This report identifies how older adults can play vital roles in the workforce, entrepreneurship and innovation, and supporting community revitalization through volunteering of time and talent. Close

• 20. [Article] Ascorbic acid, lipid peroxidation, and aging

  The role of ascorbic acid with regard to lipid peroxidation and aging has been examined. A thorough literature analysis indicates that free radical-induced lipid peroxidation is a plausible biochemical ...

  Citation × Citation Citation Abstract Copy Title: Ascorbic acid, lipid peroxidation, and aging Author: Leibovitz, Brian Evan Year: 1979 The role of ascorbic acid with regard to lipid peroxidation and aging has been examined. A thorough literature analysis indicates that free radical-induced lipid peroxidation is a plausible biochemical explanation for aging. Lipid peroxidation causes cellular damage due to altered enzyme activities, error-prone nucleic acid metabolism, and membrane dysfunction, as well as the accumulation of aging pigments in the lysomes. Ascorbic acid, a water soluble free radical quencher, was examined with regard to carbon tetra-chloride-induced lipid peroxidation and in vivo aging. Carbon tetrachloride, a well-known free radical inducer, caused marked increases in the ration of oxidized/reduced vitamin C only in the organs which metabolize carbon tetrachloride to the free radical form. Vitamin C treatment, 250 mg% in the drinking water, reduced the extent of carbon tetrachloride=induced lipid peroxidation. Aging is associated with marked increases in the ration of oxidized/reduced vitamin C in all organs examined with the exception of thymus. In organs exposed to high oxygen tensions, or in those exposed to high levels of free radicals, the ration of oxidized/reduced vitamin C exceeded the in vitro ration, indicating extensive lipid peroxidation. Vitamin C treatment reduced the extent of lipid peroxidation in vivo as determined by the ratio of oxidized/reduced vitamin C. Title: Ascorbic acid, lipid peroxidation, and aging Author: Leibovitz, Brian Evan Year: 1979 The role of ascorbic acid with regard to lipid peroxidation and aging has been examined. A thorough literature analysis indicates that free radical-induced lipid peroxidation is a plausible biochemical explanation for aging. Lipid peroxidation causes cellular damage due to altered enzyme activities, error-prone nucleic acid metabolism, and membrane dysfunction, as well as the accumulation of aging pigments in the lysomes. Ascorbic acid, a water soluble free radical quencher, was examined with regard to carbon tetra-chloride-induced lipid peroxidation and in vivo aging. Carbon tetrachloride, a well-known free radical inducer, caused marked increases in the ration of oxidized/reduced vitamin C only in the organs which metabolize carbon tetrachloride to the free radical form. Vitamin C treatment, 250 mg% in the drinking water, reduced the extent of carbon tetrachloride=induced lipid peroxidation. Aging is associated with marked increases in the ration of oxidized/reduced vitamin C in all organs examined with the exception of thymus. In organs exposed to high oxygen tensions, or in those exposed to high levels of free radicals, the ration of oxidized/reduced vitamin C exceeded the in vitro ration, indicating extensive lipid peroxidation. Vitamin C treatment reduced the extent of lipid peroxidation in vivo as determined by the ratio of oxidized/reduced vitamin C. Close