{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,1]],"date-time":"2026-10-01T17:23:18Z","timestamp":1790875398283,"version":"4.1.0"},"reference-count":43,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2025,4,9]],"date-time":"2025-04-09T00:00:00Z","timestamp":1744156800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Pharmacol."],"abstract":"<jats:sec><jats:title>Background and Objectives<\/jats:title><jats:p>Improved detection sensitivity from combined Long-Range PCR (LR-PCR), Next-Generation Sequencing (NGS), and droplet digital PCR (ddPCR) to identify multiple large-scale mtDNA deletions (MLSMD) and quantify deletion heteroplasmy have introduced clinical interpretation challenges. We sought to evaluate clinical, biochemical, and histopathological phenotypes of a large clinical cohort harboring MLSMD in muscle to better understand their significance across a range of clinical phenotypes.<\/jats:p><\/jats:sec><jats:sec><jats:title>Methods<\/jats:title><jats:p>A single-site retrospective study was performed of 212 diagnostic muscle biopsies obtained from patients referred for Primary Mitochondrial Disease (PMD) evaluation with muscle mitochondrial (mt)DNA sequencing performed at our institution, including electronic medical record (EMR) review of symptoms, biochemical results, and Mitochondrial Myopathy Composite Assessment Tool (MM-COAST) scores.<\/jats:p><\/jats:sec><jats:sec><jats:title>Results<\/jats:title><jats:p>MLSMD were identified in 50 of 212 (24%) diagnostic tissue biopsies, and were universally present. in subjects \u226550 years (n = 18\/18). In 45 of 50 (90%) subjects with MLSMD, no definitive genetic etiology was identified, despite clinical whole exome sequencing (WES) and\/or whole genome sequencing (WGS). MLSMD heteroplasmy levels quantified by ddPCR ranged from 0% to 33%, exceeding 10% heteroplasmy in 5\/45 (11%). Subjects with MLSMD (n = 45) were more likely to demonstrate mitochondrial abnormalities on histopathology, upregulation (\u2265150% of control mean) of one or more electron transport chain (ETC) complex enzyme activities, and reduced citrate synthase indicative of mitochondrial depletion (&amp;lt;60% of control mean) relative to subjects without MLSMD (n = 155). As clinical phenotypes varied across the MLSMD cohort, Bernier diagnostic criteria major\/minor symptoms were used to discriminate 13 of 45 subjects with \u201csuspected\u201d PMD having unrevealing WES\/WGS results and 32 of 45 subjects scored as \u201cless likely\u201d to have PMD. Relative to the \u201cless likely\u201d cohort, a significantly higher frequency of biochemical and muscle histopathological abnormalities (ragged red and COX negative fibers) were observed in the \u201csuspected\u201d cohort, further supporting a higher index of suspicion for PMD, p &amp;lt; 0.05.<\/jats:p><\/jats:sec><jats:sec><jats:title>Discussion<\/jats:title><jats:p>MLSMD in skeletal muscle tissue were a common molecular finding (24%) in our cohort and consistently present in subjects \u226550\u00a0years. Among those with genetically undiagnosed MLSMD (n = 45), the \u201csuspected\u201d PMD subset (n = 13\/45) represent a promising cohort for novel gene discoveries.<\/jats:p><\/jats:sec>","DOI":"10.3389\/fphar.2025.1507493","type":"journal-article","created":{"date-parts":[[2025,4,9]],"date-time":"2025-04-09T07:31:20Z","timestamp":1744183880000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":3,"title":["Interpreting the clinical significance of multiple large-scale mitochondrial DNA deletions (MLSMD) in skeletal muscle tissue in the diagnostic evaluation of primary mitochondrial disease"],"prefix":"10.3389","volume":"16","author":[{"given":"Jing","family":"Wang","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"James T.","family":"Peterson","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Joaquim Diego D.","family":"Santos","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ada J. S.","family":"Chan","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Maria Alejandra","family":"Diaz-Miranda","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Imon","family":"Rahaman","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jean","family":"Flickinger","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Amy","family":"Goldstein","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Emily","family":"Bogush","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Elizabeth M.","family":"McCormick","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Colleen C.","family":"Muraresku","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Vernon E.","family":"Anderson","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Matthew C.","family":"Dulik","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Douglas C.","family":"Wallace","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rui","family":"Xiao","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Marni J.","family":"Falk","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Angela N.","family":"Viaene","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zarazuela","family":"Zolkipli-Cunningham","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2025,4,9]]},"reference":[{"key":"B1","doi-asserted-by":"publisher","first-page":"18054","DOI":"10.3390\/ijms160818054","article-title":"Genes and pathways involved in adult onset disorders featuring muscle mitochondrial DNA instability","volume":"16","author":"Ahmed","year":"2015","journal-title":"Int. J. Mol. Sci."},{"key":"B2","doi-asserted-by":"publisher","first-page":"812","DOI":"10.1111\/j.1442-9071.2010.02335.x","article-title":"Macular lesion resembling adult-onset vitelliform macular dystrophy in Kearns-Sayre syndrome with multiple mtDNA deletions","volume":"38","author":"Ascaso","year":"2010","journal-title":"Clin. Exp. Ophthalmol."},{"key":"B3","doi-asserted-by":"publisher","first-page":"515","DOI":"10.1038\/ng1769","article-title":"High levels of mitochondrial DNA deletions in substantia nigra neurons in aging and Parkinson disease","volume":"38","author":"Bender","year":"2006","journal-title":"Nat. Genet."},{"key":"B4","doi-asserted-by":"publisher","first-page":"1406","DOI":"10.1212\/01.wnl.0000033795.17156.00","article-title":"Diagnostic criteria for respiratory chain disorders in adults and children","volume":"59","author":"Bernier","year":"2002","journal-title":"Neurology"},{"key":"B5","doi-asserted-by":"publisher","first-page":"205","DOI":"10.1016\/j.mito.2021.01.004","article-title":"Mitochondrial DNA maintenance disorders in 102 patients from different parts of Russia: Mutational spectrum and phenotypes","volume":"57","author":"Bychkov","year":"2021","journal-title":"Mitochondrion"},{"key":"B6","doi-asserted-by":"publisher","first-page":"4612","DOI":"10.1093\/hmg\/ddu176","article-title":"Dissecting the mechanisms underlying the accumulation of mitochondrial DNA deletions in human skeletal muscle","volume":"23","author":"Campbell","year":"2014","journal-title":"Hum. Mol. Genet."},{"key":"B7","doi-asserted-by":"publisher","first-page":"481","DOI":"10.1002\/ana.22109","article-title":"Mitochondrial DNA deletions and neurodegeneration in multiple sclerosis","volume":"69","author":"Campbell","year":"2011","journal-title":"Ann. Neurol."},{"key":"B8","doi-asserted-by":"publisher","first-page":"99","DOI":"10.1212\/wnl.50.1.99","article-title":"Multiple mtDNA deletions features in autosomal dominant and recessive diseases suggest distinct pathogeneses","volume":"50","author":"Carrozzo","year":"1998","journal-title":"Neurology"},{"key":"B9","doi-asserted-by":"publisher","first-page":"530","DOI":"10.1007\/s004010050462","article-title":"Cytochrome c oxidase deficiencies in the muscle of patients with inflammatory myopathies","volume":"91","author":"Chariot","year":"1996","journal-title":"Acta Neuropathol."},{"key":"B10","article-title":"Primary mitochondrial disorders overview","author":"Chinnery","year":""},{"key":"B11","doi-asserted-by":"publisher","first-page":"131","DOI":"10.1146\/annurev.med.59.053006.104646","article-title":"Inherited mitochondrial diseases of DNA replication","volume":"59","author":"Copeland","year":"2008","journal-title":"Annu. Rev. Med."},{"key":"B14","doi-asserted-by":"publisher","first-page":"119","DOI":"10.1093\/gerona\/glp179","article-title":"The effect of aging on human skeletal muscle mitochondrial and intramyocellular lipid ultrastructure","volume":"65","author":"Crane","year":"2010","journal-title":"J. Gerontol. A Biol. Sci. Med. Sci."},{"key":"B15","article-title":"Mitochondrial DNA maintenance defects overview","author":"El-Hattab","year":""},{"key":"B16","doi-asserted-by":"publisher","first-page":"109","DOI":"10.1002\/crt2.41","article-title":"Development of a mitochondrial myopathy-composite assessment Tool","volume":"6","author":"Flickinger","year":"2021","journal-title":"JCSM Clin. Rep."},{"key":"B17","doi-asserted-by":"publisher","first-page":"11244","DOI":"10.1093\/nar\/gkaa804","article-title":"Mechanisms of replication and repair in mitochondrial DNA deletion formation","volume":"48","author":"Fontana","year":"2020","journal-title":"Nucleic Acids Res."},{"key":"B18","doi-asserted-by":"publisher","first-page":"323","DOI":"10.1093\/brain\/awt321","article-title":"Disease progression in patients with single, large-scale mitochondrial DNA deletions","volume":"137","author":"Grady","year":"2014","journal-title":"Brain"},{"key":"B19","doi-asserted-by":"publisher","first-page":"634","DOI":"10.1093\/jnen\/61.7.634","article-title":"Mitochondrial DNA deletions\/rearrangements in Parkinson disease and related neurodegenerative disorders","volume":"61","author":"Gu","year":"2002","journal-title":"J. Neuropathol. Exp. Neurol."},{"key":"B20","doi-asserted-by":"publisher","first-page":"1811","DOI":"10.1007\/s40520-020-01698-7","article-title":"Mitochondrial DNA deletion mutations increase exponentially with age in human skeletal muscle","volume":"33","author":"Herbst","year":"","journal-title":"Aging Clin. Exp. Res."},{"key":"B21","doi-asserted-by":"publisher","first-page":"1253","DOI":"10.1007\/s11357-021-00351-z","article-title":"Skeletal muscle mitochondrial DNA copy number and mitochondrial DNA deletion mutation frequency as predictors of physical performance in older men and women","volume":"43","author":"Herbst","year":"","journal-title":"Geroscience"},{"key":"B22","doi-asserted-by":"publisher","first-page":"75","DOI":"10.3390\/muscles2010007","article-title":"Diagnostic testing in suspected primary mitochondrial myopathy","volume":"2","author":"Hinojosa","year":"2023","journal-title":"Muscles"},{"key":"B23","doi-asserted-by":"publisher","first-page":"1401","DOI":"10.1016\/j.bbabio.2015.05.015","article-title":"Mitochondrial DNA mutations in neurodegeneration","volume":"1847","author":"Keogh","year":"2015","journal-title":"Biochim. Biophys. Acta"},{"key":"B24","doi-asserted-by":"publisher","first-page":"305","DOI":"10.1016\/j.pediatrneurol.2007.12.001","article-title":"Presentation and diagnosis of mitochondrial disorders in children","volume":"38","author":"Koenig","year":"2008","journal-title":"Pediatr. Neurol."},{"key":"B25","doi-asserted-by":"publisher","first-page":"518","DOI":"10.1038\/ng1778","article-title":"Mitochondrial DNA deletions are abundant and cause functional impairment in aged human substantia nigra neurons","volume":"38","author":"Kraytsberg","year":"2006","journal-title":"Nat. Genet."},{"key":"B26","doi-asserted-by":"publisher","first-page":"683","DOI":"10.1146\/annurev-biochem-060408-093701","article-title":"Somatic mitochondrial DNA mutations in mammalian aging","volume":"79","author":"Larsson","year":"2010","journal-title":"Annu. Rev. Biochem."},{"key":"B27","doi-asserted-by":"publisher","first-page":"508","DOI":"10.1093\/jnen\/nlz029","article-title":"Neurogenic muscle biopsy findings are common in mitochondrial myopathy","volume":"78","author":"Lu","year":"2019","journal-title":"J. Neuropathol. Exp. Neurol."},{"key":"B28","doi-asserted-by":"publisher","first-page":"248","DOI":"10.1186\/s13059-020-02138-5","article-title":"Ultrasensitive deletion detection links mitochondrial DNA replication, disease, and aging","volume":"21","author":"Lujan","year":"2020","journal-title":"Genome Biol."},{"key":"B29","doi-asserted-by":"publisher","first-page":"96","DOI":"10.1016\/s0168-8278(97)80286-3","article-title":"Multiple hepatic mitochondrial DNA deletions suggest premature oxidative aging in alcoholic patients","volume":"27","author":"Mansouri","year":"1997","journal-title":"J. Hepatol."},{"key":"B30","doi-asserted-by":"publisher","first-page":"714","DOI":"10.1097\/MOP.0000000000000686","article-title":"Mitochondrial disease genetics update: recent insights into the molecular diagnosis and expanding phenotype of primary mitochondrial disease","volume":"30","author":"Mccormick","year":"2018","journal-title":"Curr. Opin. Pediatr."},{"key":"B31","doi-asserted-by":"publisher","first-page":"573","DOI":"10.1016\/S1474-4422(21)00098-3","article-title":"Mitochondrial disease in adults: recent advances and future promise","volume":"20","author":"Ng","year":"2021","journal-title":"Lancet Neurol."},{"key":"B32","doi-asserted-by":"publisher","first-page":"603","DOI":"10.1001\/jamaneurol.2015.0150","article-title":"Clinical and pathological features of mitochondrial DNA deletion disease following antiretroviral treatment","volume":"72","author":"Payne","year":"2015","journal-title":"JAMA Neurol."},{"key":"B33","doi-asserted-by":"publisher","first-page":"1223","DOI":"10.1016\/s0891-5849(01)00517-2","article-title":"Age-related mitochondrial genotypic and phenotypic alterations in human skeletal muscle","volume":"30","author":"Pesce","year":"2001","journal-title":"Free Radic. Biol. Med."},{"key":"B34","doi-asserted-by":"publisher","first-page":"35","DOI":"10.1016\/j.mito.2019.10.005","article-title":"Growth and differentiation factor 15 as a biomarker for mitochondrial myopathy","volume":"50","author":"Poulsen","year":"2020","journal-title":"Mitochondrion"},{"key":"B35","doi-asserted-by":"publisher","first-page":"24","DOI":"10.1002\/ana.410370107","article-title":"Ragged red fibers in normal aging and inflammatory myopathy","volume":"37","author":"Rifai","year":"1995","journal-title":"Ann. Neurol."},{"key":"B36","doi-asserted-by":"publisher","first-page":"18","DOI":"10.1159\/000213303","article-title":"Increased mitochondrial DNA deletions in the skeletal muscle of myotonic dystrophy","volume":"38","author":"Sahashi","year":"1992","journal-title":"Gerontology"},{"key":"B37","doi-asserted-by":"publisher","first-page":"1039","DOI":"10.1001\/archotol.1997.01900100009001","article-title":"Mitochondrial DNA deletions associated with aging and presbyacusis","volume":"123","author":"Seidman","year":"1997","journal-title":"Arch. Otolaryngol. Head. Neck Surg."},{"key":"B38","doi-asserted-by":"publisher","first-page":"530","DOI":"10.1038\/nrg3966","article-title":"The dynamics of mitochondrial DNA heteroplasmy: implications for human health and disease","volume":"16","author":"Stewart","year":"2015","journal-title":"Nat. Rev. Genet."},{"key":"B39","doi-asserted-by":"publisher","first-page":"32","DOI":"10.1111\/j.1600-0404.1993.tb04071.x","article-title":"Significance of mitochondrial DNA deletions in myotonic dystrophy","volume":"87","author":"Thyagarajan","year":"1993","journal-title":"Acta Neurol. Scand."},{"key":"B40","doi-asserted-by":"publisher","first-page":"12S","DOI":"10.1111\/j.1530-0277.2000.tb00004.x","article-title":"Study of mitochondrial DNA deletion in alcoholics","volume":"24","author":"Tsuchishima","year":"2000","journal-title":"Alcohol Clin. Exp. Res."},{"key":"B41","first-page":"84","article-title":"The relationship between skeletal muscle mitochondrial citrate synthase activity and whole body oxygen uptake adaptations in response to exercise training","volume":"6","author":"Vigels\u00f8","year":"2014","journal-title":"Int. J. Physiol. Pathophysiol. Pharmacol."},{"key":"B42","doi-asserted-by":"publisher","first-page":"260","DOI":"10.1159\/000117269","article-title":"Respiratory chain encephalomyopathies: a diagnostic classification","volume":"36","author":"Walker","year":"1996","journal-title":"Eur. Neurol."},{"key":"B43","doi-asserted-by":"publisher","first-page":"93","DOI":"10.1016\/j.ymgme.2021.12.006","article-title":"Advanced approach for comprehensive mtDNA genome testing in mitochondrial disease","volume":"135","author":"Wang","year":"2022","journal-title":"Mol. Genet. Metab."},{"key":"B44","doi-asserted-by":"publisher","first-page":"e003559","DOI":"10.1161\/CIRCGEN.121.003559","article-title":"End stage mitochondrial cardiomyopathy and heart transplantation due to biallelic pathogenic C1QBP variants","volume":"15","author":"Wilcox","year":"2022","journal-title":"Circ. Genom Precis. Med."},{"key":"B45","doi-asserted-by":"publisher","first-page":"34","DOI":"10.1016\/0014-5793(92)80321-7","article-title":"Multiple mitochondrial DNA deletions in an elderly human individual","volume":"297","author":"Zhang","year":"1992","journal-title":"FEBS Lett."}],"container-title":["Frontiers in Pharmacology"],"original-title":[],"link":[{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fphar.2025.1507493\/full","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,4,9]],"date-time":"2025-04-09T07:31:31Z","timestamp":1744183891000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fphar.2025.1507493\/full"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,4,9]]},"references-count":43,"alternative-id":["10.3389\/fphar.2025.1507493"],"URL":"https:\/\/doi.org\/10.3389\/fphar.2025.1507493","relation":{},"ISSN":["1663-9812"],"issn-type":[{"value":"1663-9812","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,4,9]]},"article-number":"1507493"}}