bims-mitmed 2023-12-03 papers

bims-mitmed

Biomed News

on Mitochondrial medicine

Issue of 2023–12–03
twenty-six papers selected by
Dario Brunetti, Fondazione IRCCS Istituto Neurologico

Combined RNA interference and gene replacement therapy targeting MFN2 as proof of principle for the treatment of Charcot-Marie-Tooth type 2A.
Mitochondrial DNA editing with mitoARCUS.
Efficient elimination of MELAS-associated m.3243G mutant mitochondrial DNA by an engineered mitoARCUS nuclease.
Powering prescription: Mitochondria as "Living Drugs" - Definition, clinical applications, and industry advancements.
A new family with a case of severe early-onset muscle fatigue and a peculiar maternally inherited painful swelling in chewing muscles associated with homoplasmic m.15992A>T mutation in mitochondrial tRNAPro.
Mitochondrial quality control in health and cardiovascular diseases.
Ataxia Syndrome With Hearing Loss and Nephronophthisis Associated With a Novel Homozygous Variant in XPNPEP3.
Mitophagy in human health, ageing and disease.
Multifaceted Roles of AFG3L2, a Mitochondrial ATPase in Relation to Neurological Disorders.
The clinical, myopathological, and genetic analysis of 155 Chinese mitochondrial ophthalmoplegia patients with mitochondrial DNA single large deletions.
Skeletal muscle proteome analysis underpins multifaceted mitochondrial dysfunction in Friedreich's ataxia.
TFAM mislocalization during spermatogenesis.
Basal activity of PINK1 and PRKN in cell models and rodent brain.
Analyzing Mitochondrial Function in a Drosophila melanogaster PINK1B9-Null Mutant Using High-resolution Respirometry.
Effectiveness of rehabilitation intervention in persons with Friedreich ataxia.
Metabolic control of mitophagy.
Supercomplex formation of mitochondrial respiratory chain complexes in leukocytes from patients with neurodegenerative diseases.
Case report: Two unexpected cases of DGUOK-related mitochondrial DNA depletion syndrome presenting with hyperinsulinemic hypoglycemia.
A modified mouse model of Friedreich's ataxia with conditional Fxn allele homozygosity delays onset of cardiomyopathy.
Biased placement of Mitochondria fission facilitates asymmetric inheritance of protein aggregates during yeast cell division.
Protocol for evaluating mitochondrial morphology changes in response to CCCP-induced stress through open-source image processing software.
mTOR hypoactivity leads to trophectoderm cell failure by enhancing lysosomal activation and disrupting the cytoskeleton in preimplantation embryo.
CARM1 drives mitophagy and autophagy flux during fasting-induced skeletal muscle atrophy.
Phase I clinical trial of intracerebroventricular transplantation of allogeneic neural stem cells in people with progressive multiple sclerosis.
Decoding mitochondrial-nuclear (epi)genome interactions: the emerging role of ncRNAs.
PITX2 gain-of-function mutation associated with atrial fibrillation alters mitochondrial activity in human iPSC atrial-like cardiomyocytes.

Cell Mol Life Sci. 2023 Nov 25. 80(12): 373

Combined RNA interference and gene replacement therapy targeting MFN2 as proof of principle for the treatment of Charcot-Marie-Tooth type 2A.

Federica Rizzo, Silvia Bono, Marc David Ruepp, Sabrina Salani, Linda Ottoboni, Elena Abati, Valentina Melzi, Chiara Cordiglieri, Serena Pagliarani, Roberta De Gioia, Alessia Anastasia, Michela Taiana, Manuela Garbellini, Simona Lodato, Paolo Kunderfranco, Daniele Cazzato, Daniele Cartelli, Caterina Lonati, Nereo Bresolin, Giacomo Comi, Monica Nizzardo, Stefania Corti.

  Mitofusin-2 (MFN2) is an outer mitochondrial membrane protein essential for mitochondrial networking in most cells. Autosomal dominant mutations in the MFN2 gene cause Charcot-Marie-Tooth type 2A disease (CMT2A), a severe and disabling sensory-motor neuropathy that impacts the entire nervous system. Here, we propose a novel therapeutic strategy tailored to correcting the root genetic defect of CMT2A. Though mutant and wild-type MFN2 mRNA are inhibited by RNA interference (RNAi), the wild-type protein is restored by overexpressing cDNA encoding functional MFN2 modified to be resistant to RNAi. We tested this strategy in CMT2A patient-specific human induced pluripotent stem cell (iPSC)-differentiated motor neurons (MNs), demonstrating the correct silencing of endogenous MFN2 and replacement with an exogenous copy of the functional wild-type gene. This approach significantly rescues the CMT2A MN phenotype in vitro, stabilizing the altered axonal mitochondrial distribution and correcting abnormal mitophagic processes. The MFN2 molecular correction was also properly confirmed in vivo in the MitoCharc1 CMT2A transgenic mouse model after cerebrospinal fluid (CSF) delivery of the constructs into newborn mice using adeno-associated virus 9 (AAV9). Altogether, our data support the feasibility of a combined RNAi and gene therapy strategy for treating the broad spectrum of human diseases associated with MFN2 mutations.

Keywords:  CMT2A; Gene therapy; MFN2; MitoCharc1; Motor neuron; RNA interfering

DOI:  https://doi.org/10.1007/s00018-023-05018-w
Nat Metab. 2023 Nov 30.

Mitochondrial DNA editing with mitoARCUS.

Denisa Hathazi, Rita Horvath.

DOI: https://doi.org/10.1038/s42255-023-00933-5
Nat Metab. 2023 Nov 30.

Efficient elimination of MELAS-associated m.3243G mutant mitochondrial DNA by an engineered mitoARCUS nuclease.

Wendy K Shoop, Janel Lape, Megan Trum, Alea Powell, Emma Sevigny, Adam Mischler, Sandra R Bacman, Flavia Fontanesi, Jeff Smith, Derek Jantz, Cassandra L Gorsuch, Carlos T Moraes.

Nuclease-mediated editing of heteroplasmic mitochondrial DNA (mtDNA) seeks to preferentially cleave and eliminate mutant mtDNA, leaving wild-type genomes to repopulate the cell and shift mtDNA heteroplasmy. Various technologies are available, but many suffer from limitations based on size and/or specificity. The use of ARCUS nucleases, derived from naturally occurring I-CreI, avoids these pitfalls due to their small size, single-component protein structure and high specificity resulting from a robust protein-engineering process. Here we describe the development of a mitochondrial-targeted ARCUS (mitoARCUS) nuclease designed to target one of the most common pathogenic mtDNA mutations, m.3243A>G. mitoARCUS robustly eliminated mutant mtDNA without cutting wild-type mtDNA, allowing for shifts in heteroplasmy and concomitant improvements in mitochondrial protein steady-state levels and respiration. In vivo efficacy was demonstrated using a m.3243A>G xenograft mouse model with mitoARCUS delivered systemically by adeno-associated virus. Together, these data support the development of mitoARCUS as an in vivo gene-editing therapeutic for m.3243A>G-associated diseases.

DOI: https://doi.org/10.1038/s42255-023-00932-6
Pharmacol Res. 2023 Nov 25. pii: S1043-6618(23)00374-2. [Epub ahead of print]199 107018

Powering prescription: Mitochondria as "Living Drugs" - Definition, clinical applications, and industry advancements.

Andrés Caicedo, Emilia Morales, Aldana Moyano, Sebastian Peñaherrera, José Peña-Cisneros, Abigail Benavides-Almeida, Álvaro A Pérez-Meza, Alissen Haro-Vinueza, Cristina Ruiz, Paola Robayo, Doménica Tenesaca, Diego Barba, Kevin Zambrano, Verónica Castañeda, Keshav K Singh.

  Mitochondria's role as engines and beacons of metabolism and determinants of cellular health is being redefined through their therapeutic application as "Living Drugs" (LDs). Artificial mitochondrial transfer/transplant (AMT/T), encompassing various techniques to modify, enrich, or restore mitochondria in cells and tissues, is revolutionizing acellular therapies and the future of medicine. This article proposes a necessary definition for LDs within the Advanced Therapeutic Medicinal Products (ATMPs) framework. While recognizing different types of LDs as ATMPs, such as mesenchymal stem cells (MSCs) and chimeric antigen receptor T (CAR T) cells, we focus on mitochondria due to their unique attributes that distinguish them from traditional cell therapies. These attributes include their inherent living nature, diverse sources, industry applicability, validation, customizability for therapeutic needs, and their capability to adapt and respond within recipient cells. We trace the journey from initial breakthroughs in AMT/T to the current state-of-the-art applications by emerging innovative companies, highlighting the need for manufacturing standards to navigate the transition of mitochondrial therapies from concept to clinical practice. By providing a comprehensive overview of the scientific, clinical, and commercial landscape of mitochondria as LDs, this article contributes to the essential dialogue among regulatory agencies, academia, and industry to shape their future in medicine.

Keywords:  Acellular therapy; Advanced therapeutic medicinal products (ATMPs); Artificial mitochondrial transfer transplant (AMT/T); Cell therapy; Industry; Innovation; Living drugs (LDs); Mitochondria

DOI:  https://doi.org/10.1016/j.phrs.2023.107018
Neuromuscul Disord. 2023 Nov 04. pii: S0960-8966(23)00774-5. [Epub ahead of print]

A new family with a case of severe early-onset muscle fatigue and a peculiar maternally inherited painful swelling in chewing muscles associated with homoplasmic m.15992A>T mutation in mitochondrial tRNAPro.

Elena Ghirigato, Francesca Terenzi, Mirko Baglivo, Nadia Zanetti, Francesco Baldo, Flora Maria Murru, Marco Bobbo, Egidio Barbi, Massimo Zeviani, Irene Bruno, Eleonora Lamantea.

  A 16-year-old boy was evaluated for a history of exercise-induced fatigability associated with nausea even after minimal effort, lower limbs muscle hypotrophy, and swelling of the masseter muscles after chewing. Laboratory tests were remarkable for hyperlactatemia and metabolic acidosis after short physical activity. The muscle biopsy showed non-specific mitochondrial alterations and an increase in intrafibral lipids. Biochemical analysis showed reduced activity of the respiratory chain complexes. Mitochondrial DNA sequencing revealed the presence of a homoplasmic variant m.15992A>T in the MT-TP gene, coding for the mt-tRNAPro in the patient, in his mother and in his brother. Pathogenic or likely pathogenic variants in MT-TP gene are rare. They are responsible for different clinical presentation, almost ever involving the muscle tissue. We report the first family with exercise-induced muscle weakness and swelling of the chewing muscles due to m.15992A>T variant in absence of J1c10 haplogroup, confirming its pathogenicity.

Keywords:  MT-TP; Masseter muscle swelling; Mitochondrial myopathy; Mitochondrial tRNA

DOI:  https://doi.org/10.1016/j.nmd.2023.11.001
Front Cell Dev Biol. 2023 ;11 1290046

Mitochondrial quality control in health and cardiovascular diseases.

Asli E Atici, Timothy R Crother, Magali Noval Rivas.

  Cardiovascular diseases (CVDs) are one of the primary causes of mortality worldwide. An optimal mitochondrial function is central to supplying tissues with high energy demand, such as the cardiovascular system. In addition to producing ATP as a power source, mitochondria are also heavily involved in adaptation to environmental stress and fine-tuning tissue functions. Mitochondrial quality control (MQC) through fission, fusion, mitophagy, and biogenesis ensures the clearance of dysfunctional mitochondria and preserves mitochondrial homeostasis in cardiovascular tissues. Furthermore, mitochondria generate reactive oxygen species (ROS), which trigger the production of pro-inflammatory cytokines and regulate cell survival. Mitochondrial dysfunction has been implicated in multiple CVDs, including ischemia-reperfusion (I/R), atherosclerosis, heart failure, cardiac hypertrophy, hypertension, diabetic and genetic cardiomyopathies, and Kawasaki Disease (KD). Thus, MQC is pivotal in promoting cardiovascular health. Here, we outline the mechanisms of MQC and discuss the current literature on mitochondrial adaptation in CVDs.

Keywords:  ROS; cardiovascular disease; mitobiogenesis; mitochondria; mitochondrial dynamics; mitophagy

DOI:  https://doi.org/10.3389/fcell.2023.1290046
Neurol Genet. 2023 Dec;9(6): e200100

Ataxia Syndrome With Hearing Loss and Nephronophthisis Associated With a Novel Homozygous Variant in XPNPEP3.

Ilan Ben-Shabat, Malin Kvarnung, Wolfgang Sperker, Helene Bruhn, Anna Wredenberg, Rolf Wibom, Inger Nennesmo, Martin Engvall, Martin Paucar.

Objectives: Biallelic variants in XPNPEP3 are associated with a rare mitochondrial syndrome characterized by nephronophthisis leading to kidney failure, essential tremor, hearing loss, seizures, and intellectual disability. Only 2 publications on this condition are available. We report a man with a complex ataxia syndrome, hearing loss, and kidney failure associated with a new biallelic variant in XPNPEP3.
Methods: Clinical evaluation, neuroimaging studies, a kidney biopsy, and whole genome sequencing (WGS) were applied. Since the phenotype was compatible with a mitochondrial disease, a muscle biopsy with morphological and mitochondrial biochemical investigations was performed.
Results: Axial ataxia, cerebellar atrophy, hearing loss, myopathy, ptosis, supranuclear palsy, and kidney failure because of nephronophthisis were the prominent features in this case. WGS revealed the novel biallelic variant c.766C>T (p.Gln256*) in XPNPEP3. A muscle biopsy revealed COX negative fibers, a few ragged red fibers, and ultrastructural mitochondrial changes. Enzyme activity in respiratory chain complex IV was reduced in muscle and fibroblasts.
Discussion: This is the first report of a slowly progressive cerebellar ataxia associated with a novel biallelic variant in XPNPEP3. Abnormalities typical for mitochondrial disease and the slow progression of kidney disease are also striking. Our report expands the spectrum of XPNPEP3-related diseases.

DOI: https://doi.org/10.1212/NXG.0000000000200100
Nat Metab. 2023 Nov 30.

Mitophagy in human health, ageing and disease.

Anna Picca, Julie Faitg, Johan Auwerx, Luigi Ferrucci, Davide D'Amico.

Maintaining optimal mitochondrial function is a feature of health. Mitophagy removes and recycles damaged mitochondria and regulates the biogenesis of new, fully functional ones preserving healthy mitochondrial functions and activities. Preclinical and clinical studies have shown that impaired mitophagy negatively affects cellular health and contributes to age-related chronic diseases. Strategies to boost mitophagy have been successfully tested in model organisms, and, recently, some have been translated into clinics. In this Review, we describe the basic mechanisms of mitophagy and how mitophagy can be assessed in human blood, the immune system and tissues, including muscle, brain and liver. We outline mitophagy's role in specific diseases and describe mitophagy-activating approaches successfully tested in humans, including exercise and nutritional and pharmacological interventions. We describe how mitophagy is connected to other features of ageing through general mechanisms such as inflammation and oxidative stress and forecast how strengthening research on mitophagy and mitophagy interventions may strongly support human health.

DOI: https://doi.org/10.1038/s42255-023-00930-8
Mol Neurobiol. 2023 Nov 28.

Multifaceted Roles of AFG3L2, a Mitochondrial ATPase in Relation to Neurological Disorders.

Ranita Ghosh Dastidar, Saradindu Banerjee, Piyush Behari Lal, Somasish Ghosh Dastidar.

  AFG3L2 is a zinc metalloprotease and an ATPase localized in an inner mitochondrial membrane involved in mitochondrial quality control of several nuclear- and mitochondrial-encoded proteins. Mutations in AFG3L2 lead to diseases like slow progressive ataxia, which is a neurological disorder. This review delineates the cellular functions of AFG3L2 and its dysfunction that leads to major clinical outcomes, which include spinocerebellar ataxia type 28, spastic ataxia type 5, and optic atrophy type 12. It summarizes all relevant AFG3L2 mutations associated with the clinical outcomes to understand the detailed mechanisms attributable to its structure-related multifaceted roles in proteostasis and quality control. We face early diagnostic challenges of ataxia and optic neuropathy due to asymptomatic parents and variable clinical manifestations due to heterozygosity/homozygosity of AFG3L2 mutations. This review intends to promote AFG3L2 as a putative prognostic or diagnostic marker. Functions, mutations, and clinical manifestations in AFG3L2, a mitochondrial AAA + ATPases.

Keywords:  AFG3L2; Ataxia; Mitochondria; Neurological disorders; SCA28; Zinc metalloprotease

DOI:  https://doi.org/10.1007/s12035-023-03768-z
Mol Genet Genomic Med. 2023 Nov 28. e2328

The clinical, myopathological, and genetic analysis of 155 Chinese mitochondrial ophthalmoplegia patients with mitochondrial DNA single large deletions.

Yang Zhao, Yue Hou, Xutong Zhao, Tongling Liufu, Meng Yu, Wei Zhang, Zhiying Xie, Victor Wei Zhang, Yun Yuan, Zhaoxia Wang.

   BACKGROUND: Progressive external ophthalmoplegia (PEO) is a common subtype of mitochondrial encephalomyopathy.
OBJECTIVE: The study aimed to investigate the relationship between mitochondrial DNA (mtDNA) abnormalities, muscle pathology, and clinical manifestations in Chinese patients with single large-scale mtDNA deletion presenting with PEO.
METHODS: This is a retrospective single-center study. Patients with PEO who had a single large deletion in mitochondrial DNA were included in this study. The associations were analyzed between mtDNA deletion patterns, myopathological changes, and clinical characteristics.
RESULTS: In total, 155 patients with mitochondrial PEO carrying single large-scale mtDNA mutations were enrolled, including 137 chronic progressive external ophthalmoplegia (CPEO) and 18 Kearns-Sayre syndrome (KSS) patients. The onset ages were 9.61 ± 4.12 in KSS and 20.15 ± 9.06 in CPEO. The mtDNA deletions ranged from 2225 bp to 9131 bp, with m.8470_13446del being the most common. The KSS group showed longer deletions than the CPEO group (p = 0.004). Additionally, a higher number of deleted genes encoding respiratory chain complex subunits (p = 0.001) and tRNA genes (p = 0.009) were also observed in the KSS group. A weak negative correlation between the mtDNA deletion size and ages of onset (p < 0.001, r = -0.369) was observed. The proportion of ragged red fibers, ragged blue fibers, and cytochrome c negative fibers did not correlate significantly with onset ages (p > 0.05). However, a higher percentage of abnormal muscle fibers corresponds to an increased prevalence of exercise intolerance, limb muscle weakness, dysphagia, and cerebellar ataxia.
CONCLUSION: We reported a large Chinese cohort consisting of mitochondrial PEO patients with single large-scale mtDNA deletions. Our results demonstrated that the length and locations of mtDNA deletions may influence onset ages and clinical phenotypes. The severity of muscle pathology could not only indicate diagnosis but also may be associated with clinical manifestations beyond the extraocular muscles.

Keywords:  Kearns-Sayre syndrome; chronic progressive external ophthalmoplegia; mitochondrial DNA; single large-scale deletion

DOI:  https://doi.org/10.1002/mgg3.2328
Front Neurosci. 2023 ;17 1289027

Skeletal muscle proteome analysis underpins multifaceted mitochondrial dysfunction in Friedreich's ataxia.

Elisabetta Indelicato, Klaus Faserl, Matthias Amprosi, Wolfgang Nachbauer, Rainer Schneider, Julia Wanschitz, Bettina Sarg, Sylvia Boesch.

  Friedreich's ataxia (FRDA) is a severe multisystemic disorder caused by a deficiency of the mitochondrial protein frataxin. While some aspects of FRDA pathology are developmental, the causes underlying the steady progression are unclear. The inaccessibility of key affected tissues to sampling is a main hurdle. Skeletal muscle displays a disease phenotype and may be sampled in vivo to address open questions on FRDA pathophysiology. Thus, we performed a quantitative mass spectrometry-based proteomics analysis in gastrocnemius skeletal muscle biopsies from genetically confirmed FRDA patients (n = 5) and controls. Obtained data files were processed using Proteome Discoverer and searched by Sequest HT engine against a UniProt human reference proteome database. Comparing skeletal muscle proteomics profiles between FRDA and controls, we identified 228 significant differentially expressed (DE) proteins, of which 227 were downregulated in FRDA. Principal component analysis showed a clear separation between FRDA and control samples. Interactome analysis revealed clustering of DE proteins in oxidative phosphorylation, ribosomal elements, mitochondrial architecture control, and fission/fusion pathways. DE findings in the muscle-specific proteomics suggested a shift toward fast-twitching glycolytic fibers. Notably, most DE proteins (169/228, 74%) are target of the transcription factor nuclear factor-erythroid 2. Our data corroborate a mitochondrial biosignature of FRDA, which extends beyond a mere oxidative phosphorylation failure. Skeletal muscle proteomics highlighted a derangement of mitochondrial architecture and maintenance pathways and a likely adaptive metabolic shift of contractile proteins. The present findings are relevant for the design of future therapeutic strategies and highlight the value of skeletal muscle-omics as disease state readout in FRDA.

Keywords:  Friedreich’s ataxia; frataxin; mass spectrometry; mitochondria; proteomics; skeletal muscle

DOI:  https://doi.org/10.3389/fnins.2023.1289027
Trends Genet. 2023 Nov 29. pii: S0168-9525(23)00257-3. [Epub ahead of print]

TFAM mislocalization during spermatogenesis.

Sam Kavoosi, Martin Picard, Brett A Kaufman.

  Mitochondrial DNA (mtDNA) is inherited almost exclusively from the maternal lineage. Paternal destruction of either mtDNA or whole mitochondria has been the dominant model for mtDNA transmission. Recently, Lee et al. provided evidence for mitochondrial transcription factor A (TFAM) import sequence regulation as a potential cause for mtDNA depletion in human sperm before fertilization.

Keywords:  TFAM; mtDNA; spermatogenesis; uniparental inheritance

DOI:  https://doi.org/10.1016/j.tig.2023.11.002
Autophagy. 2023 Dec 02. 1-12

Basal activity of PINK1 and PRKN in cell models and rodent brain.

Jens O Watzlawik, Fabienne C Fiesel, Gabriella Fiorino, Bernardo A Bustillos, Zahra Baninameh, Briana N Markham, Xu Hou, Caleb S Hayes, Jenny M Bredenberg, Nicholas W Kurchaba, Dominika Fričová, Joanna Siuda, Zbigniew K Wszolek, Sachiko Noda, Shigeto Sato, Nobutaka Hattori, Asheeta A Prasad, Deniz Kirik, Howard S Fox, Kelly L Stauch, Matthew S Goldberg, Wolfdieter Springer.

  The ubiquitin kinase-ligase pair PINK1-PRKN recognizes and transiently labels damaged mitochondria with ubiquitin phosphorylated at Ser65 (p-S65-Ub) to mediate their selective degradation (mitophagy). Complete loss of PINK1 or PRKN function unequivocally leads to early-onset Parkinson disease, but it is debated whether impairments in mitophagy contribute to disease later in life. While the pathway has been extensively studied in cell culture upon acute and massive mitochondrial stress, basal levels of activation under endogenous conditions and especially in vivo in the brain remain undetermined. Using rodent samples, patient-derived cells, and isogenic neurons, we here identified age-dependent, brain region-, and cell type-specific effects and determined expression levels and extent of basal and maximal activation of PINK1 and PRKN. Our work highlights the importance of defining critical risk and therapeutically relevant levels of PINK1-PRKN signaling which will further improve diagnosis and prognosis and will lead to better stratification of patients for future clinical trials.

Keywords:  Mitophagy; PINK1; PRKN; Parkinson disease; parkin; ubiquitin

DOI:  https://doi.org/10.1080/15548627.2023.2286414
J Vis Exp. 2023 Nov 10.

Analyzing Mitochondrial Function in a Drosophila melanogaster PINK1B9-Null Mutant Using High-resolution Respirometry.

Paula Michelotti, Tâmie Duarte, Cristiane L Dalla Corte.

Neurodegenerative diseases, including Parkinson's Disease (PD), and cellular disturbances such as cancer are some of the disorders that disrupt energy metabolism with impairment of mitochondrial functions. Mitochondria are organelles that control both energy metabolism and cellular processes involved in cell survival and death. For this reason, approaches to evaluate mitochondrial function can offer important insights into cellular conditions in pathological and physiological processes. In this regard, high-resolution respirometry (HRR) protocols allow evaluation of the whole mitochondrial respiratory chain function or the activity of specific mitochondrial complexes. Furthermore, studying mitochondrial physiology and bioenergetics requires genetically and experimentally tractable models such as Drosophila melanogaster. This model presents several advantages, such as its similarity to human physiology, its rapid life cycle, easy maintenance, cost-effectiveness, high throughput capabilities, and a minimized number of ethical concerns. These attributes collectively establish it as an invaluable tool for dissecting complex cellular processes. The present work explains how to analyze mitochondrial function using the Drosophila melanogaster PINK1B9-null mutant. The pink1 gene is responsible for encoding PTEN-induced putative kinase 1, through a process recognized as mitophagy, which is crucial for the removal of dysfunctional mitochondria from the mitochondrial network. Mutations in this gene have been associated with an autosomal recessive early-onset familial form of PD. This model can be used to study mitochondrial dysfunction involved in the pathophysiology of PD.

DOI: https://doi.org/10.3791/65664
Front Neurol. 2023 ;14 1270296

Effectiveness of rehabilitation intervention in persons with Friedreich ataxia.

Gabriella Paparella, Cristina Stragà, Marinela Vavla, Nicola Pesenti, Vasco Merotto, Gian A Martorel, Sara Zalunardo, Maria Armellin, Jimmy Comiotto, Andrea Martinuzzi.

   Introduction: The relevance of rehabilitation in progressive neurological disorders, such as Friedreich's Ataxia (FRDA), has yet to be convincingly proven. FRDA is characterized by ataxia, loss of gait, scoliosis, cardiomyopathy, dysarthria and dysphagia, with reduced life expectancy. The disease onset is usually in adolescence, leading to progressive disability. Omaveloxolone has been recently approved as the first pharmacological treatment for FRDA in adults and adolescents aged 16 years and older. Regarding non-pharmacological therapies, neurorehabilitation is a valuable aid in addressing the symptoms and in maintaining the residual functioning. We performed a prospective observational cohort study to evaluate the efficacy of inpatient rehabilitation (IR) for people with FRDA.
Methods: A total of 42 individuals (29 adults and 13 children) with FRDA were recruited. There were 27 ambulant and 15 non-ambulant participants. The patients underwent IR of 3 and 4 weeks in children and adults, respectively. The IR treatment was designed to be applied within a multidisciplinary setting, so FRDA patients underwent, in addition to physiotherapy, also occupational therapy, practical manual activities and psychological support aiming to enhance transferable skills useful in the activities of daily living. The primary outcome was the Scale for the Assessment and Rating of Ataxia (SARA). Other measures were: Friedreich Ataxia Rating Scale (FARS) and Nine Hole Peg Test (NHPT). Furthermore, we used the 6 Minute Walk Test (6MWT), the Timed Up and Go (TUG) and the Berg Balance Scale (BBS) only on ambulant subjects. Outcomes were evaluated at baseline and at the end of the treatment.
Results: We report that the IR significantly improves motor performance and ataxia symptoms in patients with FRDA. Our study shows significant functional improvement in all the outcome measures used, except for NHPT bilaterally. FARS and SARA scores post-IR are significatively reduced when compared (p < 0.001).
Discussion: We demonstrate that IR programs in FRDA can provide a meaningful clinical improvement in terms of outcome measures. These findings could be useful when approaching progressive neurological disorders.

Keywords:  Friedreich ataxia; adults; children; inpatients; rehabilitation; treatment outcome

DOI:  https://doi.org/10.3389/fneur.2023.1270296
Eur J Clin Invest. 2023 Dec 01. e14138

Metabolic control of mitophagy.

Andreas Zimmermann, Frank Madeo, Abhinav Diwan, Junichi Sadoshima, Simon Sedej, Guido Kroemer, Mahmoud Abdellatif.

  Mitochondrial dysfunction is a major hallmark of ageing and related chronic disorders. Controlled removal of damaged mitochondria by the autophagic machinery, a process known as mitophagy, is vital for mitochondrial homeostasis and cell survival. The central role of mitochondria in cellular metabolism places mitochondrial removal at the interface of key metabolic pathways affecting the biosynthesis or catabolism of acetyl-coenzyme A, nicotinamide adenine dinucleotide, polyamines, as well as fatty acids and amino acids. Molecular switches that integrate the metabolic status of the cell, like AMP-dependent protein kinase, protein kinase A, mechanistic target of rapamycin and sirtuins, have also emerged as important regulators of mitophagy. In this review, we discuss how metabolic regulation intersects with mitophagy. We place special emphasis on the metabolic regulatory circuits that may be therapeutically targeted to delay ageing and mitochondria-associated chronic diseases. Moreover, we identify outstanding knowledge gaps, such as the ill-defined distinction between basal and damage-induced mitophagy, which must be resolved to boost progress in this area.

Keywords:  AMPK; NAD; acetyl-CoA; ageing; ageing-related disease; metabolism; mitophagy; spermidine

DOI:  https://doi.org/10.1111/eci.14138
J Biochem. 2023 Nov 28. pii: mvad100. [Epub ahead of print]

Supercomplex formation of mitochondrial respiratory chain complexes in leukocytes from patients with neurodegenerative diseases.

Tsukasa Hara, Ryosuke Amagai, Ryuji Sakakibara, Ayako Okado-Matsumoto.

  With population aging, cognitive impairments and movement disorders due to neurodegenerative diseases, such as Alzheimer's disease (AD), Parkinson's disease (PD), and dementia with Lewy bodies (DLB), are increasingly considered as key social issues. Clinically, it has remained challenging to diagnose them before the onset of symptoms because of difficulty to observe the progressive loss of neurons in the brain. Therefore, with exploratory research into biomarkers, a number of candidates have previously been proposed, such as activities of mitochondrial respiratory chain complexes in blood in AD and PD. In this study, we focused on the formation of mitochondrial respiratory chain supercomplexes (SCs) because the formation of SC itself modulates the activity of each complex. Here we investigated the SC formation in leukocytes from patients with AD, PD, and DLB. Our results showed that SCs were well formed in AD and PD compared with controls, while poorly formed in DLB. We highlighted that the disruption of the SC formation correlated with the progression of PD and DLB. Taking our findings together, we propose that pronounced SC formation would already have occurred before the onset of AD, PD, and DLB and, with the progression of neurodegeneration, the SC formation would gradually be disrupted.

Keywords:  Alzheimer’s disease; Parkinson’s disease; dementia with Lewy bodies; high-resolution clear native polyacrylamide gel electrophoresis; in-gel activity assay; mitochondrial respiratory chain complexes

DOI:  https://doi.org/10.1093/jb/mvad100
Front Endocrinol (Lausanne). 2023 ;14 1268135

Case report: Two unexpected cases of DGUOK-related mitochondrial DNA depletion syndrome presenting with hyperinsulinemic hypoglycemia.

Herodes Guzman, Sahr Yazdani, Jennifer L Harmon, Kimberly A Chapman, Bernadette Vitola, Louise Pyle, Heather McKnight, Winnie Sigal, Katherine Lord, Diva D De Leon, Nadia Merchant, Rebecca Ganetzky.

  Timely diagnosis of persistent neonatal hypoglycemia is critical to prevent neurological sequelae, but diagnosis is complicated by the heterogenicity of the causes. We discuss two cases at separate institutions in which clinical management was fundamentally altered by the results of molecular genetic testing. In both patients, critical samples demonstrated hypoketotic hypoglycemia and a partial glycemic response to glucagon stimulation, thereby suggesting hyperinsulinism (HI). However, due to rapid genetic testing, both patients were found to have deoxyguanosine kinase (DGUOK)-related mitochondrial DNA depletion syndrome, an unexpected diagnosis. Patients with this disease typically present with either hepatocerebral disease in the neonatal period or isolated hepatic failure in infancy. The characteristic features involved in the hepatocerebral form of the disease include lactic acidosis, hypoglycemia, cholestasis, progressive liver failure, and increasing neurologic dysfunction. Those with isolated liver involvement experience hepatomegaly, cholestasis, and liver failure. Although liver transplantation is considered, research has demonstrated that for patients with DGUOK-related mitochondrial DNA depletion syndrome and neurologic symptoms, early demise occurs. Our report advocates for the prompt initiation of genetic testing in patients presenting with persistent neonatal hypoglycemia and for the incorporation of mitochondrial DNA depletion syndromes in the differential diagnosis of HI.

Keywords:  DGUOK; exome; hyperinsulinism; hypoglycemia; mitochondria

DOI:  https://doi.org/10.3389/fendo.2023.1268135
Am J Physiol Heart Circ Physiol. 2023 Dec 01.

A modified mouse model of Friedreich's ataxia with conditional Fxn allele homozygosity delays onset of cardiomyopathy.

Tyler L Perfitt, Claudia Huichalaf, Renea Gooch, Anna Kuperman, Youngwook Ahn, Xian Chen, Soumya Ullas, Dinesh Hirenallur-Shanthappa, Yutian Zhan, Diana Otis, Laurence O Whiteley, Christine Bulawa, Alain Martelli.

  Friedreich's ataxia (FA) is an autosomal recessive disorder caused by a deficiency in frataxin (FXN), a mitochondrial protein that plays a critical role in the synthesis of iron sulfur clusters (Fe-S), vital inorganic cofactors necessary for numerous cellular processes. FA is characterized by progressive ataxia and hypertrophic cardiomyopathy, with cardiac dysfunction as the most common cause of mortality in patients. Commonly used cardiac-specific mouse models of FA utilize the muscle creatine kinase (MCK) promoter to express Cre recombinase in cardiomyocytes and striated muscle cells in mice with one conditional Fxn allele and one floxed-out/null allele. These mice quickly develop cardiomyopathy that becomes fatal by 9-11 weeks of age. Here, we generated a cardiac-specific model with floxed Fxn allele homozygosity (MCK-Fxnflox/flox). MCK-Fxnflox/flox mice were phenotypically normal at 9 weeks of age, despite no detectable FXN protein expression. Between 13 and 15 weeks of age, these mice began to display progressive cardiomyopathy, including decreased ejection fraction and fractional shortening, and increased left ventricular mass. MCK-Fxnflox/flox mice began to lose weight around 16 weeks of age, characteristically associated with heart failure in other cardiac-specific FA models. By 18 weeks of age, MCK-Fxnflox/flox mice displayed elevated markers of Fe-S deficiency, cardiac stress and injury, and cardiac fibrosis. This modified model reproduced important pathophysiological and biochemical features of FA over a longer timescale than previous cardiac-specific mouse models, offering a larger window for studying potential therapeutics.

Keywords:  Friedreich's ataxia; animal models; cardiac diseases; genetic diseases

DOI:  https://doi.org/10.1152/ajpheart.00496.2023
PLoS Comput Biol. 2023 Nov 27. 19(11): e1011588

Biased placement of Mitochondria fission facilitates asymmetric inheritance of protein aggregates during yeast cell division.

Gordon Sun, Christine Hwang, Tony Jung, Jian Liu, Rong Li.

Mitochondria are essential and dynamic eukaryotic organelles that must be inherited during cell division. In yeast, mitochondria are inherited asymmetrically based on quality, which is thought to be vital for maintaining a rejuvenated cell population; however, the mechanisms underlying mitochondrial remodeling and segregation during this process are not understood. We used high spatiotemporal imaging to quantify the key aspects of mitochondrial dynamics, including motility, fission, and fusion characteristics, upon aggregation of misfolded proteins in the mitochondrial matrix. Using these measured parameters, we developed an agent-based stochastic model of dynamics of mitochondrial inheritance. Our model predicts that biased mitochondrial fission near the protein aggregates facilitates the clustering of protein aggregates in the mitochondrial matrix, and this process underlies asymmetric mitochondria inheritance. These predictions are supported by live-cell imaging experiments where mitochondrial fission was perturbed. Our findings therefore uncover an unexpected role of mitochondrial dynamics in asymmetric mitochondrial inheritance.

DOI: https://doi.org/10.1371/journal.pcbi.1011588
STAR Protoc. 2023 Nov 29. pii: S2666-1667(23)00712-8. [Epub ahead of print]4(4): 102745

Protocol for evaluating mitochondrial morphology changes in response to CCCP-induced stress through open-source image processing software.

Sudeshna Nag, Kaitlin Szederkenyi, Christopher M Yip, G Angus McQuibban.

  Mitochondrial morphology is an indicator of cellular health and function; however, its quantification and categorization into different subclasses is a complicated process. Here, we present a protocol for mitochondrial morphology quantification in the presence and absence of carbonyl cyanide m-chlorophenyl hydrazone stress. We describe steps for the preparation of cells for immunofluorescence microscopy, staining, and morphology quantification. The quantification protocol generates an aspect ratio that helps to categorize mitochondria into two clear subclasses. For complete details on the use and execution of this protocol, please refer to Nag et al.1.

Keywords:  Cell Biology; Cell culture; Metabolism; Microscopy; Molecular/Chemical Probes

DOI:  https://doi.org/10.1016/j.xpro.2023.102745
Cell Biosci. 2023 Nov 30. 13(1): 219

mTOR hypoactivity leads to trophectoderm cell failure by enhancing lysosomal activation and disrupting the cytoskeleton in preimplantation embryo.

Chiyuan Ma, Qin Li, Yuxin Yang, Lei Ge, Jiaxuan Cai, Juan Wang, Maoxian Zhu, Yue Xiong, Wenya Zhang, Jingtong Xie, Yujing Cao, Huashan Zhao, Qing Wei, Chen Huang, Junchao Shi, Jian V Zhang, Enkui Duan, Xiaohua Lei.

   BACKGROUND: Metabolic homeostasis is closely related to early impairment of cell fate determination and embryo development. The protein kinase mechanistic target of rapamycin (mTOR) is a key regulator of cellular metabolism in the body. Inhibition of mTOR signaling in early embryo causes postimplantation development failure, yet the mechanisms are still poorly understood.
METHODS: Pregnancy mice and preimplantation mouse embryo were treated with mTOR inhibitor in vivo and in vitro respectively, and subsequently examined the blastocyst formation, implantation, and post-implantation development. We used immunofluorescence staining, RNA-Seq smart2, and genome-wide bisulfite sequencing technologies to investigate the impact of mTOR inhibitors on the quality, cell fate determination, and molecular alterations in developing embryos.
RESULTS: We showed mTOR suppression during preimplantation decreases the rate of blastocyst formation and the competency of implantation, impairs the post implantation embryonic development. We discovered that blocking mTOR signaling negatively affected the transformation of 8-cell embryos into blastocysts and caused various deficiencies in blastocyst quality. These included problems with compromised trophectoderm cell differentiation, as well as disruptions in cell fate specification. mTOR suppression significantly affected the transcription and DNA methylation of embryos. Treatment with mTOR inhibitors increase lysosomal activation and disrupts the organization and dynamics of the actin cytoskeleton in blastocysts.
CONCLUSIONS: These results demonstrate that mTOR plays a crucial role in 8-cell to blastocyst transition and safeguards embryo quality during early embryo development.

Keywords:  Actin cytoskeleton disruption; DNA methylation; Lysosomal activation; Transcriptome; Trophectoderm cell failure; mTOR

DOI:  https://doi.org/10.1186/s13578-023-01176-3
Autophagy. 2023 Nov 29.

CARM1 drives mitophagy and autophagy flux during fasting-induced skeletal muscle atrophy.

Derek W Stouth, Tiffany L vanLieshout, Andrew I Mikhail, Sean Y Ng, Rozhin Raziee, Brittany A Edgett, Goutham Vasam, Erin K Webb, Kevin S Gilotra, Matthew Markou, Hannah C Pineda, Brianna G Bettencourt-Mora, Haleema Noor, Zachary Moll, Megan E Bittner, Brendon J Gurd, Keir J Menzies, Vladimir Ljubicic.

  CARM1 (coactivator associated arginine methyltransferase 1) has recently emerged as a powerful regulator of skeletal muscle biology. However, the molecular mechanisms by which the methyltransferase remodels muscle remain to be fully understood. In this study, carm1 skeletal muscle-specific knockout (mKO) mice exhibited lower muscle mass with dysregulated macroautophagic/autophagic and atrophic signaling, including depressed AMP-activated protein kinase (AMPK) site-specific phosphorylation of ULK1 (unc-51 like autophagy activating kinase 1; Ser555) and FOXO3 (forkhead box O3; Ser588), as well as MTOR (mechanistic target of rapamycin kinase)-induced inhibition of ULK1 (Ser757), along with AKT/protein kinase B site-specific suppression of FOXO1 (Ser256) and FOXO3 (Ser253). In addition to lower mitophagy and autophagy flux in skeletal muscle, carm1 mKO led to increased mitochondrial PRKN/parkin accumulation, which suggests that CARM1 is required for basal mitochondrial turnover and autophagic clearance. carm1 deletion also elicited PPARGC1A (PPARG coactivator 1 alpha) activity and a slower, more oxidative muscle phenotype. As such, these carm1 mKO-evoked adaptations disrupted mitophagy and autophagy induction during food deprivation and collectively served to mitigate fasting-induced muscle atrophy. Furthermore, at the threshold of muscle atrophy during food deprivation experiments in humans, skeletal muscle CARM1 activity decreased similarly to our observations in mice, and was accompanied by site-specific activation of ULK1 (Ser757), highlighting the translational impact of the methyltransferase in human skeletal muscle. Taken together, our results indicate that CARM1 governs mitophagic, autophagic, and atrophic processes fundamental to the maintenance and remodeling of muscle mass. Targeting the enzyme may provide new therapeutic approaches for mitigating skeletal muscle atrophy.

Keywords:  Atrophy; autophagy; coactivator-associated arginine methyltransferase 1; fasting; mitophagy; skeletal muscle

DOI:  https://doi.org/10.1080/15548627.2023.2288528
Cell Stem Cell. 2023 Nov 21. pii: S1934-5909(23)00393-4. [Epub ahead of print]

Phase I clinical trial of intracerebroventricular transplantation of allogeneic neural stem cells in people with progressive multiple sclerosis.

Maurizio A Leone, Maurizio Gelati, Daniela C Profico, Claudio Gobbi, Emanuele Pravatà, Massimiliano Copetti, Carlo Conti, Lucrezia Abate, Luigi Amoruso, Francesco Apollo, Rosario F Balzano, Ilaria Bicchi, Massimo Carella, Alessandro Ciampini, Carlo Colosimo, Paola Crociani, Giada D'Aloisio, Pietro Di Viesti, Daniela Ferrari, Danilo Fogli, Andrea Fontana, Domenico Frondizi, Valentina Grespi, Jens Kuhle, Antonio Laborante, Ivan Lombardi, Gianmarco Muzi, Francesca Paci, Giuliana Placentino, Teresa Popolizio, Claudia Ricciolini, Simonetta Sabatini, Giada Silveri, Cristina Spera, Daniel Stephenson, Giuseppe Stipa, Elettra Tinella, Michele Zarrelli, Chiara Zecca, Yendri Ventura, Angelo D'Alessandro, Luca Peruzzotti-Jametti, Stefano Pluchino, Angelo L Vescovi.

  We report the analysis of 1 year of data from the first cohort of 15 patients enrolled in an open-label, first-in-human, dose-escalation phase I study (ClinicalTrials.gov: NCT03282760, EudraCT2015-004855-37) to determine the feasibility, safety, and tolerability of the transplantation of allogeneic human neural stem/progenitor cells (hNSCs) for the treatment of secondary progressive multiple sclerosis. Participants were treated with hNSCs delivered via intracerebroventricular injection in combination with an immunosuppressive regimen. No treatment-related deaths nor serious adverse events (AEs) were observed. All participants displayed stability of clinical and laboratory outcomes, as well as lesion load and brain activity (MRI), compared with the study entry. Longitudinal metabolomics and lipidomics of biological fluids identified time- and dose-dependent responses with increased levels of acyl-carnitines and fatty acids in the cerebrospinal fluid (CSF). The absence of AEs and the stability of functional and structural outcomes are reassuring and represent a milestone for the safe translation of stem cells into regenerative medicines.

Keywords:  ATMPs; advanced cell therapy; clinical trial; human neural progenitor cells; intracerebroventricular injection; metabolomics; neural stem cells; progressive multiple sclerosis

DOI:  https://doi.org/10.1016/j.stem.2023.11.001
Epigenomics. 2023 Nov;15(21): 1121-1136

Decoding mitochondrial-nuclear (epi)genome interactions: the emerging role of ncRNAs.

Julia Nguyen, Quinn Le, Phyo W Win, Kathleen A Hill, Shiva M Singh, Christina A Castellani.

  Bidirectional communication between the mitochondria and the nucleus is required for several physiological processes, and the nuclear epigenome is a key mediator of this relationship. ncRNAs are an emerging area of discussion for their roles in cellular function and regulation. In this review, we highlight the role of mitochondrial-encoded ncRNAs as mediators of communication between the mitochondria and the nuclear genome. We focus primarily on retrograde signaling, a process in which the mitochondrion relays ncRNAs to translate environmental stress signals to changes in nuclear gene expression, with implications on stress responses that may include disease(s). Other biological roles of mitochondrial-encoded ncRNAs, such as mitochondrial import of proteins and regulation of cell signaling, will also be discussed.

Keywords:  disease risk; gene expression; mitochondria; ncRNA; retrograde signaling

DOI:  https://doi.org/10.2217/epi-2023-0322
Front Physiol. 2023 ;14 1250951

PITX2 gain-of-function mutation associated with atrial fibrillation alters mitochondrial activity in human iPSC atrial-like cardiomyocytes.

Patrizia Benzoni, Lorenzo Da Dalt, Noemi Elia, Vera Popolizio, Alessandro Cospito, Federica Giannetti, Patrizia Dell'Era, Morten S Olesen, Annalisa Bucchi, Mirko Baruscotti, Giuseppe Danilo Norata, Andrea Barbuti.

  Atrial fibrillation (AF) is the most common cardiac arrhythmia worldwide; however, the underlying causes of AF initiation are still poorly understood, particularly because currently available models do not allow in distinguishing the initial causes from maladaptive remodeling that induces and perpetuates AF. Lately, the genetic background has been proven to be important in the AF onset. iPSC-derived cardiomyocytes, being patient- and mutation-specific, may help solve this diatribe by showing the initial cell-autonomous changes underlying the development of the disease. Transcription factor paired-like homeodomain 2 (PITX2) has been identified as a key regulator of atrial development/differentiation, and the PITX2 genomic locus has the highest association with paroxysmal AF. PITX2 influences mitochondrial activity, and alterations in either its expression or function have been widely associated with AF. In this work, we investigate the activity of mitochondria in iPSC-derived atrial cardiomyocytes (aCMs) obtained from a young patient (24 years old) with paroxysmal AF, carrying a gain-of-function mutation in PITX2 (rs138163892) and from its isogenic control (CTRL) in which the heterozygous point mutation has been reverted to WT. PITX2 aCMs show a higher mitochondrial content, increased mitochondrial activity, and superoxide production under basal conditions when compared to CTRL aCMs. However, increasing mitochondrial workload by FCCP or β-adrenergic stimulation allows us to unmask mitochondrial defects in PITX2 aCMs, which are incapable of responding efficiently to the higher energy demand, determining ATP deficiency.

Keywords:  PITX2; atrial fibrillation; iPS-derived atrial cardiomyocytes; mitochondria; oxidative phosphorylation

DOI:  https://doi.org/10.3389/fphys.2023.1250951