Decrease In Mitochondrial Density Research Articles

Exploring the aging effect of the anticancer drugs doxorubicin and mitoxantrone on cardiac mitochondrial proteome using a murine model

Current cancer therapies are successfully increasing the lifespan of cancer patients. Nevertheless, cardiotoxicity is a serious chemotherapy-induced adverse side effect. Doxorubicin (DOX) and mitoxantrone (MTX) are cardiotoxic anticancer agents, whose toxicological mechanisms are still to be identified. This study focused on DOX and MTX’s cardiac mitochondrial damage and their molecular mechanisms. As a hypothesis, we also sought to compare the cardiac modulation caused by 9 mg/kg of DOX or 6 mg/kg of MTX in young adult mice (3 months old) with old control mice (aged control, 18–20 months old) to determine if DOX- and MTX-induced damage had common links with the aging process.Cardiac homogenates and enriched mitochondrial fractions were prepared from treated and control animals and analyzed by immunoblotting and enzymatic assays. Enriched mitochondrial fractions were also characterized by mass spectrometry-based proteomics. Data obtained showed a decrease in mitochondrial density in young adults treated with DOX or MTX and aged control, as assessed by citrate synthase (CS) activity. Furthermore, aged control had increased expression of the peroxisome proliferator-activated receptor γ coactivator 1 α (PGC1α) and manganese superoxide dismutase (MnSOD). Regarding the enriched mitochondrial fractions, DOX and MTX led to downregulation of proteins related to oxidative phosphorylation, fatty acid oxidation, amino acid metabolic process, and tricarboxylic acid cycle. MTX had a greater impact on malate dehydrogenase (MDH2) and pyruvate dehydrogenase E1 component subunit α (PDHA1). No significant proteomic changes were observed in the enriched mitochondrial fractions of aged control when compared to young control. To conclude, DOX and MTX promoted changes in several mitochondrial-related proteins in young adult mice, but none resembling the aged phenotype.

Beneficial Effects of Bariatric Surgery-Induced by Weight Loss on the Proteome of Abdominal Subcutaneous Adipose Tissue

Bariatric surgery (BS) is the most effective treatment for obesity and has a positive impact on cardiometabolic risk and in the remission of type 2 diabetes. Following BS, the majority of fat mass is lost from the subcutaneous adipose tissue depot (SAT). However, the changes in this depot and functions and as well as its relative contribution to the beneficial effects of this surgery are still controversial. With the aim of studying altered proteins and molecular pathways in abdominal SAT (aSAT) after body weight normalization induced by BS, we carried out a proteomic approach sequential window acquisition of all theoretical mass spectra (SWATH-MS) analysis. These results were complemented by Western blot, electron microscopy and RT-qPCR. With all of the working tools mentioned, we confirmed that after BS, up-regulated proteins were associated with metabolism, the citric acid cycle and respiratory electron transport, triglyceride catabolism and metabolism, formation of ATP, pyruvate metabolism, glycolysis/gluconeogenesis and thermogenesis among others. In contrast, proteins with decreased values are part of the biological pathways related to the immune system. We also confirmed that obesity caused a significant decrease in mitochondrial density and coverage, which was corrected by BS. Together, these findings reveal specific molecular mechanisms, genes and proteins that improve adipose tissue function after BS characterized by lower inflammation, increased glucose uptake, higher insulin sensitivity, higher de novo lipogenesis, increased mitochondrial function and decreased adipocyte size.

Disrupted expression of genes essential for skeletal muscle fibre integrity and energy metabolism in Vitamin D deficient rats

Vitamin D, a secosteroid that regulates mineral homeostasis via its actions in intestine, bone, kidneys and parathyroid glands, has many other target tissues, including skeletal muscle. In the present study, we used rats to examine if diet-induced vitamin D deficiency or insufficiency altered protein synthesis in muscle via the mTOR pathway, and impaired skeletal muscle quality by changing expression of genes needed for its function. Vitamin D deficiency resulted in reduced levels of phosphorylated mTOR, and suppressed mTOR-dependent phosphorylation of 4E-BP1 and p70-S6K, implying a decrease in activity of the protein synthesis machinery. These changes were coupled with up regulation of genes that are negative regulators of muscle growth (Fbxo32 & Trim63), leading to a net loss of skeletal muscle mass. Vitamin D deficiency or insufficiency also led to a decrease in expression of both myosin and actin-associated proteins (Myh1, Myh2, Myh7, Tnnc1& Tnnt1), which are essential for generation of the mechanical force needed for muscle contraction. We also detected a decrease in expression of glycolytic and oxidative enzyme genes (Hk2, Pfkm, Cs, Pdk4 & βHad) and transcriptional coactivator genes (Ppargc-1α & Ppargc-1β) which indicate a low oxidative capacity of skeletal muscle in the vitamin D deficient state. Furthermore, decreased citrate synthase activity corroborates a decrease in mitochondrial density and aerobic capacity of the muscle. In conclusion, our study demonstrates that chronic vitamin D deficiency or insufficiency reduced the size of skeletal muscle fibres, altered their composition, and decreased their oxidative potential. Most of the changes observed were reversible, either partially or completely, by restoring vitamin D to the diet of the deficient rats.

Regulation of Brown Adipose Tissue Function by HuR

Stimulating the metabolic function of brown adipose tissue (BAT) and/or inducing the beiging of white adipocytes (WAT) to BAT‐like cells represent potential therapeutic strategies for increasing energy expenditure and reducing obesity. HuR (ELAVL1) is an RNA binding protein that mediates gene expression through stabilization of mRNA targets and is highly expressed in both WAT and BAT. We identified an inverse correlation between HuR expression and obesity in humans via the METabolic Syndrome In Men study of Finnish men. Thus, we hypothesized that HuR expression in adipocytes plays a critical role in maintaining metabolic homeostasis.Our results show that, compared to control littermates, adipocyte‐specific HuR‐null (adipo‐HuR−/−) mice display a lean phenotype with decreased fat mass and increased energy expenditure, independent of diet. Interestingly, adipo‐HuR−/− mice also show a disruption of BAT structure and function, with increased lipid droplet size, decreased BAT density, and an inability to thermoregulate when subjected to cold. Examination of skeletal muscle suggests that the lean phenotype in adipo‐HuR−/− mice is due to compensatory shivering to thermoregulate in the absence of functional BAT, a result consistent with UCP1−/− mice. Surprisingly, we found no difference in basal expression of UCP1 in BAT from control and adipo‐HuR−/− mice. However, HuR is activated downstream of β3 adrenergic stimulation and appears to play a role in cold/β3 induced UCP1 expression. Additionally, adipo−HuR−/− mice show a decrease in mitochondrial density in both BAT and subcutaneous (sc) WAT, as well as a decrease in brown/beige adipocyte marker genes (PRDM16, Elovl3) in scWAT.In conclusion, our results demonstrate that HuR plays a central role in the structure and function of BAT. Work is ongoing to identify the transcriptome wide changes in HuR‐dependent gene expression as well as the potential role of HuR in beiging of WAT.Support or Funding InformationThis work was partially funded by a University of Cincinnati Heart, Lung, and Vascular Institute Pilot Grant (MT, APO).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Skeletal muscle mitochondrial dysfunction during chronic obstructive pulmonary disease: central actor and therapeutic target

Muscle dysfunction is a common complication and an important prognostic factor in chronic obstructive pulmonary disease (COPD). As therapeutic strategies are still needed to treat this complication, gaining more insight into the process that leads to skeletal muscle decline in COPD appears to be an important issue. This review focuses on mitochondrial involvement in limb skeletal muscle alterations (decreased muscle mass, strength, endurance and power and increased fatigue) in COPD. Mitochondria are the main source of energy for the cells; they are involved in production of reactive oxygen species and activate an important pathway that leads to apoptosis. In COPD patients, skeletal muscles are characterized by decreased mitochondrial density and biogenesis, impaired activity and coupling of mitochondrial respiratory chain complexes, increased mitochondrial production of reactive oxygen species and, possibly, increased apoptosis. Of particular interest, a sedentary lifestyle, hypoxia, hypercapnia, tobacco smoking, corticosteroid therapy and, possibly, inflammation participate in this mitochondrial dysfunction, which is accessible to conventional therapies, such as exercise and tobacco cessation, as well as, potentially, to more innovative approaches, such as antioxidant treatment and supplementation with polyunsaturated fatty acids.

General Anesthesia Causes Long-term Impairment of Mitochondrial Morphogenesis and Synaptic Transmission in Developing Rat Brain.

Clinically used general anesthetics, alone or in combination, are damaging to the developing mammalian brain. In addition to causing widespread apoptotic neurodegeneration in vulnerable brain regions, exposure to general anesthesia at the peak of synaptogenesis causes learning and memory deficiencies later in life. In vivo rodent studies have suggested that activation of the intrinsic (mitochondria-dependent) apoptotic pathway is the earliest warning sign of neuronal damage, suggesting that a disturbance in mitochondrial integrity and function could be the earliest triggering events. Because proper and timely mitochondrial morphogenesis is critical for brain development, the authors examined the long-term effects of a commonly used anesthesia combination (isoflurane, nitrous oxide, and midazolam) on the regional distribution, ultrastructural properties, and electron transport chain function of mitochondria, as well as synaptic neurotransmission, in the subiculum of rat pups. This anesthesia, administered at the peak of synaptogenesis, causes protracted injury to mitochondria, including significant enlargement of mitochondria (more than 30%, P < 0.05), impairment of their structural integrity, an approximately 28% increase in their complex IV activity (P < 0.05), and a twofold decrease in their regional distribution in presynaptic neuronal profiles (P < 0.05), where their presence is important for the normal development and functioning of synapses. Consequently, the authors showed that impaired mitochondrial morphogenesis is accompanied by heightened autophagic activity, decrease in mitochondrial density (approximately 27%, P < 0.05), and long-lasting disturbances in inhibitory synaptic neurotransmission. The interrelation of these phenomena remains to be established. Developing mitochondria are exquisitely vulnerable to general anesthesia and may be important early target of anesthesia-induced developmental neurodegeneration.

Striatal mitochondria in subjects with chronic undifferentiated vs. chronic paranoid schizophrenia

Schizophrenia (SZ) is a heterogeneous disease with a spectrum of symptoms, risk factors, and etiology. Abnormalities in mitochondria, the energy-producing organelles of the cell, have been observed in mixed cohorts of subjects with SZ. The purpose of the present study was to determine if striatal mitochondria were differentially affected in two different DSM-IV subgroups of SZ. Postmortem striatal tissue was examined from normal controls (NC), chronic paranoid SZs (SZP), and chronic undifferentiated SZs (SZU). Tissue was processed for calbindin immunohistochemistry to identify striosomal compartments, prepared for electron microscopy and analyzed using stereological methods. In both caudate and putamen, the density of mitochondria in the neuropil was decreased in SZP compared to both NCs and SZU. In the putamen, both the SZP and the SZU subgroups had fewer mitochondria per synapse than did NCs. When examining patch matrix compartments, striatal compartments associated with different circuitry and function, only the matrix exhibited changes. In the caudate matrix, the SZP subgroup had fewer mitochondria in the neuropil than did the SZU and NCs. In the putamen matrix, the SZP had fewer mitochondria in the neuropil as compared to NCs, but not the SZU. The numbers of mitochondria per synapse in both the SZP and the SZU groups were similar to each other and fewer than that of NCs. A decrease in mitochondrial density in the neuropil distinguishes the SZP from the SZU subgroup, which could be associated with the symptoms of paranoia and/or could represent a protective mechanism against some of the symptoms that are less pronounced in this subtype than in the SZU subgroup such as cognitive and emotional deficits.

Cardiac-specific overexpression of dominant-negative CREB leads to increased mortality and mitochondrial dysfunction in female mice

Cardiac failure is associated with diminished activation of the transcription factor cyclic nucleotide regulatory element binding-protein (CREB), and heart-specific expression of a phosphorylation-deficient CREB mutant in transgenic mice [dominant negative CREB (dnCREB) mice] recapitulates the contractile phenotypes of cardiac failure (Fentzke RC, Korcarz CE, Lang RM, Lin H, Leiden JM. Dilated cardiomyopathy in transgenic mice expressing a dominant-negative CREB transcription factor in the heart. J Clin Invest 101: 2415-2426, 1998). In the present study, we demonstrated significantly elevated mortality and contractile dysfunction in female compared with male dnCREB mice. Female dnCREB mice demonstrated a 21-wk survival of only 17% compared with 67% in males (P < 0.05) and exclusively manifest decreased cardiac peroxisome proliferator-activated receptor-γ coactivator-1α and estrogen-related receptor-α content, suggesting sex-related effects on cardiac mitochondrial function. Hearts from 4-wk-old dnCREB mice of both sexes demonstrated diminished mitochondrial respiratory capacity compared with nontransgenic controls. However, by 12 wk of age, there was a significant decrease in mitochondrial density (citrate synthase activity) and deterioration of mitochondrial structure, as demonstrated by transmission electron microscopy, in female dnCREB mice, which were not found in male transgenic littermates. Subsarcolemmal mitochondria isolated from hearts of female, but not male, dnCREB mice demonstrated increased ROS accompanied by decreases in the expression/activity of the mitochondrial antioxidants MnSOD and glutathione peroxidase. These results demonstrate that heart-specific dnCREB expression results in mitochondrial respiratory dysfunction in both sexes; however, increased oxidant burden, reduced antioxidant expression, and disrupted mitochondrial structure are exacerbated by the female sex, preceding and contributing to the greater contractile morbidity and mortality. These results provide further support for the role of the CREB transcription factor in regulating mitochondrial integrity and identify a critical pathway that may contribute to sex differences in heart failure.

The ontogeny of aerobic and diving capacity in the skeletal muscles of Weddell seals

Our objective was to determine the ontogenetic changes in the skeletal muscles of Weddell seals that transform a non-diving pup into an elite diving adult. Muscle biopsies were collected from pups, juveniles and adults and analyzed for changes in fiber type, mitochondrial density, myoglobin concentrations and aerobic, lipolytic and anaerobic enzyme activities. The fiber type results demonstrated a decrease in slow-twitch oxidative (Type I) fibers and a significant increase in fast-twitch oxidative (Type IIA) fibers as the animals mature. In addition, the volume density of mitochondria and the activity of lipolytic enzymes significantly decreased as the seals matured. To our knowledge, this is the first quantitative account describing a decrease in aerobic fibers shifting towards an increase in fast-twitch oxidative fibers with a significant decrease in mitochondrial density as animals mature. These differences in the muscle physiology of Weddell seals are potentially due to their three very distinct stages of life history: non-diving pup, novice diving juvenile, and elite deep diving adult. During the first few weeks of life, pups are a non-diving terrestrial mammal that must rely on lanugo (natal fur) for thermoregulation in the harsh conditions of Antarctica. The increased aerobic capacity of pups, associated with increased mitochondrial volumes, acts to provide additional thermogenesis. As these future elite divers mature, their skeletal muscles transform to a more sedentary state in order to maintain the low levels of aerobic metabolism associated with long-duration diving.

Mitochondrial dysfunction, insulin resistance, and type 2 diabetes mellitus

Insulin resistance is a characteristic feature of type 2 diabetes mellitus, obesity, and the metabolic syndrome. Increased intracellular fat content in skeletal muscle and liver associated with insulin resistance has led to the hypothesis that a mitochondrial defect in substrate oxidation exists in disorders of insulin resistance. In vivo measurements of metabolic fluxes through the tricarboxylic acid and oxidative phosphorylation with magnetic resonance spectroscopy have demonstrated multiple defects in mitochondrial function in skeletal muscle. A decrease in mitochondrial density and mitochondrial copy number has been reported in insulin-resistant individuals. However, these findings have not been a consistent observation in all studies. Similarly, an intrinsic functional defect in mitochondrial adenosine triphosphate production synthesis has been reported in some but not all studies. This review summarizes evidence that implicates a defect in mitochondrial oxidative phosphorylation and its relationship to insulin resistance in common metabolic diseases characterized by impaired insulin action.

The ontogeny of skeletal muscle adaptations that enable long deep dives in Weddell seals

Our objective was to determine the ontogenetic changes in the skeletal muscles of Weddell seals that transform a non‐diving pup into an elite diving adult. Muscle biopsies were collected from pups, juveniles and adults and analyzed for changes in fiber type, mitochondrial density, myoglobin concentrations, and aerobic, lipolytic and anaerobic enzyme activities. The fiber type results showed a decrease in type I fibers with a significant increase in type IIA fibers as the animals mature. In addition, the volume density of mitochondria and the activity of lipolytic enzymes significantly decreased as the seals matured. To our knowledge, this is the first quantitative account describing a decrease in aerobic fibers shifting towards an increase in fast‐twitch oxidative fibers with a significant decrease in mitochondrial density as the animals mature. These differences in muscle physiology are due to their three stages of life history. During the first few weeks of life, pups are a non‐diving terrestrial mammal that must rely on natal fur for thermoregulation in the harsh conditions of Antarctica. The increased aerobic capacity of pups, associated with increased mitochondrial volumes, acts to provide additional thermogenesis. As these future elite divers mature, their skeletal muscles transform to a more sedentary state in order to maintain the low levels of aerobic metabolism associated with long duration diving.

Hypoxia causes perinuclear mitochondrial clustering and nuclear oxidant stress in pulmonary artery endothelial cells (PAECS) via a dynein‐dependent molecular motor

Accumulating evidence suggests that mitochondria are motile, and may be recruited to subcellular domains where they participate in signal transduction. Mitochondria respond to hypoxia in PAECs by increasing oxygen radical generation, but whether hypoxia alters the disposition of mitochondria in PAECs is unknown, and if so, the molecular mechanism remains to be clarified. Accordingly, we mapped the disposition of Mitotracker‐stained mitochondria in cultured PAECs, and found that hypoxia caused a time‐dependent decrease in mitochondrial density near the plasma membrane and an increase in the perinuclear region. Interruption of the dynein‐dependent molecular motor using a specific siRNA prevented both the perinuclear mitochondrial clustering and, importantly, a hypoxia‐induced nuclear oxidant stress as detected by nuclear DCF fluorescence and by the level of oxidative base modifications in the VEGF promoter. Mitochondria were the source of the DNA‐modifying oxidants as evidenced by the finding that the respiratory chain inhibitor, myxothiazol, prevented the hypoxia‐induced nuclear DCF fluorescence without altering mitochondrial perinuclear clustering. These data suggest that hypoxia causes dynein‐dependent mitochondrial translocation to the perinuclear domain in PAECs that serves to create a nuclear oxidant stress important for hypoxic signaling.Supported by NIH

Regional absence of mitochondria causing energy depletion in the myocardium of muscle LIM protein knockout mice

Defects in myocardial mitochondrial structure and function have been associated with heart failure in humans and animal models. Mice lacking the muscle LIM protein (MLP) develop morphological and clinical signs resembling human dilated cardiomyopathy and heart failure. We tested the hypothesis that defects in the cytoskeleton lead to dilated cardiomyopathy through mitochondrial dysfunction in the MLP mouse model. Oxidative phosphorylation activity was determined in left ventricles of MLP knockout (KO) mice and control littermates by measuring complex activities of the electron transport chain (I-IV) and ATP synthase (complex V). All complexes and citrate synthase (CS) showed decreased activities in the KO mice, although activity per amount of CS, a measure for mitochondrial density, was normal. Light and electron microscopy revealed a disorganization of mitochondria and a dramatic decrease in mitochondrial density, even revealing regions completely lacking mitochondria in the KO hearts. Real-time PCR analysis showed decreased transcript levels of mtDNA and nuclear encoded mitochondrial genes and of peroxisome proliferator activated receptor gamma co-activator 1alpha (PGC-1alpha), a key regulator of mitochondrial biogenesis. MtDNA copy number (ratio mtDNA/nuclear DNA) was slightly increased in the MLP KO mice. Our results show that the absence of MLP causes a local loss of mitochondria. We hypothesize that this is caused by a disturbed interaction between cytoskeleton and mitochondria, which interferes with energy sensing and energy transfer. Recovery of energy depletion by stimulating mitochondrial biogenesis might be a useful therapeutic strategy for improving the energy imbalance in heart failure.

The combined effects of enhanced UV-B radiation and selenium on growth, chlorophyll fluorescence and ultrastructure in strawberry ( Fragaria × ananassa) and barley ( Hordeum vulgare) treated in the field

The possible ameliorative effects of selenium (Se) addition to soil on the detrimental effects of enhanced UV-B radiation were tested on strawberry and barley during 4 months of field experiment in Kuopio, Central Finland. Control plants were exposed to ambient levels of UV radiation, using arrays of unenergized lamps. A control for UV-A radiation was also included in the experiment. Added Se, applied as H 2SeO 4, at the level of 0.1 mg kg −1 soil (low dosage) and 1 mg kg −1 soil (high dosage) increased Se concentrations in plants more than 10 and 100 times, respectively. After 4 months of exposure, strawberry and barley plants were harvested for biomass analysis. Chlorophyll fluorescence was measured using the Hansatech FMS2 fluorescence monitoring system. Leaf anatomy and ultrastructure were observed by light and transmission electron microscope. Several effects of UV and Se as well as their interaction were found, mostly for strawberry, but not for barley, indicating species-specific responses. Our results provided evidence that the high Se concentration in soil had no ameliorative effect but increased the sensitivity of strawberry to enhanced UV-B radiation in the field. Under ambient radiation, Se did not alter leaf growth of strawberry, whereas under UV-B radiation, the high Se addition significantly decreased leaf growth. Strawberry runner biomass was affected by the interaction of Se and UV. Under ambient radiation Se did not change dry weight of runners, but in combination with UV-A or UV-B radiation the high Se dosage decreased dry weight of runners by about 30%. Although the high Se concentration positively influenced on quantum efficiency of photosystem II (PSII) in strawberry leaves, it reduced runner biomass, leaf number and ratio of starch to chloroplast area. This suggests that the harmful effects of the high Se dosage on photosynthetic processes occurred as a result of changes in activity or/and biosynthesis of enzymes, rather than alteration of PSII. At the low concentration, Se effects were slight and variable. Although barley leaves accumulated higher Se concentrations than strawberry, there were no apparent changes in their growth, biomass or chlorophyll fluorescence due to Se effect either alone or in combination with UV-B. However, at the ultrastructural level, an enlargement in the peroxisome area was found due to combination of UV radiation with Se, suggesting the activation of antioxidative enzymes, possibly catalase. Decrease in mitochondrial density in barley cells in response to Se might be attributed to alteration of mitochondrial division. Increase in the proportion of cells with cytoplasmic lipid bodies due to combined effect of UV-B and Se indicated the alteration of lipid metabolism and the acceleration of cell senescence in barley. Main UV-B effects were found, mostly at the tissue and ultrastructural level in strawberry, but not in barley, indicating species-specific susceptibility to enhanced UV-B radiation. UV-B-treated strawberry plants developed marginally thinner leaves with reduced ratio of starch to chloroplast area in their cells, suggesting negative influence of UV-B on photosynthetic processes.