bims-mitdis 2023-06-04 papers

bims-mitdis

Biomed News

on Mitochondrial disorders

Issue of 2023–06–04
34 papers selected by
Catalina Vasilescu, Helmholz Munich

Healthcare resource utilization of patients with mitochondrial disease in an outpatient hospital setting.
Fluorescence-Based Quantification of Mitochondrial Membrane Potential and Superoxide Levels using Live Imaging in HeLa Cells.
Expert panel curation of 113 primary mitochondrial disease genes for the Leigh syndrome spectrum.
Cytochrome c oxidase biogenesis - from translation to early assembly of the core subunit COX1.
Mitochondrial Replication Assay (MIRA) for Efficient in situ Quantification of Nascent mtDNA and Protein Interactions with Nascent mtDNA (mitoSIRF).
Astrocytic pathology in Alpers' syndrome.
Structural basis of purine nucleotide inhibition of human uncoupling protein 1.
Efficacy and Safety of Elamipretide in Individuals With Primary Mitochondrial Myopathy: The MMPOWER-3 Randomized Clinical Trial.
Progressive mitochondrial dysfunction in cerebellar synaptosomes of cystatin B-deficient mice.
PINK1-Dependent Mitophagy Inhibits Elevated Ubiquitin Phosphorylation Caused by Mitochondrial Damage.
Isolation of Mitochondria from Mouse Tissues for Functional Analysis.
Lipid droplet-associated lncRNA LIPTER preserves cardiac lipid metabolism.
Effectors enabling adaptation to mitochondrial complex I loss in Hurthle cell carcinoma.
Rare penetrant mutations confer severe risk of common diseases.
Transitioning single-cell genomics into the clinic.
Mitochondrial pyruvate metabolism and glutaminolysis toggle steady-state and emergency myelopoiesis.
Parkinsonism in spinocerebellar ataxia with axonal neuropathy caused by adult-onset COA7 variants: a case report.
The E3 ubiquitin ligase FBXL6 controls the quality of newly synthesized mitochondrial ribosomal proteins.
M1BP is an essential transcriptional activator of oxidative metabolism during Drosophila development.
SETD5 haploinsufficiency affects mitochondrial compartment in neural cells.
Functional and structural impact of 10 ACADM missense mutations on human medium chain acyl-Coa dehydrogenase.
Measurement of Fatty Acid Oxidation by High-Resolution Respirometry: Special Considerations for Analysis of Skeletal and Cardiac Muscle and Adipose Tissue.
Mitochondria-associated endoplasmic reticulum membranes (MAMs) and their role in glaucomatous retinal ganglion cell degeneration-a mini review.
The genetic and phenotypic correlates of mtDNA copy number in a multi-ancestry cohort.
When and why are mitochondria paternally inherited?
Method for detecting CoQ10 incorporation in the mitochondrial respiratory chain supercomplex.
Challenges and strategies for clinical trials in propionic and methylmalonic acidemias.
Genomics of aging: Reactive oxidation and inefficient mitochondria.
Targeting NAD Metabolism for the Therapy of Age-Related Neurodegenerative Diseases.
Machine learning in rare disease.
Fibroblast growth factor 21 as a candidate of a novel serum biomarker for mitochondrial diabetes.
The contributions of mitochondrial and nuclear mitochondrial genetic variation to neuroticism.
Low Plasma Citrulline Guiding the Diagnosis of a Mitochondrial Disorder.
Novel ITPA variants identified by whole genome sequencing and RNA sequencing.

Orphanet J Rare Dis. 2023 05 29. 18(1): 129

Healthcare resource utilization of patients with mitochondrial disease in an outpatient hospital setting.

Sameen Haque, Karen Crawley, Rupendra Shrestha, Deborah Schofield, Carolyn M Sue.

   BACKGROUND AND OBJECTIVES: Mitochondrial diseases present as multi-system disorders requiring a comprehensive multidisciplinary approach. The data on healthcare resource utilization associated with mitochondrial diseases and the clinical drivers of these costs are limited including for the out-patient setting where the majority of the clinical care for mitochondrial disease patients occurs. We performed a cross-sectional retrospective study of out-patient healthcare resource utilization and costs for patients with a confirmed diagnosis of mitochondrial disease.
METHODS: We recruited participants from the Mitochondrial Disease Clinic in Sydney and stratified them into three groups: those with mitochondrial DNA (mtDNA) mutations (Group 1), those with nuclear DNA (nDNA) mutations and the predominant phenotype of chronic progressive external ophthalmoplegia (CPEO) or optic atrophy (Group 2) and those without a confirmed genetic diagnosis but clinical criteria and muscle biopsy findings supportive of a diagnosis of mitochondrial disease (Group 3). Data was collected through retrospective chart review and out-patient costs were calculated using the Medicare Benefits Schedule.
RESULTS: We analyzed the data from 91 participants and found that Group 1 had the greatest average out-patient costs per person per annum ($838.02; SD 809.72). Neurological investigations were the largest driver of outpatient healthcare costs in all groups (average costs per person per annum:-Group 1: $364.11; SD 340.93, Group 2: $247.83; SD 113.86 and Group 3: $239.57; SD 145.69) consistent with the high frequency (94.5%) of neurological symptoms. Gastroenterological and cardiac-related out-patient costs were also major contributors to out-patient healthcare resource utilization in Groups 1 and 3. In Group 2, ophthalmology was the second-most resource intensive specialty ($136.85; SD 173.35). The Group 3 had the greatest average healthcare resource utilization per person over the entire duration of out-patient clinic care ($5815.86; SD 3520.40) most likely due to the lack of a molecular diagnosis and a less customized management approach.
CONCLUSION: The drivers of healthcare resource utilization are dependent on the phenotype-genotype characteristics. Neurological, cardiac, and gastroenterological costs were the top three drivers in the out-patient clinics unless the patient had nDNA mutations with predominant phenotype of CPEO and/or optic atrophy wherein ophthalmological-related costs were the second most resource intensive driver.

Keywords:  Health care costs; Health resources; Health services; Mitochondrial diseases; Mitochondrial disorders; Outpatients

DOI:  https://doi.org/10.1186/s13023-023-02746-x
J Vis Exp. 2023 05 12.

Fluorescence-Based Quantification of Mitochondrial Membrane Potential and Superoxide Levels using Live Imaging in HeLa Cells.

Mohammad Fazli, Chantell S Evans.

Mitochondria are dynamic organelles critical for metabolic homeostasis by controlling energy production via ATP synthesis. To support cellular metabolism, various mitochondrial quality control mechanisms cooperate to maintain a healthy mitochondrial network. One such pathway is mitophagy, where PTEN-induced kinase 1 (PINK1) and Parkin phospho-ubiquitination of damaged mitochondria facilitate autophagosome sequestration and subsequent removal from the cell via lysosome fusion. Mitophagy is important for cellular homeostasis, and mutations in Parkin are linked to Parkinson's disease (PD). Due to these findings, there has been a significant emphasis on investigating mitochondrial damage and turnover to understand the molecular mechanisms and dynamics of mitochondrial quality control. Here, live-cell imaging was used to visualize the mitochondrial network of HeLa cells, to quantify the mitochondrial membrane potential and superoxide levels following treatment with carbonyl cyanide m-chlorophenyl hydrazone (CCCP), a mitochondrial uncoupling agent. In addition, a PD-linked mutation of Parkin (ParkinT240R) that inhibits Parkin-dependent mitophagy was expressed to determine how mutant expression impacts the mitochondrial network compared to cells expressing wild-type Parkin. The protocol outlined here describes a simple workflow using fluorescence-based approaches to quantify mitochondrial membrane potential and superoxide levels effectively.

DOI: https://doi.org/10.3791/65304
Ann Neurol. 2023 May 31.

Expert panel curation of 113 primary mitochondrial disease genes for the Leigh syndrome spectrum.

NICHD ClinGen U24 Mitochondrial Disease Gene Curation Expert Panel

OBJECTIVE: Primary mitochondrial diseases (PMDs) are heterogeneous disorders caused by inherited mitochondrial dysfunction. Classically defined neuropathologically as subacute necrotizing encephalomyelopathy, Leigh syndrome spectrum (LSS) is the most frequent manifestation of PMD in children, but may also present in adults. A major challenge for accurate diagnosis of LSS in the genomic medicine era is establishing gene-disease relationships (GDRs) for this syndrome with >100 monogenic causes across both nuclear and mitochondrial genomes.
METHODS: The Clinical Genome Resource (ClinGen) Mitochondrial Disease Gene Curation Expert Panel (GCEP), comprising 40 international PMD experts, met monthly for 4 years to review GDRs for LSS. The GCEP standardized gene curation for LSS by refining the phenotypic definition, modifying the ClinGen Gene-Disease Clinical Validity Curation Framework to improve interpretation for LSS, and establishing a scoring rubric for LSS.
RESULTS: The GDR with LSS across the nuclear and mitochondrial genomes was classified as definitive for 31/114 gene-disease relationships curated (27%); moderate for 38 (33%); limited for 43 (38%); and 2 as disputed (2%). Ninety genes were associated with autosomal recessive inheritance, 16 were maternally inherited, 5 autosomal dominant, and 3 X-linked.
INTERPRETATION: GDRs for LSS were established for genes across both nuclear and mitochondrial genomes. Establishing these GDRs will allow accurate variant interpretation, expedite genetic diagnosis of LSS, and facilitate precision medicine, multi-system organ surveillance, recurrence risk counselling, reproductive choice, natural history studies and eligibility for interventional clinical trials. This article is protected by copyright. All rights reserved.

DOI: https://doi.org/10.1002/ana.26716
FEBS Lett. 2023 May 29.

Cytochrome c oxidase biogenesis - from translation to early assembly of the core subunit COX1.

Sven Dennerlein, Peter Rehling, Ricarda Richter-Dennerlein.

  Mitochondria are the powerhouses of the cell as they produce the majority of ATP with their oxidative phosphorylation (OXPHOS) machinery. The OXPHOS system is composed of the F1 Fo ATP synthase and four mitochondrial respiratory chain complexes, the terminal enzyme of which is the cytochrome c oxidase (complex IV) that transfers electrons to oxygen, generating water. Complex IV comprises of 14 structural subunits of dual genetic origin: while the three core subunits are mitochondrial encoded, the remaining constituents are encoded by the nuclear genome. Hence, the assembly of complex IV requires the coordination of two spatially separated gene expression machinery. Recent efforts elucidated an increasing number of proteins involved in mitochondrial gene expression, which are linked to complex IV assembly. Additionally, several COX1 biogenesis factors have been intensively biochemically investigated and an increasing number of structural snapshots shed light on the organization of macromolecular complexes such as the mitoribosome or the cytochrome c oxidase. Here, we focus on COX1 translation regulation and highlight the advanced understanding of early steps during COX1 assembly and its link to mitochondrial translation regulation.

Keywords:  COX1; OXPHOS; complex IV; cytochrome c oxidase; mitochondria

DOI:  https://doi.org/10.1002/1873-3468.14671
Bio Protoc. 2023 May 20. 13(10): e4680

Mitochondrial Replication Assay (MIRA) for Efficient in situ Quantification of Nascent mtDNA and Protein Interactions with Nascent mtDNA (mitoSIRF).

Macy Lozen, Yue Chen, Rebecca A Boisvert, Katharina Schlacher.

  Mitochondria play decisive roles in bioenergetics and intracellular communication. These organelles contain a circular mitochondrial DNA (mtDNA) genome that is duplicated within one to two hours by a mitochondrial replisome, independently from the nuclear replisome. mtDNA stability is regulated in part at the level of mtDNA replication. Consequently, mutations in mitochondrial replisome components result in mtDNA instability and are associated with diverse disease phenotypes, including premature aging, aberrant cellular energetics, and developmental defects. The mechanisms ensuring mtDNA replication stability are not completely understood. Thus, there remains a need to develop tools to specifically and quantifiably examine mtDNA replication. To date, methods for labeling mtDNA have relied on prolonged exposures of 5'-bromo-2'-deoxyuridine (BrdU) or 5'-ethynyl-2'-deoxyuridine (EdU). However, labeling with these nucleoside analogs for a sufficiently short time in order to monitor nascent mtDNA replication, such as under two hours, does not produce signals suited for efficient or accurate quantitative analysis. The assay system described here, termed Mitochondrial Replication Assay (MIRA), utilizes proximity ligation assay (PLA) combined with EdU-coupled Click-IT chemistry to address this limitation, thereby enabling sensitive and quantitative analysis of nascent in situ mtDNA replication with single-cell resolution. This method can be further paired with conventional immunofluorescence (IF) for multi-parameter cell analysis. By enabling monitoring nascent mtDNA prior to the complete replication of the entire mtDNA genome, this new assay system allowed the discovery of a new mitochondrial stability pathway, mtDNA fork protection. Moreover, a modification in primary antibodies application allows the adaptation of our previously described in situ protein Interactions with nascent DNA Replication Forks (SIRF) for the detection of proteins of interest to nascent mtDNA replication forks on a single molecule level (mitoSIRF). Graphical overview Schematic overview of Mitochondrial Replication Assay (MIRA). 5'-ethynyl-2'-deoxyuridine (EdU; green) incorporated in DNA is tagged with biotin (blue) using Click-IT chemistry. Subsequent proximity ligation assay (PLA, pink circles) using antibodies against biotin allows the fluorescent tagging of the nascent EdU and amplification of the signal sufficient for visualization by standard immunofluorescence. PLA signals outside the nucleus denote mitochondrial DNA (mtDNA) signals. Ab, antibody. In in situ protein interactions with nascent DNA replication forks (mitoSIRF), one of the primary antibodies is directed against a protein of interest, while the other detects nascent biotinylated EdU, thus enabling in situ protein interactions with nascent mtDNA.

Keywords:  BRCA2; Fanconi anemia; MIRA; Mitochondria; Mitochondrial DNA; Proximity ligation assay; mtDNA instability; mtDNA replication

DOI:  https://doi.org/10.21769/BioProtoc.4680
Acta Neuropathol Commun. 2023 May 31. 11(1): 86

Astrocytic pathology in Alpers' syndrome.

Laura A Smith, Chun Chen, Nichola Z Lax, Robert W Taylor, Daniel Erskine, Robert McFarland.

  Refractory epilepsy is the main neurological manifestation of Alpers' syndrome, a severe childhood-onset mitochondrial disease caused by bi-allelic pathogenic variants in the mitochondrial DNA (mtDNA) polymerase gamma gene (POLG). The pathophysiological mechanisms underpinning neuronal hyperexcitabilty leading to seizures in Alpers' syndrome remain unknown. However, pathological changes to reactive astrocytes are hypothesised to exacerbate neural dysfunction and seizure-associated cortical activity in POLG-related disease. Therefore, we sought to phenotypically characterise astrocytic pathology in Alpers' syndrome. We performed a detailed quantitative investigation of reactive astrocytes in post-mortem neocortical tissues from thirteen patients with Alpers' syndrome, eight neurologically normal controls and five sudden unexpected death in epilepsy (SUDEP) patients, to control for generalised epilepsy-associated astrocytic pathology. Immunohistochemistry to identify glial fibrillary acidic protein (GFAP)-reactive astrocytes revealed striking reactive astrogliosis localised to the primary visual cortex of Alpers' syndrome tissues, characterised by abnormal-appearing hypertrophic astrocytes. Phenotypic characterisation of individual GFAP-reactive astrocytes demonstrated decreased abundance of mitochondrial oxidative phosphorylation (OXPHOS) proteins and altered expression of key astrocytic proteins including Kir4.1 (subunit of the inwardly rectifying K+ ion channel), AQP4 (astrocytic water channel) and glutamine synthetase (enzyme that metabolises glutamate). These phenotypic astrocytic changes were typically different from the pathology observed in SUDEP tissues, suggesting alternative mechanisms of astrocytic dysfunction between these epilepsies. Crucially, our findings provide further evidence of occipital lobe involvement in Alpers' syndrome and support the involvement of reactive astrocytes in the pathogenesis of POLG-related disease.

Keywords:  Alpers’ syndrome; Aquaporin 4 (AQP4); GFAP; Glutamine synthetase (GS); Kir4.1; Mitochondrial Epilepsy; POLG; Reactive astrogliosis

DOI:  https://doi.org/10.1186/s40478-023-01579-w
Sci Adv. 2023 Jun 02. 9(22): eadh4251

Structural basis of purine nucleotide inhibition of human uncoupling protein 1.

Scott A Jones, Prerana Gogoi, Jonathan J Ruprecht, Martin S King, Yang Lee, Thomas Zögg, Els Pardon, Deepak Chand, Stefan Steimle, Danielle M Copeman, Camila A Cotrim, Jan Steyaert, Paul G Crichton, Vera Moiseenkova-Bell, Edmund R S Kunji.

Mitochondrial uncoupling protein 1 (UCP1) gives brown adipose tissue of mammals its specialized ability to burn calories as heat for thermoregulation. When activated by fatty acids, UCP1 catalyzes the leak of protons across the mitochondrial inner membrane, short-circuiting the mitochondrion to generate heat, bypassing ATP synthesis. In contrast, purine nucleotides bind and inhibit UCP1, regulating proton leak by a molecular mechanism that is unclear. We present the cryo-electron microscopy structure of the GTP-inhibited state of UCP1, which is consistent with its nonconducting state. The purine nucleotide cross-links the transmembrane helices of UCP1 with an extensive interaction network. Our results provide a structural basis for understanding the specificity and pH dependency of the regulatory mechanism. UCP1 has retained all of the key functional and structural features required for a mitochondrial carrier-like transport mechanism. The analysis shows that inhibitor binding prevents the conformational changes that UCP1 uses to facilitate proton leak.

DOI: https://doi.org/10.1126/sciadv.adh4251
Neurology. 2023 Jun 02. pii: 10.1212/WNL.0000000000207402. [Epub ahead of print]

Efficacy and Safety of Elamipretide in Individuals With Primary Mitochondrial Myopathy: The MMPOWER-3 Randomized Clinical Trial.

MMPOWER-3 Trial Investigators

   BACKGROUND AND OBJECTIVES: Primary Mitochondrial Myopathies (PMMs) encompass a group of genetic disorders that impair mitochondrial oxidative phosphorylation, adversely impacting physical function, exercise capacity, and quality of life (QoL). Current PMM standards-of-care address symptoms, with limited clinical impact, constituting a significant therapeutic unmet need. We present data from MMPOWER-3, a pivotal, phase-3, randomized, double-blind, placebo-controlled clinical trial that evaluated the efficacy and safety of elamipretide in participants with genetically-confirmed PMM.
METHODS: Following screening, eligible participants were randomized 1:1 to receive either 24weeks of elamipretide 40mg/day or placebo subcutaneously. Primary efficacy endpoints included change from baseline to Week 24 on the distance walked on the 6-minute Walk Test (6MWT), and Total Fatigue on the Primary Mitochondrial Myopathy Symptom Assessment (PMMSA). Secondary endpoints included Most Bothersome Symptom Score on the PMMSA, NeuroQoL Fatigue Short Form scores, and the Patient- and Clinician-Global Impression of PMM Symptoms.
RESULTS: Participants (N=218) were randomized (n=109 elamipretide; n=109 placebo). Mean age was 45.6 year (64% women; 94% white). The majority of participants (n=162 [74%]) had mitochondrial DNA (mtDNA) mutations, with the remainder having nuclear DNA (nDNA) defects. At screening, the most frequent bothersome PMM symptom on the PMMSA was tiredness during activities (28.9%). At baseline, mean distance walked on the 6MWT was 336.7±81.2 meters, mean score for Total Fatigue on the PMMSA was 10.6±2.5, and mean T-score for the Neuro-QoL Fatigue Short Form was 54.7±7.5. The study did not meet its primary endpoints assessing changes in the 6MWT and PMMSA Total Fatigue Score (TFS). Between the participants receiving elamipretide versus placebo, the difference in the Least Squares Mean (SE) from baseline to Week 24 on distance walked on the 6MWT was -3.2 (95% confidence interval,-18.7,12.3; p=0.69) meters and on the PMMSA Total Fatigue Score was -0.07 (95% confidence interval,-0.10,0.26; p=0.37). Elamipretide treatment was well-tolerated with most adverse events being mild to moderate in severity.
DISCUSSION: Subcutaneous elamipretide treatment did not improve outcomes in the 6MWT and PMMSA TFS in patients with PMM. However, this phase-3 study demonstrated that subcutaneous elamipretide is well-tolerated.
TRIAL REGISTRATION INFORMATION: Trial registered with clinicaltrials.gov, Clinical Trials Identifier: NCT03323749; submitted on October 12, 2017;first patient enrolled October 9, 2017. https://clinicaltrials.gov/ct2/show/NCT03323749?term=elamipretide&draw=2&rank=9 CLASSIFICATION OF EVIDENCE: This study provides Class I evidence that elamipretide does not improve the 6 minute walk test or fatigue at 24 weeks compared to placebo in patients with primary mitochondrial myopathy.

Keywords:  elamipretide; exercise intolerance; myopathy; primary mitochondrial disease; primary mitochondrial myopathy

DOI:  https://doi.org/10.1212/WNL.0000000000207402
Front Mol Neurosci. 2023 ;16 1175851

Progressive mitochondrial dysfunction in cerebellar synaptosomes of cystatin B-deficient mice.

Katarin Gorski, Christopher B Jackson, Tuula A Nyman, Veronika Rezov, Brendan J Battersby, Anna-Elina Lehesjoki.

  The involvement of mitochondrial dysfunction in cystatin B (CSTB) deficiency has been suggested, but its role in the onset of neurodegeneration, myoclonus, and ataxia in the CSTB-deficient mouse model (Cstb-/-) is yet unknown. CSTB is an inhibitor of lysosomal and nuclear cysteine cathepsins. In humans, partial loss-of-function mutations cause the progressive myoclonus epilepsy neurodegenerative disorder, EPM1. Here we applied proteome analysis and respirometry on cerebellar synaptosomes from early symptomatic (Cstb-/-) mice to identify the molecular mechanisms involved in the onset of CSTB-deficiency associated neural pathogenesis. Proteome analysis showed that CSTB deficiency is associated with differential expression of mitochondrial and synaptic proteins, and respirometry revealed a progressive impairment in mitochondrial function coinciding with the onset of myoclonus and neurodegeneration in (Cstb-/-) mice. This mitochondrial dysfunction was not associated with alterations in mitochondrial DNA copy number or membrane ultrastructure. Collectively, our results show that CSTB deficiency generates a defect in synaptic mitochondrial bioenergetics that coincides with the onset and progression of the clinical phenotypes, and thus is likely a contributor to the pathogenesis of EPM1.

Keywords:  OXPHOS; mitochondria; myoclonus; neurodegeneration; proteomics; respiration; synaptosome

DOI:  https://doi.org/10.3389/fnmol.2023.1175851
J Med Chem. 2023 May 29.

PINK1-Dependent Mitophagy Inhibits Elevated Ubiquitin Phosphorylation Caused by Mitochondrial Damage.

Olivia A Lambourne, Shane Bell, Léa P Wilhelm, Erika B Yarbrough, Gabriel G Holly, Oliver M Russell, Arwa M Alghamdi, Azeza M Fdel, Carmine Varricchio, Emma L Lane, Ian G Ganley, Arwyn T Jones, Matthew S Goldberg, Youcef Mehellou.

Ubiquitin phosphorylation by the mitochondrial protein kinase PTEN-induced kinase 1 (PINK1), upon mitochondrial depolarization, is an important intermediate step in the recycling of damaged mitochondria via mitophagy. As mutations in PINK1 can cause early-onset Parkinson's disease (PD), there has been a growing interest in small-molecule activators of PINK1-mediated mitophagy as potential PD treatments. Herein, we show that N6-substituted adenosines, such as N6-(2-furanylmethyl)adenosine (known as kinetin riboside) and N6-benzyladenosine, activate PINK1 in HeLa cells and induce PINK1-dependent mitophagy in primary mouse fibroblasts. Interestingly, pre-treatment of HeLa cells and astrocytes with these compounds inhibited elevated ubiquitin phosphorylation that is induced by established mitochondrial depolarizing agents, carbonyl cyanide m-chlorophenyl-hydrazine and niclosamide. Together, this highlights N6-substituted adenosines as progenitor PINK1 activators that could potentially be developed, in the future, as treatments for aged and sporadic PD patients who have elevated phosphorylated ubiquitin levels in the brain.

DOI: https://doi.org/10.1021/acs.jmedchem.3c00555
Methods Mol Biol. 2023 ;2675 77-96

Isolation of Mitochondria from Mouse Tissues for Functional Analysis.

Rebeca Acín-Pérez, Katrina P Montales, Kaitlyn B Nguyen, Alexandra J Brownstein, Linsey Stiles, Ajit S Divakaruni.

  Methods for isolating mitochondria from different rodent tissues have been established for decades. Although the general principles for crude mitochondrial preparations are largely shared across tissues - tissue disruption followed by differential centrifugation - critical differences exist for isolation from different tissues to optimize mitochondrial yield and function. This protocol offers a unified resource for preparations of isolated mitochondria from mouse liver, kidney, heart, brain, skeletal muscle, and brown and white adipose tissue suitable for functional analysis.

Keywords:  Bioenergetics; Brain; Brown adipose tissue; Heart; Kidney; Liver; Mitochondria; Oxidative phosphorylation; Skeletal muscle; White adipose tissue

DOI:  https://doi.org/10.1007/978-1-0716-3247-5_7
Nat Cell Biol. 2023 Jun 01.

Lipid droplet-associated lncRNA LIPTER preserves cardiac lipid metabolism.

Lei Han, Dayang Huang, Shiyong Wu, Sheng Liu, Cheng Wang, Yi Sheng, Xiongbin Lu, Hal E Broxmeyer, Jun Wan, Lei Yang.

Lipid droplets (LDs) are cellular organelles critical for lipid homeostasis, with intramyocyte LD accumulation implicated in metabolic disorder-associated heart diseases. Here we identify a human long non-coding RNA, Lipid-Droplet Transporter (LIPTER), essential for LD transport in human cardiomyocytes. LIPTER binds phosphatidic acid and phosphatidylinositol 4-phosphate on LD surface membranes and the MYH10 protein, connecting LDs to the MYH10-ACTIN cytoskeleton and facilitating LD transport. LIPTER and MYH10 deficiencies impair LD trafficking, mitochondrial function and survival of human induced pluripotent stem cell-derived cardiomyocytes. Conditional Myh10 deletion in mouse cardiomyocytes leads to LD accumulation, reduced fatty acid oxidation and compromised cardiac function. We identify NKX2.5 as the primary regulator of cardiomyocyte-specific LIPTER transcription. Notably, LIPTER transgenic expression mitigates cardiac lipotoxicity, preserves cardiac function and alleviates cardiomyopathies in high-fat-diet-fed and Leprdb/db mice. Our findings unveil a molecular connector role of LIPTER in intramyocyte LD transport, crucial for lipid metabolism of the human heart, and hold significant clinical implications for treating metabolic syndrome-associated heart diseases.

DOI: https://doi.org/10.1038/s41556-023-01162-4
Cancer Discov. 2023 Jun 01. pii: CD-22-0976. [Epub ahead of print]

Effectors enabling adaptation to mitochondrial complex I loss in Hurthle cell carcinoma.

Raj K Gopal, Venkata R Vantaku, Apekshya Panda, Bryn Reimer, Sneha Rath, Tsz-Leung To, Adam S Fisch, Murat Cetinbas, Maia Livneh, Michael J Calcaterra, Benjamin J Gigliotti, Kerry A Pierce, Clary B Clish, Dora Dias-Santagata, Peter M Sadow, Lori J Wirth, Gilbert H Daniels, Ruslan I Sadreyev, Sarah E Calvo, Sareh Parangi, Vamsi K Mootha.

Oncocytic (Hurthle cell) carcinoma of the thyroid (HCC) is genetically characterized by complex I mitochondrial DNA mutations and widespread chromosomal losses. Here, we utilize RNA-seq and metabolomics to identify candidate molecular effectors activated by these genetic drivers. We find glutathione biosynthesis, amino acid metabolism, mitochondrial unfolded protein response, and lipid peroxide scavenging to be increased in HCC. A CRISPR-Cas9 knockout screen in a new HCC model reveals which pathways are key for fitness, and highlights loss of GPX4, a defense against lipid peroxides and ferroptosis, as a strong liability. Rescuing complex I redox activity with the yeast NADH dehydrogenase (NDI1) in HCC cells diminishes ferroptosis sensitivity, while inhibiting complex I in normal thyroid cells augments ferroptosis induction. Our work demonstrates unmitigated lipid peroxide stress to be an HCC vulnerability that is mechanistically coupled to the genetic loss of mitochondrial complex I activity.

DOI: https://doi.org/10.1158/2159-8290.CD-22-0976
Science. 2023 Jun 02. 380(6648): eabo1131

Rare penetrant mutations confer severe risk of common diseases.

Petko P Fiziev, Jeremy McRae, Jacob C Ulirsch, Jacqueline S Dron, Tobias Hamp, Yanshen Yang, Pierrick Wainschtein, Zijian Ni, Joshua G Schraiber, Hong Gao, Dylan Cable, Yair Field, Francois Aguet, Marc Fasnacht, Ahmed Metwally, Jeffrey Rogers, Tomas Marques-Bonet, Heidi L Rehm, Anne O'Donnell-Luria, Amit V Khera, Kyle Kai-How Farh.

We examined 454,712 exomes for genes associated with a wide spectrum of complex traits and common diseases and observed that rare, penetrant mutations in genes implicated by genome-wide association studies confer ~10-fold larger effects than common variants in the same genes. Consequently, an individual at the phenotypic extreme and at the greatest risk for severe, early-onset disease is better identified by a few rare penetrant variants than by the collective action of many common variants with weak effects. By combining rare variants across phenotype-associated genes into a unified genetic risk model, we demonstrate superior portability across diverse global populations compared with common-variant polygenic risk scores, greatly improving the clinical utility of genetic-based risk prediction.

DOI: https://doi.org/10.1126/science.abo1131
Nat Rev Genet. 2023 May 31.

Transitioning single-cell genomics into the clinic.

Jennifer Lim, Venessa Chin, Kirsten Fairfax, Catia Moutinho, Dan Suan, Hanlee Ji, Joseph E Powell.

The use of genomics is firmly established in clinical practice, resulting in innovations across a wide range of disciplines such as genetic screening, rare disease diagnosis and molecularly guided therapy choice. This new field of genomic medicine has led to improvements in patient outcomes. However, most clinical applications of genomics rely on information generated from bulk approaches, which do not directly capture the genomic variation that underlies cellular heterogeneity. With the advent of single-cell technologies, research is rapidly uncovering how genomic data at cellular resolution can be used to understand disease pathology and mechanisms. Both DNA-based and RNA-based single-cell technologies have the potential to improve existing clinical applications and open new application spaces for genomics in clinical practice, with oncology, immunology and haematology poised for initial adoption. However, challenges in translating cellular genomics from research to a clinical setting must first be overcome.

DOI: https://doi.org/10.1038/s41576-023-00613-w
J Exp Med. 2023 Sep 04. pii: e20221373. [Epub ahead of print]220(9):

Mitochondrial pyruvate metabolism and glutaminolysis toggle steady-state and emergency myelopoiesis.

Hannah A Pizzato, Yahui Wang, Michael J Wolfgang, Brian N Finck, Gary J Patti, Deepta Bhattacharya.

To define the metabolic requirements of hematopoiesis, we examined blood lineages in mice conditionally deficient in genes required for long-chain fatty acid oxidation (Cpt2), glutaminolysis (Gls), or mitochondrial pyruvate import (Mpc2). Genetic ablation of Cpt2 or Gls minimally impacted most blood lineages. In contrast, deletion of Mpc2 led to a sharp decline in mature myeloid cells and a slower reduction in T cells, whereas other hematopoietic lineages were unaffected. Yet MPC2-deficient monocytes and neutrophils rapidly recovered due to a transient and specific increase in myeloid progenitor proliferation. Competitive bone marrow chimera and stable isotope tracing experiments demonstrated that this proliferative burst was progenitor intrinsic and accompanied by a metabolic switch to glutaminolysis. Myeloid recovery after loss of MPC2 or cyclophosphamide treatment was delayed in the absence of GLS. Reciprocally, MPC2 was not required for myeloid recovery after cyclophosphamide treatment. Thus, mitochondrial pyruvate metabolism maintains myelopoiesis under steady-state conditions, while glutaminolysis in progenitors promotes emergency myelopoiesis.

DOI: https://doi.org/10.1084/jem.20221373
BMC Neurol. 2023 Jun 01. 23(1): 211

Parkinsonism in spinocerebellar ataxia with axonal neuropathy caused by adult-onset COA7 variants: a case report.

Shogo Ouchi, Kazuhiro Ishii, Kenjiro Kosaki, Hisato Suzuki, Mamiko Yamada, Toshiki Takenouchi, Akira Tamaoka.

   BACKGROUND: Individuals with variants of cytochrome c oxidase assembly factor 7 (COA7), a mitochondrial functional-related gene, exhibit symptoms of spinocerebellar ataxia with axonal neuropathy before the age of 20. However, COA7 variants with parkinsonism or adult-onset type cases have not been described.
CASE PRESENTATION: We report the case of a patient who developed cerebellar symptoms and slowly progressive sensory and motor neuropathy in the extremities, similar to Charcot-Marie-Tooth disease, at age 30, followed by parkinsonism at age 58. Exome analysis revealed COA7 missense mutation in homozygotes (NM_023077.2:c.17A > G, NP_075565.2: p.Asp6Gly). Dopamine transporter single-photon emission computed tomography using a 123I-Ioflupane revealed clear hypo-accumulation in the bilateral striatum. However, 123I-metaiodobenzylguanidine myocardial scintigraphy showed normal sympathetic nerve function. Levodopa administration improved parkinsonism in this patient.
CONCLUSIONS: COA7 gene variants may have caused parkinsonism in this case because mitochondrial function-related genes, such as parkin and PINK1, are known causative genes in some familial Parkinson's diseases.

Keywords:  COA7; Charcot-Marie-Tooth disease; Parkinsonism; Spinocerebellar ataxia

DOI:  https://doi.org/10.1186/s12883-023-03202-w
Cell Rep. 2023 May 31. pii: S2211-1247(23)00590-9. [Epub ahead of print]42(6): 112579

The E3 ubiquitin ligase FBXL6 controls the quality of newly synthesized mitochondrial ribosomal proteins.

Julie Lavie, Claude Lalou, Walid Mahfouf, Jean-William Dupuy, Aurélie Lacaule, Agata Ars Cywinska, Didier Lacombe, Anne-Marie Duchêne, Anne-Aurélie Raymond, Hamid Reza Rezvani, Richard Patryk Ngondo, Giovanni Bénard.

  In mammals, about 99% of mitochondrial proteins are synthesized in the cytosol as precursors that are subsequently imported into the organelle. The mitochondrial health and functions rely on an accurate quality control of these imported proteins. Here, we show that the E3 ubiquitin ligase F box/leucine-rich-repeat protein 6 (FBXL6) regulates the quality of cytosolically translated mitochondrial proteins. Indeed, we found that FBXL6 substrates are newly synthesized mitochondrial ribosomal proteins. This E3 binds to chaperones involved in the folding and trafficking of newly synthesized peptide and to ribosomal-associated quality control proteins. Deletion of these interacting partners is sufficient to hamper interactions between FBXL6 and its substrate. Furthermore, we show that cells lacking FBXL6 fail to degrade specifically mistranslated mitochondrial ribosomal proteins. Finally, showing the role of FBXL6-dependent mechanism, FBXL6-knockout (KO) cells display mitochondrial ribosomal protein aggregations, altered mitochondrial metabolism, and inhibited cell cycle in oxidative conditions.

Keywords:  CP: Cell biology; F box leucin-rich repeat E3 ubiquitin ligase; FBXL6; mitochondria; protein quality control; ribosomal proteins

DOI:  https://doi.org/10.1016/j.celrep.2023.112579
Nat Commun. 2023 Jun 02. 14(1): 3187

M1BP is an essential transcriptional activator of oxidative metabolism during Drosophila development.

Gabriela Poliacikova, Marine Barthez, Thomas Rival, Aïcha Aouane, Nuno Miguel Luis, Fabrice Richard, Fabrice Daian, Nicolas Brouilly, Frank Schnorrer, Corinne Maurel-Zaffran, Yacine Graba, Andrew J Saurin.

Oxidative metabolism is the predominant energy source for aerobic muscle contraction in adult animals. How the cellular and molecular components that support aerobic muscle physiology are put in place during development through their transcriptional regulation is not well understood. Using the Drosophila flight muscle model, we show that the formation of mitochondria cristae harbouring the respiratory chain is concomitant with a large-scale transcriptional upregulation of genes linked with oxidative phosphorylation (OXPHOS) during specific stages of flight muscle development. We further demonstrate using high-resolution imaging, transcriptomic and biochemical analyses that Motif-1-binding protein (M1BP) transcriptionally regulates the expression of genes encoding critical components for OXPHOS complex assembly and integrity. In the absence of M1BP function, the quantity of assembled mitochondrial respiratory complexes is reduced and OXPHOS proteins aggregate in the mitochondrial matrix, triggering a strong protein quality control response. This results in isolation of the aggregate from the rest of the matrix by multiple layers of the inner mitochondrial membrane, representing a previously undocumented mitochondrial stress response mechanism. Together, this study provides mechanistic insight into the transcriptional regulation of oxidative metabolism during Drosophila development and identifies M1BP as a critical player in this process.

DOI: https://doi.org/10.1038/s41467-023-38986-5
Mol Autism. 2023 Jun 01. 14(1): 20

SETD5 haploinsufficiency affects mitochondrial compartment in neural cells.

Mattia Zaghi, Fabiana Longo, Luca Massimino, Alicia Rubio, Simone Bido, Pietro Giuseppe Mazzara, Edoardo Bellini, Federica Banfi, Paola Podini, Francesca Maltecca, Alessio Zippo, Vania Broccoli, Alessandro Sessa.

BACKGROUND: Neurodevelopmental disorders (NDDs) are heterogeneous conditions due to alterations of a variety of molecular mechanisms and cell dysfunctions. SETD5 haploinsufficiency leads to NDDs due to chromatin defects. Epigenetic basis of NDDs has been reported in an increasing number of cases while mitochondrial dysfunctions are more common within NDD patients than in the general population.
METHODS: We investigated in vitro neural stem cells as well as the brain of the Setd5 haploinsufficiency mouse model interrogating its transcriptome, analyzing mitochondrial structure, biochemical composition, and dynamics, as well as mitochondrial functionality.
RESULTS: Mitochondrial impairment is facilitated by transcriptional aberrations originated by the decrease of the SETD5 enzyme. Low levels of SETD5 resulted in fragmented mitochondria, reduced mitochondrial membrane potential, and ATP production both in neural precursors and neurons. Mitochondria were also mislocalized in mutant neurons, with reduced organelles within neurites and synapses.
LIMITATIONS: We found several defects in the mitochondrial compartment; however, we can only speculate about their position in the hierarchy of the pathological mechanisms at the basis of the disease.
CONCLUSIONS: Our study explores the interplay between chromatin regulation and mitochondria functions as a possible important aspect of SETD5-associated NDD pathophysiology. Our data, if confirmed in patient context, suggest that the mitochondrial activity and dynamics may represent new therapeutic targets for disorders associated with the loss of SETD5.

DOI: https://doi.org/10.1186/s13229-023-00550-9
Biochim Biophys Acta Mol Basis Dis. 2023 May 29. pii: S0925-4439(23)00132-1. [Epub ahead of print] 166766

Functional and structural impact of 10 ACADM missense mutations on human medium chain acyl-Coa dehydrogenase.

Catarina A Madeira, Carolina Anselmo, João M Costa, Cátia A Bonito, Ricardo J Ferreira, Daniel J V A Santos, Ronald Wanders, João B Vicente, Fátima V Ventura, Paula Leandro.

  Medium chain acyl-CoA dehydrogenase (MCAD) deficiency (MCADD) is associated with ACADM gene mutations, leading to an impaired function and/or structure of MCAD. Importantly, after import into the mitochondria, MCAD must incorporate a molecule of flavin adenine dinucleotide (FAD) per subunit and assemble into tetramers. However, the effect of MCAD amino acid substitutions on FAD incorporation has not been investigated. Herein, the commonest MCAD variant (p.K304E) and 11 additional rare variants (p.Y48C, p.R55G, p.A88P, p.Y133C, p.A140T, p.D143V, p.G224R, p.L238F, p.V264I, p.Y372N, and p.G377V) were functionally and structurally characterized. Half of the studied variants presented a FAD content <65 % compared to the wild-type. Most of them were recovered as tetramers, except the p.Y372N (mainly as dimers). No correlation was found between the levels of tetramers and FAD content. However, a correlation between FAD content and the cofactor's affinity, proteolytic stability, thermostability, and thermal inactivation was established. We showed that the studied amino acid changes in MCAD may alter the substrate chain-length dependence and the interaction with electron-transferring-flavoprotein (ETF) necessary for a proper functioning electron transfer thus adding additional layers of complexity to the pathological effect of ACADM missense mutations. Although the majority of the variant MCADs presented an impaired capacity to retain FAD during their synthesis, some of them were structurally rescued by cofactor supplementation, suggesting that in the mitochondrial environment the levels and activity of those variants may be dependent of FAD's availability thus contributing for the heterogeneity of the MCADD phenotype found in patients presenting the same genotype.

Keywords:  Disease-causing mutations; Electron transferring flavoprotein; Flavin adenine dinucleotide; Inborn metabolic disorders; Medium chain acyl-CoA dehydrogenase deficiency; Protein misfolding

DOI:  https://doi.org/10.1016/j.bbadis.2023.166766
Methods Mol Biol. 2023 ;2675 27-41

Measurement of Fatty Acid Oxidation by High-Resolution Respirometry: Special Considerations for Analysis of Skeletal and Cardiac Muscle and Adipose Tissue.

Nicole T Watt, Amanda D V MacCannell, Lee D Roberts.

  High-resolution respirometry is a state-of-the-art approach for the quantitation of mitochondrial function. Isolated mitochondria, cultured cells, or tissues/fibers are suspended in oxygenated respiration medium within a closed chamber and substrates or inhibitors added in a stepwise manner. The dissolved oxygen concentration decreases as aerobic metabolism in the specimen proceeds, recorded by an oxygen sensor within the chamber to give a quantifiable measure of oxygen consumption by the sample. Measuring oxygen consumption using a variety of respiratory substrates or respiratory complex-targeted inhibitors enables multiple respiratory pathways to be interrogated to determine the functional capacity of the mitochondria in real time. Using a substrate-uncoupler-inhibitor titration (SUIT) protocol, we have developed a method which makes use of differing chain length fatty acids to derive a measure of fatty acid-stimulated respiration through β-oxidation in a variety of tissue types including skeletal and cardiac muscles and brown and white adipose tissues. This report provides technical details of the protocol, and the adaptations employed, to generate robust analysis of mitochondrial fatty acid β-oxidation.

Keywords:  Carnitine palmitoyltransferase 1 activity; Fatty acid oxidation; High-resolution respirometry; Metabolism; Mitochondrial respiration; OROBOROS; Respiration; SUIT protocol; Uncoupling protein 1

DOI:  https://doi.org/10.1007/978-1-0716-3247-5_3
Front Neurosci. 2023 ;17 1198343

Mitochondria-associated endoplasmic reticulum membranes (MAMs) and their role in glaucomatous retinal ganglion cell degeneration-a mini review.

Jennifer H Pham, Dorota L Stankowska.

  Glaucoma is a leading cause of blindness worldwide, commonly associated with elevated intraocular pressure (IOP), leading to degeneration of the optic nerve and death of retinal ganglion cells, the output neurons in the eye. In recent years, many studies have implicated mitochondrial dysfunction as a crucial player in glaucomatous neurodegeneration. Mitochondrial function has been an increasingly researched topic in glaucoma, given its vital role in bioenergetics and propagation of action potentials. One of the most metabolically active tissues in the body characterized by high oxygen consumption is the retina, particularly the retinal ganglion cells (RGCs). RGCs, which have long axons that extend from the eyes to the brain, rely heavily on the energy generated by oxidative phosphorylation for signal transduction, rendering them more vulnerable to oxidative damage. In various glaucoma models, mitochondrial dysfunction and stress from protein aggregates in the endoplasmic reticulum (ER) have been observed in the RGCs. However, it has been shown that the two organelles are connected through a network called mitochondria-associated ER membranes (MAMs); hence this crosstalk in a pathophysiological condition such as glaucoma should be evaluated. Here, we review the current literature suggestive of mitochondrial and ER stress related to glaucoma, indicating potential cross-signaling and the potential roles of MAMs.

Keywords:  ER stress; MAMs; endoplasmic reticulum; glaucoma; inflammation; mitochondria; oxidative stress; retinal ganglion cells

DOI:  https://doi.org/10.3389/fnins.2023.1198343
HGG Adv. 2023 Jul 13. 4(3): 100202

The genetic and phenotypic correlates of mtDNA copy number in a multi-ancestry cohort.

Penn Medicine BioBank

  Mitochondrial DNA copy number (mtCN) is often treated as a proxy for mitochondrial (dys-) function and disease risk. Pathological changes in mtCN are common symptoms of rare mitochondrial disorders, but reported associations between mtCN and common diseases vary across studies. To understand the biology of mtCN, we carried out genome- and phenome-wide association studies of mtCN in 30,666 individuals from the Penn Medicine BioBank (PMBB)-a diverse cohort of largely African and European ancestry. We estimated mtCN in peripheral blood using exome sequence data, taking cell composition into account. We replicated known genetic associations of mtCN in the PMBB and found that their effects are highly correlated between individuals of European and African ancestry. However, the heritability of mtCN was much higher among individuals of largely African ancestry (h2=0.3) compared with European ancestry individuals(h2=0.1). Admixture mapping suggests that there are undiscovered variants underlying mtCN that are differentiated in frequency between individuals with African and European ancestry. We show that mtCN is associated with many health-related phenotypes. We discovered robust associations between mtDNA copy number and diseases of metabolically active tissues, such as cardiovascular disease and liver damage, that were consistent across African and European ancestry individuals. Other associations, such as epilepsy and prostate cancer, were only discovered in either individuals with European or African ancestry but not both. We show that mtCN-phenotype associations can be sensitive to blood cell composition and environmental modifiers, explaining why such associations are inconsistent across studies. Thus, mtCN-phenotype associations must be interpreted with care.

Keywords:  GWAS; Mitochondrial DNA; Penn Medicine Biobank; PheWAS; heritability; mtDNA copy number; multi-ancestry

DOI:  https://doi.org/10.1016/j.xhgg.2023.100202
Curr Opin Genet Dev. 2023 May 26. pii: S0959-437X(23)00033-3. [Epub ahead of print]80 102053

When and why are mitochondria paternally inherited?

Manisha Munasinghe, J Arvid Ågren.

In contrast with nuclear genes that are passed on through both parents, mitochondrial genes are maternally inherited in most species, most of the time. The genetic conflict stemming from this transmission asymmetry is well-documented, and there is an abundance of population-genetic theory associated with it. While occasional or aberrant paternal inheritance occurs, there are only a few cases where exclusive paternal inheritance of mitochondrial genomes is the evolved state. Why this is remains poorly understood. By examining commonalities between species with exclusive paternal inheritance, we discuss what they may tell us about the evolutionary forces influencing mitochondrial inheritance patterns. We end by discussing recent technological advances that make exploring the causes and consequences of paternal inheritance feasible.

DOI: https://doi.org/10.1016/j.gde.2023.102053
J Clin Biochem Nutr. 2023 May;72(3): 207-214

Method for detecting CoQ10 incorporation in the mitochondrial respiratory chain supercomplex.

Kyousuke Sugawara, Seiji Sato, Yuto Tanaka, Akari Nakamura, Akio Fujisawa, Yorihiro Yamamoto, Misato Kashiba.

  Coenzyme Q10 is an important component of the mitochondrial electron transfer chain. A supercomplex of mitochondrial electron transfer system proteins exists. This complex also contains coenzyme Q10. The concentrations of coenzyme Q10 in tissues decrease with age and pathology. Coenzyme Q10 is given as a supplement. It is unknown whether coenzyme Q10 is transported to the supercomplex. We develop a method for measuring coenzyme Q10 in the mitochondrial respiratory chain supercomplex in this study. Blue native electrophoresis was used to separate mitochondrial membranes. Electrophoresis gels were cut into 3 mm slices. Hexane was used to extract coenzyme Q10 from this slice, and HPLC-ECD was used to analyze coenzyme Q10. Coenzyme Q10 was found in the gel at the same site as the supercomplex. Coenzyme Q10 at this location was thought to be coenzyme Q10 in the supercomplex. We discovered that 4-nitrobenzoate, a coenzyme Q10 biosynthesis inhibitor, reduced the amount of coenzyme Q10 both within and outside the supercomplex. We also observed that the addition of coenzyme Q10 to cells increased the amount of coenzyme Q10 in the supercomplex. It is expected to analysis coenzyme Q10 level in supercomplex in various samples by using this novel method.

Keywords:  HPLC-ECD; blue native electrophoresis; coenzyme Q10; mitochondria; respiratory chain supercomplex

DOI:  https://doi.org/10.3164/jcbn.22-137
Mol Genet Metab. 2023 May 21. pii: S1096-7192(23)00242-1. [Epub ahead of print]139(3): 107612

Challenges and strategies for clinical trials in propionic and methylmalonic acidemias.

Jerry Vockley, Barbara Burton, Agnieszka Jurecka, Jitendra Ganju, Beth Leiro, Roberto Zori, Nicola Longo.

  Clinical trial development in rare diseases poses significant study design and methodology challenges, such as disease heterogeneity and appropriate patient selection, identification and selection of key endpoints, decisions on study duration, choice of control groups, selection of appropriate statistical analyses, and patient recruitment. Therapeutic development in organic acidemias (OAs) shares many challenges with other inborn errors of metabolism, such as incomplete understanding of natural history, heterogenous disease presentations, requirement for sensitive outcome measures and difficulties recruiting a small sample of participants. Here, we review strategies for the successful development of a clinical trial to evaluate treatment response in propionic and methylmalonic acidemias. Specifically, we discuss crucial decisions that may significantly impact success of the study, including patient selection, identification and selection of endpoints, determination of the study duration, consideration of control groups including natural history controls, and selection of appropriate statistical analyses. The significant challenges associated with designing a clinical trial in rare disease can sometimes be successfully met through strategic engagement with experts in the rare disease, seeking regulatory and biostatistical guidance, and early involvement of patients and families.

Keywords:  Clinical trial design; Inborn errors of metabolism; Methylmalonic acidemia; Orphan disease; Propionic acidemia; Ultra-rare disease

DOI:  https://doi.org/10.1016/j.ymgme.2023.107612
J Am Assoc Nurse Pract. 2023 Jun 01. 35(6): 334-336

Genomics of aging: Reactive oxidation and inefficient mitochondria.

Kenneth Wysocki, Beth Heuer.

ABSTRACT: Many things are associated with decreased health and lifespan, including cancer, diabetes, atherosclerosis, high blood pressure, and chronic inflammatory conditions. Clinicians may not be familiar with the role that mitochondrial mutations and associated mitochondrial dysfunction play in a shortened lifespan. This article, the fifth in the JAANP Genomics of Aging series, describes the role that mitochondrial dysfunction plays in the development of age-related diseases such as Alzheimer disease, Parkinson disease, cancer, heart disease, and stroke.

DOI: https://doi.org/10.1097/JXX.0000000000000880
Neurosci Bull. 2023 May 31.

Targeting NAD Metabolism for the Therapy of Age-Related Neurodegenerative Diseases.

Feifei Li, Chou Wu, Gelin Wang.

  As the aging population continues to grow rapidly, age-related diseases are becoming an increasing burden on the healthcare system and a major concern for the well-being of elderly individuals. While aging is an inevitable process for all humans, it can be slowed down and age-related diseases can be treated or alleviated. Nicotinamide adenine dinucleotide (NAD) is a critical coenzyme or cofactor that plays a central role in metabolism and is involved in various cellular processes including the maintenance of metabolic homeostasis, post-translational protein modifications, DNA repair, and immune responses. As individuals age, their NAD levels decline, and this decrease has been suggested to be a contributing factor to the development of numerous age-related diseases, such as cancer, diabetes, cardiovascular diseases, and neurodegenerative diseases. In pursuit of healthy aging, researchers have investigated approaches to boost or maintain NAD levels. Here, we provide an overview of NAD metabolism and the role of NAD in age-related diseases and summarize recent progress in the development of strategies that target NAD metabolism for the treatment of age-related diseases, particularly neurodegenerative diseases.

Keywords:  Ageing; NAD metabolism; Neurodegenerative diseases; Neuroprotection; Therapeutic strategy

DOI:  https://doi.org/10.1007/s12264-023-01072-3
Nat Methods. 2023 May 29.

Machine learning in rare disease.

Jineta Banerjee, Jaclyn N Taroni, Robert J Allaway, Deepashree Venkatesh Prasad, Justin Guinney, Casey Greene.

High-throughput profiling methods (such as genomics or imaging) have accelerated basic research and made deep molecular characterization of patient samples routine. These approaches provide a rich portrait of genes, molecular pathways and cell types involved in disease phenotypes. Machine learning (ML) can be a useful tool for extracting disease-relevant patterns from high-dimensional datasets. However, depending upon the complexity of the biological question, machine learning often requires many samples to identify recurrent and biologically meaningful patterns. Rare diseases are inherently limited in clinical cases, leading to few samples to study. In this Perspective, we outline the challenges and emerging solutions for using ML for small sample sets, specifically in rare diseases. Advances in ML methods for rare diseases are likely to be informative for applications beyond rare diseases for which few samples exist with high-dimensional data. We propose that the method community prioritize the development of ML techniques for rare disease research.

DOI: https://doi.org/10.1038/s41592-023-01886-z
J Diabetes Investig. 2023 May 29.

Fibroblast growth factor 21 as a candidate of a novel serum biomarker for mitochondrial diabetes.

Yuji Komorita, Akiyo Shiroozu, Hidetoshi Nakamura.

Serum fibroblast growth factor 21 levels in patients with mitochondrial diabetes might be much higher than in those with other diabetes types. The results of this study could lead to the establishment of a simple method for screening mitochondrial diabetes using peripheral blood serum.

DOI: https://doi.org/10.1111/jdi.14030
Nat Commun. 2023 May 30. 14(1): 3146

The contributions of mitochondrial and nuclear mitochondrial genetic variation to neuroticism.

Charley Xia, Sarah J Pickett, David C M Liewald, Alexander Weiss, Gavin Hudson, W David Hill.

Neuroticism is a heritable trait composed of separate facets, each conferring different levels of protection or risk, to health. By examining mitochondrial DNA in 269,506 individuals, we show mitochondrial haplogroups explain 0.07-0.01% of variance in neuroticism and identify five haplogroup and 15 mitochondria-marker associations across a general factor of neuroticism, and two special factors of anxiety/tension, and worry/vulnerability with effect sizes of the same magnitude as autosomal variants. Within-haplogroup genome-wide association studies identified H-haplogroup-specific autosomal effects explaining 1.4% variance of worry/vulnerability. These H-haplogroup-specific autosomal effects show a pleiotropic relationship with cognitive, physical and mental health that differs from that found when assessing autosomal effects across haplogroups. We identify interactions between chromosome 9 regions and mitochondrial haplogroups at P < 5 × 10-8, revealing associations between general neuroticism and anxiety/tension with brain-specific gene co-expression networks. These results indicate that the mitochondrial genome contributes toward neuroticism and the autosomal links between neuroticism and health.

DOI: https://doi.org/10.1038/s41467-023-38480-y
Clin Chem. 2023 Jun 01. 69(6): 661-664

Low Plasma Citrulline Guiding the Diagnosis of a Mitochondrial Disorder.

Parith Wongkittichote, Rebecca D Ganetzky, Matthew M Demczko, Xinying Hong, Miao He, Stephen R Master.

DOI: https://doi.org/10.1093/clinchem/hvad039
J Hum Genet. 2023 May 29.

Novel ITPA variants identified by whole genome sequencing and RNA sequencing.

Nanako Omichi, Yoshihito Kishita, Mina Nakama, Hideo Sasai, Atsushi Terazawa, Emiko Kobayashi, Takuya Fushimi, Yohei Sugiyama, Keiko Ichimoto, Kazuhiro R Nitta, Yukiko Yatsuka, Akira Ohtake, Kei Murayama, Yasushi Okazaki.

Approximately 80% of rare diseases have a genetic cause, and an accurate genetic diagnosis is necessary for disease management, prognosis prediction, and genetic counseling. Whole-exome sequencing (WES) is a cost-effective approach for exploring the genetic cause, but several cases often remain undiagnosed. We combined whole genome sequencing (WGS) and RNA sequencing (RNA-seq) to identify the pathogenic variants in an unsolved case using WES. RNA-seq revealed aberrant exon 4 and exon 6 splicing of ITPA. WGS showed a previously unreported splicing donor variant, c.263+1G>A, and a novel heterozygous deletion, including exon 6. Detailed examination of the breakpoint indicated the deletion caused by recombination between Alu elements in different introns. The proband was found to have developmental and epileptic encephalopathies caused by variants in the ITPA gene. The combination of WGS and RNA-seq may be effective in diagnosing conditions in proband who could not be diagnosed using WES.

DOI: https://doi.org/10.1038/s10038-023-01156-y