Decline In Mitochondrial Function Research Articles

A Ketogenic Diet Increases Markers of Mitochondrial Content in a Tissue Specific Manner in Adult Mice

Dysregulation of mitochondrial energy metabolism is a major hallmark of mammalian aging. Calorie restriction (CR) has been shown to effectively increase lifespan and is associated with a metabolic shift toward a decrease in glycolysis and an increase in fatty acid oxidation. This shift in fuel utilization that occurs with CR relies heavily on mitochondrial energy metabolism. Thus, a diet that can mimic the metabolic reprogramming that occurs with CR may reduce age‐related decline in mitochondrial function and energy metabolism, and contribute to healthy aging and longevity. A ketogenic diet, which is depleted in carbohydrates, would drive the utilization of fat as a fuel and increase mitochondrial fatty acid oxidation. By chronically stimulating pathways in mitochondrial oxidative metabolism, the ketogenic diet could influence mitochondrial function and biogenesis, and may prevent age‐related decreases in mitochondrial content. In this study, mice were fed an isocaloric control (CLT), low carbohydrate (low‐CHO), or ketogenic diet (KD) from the age of 12 months. Tissues were collected after 1 month and 14 months of dietary treatment for evaluation of mitochondrial content. The results showed that the low‐CHO and KD increase markers of mitochondrial content in skeletal muscle in old mice (age of 26 months). The activities of both Complex I and IV were increased (p<0.05) in the low CHO and KD diet compared to the CTL diet. Old mice fed a KD also maintained activity of Complex I compared to the young mice, while mice fed the CLT and low‐CHO diets showed a decline (p<0.05) in Complex I activity with aging. Mice fed a KD also showed increased muscle citrate synthase activity with age. However, these changes are tissue specific with liver showing only an increase in Complex IV activity with aging in the KD group. In summary, the study demonstrates that long‐term ketogenic diet may prevent age related decline in mitochondrial content in some tissues in adult mice.Support or Funding InformationNIH Grant PO1AGO25532

Sirtuin 3‐ and Diet‐Mediated Regulation of Mitochondrial Function During Aging

Aging is a multifactorial process that results in the gradual damage of a number of cellular systems. Specifically, the decline of mitochondrial function due to hyperacetylation has been implicated in numerous metabolic defects, including senescence. The NAD+‐dependent mitochondrial deacetylase, sirtuin 3 (SIRT3) has a marked impact on mitochondrial acetylation status and may be responsible for attenuating oxidative stress. Furthermore, caloric restriction, which induces SIRT3 expression, has been linked to increased longevity. However, the impact of acetylation on mitochondrial function over the lifespan of an organism remains largely unknown. Here, we used a multi‐tissue approach to establish the role of SIRT3 and acetylation status in governing aging phenotypes by comparing mitochondrial function and structure in aging mice.C57BL/6 mice in the SIRT3+/+ and −/− condition were fed a control or calorically‐restricted diet, and sampled at 5, 15, and 25 months of age. Gastrocnemius, brain, liver and heart were measured for respiratory capacity, complex enzyme kinetics, mitochondrial morphology, mtDNA content, and the protein expression of key SIRT3 targets. Metabolomic profiling of the liver was used to assess the impact of electron transport impairments on substrate utilization and accumulation. To determine possible mechanisms for the aging phenotypes observed in the present study, we examined expression levels of ATP‐dependent mitochondrial proteases. Generally, all tissues had markedly different respiratory responses to the treatment condition. However, oxygen flux through complex II revealed a significant SIRT3‐CR “rescue” effect at old age in most tissues. These results suggest that the deficiencies in oxidative phosphorylation may be driven by an impaired function at respiratory complexes caused by reduced protein content or complex activity. Mitochondrial structure, as determined by electron microscopy, revealed differences amongst groups, dependent on both genotype and diet. Overall, our findings suggest that SIRT3 in conjunction with caloric restriction may play a pivotal role in the age‐related response of the mitochondrial respiratory chain to aging in a tissue‐specific manner.Support or Funding InformationNational Institute of Health, National Institute on Aging

Mitochondrial adaptations in aged skeletal muscle: effect of exercise training.

The aging process is associated with a decline in mitochondrial functions. Mitochondria dysfunction is involved in initiation and progression of many health problems including neuromuscular, metabolic and cardiovascular diseases. It is well known that endurance exercise improves mitochondrial function, especially in the elderly. However, recent studies have demonstrated that resistance training lead also to substantial increases in mitochondrial function in skeletal muscle. A comprehensive understanding of the cellular mechanisms involved in the skeletal muscle mitochondrial adaptations to exercise training in healthy elderly subjects, can help practitioners to design and prescribe more effective exercise trainings.

Cardioprotective kinase signaling to subsarcolemmal and interfibrillar mitochondria is mediated by caveolar structures

The demonstration that caveolin-3 overexpression reduces myocardial ischemia/reperfusion injury and our own finding that multiprotein signaling complexes increase in mitochondria in association with caveolin-3 levels, led us to investigate the contribution of caveolae-driven extracellular signal-regulated kinases 1/2 (ERK1/2) on maintaining the function of cardiac mitochondrial subpopulations from reperfused hearts subjected to postconditioning (PostC). Rat hearts were isolated and subjected to ischemia/reperfusion and to PostC. Enhanced cardiac function, reduced infarct size and preserved ultrastructure of cardiomyocytes were associated with increased formation of caveolar structures, augmented levels of caveolin-3 and mitochondrial ERK1/2 activation in PostC hearts in both subsarcolemmal (SSM) and interfibrillar (IFM) subpopulations. Disruption of caveolae with methyl-β-cyclodextrin abolished cardioprotection in PostC hearts and diminished pho-ERK1/2 gold-labeling in both mitochondrial subpopulations in correlation with suppression of resistance to permeability transition pore opening. Also, differences between the mitochondrial subpopulations in the setting of PostC were evaluated. Caveolae disruption with methyl-β-cyclodextrin abolished the cardioprotective effect of postconditioning by inhibiting the interaction of ERK1/2 with mitochondria and promoted decline in mitochondrial function. SSM, which are particularly sensitive to reperfusion damage, take advantage of their location in cardiomyocyte boundary and benefit from the cardioprotective signaling driven by caveolae, avoiding injury propagation.

The Role of Mitochondrial Non-Enzymatic Protein Acylation in Ageing.

In recent years, various large-scale proteomic studies have demonstrated that mitochondrial proteins are highly acylated, most commonly by addition of acetyl and succinyl groups. These acyl modifications may be enzyme catalysed but can also be driven non-enzymatically. The latter mechanism is promoted in mitochondria due to the nature of the mitochondrial microenvironment, which is alkaline and contains high concentrations of acyl-CoA species. Protein acylation may modify enzyme activity, typically inhibiting it. We posited that organismal ageing might be accompanied by an accumulation of acylated proteins, especially in mitochondria, and that this might compromise mitochondrial function and contribute to ageing. In this study, we used R. norvegicus, C. elegans and D. melanogaster to compare the acylation status of mitochondrial proteins between young and old animals. We observed a specific age-dependent increase in protein succinylation in worms and flies but not in rat. Rats have two substrate-specific mitochondrial deacylases, SIRT3 and SIRT5 while both flies and worms lack these enzymes. We propose that accumulation of mitochondrial protein acylation contributes to age-dependent mitochondrial functional decline and that SIRT3 and SIRT5 enzymes may promote longevity through regulation of mitochondrial protein acylation during ageing.

Mitochondrial Dysfunction Prevents Repolarization of Inflammatory Macrophages

Macrophages are innate immune cells that adopt diverse activation states in response to their microenvironment. Editing macrophage activation to dampen inflammatory diseases by promoting the repolarization of inflammatory (M1) macrophages to anti-inflammatory (M2) macrophages is of high interest. Here, we find that mouse and human M1 macrophages fail to convert into M2 cells upon IL-4 exposure invitro and invivo. In sharp contrast, M2 macrophages are more plastic and readily repolarized into an inflammatory M1 state. We identify M1-associated inhibition of mitochondrial oxidative phosphorylation as the factor responsible for preventing M1→M2 repolarization. Inhibiting nitric oxide production, a key effector molecule in M1 cells, dampens the decline in mitochondrial function to improve metabolic and phenotypic reprogramming to M2 macrophages. Thus, inflammatory macrophage activation blunts oxidative phosphorylation, thereby preventing repolarization. Therapeutically restoring mitochondrial function might be useful toimprove the reprogramming of inflammatory macrophages into anti-inflammatory cells to control disease.

OPA1 gene and mitochondrial optic neuropathy: disease mechanisms and potential therapies

SummaryPrimary inherited optic neuropathies are a group of blinding genetic disorders in which optic atrophy secondary to loss of retinal ganglion cells is a key clinical feature. The commonest causes world‐wide are mutation in mitochondrial DNA (causing Leber's Hereditary Optic Neuropathy) and mutation in the nuclear gene, OPA1 (causing Autosomal Dominant Optic Atrophy: ADOA). 60–75% of patients with autosomal dominant optic atrophy have mutations in the OPA1 gene. The OPA1 protein is targeted to the mitochondria and is involved in regulation of mitochondrial fusion. A better understanding of mitochondrial function, including dynamics, is revealing that functional and structural changes in mitochondrial morphology are important factors in diseases of ageing in the eye and visual system. Key proteins have been discovered which control the balance of mitochondrial fusion and fission and have a range of other functions, such as controlling maintenance of mitochondrial DNA, cell death, autophagy, mitochondrial metabolism and redox signalling. A decline in mitochondrial function plays a role in the ageing process and increases the incidence of age‐related disorders. Mitochondrial optic neuropathies are ‘orphan’ diseases but with the advent of recent trials of novel therapies in patients with the mitochondrial optic neuropathy, Leber's hereditary optic neuropathy, there is the first glimmer of hope for the treatment of this group of patients.

In EXOG-depleted cardiomyocytes cell death is marked by a decreased mitochondrial reserve capacity of the electron transport chain.

Depletion of mitochondrial endo/exonuclease G-like (EXOG) in cultured neonatal cardiomyocytes stimulates mitochondrial oxygen consumption rate (OCR) and induces hypertrophy via reactive oxygen species (ROS). Here, we show that neurohormonal stress triggers cell death in endo/exonuclease G-like-depleted cells, and this is marked by a decrease in mitochondrial reserve capacity. Neurohormonal stimulation with phenylephrine (PE) did not have an additive effect on the hypertrophic response induced by endo/exonuclease G-like depletion. Interestingly, PE-induced atrial natriuretic peptide (ANP) gene expression was completely abolished in endo/exonuclease G-like-depleted cells, suggesting a reverse signaling function of endo/exonuclease G-like. Endo/exonuclease G-like depletion initially resulted in increased mitochondrial OCR, but this declined upon PE stimulation. In particular, the reserve capacity of the mitochondrial respiratory chain and maximal respiration were the first indicators of perturbations in mitochondrial respiration, and these marked the subsequent decline in mitochondrial function. Although pathological stimulation accelerated these processes, prolonged EXOG depletion also resulted in a decline in mitochondrial function. At early stages of endo/exonuclease G-like depletion, mitochondrial ROS production was increased, but this did not affect mitochondrial DNA (mtDNA) integrity. After prolonged depletion, ROS levels returned to control values, despite hyperpolarization of the mitochondrial membrane. The mitochondrial dysfunction finally resulted in cell death, which appears to be mainly a form of necrosis. In conclusion, endo/exonuclease G-like plays an essential role in cardiomyocyte physiology. Loss of endo/exonuclease G-like results in diminished adaptation to pathological stress. The decline in maximal respiration and reserve capacity is the first sign of mitochondrial dysfunction that determines subsequent cell death.

Pancreatic β-Cells From Mice Offset Age-Associated Mitochondrial Deficiency With Reduced KATP Channel Activity.

Aging is accompanied by impaired glucose homeostasis and an increased risk of type 2 diabetes, culminating in the failure of insulin secretion from pancreatic β-cells. To investigate the effects of age on β-cell metabolism, we established a novel assay to directly image islet metabolism with NAD(P)H fluorescence lifetime imaging (FLIM). We determined that impaired mitochondrial activity underlies an age-dependent loss of insulin secretion in human islets. NAD(P)H FLIM revealed a comparable decline in mitochondrial function in the pancreatic islets of aged mice (≥24 months), the result of 52% and 57% defects in flux through complex I and II, respectively, of the electron transport chain. However, insulin secretion and glucose tolerance are preserved in aged mouse islets by the heightened metabolic sensitivity of the β-cell triggering pathway, an adaptation clearly encoded in the metabolic and Ca2+ oscillations that trigger insulin release (Ca2+ plateau fraction: young 0.211 ± 0.006, aged 0.380 ± 0.007, P < 0.0001). This enhanced sensitivity is driven by a reduction in KATP channel conductance (diazoxide: young 5.1 ± 0.2 nS; aged 3.5 ± 0.5 nS, P < 0.01), resulting in an ∼2.8 mmol/L left shift in the β-cell glucose threshold. The results demonstrate how mice but not humans are able to successfully compensate for age-associated metabolic dysfunction by adjusting β-cell glucose sensitivity and highlight an essential mechanism for ensuring the maintenance of insulin secretion.

Specific protein changes contribute to the differential muscle mass loss during ageing.

In the skeletal muscle, the ageing process is characterized by a loss of muscle mass and strength, coupled with a decline of mitochondrial function and a decrease of satellite cells. This profile is more pronounced in hindlimb than in forelimb muscles, both in humans and in rodents. Utilizing light and electron microscopy, myosin heavy chain isoform distribution, proteomic analysis by 2D-DIGE, MALDI-TOF MS and quantitative immunoblotting, this study analyzes the protein levels and the nuclear localization of specific molecules, which can contribute to a preferential muscle loss. Our results identify the molecular changes in the hindlimb (gastrocnemius) and forelimb (triceps) muscles during ageing in rats (3- and 22-month-old). Specifically, the oxidative metabolism contributes to tissue homeostasis in triceps, whereas respiratory chain disruption and oxidative-stress-induced damage imbalance the homeostasis in gastrocnemius muscle. High levels of dihydrolipoyllysine-residue acetyltransferase (Dlat) and ATP synthase subunit alpha (Atp5a1) are detected in triceps and gastrocnemius, respectively. Interestingly, in triceps, both molecules are increased in the nucleus in aged rats and are associated to an increased protein acetylation and myoglobin availability. Furthermore, autophagy is retained in triceps whereas an enhanced fusion, decrement of mitophagy and of regenerative potential is observed in aged gastrocnemius muscle.

Life span extension by targeting a link between metabolism and histone acetylation in Drosophila.

Old age is associated with a progressive decline of mitochondrial function and changes in nuclear chromatin. However, little is known about how metabolic activity and epigenetic modifications change as organisms reach their midlife. Here, we assessed how cellular metabolism and protein acetylation change during early aging in Drosophila melanogaster. Contrary to common assumptions, we find that flies increase oxygen consumption and become less sensitive to histone deacetylase inhibitors as they reach midlife. Further, midlife flies show changes in the metabolome, elevated acetyl‐CoA levels, alterations in protein—notably histone—acetylation, as well as associated transcriptome changes. Based on these observations, we decreased the activity of the acetyl‐CoA‐synthesizing enzyme ATP citrate lyase (ATPCL) or the levels of the histone H4 K12‐specific acetyltransferase Chameau. We find that these targeted interventions both alleviate the observed aging‐associated changes and promote longevity. Our findings reveal a pathway that couples changes of intermediate metabolism during aging with the chromatin‐mediated regulation of transcription and changes in the activity of associated enzymes that modulate organismal life span.

Peculiarities and pitfalls of quantifying mitochondrial energy metabolism in the skin.

Peculiarities and pitfalls of quantifying mitochondrial energy metabolism in the skin.

TNF-α and Beyond: Rapid Mitochondrial Dysfunction Mediates TNF-α-Induced Neurotoxicity.

This short communication describes our research which demonstrates that TNF-α causes a rapid decline in mitochondrial function, leading to neuronal cell death. As such, this neurotoxic proinflammatory cytokine may play a role in brain damage from stroke and neurodegeneration in chronic conditions such as Alzheimer’s disease (AD) and Parkinson’s disease. We have extended this initial observation by demonstrating that TNF-α stimulates a microRNA (miR-34a) which we have shown reduces five key proteins in the mitochondrial electron transport chain through base-pair complementarity. miR-34a is increased in affected brain regions of Alzheimer’s patients and transgenic AD mouse models. We have further shown that oligomeric amyloid beta 42 (oAβ42) stimulates miR-34a. Collectively, these data suggest that TNF-α, oAβ42, and miR-34a participate in a vicious cycle, resulting in mitochondrial dysfunction, which is critical to the neuropathology of AD.

Role of Mitochondrial Protein Quality Control in Oxidative Stress-induced Neurodegenerative Diseases.

Proteins are constantly exposed to environmental stressors such as free radicals and heat shock leading to their misfolding and later to aggregation. In particular mitochondrial proteins are challenged by reactive oxygen species (ROS) due to the oxidative metabolism of the organelle. Protein aggregation has been associated with a wide variety of pathological conditions called proteopathies. However, for the maintenance of protein and cellular homeostasis, mitochondria have developed an elaborate protein quality control system consisting of chaperones and ATP-dependent proteases, specifically employed to rescue this organelle from damage due to the accumulation of misfolded proteins and toxic aggregates. Aging is characterized by a general decline of mitochondrial functions, correlating with a decrease in mitochondrial protein quality control activity and an increase of free radical production. In particular in age-related diseases like neurodegeneration, a correlation between mitochondrial damage and disease onset has been established. In this review we summarize the current knowledge about mitochondrial protein quality control mechanisms in mammalian cells, with a special emphasis on the role in oxidative stress and in neurodegenerative diseases.

Increased expression of mitochondrial DNA-encoded genes in human renal mesangial cells in response to high glucose-induced reactive oxygen species.

Reactive oxygen species (ROS)-mediated disruption of mitochondrial respiratory function has been implicated in the complications of diabetes. The present study examined changes in the gene expression of mitochondrial DNA (mtDNA)-encoded subunits of electron transport chain complexes in response to high glucose-induced ROS overproduction in an in vitro model of diabetic nephropathy using human renal mesangial cells. Mitochondrial ROS generation was assessed by confocal microscopy and flow cytometry in the cells following culture in 5 and 25 mM glucose. The mRNA expression levels of nicotinamide adenine dinucleotide dehydrogenase 2 (ND2) of complex I, cytochrome b (CYTB) of complex III, cytochrome c oxidase (COI) of complex IV and ATPase 6 of complex V were analyzed by reverse transcription-quantitative polymerase chain reaction. The protein expression levels of ND2, CYTB, COI and ATPase 6 were analyzed by western blotting. A significant increase in mitochondrial ROS production was observed in the cells cultured in 25 mM glucose, compared with cells cultured in 5 mM glucose (P<0.05). The mRNA expression of ND2, CYTB, CO1 and ATPase 6 was significantly increased following culture in 25 mM glucose, compared with the cells cultured in 5 mM glucose (P<0.05). This increase in mRNA expression was accompanied by significant increases in protein expression following incubation in 25 mM glucose (P<0.05). The increase in mtDNA-encoded gene expression in the electron transport subunits following exposure to high glucose-induced ROS may be a compensatory response mechanism for the decline in mitochondrial function, which may be important in the development of diabetic nephropathy through enhanced ROS generation.

Molecular insight and pharmacological approaches targeting mitochondrial dynamics in skeletal muscle during obesity.

Obesity-associated insulin resistance is the major characteristic of the early stage of metabolic syndrome. A decline in mitochondrial function plays a role in the development of insulin resistance in obesity and type 2 diabetes. Accumulating data reveal that mitochondrial dynamics, the balance between mitochondrial fusion and fission, are an important factor in the maintenance of mitochondrial function. Thus, the mechanisms underlying the regulation of mitochondrial dynamics in obesity deserve further investigation. This review describes an overview of mitochondrial fusion and fission machineries, and discusses the mechanistic and functional aspects of mitochondrial dynamics, with a focus on skeletal muscle in obesity. Finally, we discuss current pharmacological approaches of targeting mitochondrial dynamics. Elucidating the role of mitochondrial dynamics in skeletal muscle afflicted by obesity may provide not only important clues in understanding muscle insulin resistance, but also new therapeutic targets.

Eicosapentaenoic acid but not docosahexaenoic acid restores skeletal muscle mitochondrial oxidative capacity in old mice.

Mitochondrial dysfunction is often observed in aging skeletal muscle and is implicated in age-related declines in physical function. Early evidence suggests that dietary omega-3 polyunsaturated fatty acids (n-3 PUFAs) improve mitochondrial function. Here, we show that 10 weeks of dietary eicosapentaenoic acid (EPA) supplementation partially attenuated the age-related decline in mitochondrial function in mice, but this effect was not observed with docosahexaenoic acid (DHA). The improvement in mitochondrial function with EPA occurred in the absence of any changes in mitochondrial abundance or biogenesis, which was evaluated from RNA sequencing, large-scale proteomics, and direct measurements of muscle mitochondrial protein synthesis rates. We find that EPA improves muscle protein quality, specifically by decreasing mitochondrial protein carbamylation, a post-translational modification that is driven by inflammation. These results demonstrate that EPA attenuated the age-related loss of mitochondrial function and improved mitochondrial protein quality through a mechanism that is likely linked with anti-inflammatory properties of n-3 PUFAs. Furthermore, we demonstrate that EPA and DHA exert some common biological effects (anticoagulation, anti-inflammatory, reduced FXR/RXR activation), but also exhibit many distinct biological effects, a finding that underscores the importance of evaluating the therapeutic potential of individual n-3 PUFAs.

Mechanisms by which different functional states of mitochondria define yeast longevity.

Mitochondrial functionality is vital to organismal physiology. A body of evidence supports the notion that an age-related progressive decline in mitochondrial function is a hallmark of cellular and organismal aging in evolutionarily distant eukaryotes. Studies of the baker’s yeast Saccharomyces cerevisiae, a unicellular eukaryote, have led to discoveries of genes, signaling pathways and chemical compounds that modulate longevity-defining cellular processes in eukaryotic organisms across phyla. These studies have provided deep insights into mechanistic links that exist between different traits of mitochondrial functionality and cellular aging. The molecular mechanisms underlying the essential role of mitochondria as signaling organelles in yeast aging have begun to emerge. In this review, we discuss recent progress in understanding mechanisms by which different functional states of mitochondria define yeast longevity, outline the most important unanswered questions and suggest directions for future research.

Enhanced cardiac Akt/protein kinase B signaling contributes to pathological cardiac hypertrophy in part by impairing mitochondrial function via transcriptional repression of mitochondrion-targeted nuclear genes.

Sustained Akt activation induces cardiac hypertrophy (LVH), which may lead to heart failure. This study tested the hypothesis that Akt activation contributes to mitochondrial dysfunction in pathological LVH. Akt activation induced LVH and progressive repression of mitochondrial fatty acid oxidation (FAO) pathways. Preventing LVH by inhibiting mTOR failed to prevent the decline in mitochondrial function, but glucose utilization was maintained. Akt activation represses expression of mitochondrial regulatory, FAO, and oxidative phosphorylation genes in vivo that correlate with the duration of Akt activation in part by reducing FOXO-mediated transcriptional activation of mitochondrion-targeted nuclear genes in concert with reduced signaling via peroxisome proliferator-activated receptor α (PPARα)/PGC-1α and other transcriptional regulators. In cultured myocytes, Akt activation disrupted mitochondrial bioenergetics, which could be partially reversed by maintaining nuclear FOXO but not by increasing PGC-1α. Thus, although short-term Akt activation may be cardioprotective during ischemia by reducing mitochondrial metabolism and increasing glycolysis, long-term Akt activation in the adult heart contributes to pathological LVH in part by reducing mitochondrial oxidative capacity.

Decreased mitochondrial bioenergetics and calcium buffering capacity in the basal ganglia correlates with motor deficits in a nonhuman primate model of aging

Altered mitochondrial function in the basal ganglia has been hypothesized to underlie cellular senescence and promote age-related motor decline. We tested this hypothesis in a nonhuman primate model of human aging. Six young (6–8 years old) and 6 aged (20–25 years old) female Rhesus monkeys (Macaca mulatta) were behaviorally characterized from standardized video records. Additionally, we measured mitochondrial bioenergetics along with calcium buffering capacity in the substantia nigra and putamen (PUT) from both age groups. Our results demonstrate that the aged animals had significantly reduced locomotor activity and movement speed compared with younger animals. Moreover, aged monkeys had significantly reduced ATP synthesis capacity (in substantia nigra and PUT), reduced pyruvate dehydrogenase activity (in PUT), and reduced calcium buffering capacity (in PUT) compared with younger animals. Furthermore, this age-related decline in mitochondrial function in the basal ganglia correlated with decline in motor function. Overall, our results suggest that drug therapies designed to enhance altered mitochondrial function may help improve motor deficits in the elderly.