Nanoscopic quantification of sub-mitochondrial morphology, mitophagy and mitochondrial dynamics in living cells derived from patients with mitochondrial diseases

AbstractSLC25A46 mutations have been found to lead to mitochondrial hyper-fusion and reduced mitochondrial respiratory function, which results in optic atrophy, cerebellar atrophy, and other clinical symptoms of mitochondrial disease. However, it is generally believed that mitochondrial fusion is attributable to increased mitochondrial oxidative phosphorylation (OXPHOS), which is inconsistent with the decreased OXPHOS of highly-fused mitochondria observed in previous studies. In this paper, we have used the live-cell nanoscope to observe and quantify the structure of mitochondrial cristae, and the behavior of mitochondria and lysosomes in patient-derived SLC25A46 mutant fibroblasts. The results show that the cristae have been markedly damaged in the mutant fibroblasts, but there is no corresponding increase in mitophagy. This study suggests that severely damaged mitochondrial cristae might be the predominant cause of reduced OXPHOS in SLC25A46 mutant fibroblasts. This study demonstrates the utility of nanoscope-based imaging for realizing the sub-mitochondrial morphology, mitophagy and mitochondrial dynamics in living cells, which may be particularly valuable for the quick evaluation of pathogenesis of mitochondrial morphological abnormalities.

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Nanoscopic Quantification of Sub-mitochondrial Morphology, Mitophagy and Mitochondrial Dynamics in Patients With Mitochondrial Disease

10.21203/rs.3.rs-105738/v1 ◽

2020 ◽

Author(s):

Weiwei Zou ◽

Qixin Chen ◽

Jesse Slone ◽

Li Yang ◽

Xiaoting Lou ◽

...

Keyword(s):

Oxidative Phosphorylation ◽

Mitochondrial Disease ◽

Optic Atrophy ◽

Respiratory Function ◽

Clinical Symptoms ◽

Mitochondrial Dynamics ◽

Cerebellar Atrophy ◽

Live Cells ◽

Mitochondrial Morphology ◽

Mitochondrial Oxidative Phosphorylation

Abstract SLC25A46 mutations have been found to lead to mitochondrial hyper-fusion and reduced mitochondrial respiratory function, which results in optic atrophy, cerebellar atrophy, and other clinical symptoms of mitochondrial disease. However, it is generally believed that mitochondrial fusion is attributable to increased mitochondrial oxidative phosphorylation (OXPHOS)[1], which is inconsistent with the decreased OXPHOS of highly-fused mitochondria observed in previous studies. In this paper, we have used the live-cell nanoscope to observe and quantify the structure of mitochondrial cristae, and the behavior of mitochondria and lysosomes in patient-derived SLC25A46 mutant fibroblasts. The results show that the crista have been markedly damaged in the mutant fibroblasts, but that there is no corresponding increase in mitophagy. This study suggests that severely damaged mitochondrial cristae might be the predominant cause of reduced OXPHOS in SLC25A46 mutant fibroblasts. This study demonstrates the utility of nanoscope-based imaging for realizing the sub-mitochondrial morphology, mitophagy and mitochondrial dynamics in live cells, which may be particularly helpful for the quick assessment and diagnosis of mitochondrial abnormalities.

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Abstract 13320: Direct Cardiac Actions of Empagliflozin Improve Mitochondrial Oxidative Phosphorylation and Attenuate Pressure-overload Heart Failure

Circulation ◽

10.1161/circ.142.suppl_3.13320 ◽

2020 ◽

Vol 142 (Suppl_3) ◽

Author(s):

Xuan Li ◽

Jussara M do Carmo ◽

Zhen Wang ◽

Alexandre A da Sivla ◽

Alan J Mouton ◽

...

Keyword(s):

Heart Failure ◽

Molecular Docking ◽

Oxidative Phosphorylation ◽

Mitochondrial Biogenesis ◽

Glucose Transporters ◽

Left Ventricular ◽

Mitochondrial Morphology ◽

Mitochondrial Oxidative Phosphorylation ◽

Docking Analysis ◽

Molecular Docking Analysis

Introduction: The underlying mechanisms by which empagliflozin (EMPA) and other sodium glucose co-transporter 2 (SGLT2) inhibitors attenuate heart failure (HF) are still poorly understood. However, this protection does not appear to be fully explained by their antihyperglycemic or diuretic effects. Hypothesis: EMPA attenuates HF by direct effects on the heart to improve its metabolism and function. Methods: C57BL/6J mice (4-6 months) were subjected to transverse aortic constriction (TAC) or sham surgeries. Two weeks after TAC, EMPA (10 mg/kg/day) or vehicle was administered daily for 4 additional weeks. Cardiac function was assessed by echocardiography and cardiac substrate metabolism measured in isolated perfused hearts. Transmission electron microscopy was used to evaluate mitochondrial morphology and molecular docking analysis to predict potential cardiac targets of EMPA. Results: EMPA increased survival and attenuated adverse left ventricle remodeling and cardiac fibrosis after TAC. EMPA also attenuated left ventricular systolic dysfunction (ejection fraction 51.6 vs. 40.2% p<0.05; fraction shortening 28.8 vs 18.4% p<0.05). EMPA rescued impaired glucose and fatty acid oxidation in failing hearts, while reducing glycolysis. Molecular docking analysis and isolated perfused heart experiments indicated that EMPA can directly bind glucose transporters in the heart to reduce glycolysis, and enhance AMP-activated protein kinase. EMPA treatment enhanced mitochondrial biogenesis, restored mitochondria cristae integrity, increased expression of endogenous antioxidants, and reduced cellular apoptosis caused by leakage of cytochrome C from mitochondria into the cytosol. These beneficial cardiac effects of EMPA occurred despite no alterations in fasting blood glucose, body weight, or daily urine volume. Conclusions. Our study demonstrated that EMPA may directly bind glucose transporters and reduce excessive glycolysis in failing hearts. EMPA enhanced mitochondrial biogenesis, improved mitochondrial oxidative phosphorylation, and reduced mitochondria-mediated apoptosis, thereby attenuating cardiac dysfunction and progression of HF.

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Mitochondrial oxidative phosphorylation in autosomal dominant optic atrophy

BMC Biochemistry ◽

10.1186/1471-2091-9-22 ◽

2008 ◽

Vol 9 (1) ◽

pp. 22 ◽

Cited By ~ 25

Author(s):

Vladimir I Mayorov ◽

Angela J Lowrey ◽

Valerie Biousse ◽

Nancy J Newman ◽

Susan D Cline ◽

...

Keyword(s):

Oxidative Phosphorylation ◽

Optic Atrophy ◽

Autosomal Dominant ◽

Mitochondrial Oxidative Phosphorylation ◽

Autosomal Dominant Optic Atrophy ◽

Dominant Optic Atrophy

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Mitochondrial Oxidative Phosphorylation Compensation May Preserve Vision in Patients with OPA1-Linked Autosomal Dominant Optic Atrophy

PLoS ONE ◽

10.1371/journal.pone.0021347 ◽

2011 ◽

Vol 6 (6) ◽

pp. e21347 ◽

Cited By ~ 30

Author(s):

Nicole J. Van Bergen ◽

Jonathan G. Crowston ◽

Lisa S. Kearns ◽

Sandra E. Staffieri ◽

Alex W. Hewitt ◽

...

Keyword(s):

Oxidative Phosphorylation ◽

Optic Atrophy ◽

Autosomal Dominant ◽

Mitochondrial Oxidative Phosphorylation ◽

Autosomal Dominant Optic Atrophy ◽

Dominant Optic Atrophy

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Oxidative Phosphorylation Dysfunction Modifies the Cell Secretome

International Journal of Molecular Sciences ◽

10.3390/ijms21093374 ◽

2020 ◽

Vol 21 (9) ◽

pp. 3374

Author(s):

Nuria Garrido-Pérez ◽

Ana Vela-Sebastián ◽

Ester López-Gallardo ◽

Sonia Emperador ◽

Eldris Iglesias ◽

...

Keyword(s):

Genetic Variability ◽

Oxidative Phosphorylation ◽

Mitochondrial Diseases ◽

Mitochondrial Oxidative Phosphorylation ◽

Circulating Biomarkers ◽

New Biomarkers ◽

Oxidative Phosphorylation Disorders ◽

Selection Of

Mitochondrial oxidative phosphorylation disorders are extremely heterogeneous conditions. Their clinical and genetic variability makes the identification of reliable and specific biomarkers very challenging. Until now, only a few studies have focused on the effect of a defective oxidative phosphorylation functioning on the cell’s secretome, although it could be a promising approach for the identification and pre-selection of potential circulating biomarkers for mitochondrial diseases. Here, we review the insights obtained from secretome studies with regard to oxidative phosphorylation dysfunction, and the biomarkers that appear, so far, to be promising to identify mitochondrial diseases. We propose two new biomarkers to be taken into account in future diagnostic trials.

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Enantiomers of flurbiprofen can distinguish key pathophysiological steps of NSAID enteropathy in the rat

Gut ◽

10.1136/gut.43.6.775 ◽

1998 ◽

Vol 43 (6) ◽

pp. 775-782 ◽

Cited By ~ 21

Author(s):

T Mahmud ◽

S Somasundaram ◽

G Sigthorsson ◽

R J Simpson ◽

S Rafi ◽

...

Keyword(s):

Oxidative Phosphorylation ◽

Intestinal Permeability ◽

Racemic Mixture ◽

Mitochondrial Morphology ◽

Intestinal Damage ◽

Mitochondrial Oxidative Phosphorylation ◽

Nsaid Enteropathy ◽

Small Intestinal

Background—Non-steroidal anti-inflammatory drugs (NSAIDs) cause gastrointestinal damage by a non-prostaglandin (PG) dependent “topical” action and by inhibiting cyclooxygenase.Aims—To discriminate between these two effects by studying some key pathophysiological steps in NSAID enteropathy following administration of (R)- and (S)-flurbiprofen, the racemic mixture, and an uncoupler, dinitrophenol.Methods—The effects of dinitrophenol, racemic, (R)-, and (S)-flurbiprofen on mitochondria were assessed in vitro and on key pathophysiological features of small intestinal damage in vivo (ultrastructure by electron microscopy, mucosal prostanoid concentrations, intestinal permeability, inflammation, and ulcer count) in rats.Results—All the drugs uncoupled mitochondrial oxidative phosphorylation in vitro, caused mitochondrial damage in vivo, and increased intestinal permeability. Dinitrophenol and (R)-flurbiprofen caused no significant decreases in mucosal prostanoid concentrations (apart from a decrease in thromboxane (TX) B2 concentrations following (R)-flurbiprofen) while racemic and (S)- flurbiprofen reduced mucosal prostanoids significantly (PGE, TXB2, and 6-keto-PGF1α concentrations by 73–95%). Intestinal inflammation was significantly greater following administration of (S)-flurbiprofen and racemate than with dinitrophenol and (R)-flurbiprofen. No small intestinal ulcers were found following dinitrophenol or (R)-flurbiprofen while both racemic and (S)-flurbiprofen caused numerous ulcers.Conclusions—Dinitrophenol and (R)-flurbiprofen show similarities in their actions to uncouple mitochondrial oxidative phosphorylation in vitro, alter mitochondrial morphology in vivo, increase intestinal permeability, and cause mild inflammation without ulcers. Concurrent severe decreases in mucosal prostanoids seem to be the driving force for the development of severe inflammation and ulcers.

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An analysis of respiratory function and mitochondrial morphology in Candida albicans

10.1101/697516 ◽

2019 ◽

Cited By ~ 1

Author(s):

Lucian Duvenage ◽

Daniel R. Pentland ◽

Carol A. Munro ◽

Campbell W. Gourlay

Keyword(s):

Candida Albicans ◽

Respiratory Function ◽

Fungal Pathogens ◽

Mitochondrial Dynamics ◽

Cell Types ◽

Mitochondrial Activity ◽

Mitochondrial Morphology ◽

Mitochondrial Network ◽

Response To Stress ◽

And Oxidative Stress

AbstractRespiratory function and mitochondrial dynamics have been well characterised in a number of cell types, including the model yeast Saccharomyces cerevisiae, but remain under-researched in fungal pathogens such as Candida albicans. An understanding of mitochondrial activity and morphology is important if we are to understand the role that this organelle plays in adaption and response to stress. Here we examine the respiratory profiles of several prominent pathogenic Candida species and present a useful GFP probe for the study of mitochondrial morphology. We examine mitochondrial morphology under a variety of conditions that Candida species may encounter within the host, such as acidic pH, respiratory and oxidative stress. The GFP probe also allowed for the visualisation of mitochondria during hyphal development, during growth following macrophage engulfment and distribution within biofilms. These data demonstrate that the mitochondrial network of C. albicans is highly responsive to both environmental conditions and developmental cues, suggesting important roles for this organelle in environmental adaption.

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High-throughput phenotypic screen for genetic modifiers in patient-derived OPA1 mutant fibroblasts identifies PGS1 as a functional suppressor of mitochondrial fragmentation

10.1101/2021.01.14.426579 ◽

2021 ◽

Author(s):

Emma Cretin ◽

Priscilla Lopes ◽

Elodie Vimont ◽

Takashi Tatsuta ◽

Thomas Langer ◽

...

Keyword(s):

High Throughput ◽

Mitochondrial Disease ◽

Optic Atrophy ◽

Mitochondrial Dynamics ◽

Mitochondrial Fission ◽

Supervised Machine Learning ◽

Mitochondrial Morphology ◽

Phenotypic Screening ◽

Genetic Modifiers ◽

Mitochondrial Fragmentation

AbstractMutations affecting the mitochondrial fusion protein Optic Atrophy 1 (OPA1) cause autosomal dominant optic atrophy (DOA) – one of the most common form of mitochondrial disease. The majority of patients develop isolated optic atrophy, but about 20% of OPA1 mutation carriers manifest more severe neurological deficits as part of a “DOA+” phenotype. OPA1 deficiency causes mitochondrial fragmentation and also disrupts cristae organization, oxidative phosphorylation, mitochondrial DNA (mtDNA) maintenance, and cell viability. It has not yet been established whether phenotypic severity can be modulated by genetic modifiers of OPA1. To better understand the genetic regulation of mitochondrial dynamics, we established a high-throughput imaging pipeline using supervised machine learning (ML) to perform unbiased, quantitative mitochondrial morphology analysis that was coupled with a bespoke siRNA library targeting the entire known mitochondrial proteome (1531 genes), providing a detailed phenotypic screening of human fibroblasts. In control fibroblasts, we identified known and novel genes whose depletion promoted elongation or fragmentation of the mitochondrial network. In DOA+ patient fibroblasts, we identified 91 candidate genes whose depletion prevents mitochondrial fragmentation, including the mitochondrial fission genes DNM1L, MIEF1, and SLC25A46, but also genes not previously linked to mitochondrial dynamics such as Phosphatidyl Glycerophosphate Synthase (PGS1), which belongs to the cardiolipin (CL) synthesis pathway. PGS1 depletion reduces CL content in mitochondria and rebalances mitochondrial dynamics in OPA1-deficient fibroblasts by inhibiting mitochondrial fission, which improves defective respiration, but does not rescue mtDNA depletion, cristae dysmorphology or apoptotic sensitivity. Our data reveal that the multifaceted roles of OPA1 in mitochondria can be functionally uncoupled by modulating mitochondrial lipid metabolism, providing novel insights into the cellular relevance of mitochondrial fragmentation. This study illustrates the power of a first-in-kind objective automated imaging approach to uncover genetic modifiers of mitochondrial disease through high-throughput phenotypic screening of patient fibroblasts.

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Dependence of Platelet Adenyl Cyclase System on Oxidative Phosphorylation

Thrombosis and Haemostasis ◽

10.1055/s-0038-1651444 ◽

1975 ◽

Vol 34 (01) ◽

pp. 042-049 ◽

Cited By ~ 1

Author(s):

Shuichi Hashimoto ◽

Sachiko Shibata ◽

Bokro Kobayashi

Keyword(s):

Cyclic Amp ◽

Oxidative Phosphorylation ◽

Sodium Succinate ◽

Potassium Cyanide ◽

Adenyl Cyclase ◽

Mitochondrial Oxidative Phosphorylation ◽

Adenyl Cyclase Activity ◽

Intact Rabbit ◽

Adenyl Cyclase System ◽

Cyclic Amp Synthesis

SummaryThe radioactive adenosine 3′,5′-monophosphate (cyclic AMP) level derived from 8-14C adenine in intact rabbit platelets decreased in the presence of mitochondrial inhibitor (potassium cyanide) or uncoupler (sodium azide), and markedly increased by the addition of NaF, monoiodoacetic acid (MIA), or 2-deoxy-D-glucose. The stimulative effect of the glycolytic inhibitors was distinctly enhanced by the simultaneous addition of sodium succinate. MIA did neither directly stimulate the adenyl cyclase activity nor inhibit the phosphodiesterase activity. These results suggest that cyclic AMP synthesis in platelets is closely linked to mitochondrial oxidative phosphorylation.

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