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University of Groningen Exploring the mechanisms underlying the phenotype of MCAD deficiency with Systems Medicine Martines, Anne-Claire

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(1)University of Groningen. Exploring the mechanisms underlying the phenotype of MCAD deficiency with Systems Medicine Martines, Anne-Claire. IMPORTANT NOTE: You are advised to consult the publisher's version (publisher's PDF) if you wish to cite from it. Please check the document version below.. Document Version Publisher's PDF, also known as Version of record. Publication date: 2019 Link to publication in University of Groningen/UMCG research database. Citation for published version (APA): Martines, A-C. (2019). Exploring the mechanisms underlying the phenotype of MCAD deficiency with Systems Medicine: from computational model to mice to man. Rijksuniversiteit Groningen.. Copyright Other than for strictly personal use, it is not permitted to download or to forward/distribute the text or part of it without the consent of the author(s) and/or copyright holder(s), unless the work is under an open content license (like Creative Commons). Take-down policy If you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediately and investigate your claim.. Downloaded from the University of Groningen/UMCG research database (Pure): http://www.rug.nl/research/portal. For technical reasons the number of authors shown on this cover page is limited to 10 maximum.. Download date: 28-06-2021.

(2) Chapter 6 Towards identifying in vitro metabolic adaptations that explain symptomatology in MCADD individuals A short report Anne-Claire M.F. Martines1,#, Emmalie A. Jager1,2,#, Albert Gerding1,3, Justina C. Wolters1,2, Dirk-Jan Reijngoud1,2, Terry G.J. Derksϭ͕Ϯ͕ϰ͕‫ۥ‬, Barbara M. Bakkerƌϭ͕‫ۥ‬. 1 Laboratory. of Pediatrics, Center of Liver, Digestive and Metabolic Diseases, University of Groningen, Groningen, University Medical Center Groningen, Groningen, The Netherlands, 2 Department of Pediatrics, University Medical Center Groningen, University of Groningen, Groningen, The Netherlands 3 Department of Laboratory Medicine, University of Groningen, University Medical Center Groningen, The Netherlands 4 Section of Metabolic Diseases, Beatrix Children's Hospital, University Medical Center of Groningen, University of Groningen. η͕‫ۥ‬These. authors contributed equally.. Journal publication in preparation.

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(4) In vitro metabolic adaptations that explain symptomatology in MCADD individuals. Abstract

(5)  –Š‹• •–—†› ™‡ ‹˜‡•–‹‰ƒ–‡† —Ž–—”‡† •‹ ˆ‹„”‘„Žƒ•–• ‘ˆ ›‘—‰ •›’–‘ƒ–‹  ȋαͶȌ ƒ† ƒ•›’–‘ƒ–‹  Ǧ†‡ˆ‹ ‹‡– ȋȌ ‹†‹˜‹†—ƒŽ• ȋα͸Ȍǡ ‹ Ž—†‹‰ ƒ ‘Ž†‡”  ‹†‹˜‹†—ƒŽȋ͵Ͳ›‡ƒ”•Ȍ™Š‘”‡ƒ‹‡†ƒ•›’–‘ƒ–‹ ǤŠ‡ ‘–”‘Ž•‹ˆ‹„”‘„Žƒ•–ȋα͸Ȍ™‡”‡ ‘„–ƒ‹‡† ˆ”‘ ‹†‹˜‹†—ƒŽ• ƒ’’ƒ”‡–Ž› ˆ”‡‡ ‘ˆ ƒ ‡–ƒ„‘Ž‹  †‹•‡ƒ•‡Ǥ †‡” „ƒ•ƒŽ —Ž–—”‹‰ ‘†‹–‹‘•ǡ ͸ǦͳͲ ƒ ›ŽǦ ƒ”‹–‹‡ •‡ ”‡–‹‘ ƒ† ‹–”ƒ ‡ŽŽ—Žƒ” ͸Ǧ ƒ† ͺǦƒ ›ŽǦ ƒ”‹–‹‡ ‘ ‡–”ƒ–‹‘•™‡”‡Š‹‰Š‡”‹ˆ‹„”‘„Žƒ•–•–Šƒ‹ ‘–”‘Žˆ‹„”‘„Žƒ•–•Ǥ

(6) ƒ††‹–‹‘ǡ–Š‡ ˆ‹„”‘„Žƒ•–• ‘ˆ –Š‡ ‡Ž†‡•– ƒ•›’–‘ƒ–‹   ‹†‹˜‹†—ƒŽ •Š‘™‡† Š‹‰Š‡” ƒ ›ŽǦ ƒ”‹–‹‡ •‡ ”‡–‹‘ –Šƒ –Š‘•‡ ‘ˆ –Š‡ ‘–Š‡”  ‹†‹˜‹†—ƒŽ•Ǥ —ƒ–‹–ƒ–‹˜‡ǡ –ƒ”‰‡–‡† ’”‘–‡‘‹ • •Š‘™‡†ƒ—’”‡‰—Žƒ–‹‘‘ˆ‹–Š‡•ƒ‡ƒ•›’–‘ƒ–‹ ‹†‹˜‹†—ƒŽǤ–Š‡”™‹•‡‘ ƒ†ƒ’–ƒ–‹‘•‹–Š‡’”‘–‡‘‡™‡”‡ˆ‘—†‹–Š‡ˆ‹„”‘„Žƒ•–•ǤŠ‡”‡•—Ž–•‹ŽŽ—•–”ƒ–‡–Š‡ ’‘–‡–‹ƒŽƒ†–Š‡ ŠƒŽŽ‡‰‡•‘ˆƒ’‡”•‘ƒŽ‹œ‡†‡†‹ ‹‡•–”ƒ–‡‰›Ǥ‡†‹• —••Š‘™–Š‡ —Ž–—”‡ ‘†‹–‹‘•ƒ›„‡‘’–‹‹œ‡†–‘”‡˜‡ƒŽ‡–ƒ„‘Ž‹ ƒ†ƒ’–ƒ–‹‘•ǡˆ‘”‹•–ƒ ‡„›‹’‘•‹‰Ž‘™ ‰Ž— ‘•‡ ‘ ‡–”ƒ–‹‘•Ǥ . ʹͲ͹. 6.

(7) Chapter 6. Introduction  †‡ˆ‹ ‹‡ › ‹• ‘‡ ‘ˆ –Š‡ ‘•– ’”‡˜ƒŽ‡– ‹Š‡”‹–‡† ‡–ƒ„‘Ž‹  †‹•‡ƒ•‡• ƒ† –Š‡ ‘•– ‘‘‹„‘”‡””‘”‘ˆ–Š‡‹–‘ Š‘†”‹ƒŽˆƒ––›ƒ ‹†‘š‹†ƒ–‹‘ȋ Ȍ’ƒ–Š™ƒ›ȏͳȂ͸ȐǤ Š‡ ’”‡˜ƒŽ‡ ‡ƒ–„‹”–Š™ƒ•ͳȀͺ͵ͲͲ‹–Š‡‡–Š‡”Žƒ†•„‡–™‡‡ʹͲͲ͹ƒ†ʹͲͳͷȏ͹ȐǤ  ’Žƒ›•ƒ‹’‘”–ƒ–”‘Ž‡‹‡‡”‰›Š‘‡‘•–ƒ•‹•†—”‹‰ˆƒ•–‹‰ƒ† ‘†‹–‹‘•‘ˆŠ‹‰ŠǦ ‡‡”‰› †‡ƒ†Ǥ Ǧ†‡ˆ‹ ‹‡– ȋȌ ‹†‹˜‹†—ƒŽ• ”— ƒ ”‹• ‘ˆ ‡–ƒ„‘Ž‹  ”‹•‹• —†‡” •— Šƒ ‘†‹–‹‘ǡ‡•’‡ ‹ƒŽŽ›‹ˆ’”‘Ž‘‰‡†ˆƒ•–‹‰‘ —”•–‘‰‡–Š‡”™‹–Š‹–‡” —””‡–‹ŽŽ‡••ǡ•— Š ƒ• ƒ‹”™ƒ› ‘” ‰ƒ•–”‘‹–‡•–‹ƒŽ ‹ˆ‡ –‹‘• ȏͺȂͳͳȐǤ  —–‡ •›’–‘• ‹ Ž—†‡ Ž‹ˆ‡Ǧ–Š”‡ƒ–‡‹‰ Š›’‘‡–‘–‹ Š›’‘‰Ž› ‡‹ƒǡ ‘ƒƒ†•‡‹œ—”‡•Ǥ

(8) –‡”‡•–‹‰Ž›ǡ‡˜‡™‹–Š‹–Š‡•—„’‘’—Žƒ–‹‘ ‘ˆ‹†‹˜‹†—ƒŽ•™‹–ŠƒŠ‘‘œ›‰‘—•Ž‘••Ǧ‘ˆǦˆ— –‹‘ Ǥͻͺͷε —–ƒ–‹‘‹–Š‡Ǧ‡ ‘†‹‰ ACADM ‰‡‡ǡ •›’–‘• ƒ ”ƒ‰‡ ˆ”‘ ˆƒ–ƒŽ Š›’‘‡–‘–‹  Š›’‘‰Ž› ‡‹ƒ ‡ƒ”Ž› ‹ Ž‹ˆ‡ –‘ ƒ ‘’Ž‡–‡Žƒ ‘ˆ•›’–‘•–Š”‘—‰Š‘—–Ž‹ˆ‡ȏͳͲǡͳʹȂʹͲȐǤ ‡‡–‹ ƒ†‡˜‹”‘‡–ƒŽ‘†‹ˆ‹‡”• ‘—Ž† ’Žƒ› ƒ ”‘Ž‡ ‹ –Š‡ Š‡–‡”‘‰‡‡‹–› ‹ ˜—Ž‡”ƒ„‹Ž‹–› –‘ ‘” ”‘„—•–‡•• ƒ‰ƒ‹•– ‡–ƒ„‘Ž‹  •–”‡••Ǥ ‘™–Š‹•˜—Ž‡”ƒ„‹Ž‹–›‘””‘„—•–‡••ƒ”‹•‡•‹•ƒ•›‡–— Ž‡ƒ”Ǥ •ƒŽ”‡ƒ†›‹†‹ ƒ–‡†‹–Š‡ ‡‡”ƒŽ

(9) –”‘†— –‹‘‹Šƒ’–‡”ͳ‘ˆ–Š‹•–Š‡•‹•ǡ‹–Š‡’ƒ•–†‡ ƒ†‡• ”‡Ž‹ƒ„Ž‡—–ƒ”‰‡–‡†ƒ†–ƒ”‰‡–‡†—Ž–‹Ǧ‘‹ •‡–Š‘†•Šƒ˜‡„‡‡†‡˜‡Ž‘’‡†ǤŠ‹•‘ˆˆ‡”•–Š‡ ‘’’‘”–—‹–›–‘•–—†›‹„‘”‡””‘”•‘ˆ‡–ƒ„‘Ž‹•‘Ž‡˜‡Ž•‘–’”‡˜‹‘—•Ž›’‘••‹„Ž‡ȏʹͳȂʹ͵ȐǤ Š‡ ƒ’’Ž‹ ƒ–‹‘ ‘ˆ –Š‡•‡ —Ž–‹Ǧ‘‹ • –‡ Š‘Ž‘‰‹‡• ‘ —Ž–—”‡† •‹ ˆ‹„”‘„Žƒ•–• ‘”‹‰‹ƒ–‹‰ ˆ”‘•›’–‘ƒ–‹ ƒ†ƒ•›’–‘ƒ–‹ ‹†‹˜‹†—ƒŽ• ƒ„‡ƒˆ‹”•–•–‡’–‘‰ƒ‹‹•‹‰Š– ‹–‘”‡‰—Žƒ–‘”›‡ Šƒ‹••—†‡”Ž›‹‰•›’–‘ƒ–‘Ž‘‰›‹†‡ˆ‹ ‹‡ ›Ǥ

(10) ƒ††‹–‹‘ǡ‹– ‹‰Š–‡˜‡ƒŽŽ‘™—•–‘‹†‡–‹ˆ›‡ƒ‹‰ˆ—Ž„‹‘ƒ”‡”•–Šƒ–’”‡†‹ –™Š‡–Š‡”ƒ’ƒ–‹‡– ‘—Ž† †‡˜‡Ž‘’ •›’–‘• ƒ† ™Š‹ Š ƒ†ƒ’–ƒ–‹‘• ‹ ‡‡”‰› ‡–ƒ„‘Ž‹• ‘—Ž† —†‡”Ž‹‡ –Š‡‹” ”‘„—•–‡•• ‘” ˜—Ž‡”ƒ„‹Ž‹–›Ǥ ”‡˜‹‘—•Ž›ǡ ™‡ ‘„•‡”˜‡† ƒ†ƒ’–ƒ–‹‘• ‹ –Š‡ ’”‘–‡‘‡ ‘ˆ ‹–‘ Š‘†”‹ƒŽ ‡‡”‰› ‡–ƒ„‘Ž‹• ‹ —Ž–—”‡† •‹ ˆ‹„”‘„Žƒ•–• ‘ˆ  ‹†‹˜‹†—ƒŽ• ‘’ƒ”‡†–‘ ‘–”‘Ž•ǤŠ‡Ž‡˜‡Ž•‘ˆ•‡˜‡”ƒŽ ’”‘–‡‹•ǡ‹ Ž—†‹‰ǡǡ •—„—‹–ƒ† •—„—‹–ǡ™‡”‡Š‹‰Š‡”‹‹–‘ Š‘†”‹ƒ‹•‘Žƒ–‡†ˆ”‘•‹ˆ‹„”‘„Žƒ•–•‘ˆ –Š”‡‡’ƒ–‹‡–•™‹–Š–Š‡Š‘‘œ›‰‘—• Ǥͻͺͷε ACADM—–ƒ–‹‘–Šƒ‹‹–‘ Š‘†”‹ƒ ˆ”‘ ‘–”‘Žˆ‹„”‘„Žƒ•–•ȏʹͶȐǤŠ‡Žƒ––‡”•–—†›ˆ‘ —•‡†‘–Š‡†‡˜‡Ž‘’‡–‘ˆ–Š‡ –ƒ”‰‡–‡† ’”‘–‡‘‹ •‡–Š‘†‘Ž‘‰›Ǥ Š‡ ’ƒ–‹‡– ˆ‹„”‘„Žƒ•–•‹ŽŽ—•–”ƒ–‡† –Š‡ ’‘–‡–‹ƒŽ ‘ˆ –Š‡ –‡ Š‹“—‡Ǥ —‡–‘–Š‡˜‡”›Ž‘™—„‡”‘ˆ‹†‹˜‹†—ƒŽ•ǡŠ‘™‡˜‡”ǡ‘ ‘ Ž—•‹‘•‘ˆƒ› Ž‹‹ ƒŽ ”‡Ž‡˜ƒ ‡ ‘—Ž†„‡†”ƒ™Ǥ

(11) ‘”†‡”–‘•–—†›–Š‡‡ˆˆ‡ –‘ˆ‘†‹ˆ‹‡”•‘–Š‡•›’–‘ƒ–‘Ž‘‰›‹ ǡ ƒ Žƒ”‰‡” —„‡” ‘ˆ „‘–Š •›’–‘ƒ–‹  ƒ† ƒ•›’–‘ƒ–‹   ‹†‹˜‹†—ƒŽ• ‹• ”‡“—‹”‡†Ǥ

(12) –Š‡•–—†›’”‡•‡–‡†Š‡”‡ǡ™‡‡š’Ž‘”‡†™Š‡–Š‡”•›’–‘ƒ–‹ ƒ†ƒ•›’–‘ƒ–‹  ˆ‹„”‘„Žƒ•–• ƒ† ‘–”‘Ž ˆ‹„”‘„Žƒ•–• •Š‘™ †‹ˆˆ‡”‡ ‡• ‹ –Š‡‹” ‹–‘ Š‘†”‹ƒŽ ‡‡”‰› ‡–ƒ„‘Ž‹•—†‡”„ƒ•‡Ž‹‡ ‘†‹–‹‘•ȋ•–ƒ†ƒ”† —Ž–—”‡‡†‹—™‹–Š͸Ǥͳ‰Ž— ‘•‡ȌǤ‡ ‡ƒ•—”‡†–Š‡’”‘–‡‘‡‘ˆ‹–‘ Š‘†”‹ƒŽ‡‡”‰›‡–ƒ„‘Ž‹•ȏʹͶȐƒ†–Š‡‹–”ƒ ‡ŽŽ—Žƒ”ƒ† ‡š–”ƒ ‡ŽŽ—Žƒ” ƒ ›ŽǦ ƒ”‹–‹‡ ’”‘ˆ‹Ž‡• ‘ˆ ˆ‹„”‘„Žƒ•–•‘ˆ Ͷ •›’–‘ƒ–‹ ǡ ͸ ƒ•›’–‘ƒ–‹  ƒ† ͸ ‹†‹˜‹†—ƒŽ•ƒ’’ƒ”‡–Ž›ˆ”‡‡‘ˆƒ‡–ƒ„‘Ž‹ †‹•‡ƒ•‡Ǥ

(13) –Š‡ˆ‹”•–’ƒ”–‘ˆ–Š‹••–—†›ǡ‘‡„‹‘Ž‘‰‹ ƒŽ ”‡’Ž‹ ƒ–‡ ˆ‘” ‡ƒ Š •‹ ˆ‹„”‘„Žƒ•– ‡ŽŽ Ž‹‡ ™ƒ• ‰‡‡”ƒ–‡†ǡ ƒ• ™‡ •‡ƒ” Š‡† ˆ‘” †‹ˆˆ‡”‡ ‡• „‡–™‡‡ –Š‡ ‰”‘—’•Ǥ Š‡ ”‡•—Ž–• ‘ˆ –Š‡ ƒƒŽ›•‹• •—‰‰‡•–‡† ƒŒ‘” ƒ†ƒ’–ƒ–‹‘• ‹ ‘Ž› ‘‡ ‹†‹˜‹†—ƒŽǣ „‘–Š –Š‡ ’”‘–‡‘‡ ƒ† –Š‡ ƒ ›ŽǦ ƒ”‹–‹‡ ’”‘ˆ‹Ž‡ ‘ˆ –Š‡ ‡Ž†‡•– ƒ•›’–‘ƒ–‹   ‹†‹˜‹†—ƒŽ †‹ˆˆ‡”‡† •—„•–ƒ–‹ƒŽŽ› ˆ”‘ –Šƒ– ‘ˆ ƒŽŽ ‘–Š‡”  ‹†‹˜‹†—ƒŽ•Ǥ Š‡•‡ ’‘–‡–‹ƒŽŽ› ˆ— –‹‘ƒŽ ƒ†ƒ’–ƒ–‹‘• ‘—Ž† —†‡”Ž‹‡ –Š‡ Žƒ  ‘ˆ •›’–‘• ‹ –Š‹• ‹†‹˜‹†—ƒŽǤ Š‡”‡ˆ‘”‡ǡ ™‡ ‡š–‡†‡† –Š‡ •–—†› ƒ† ”‡’‡ƒ–‡† –Š‡ ƒƒŽ›•‹• ‘ˆ –Š‡ ’”‘–‡‘‡ ƒ† ƒ ›ŽǦ ʹͲͺ.

(14) In vitro metabolic adaptations that explain symptomatology in MCADD individuals ƒ”‹–‹‡’”‘ˆ‹Ž‡‹ƒ–‘–ƒŽ‘ˆ͵ǦͶ„‹‘Ž‘‰‹ ƒŽ”‡’Ž‹ ƒ–‡•ˆ‹„”‘„Žƒ•– —Ž–—”‡•‘ˆ‡ƒ Š‹†‹˜‹†—ƒŽǡ–‘ ˜ƒŽ‹†ƒ–‡–Š‡‹‹–‹ƒŽ‡š’‡”‹‡–ƒŽˆ‹†‹‰•ǤŠ‡ƒ‹”‡•—Ž–•ƒ”‡’”‡•‡–‡†‹–Š‹• Šƒ’–‡”Ǥ. Results Cohort and their characteristics ‡—•‡†’”‹ƒ”›Š—ƒ•‹ˆ‹„”‘„Žƒ•–•ˆ”‘‹†‹˜‹†—ƒŽ•Š‘‘œ›‰‘—•ˆ‘”–Š‡Ž‘••Ǧ ‘ˆǦˆ— –‹‘ Ǥͻͺͷε —–ƒ–‹‘ȋƒ„Ž‡ͳȌǤ  Table 1: Summary of patient characteristics. Patient ID. Gender. Gene mutation. Manifestation. ͳ. . Ǥͻͺͷε . ›’–. Age of onset/ Diagnosis (years) ʹ. ʹ. . Ǥͻͺͷε . •›’–. ͳ. ͵. . Ǥͻͺͷε . •›’–. ͳǤͷ. Ͷ. . Ǥͻͺͷε . •›’–. ͳͳ. ͷ. . Ǥͻͺͷε . ›’–. ʹ. ͸. . Ǥͻͺͷε . •›’–. Ͳ. ͹. . Ǥͻͺͷε . ›’–. ʹ. ͺ. . Ǥͻͺͷε . ›’–. ͳ. ͻ. . Ǥͻͺͷε . •›’–. Ͳ. ͳͲ. . Ǥͻͺͷε . •›’–. ͵Ͳ. Clinical manifestation. ‘—† •—„Ǧ ‘ƒ–‘•‡ ‹ „‡†ǡ ™‹–Š Š›’‘‡–‘–‹  Š›’‘‰Ž› ‡‹ƒǡ ƒˆ–‡” ƒ ‡’‹•‘†‡‘ˆ†‹ƒ””Š‡ƒǤ Ž†‡” •‹„Ž‹‰ ’”‡•‡–‡† ƒ– ƒ‰‡ ͳ ™‹–Š Ǧ”‡Žƒ–‡† ‡–ƒ„‘Ž‹  ”‹•‹•Ǥ Š‡”‡ƒˆ–‡”ǡ–ʹ™ƒ•–”‡ƒ–‡†™‹–Šˆ”‡“—‡– ˆ‡‡†‹‰• ƒ† ƒ ˆƒ–Ǧ”‡•–”‹ –‡† †‹‡–Ǥ †‹––‡† •‡˜‡”ƒŽ –‹‡• ƒ• ƒ ’”‡ ƒ—–‹‘ †—”‹‰ ‹–‡” —””‡– ‹ŽŽ‡••ǡ „—– ™‹–Š‘—– †‘ —‡–‡† Š›’‘‰Ž› ‡‹ƒ ‘” ‘–Š‡” •‡˜‡”‡•›’–‘•Ǥ ‹ƒ‰‘•‡†ƒˆ–‡”–™‹•‹„Ž‹‰†‹‡†ƒ–ͳǤͷ› †—”‹‰ ƒ ˆŽ— ‡’‹•‘†‡Ǥ ”‡ƒ–‡† ™‹–Š ˆ”‡“—‡–ˆ‡‡†‹‰•ƒ†Ǧ ƒ”Ǥ Ž†‡” •‹„Ž‹‰ ‘ˆ –͵Ǥ ‘ †‘ —‡–‡† †‡”ƒ‰‡‡–•Ǥ ”‡•‡–‡† ™‹–Š ’‘•–Ǧ‘’‡”ƒ–‹˜‡ Š›’‘‰Ž› ‡‹  ‘ƒǡ Ž‡ƒ†‹‰ –‘ ”‡ƒ‹ƒ–‹‘ƒ†•‡˜‡”‡†‹•ƒ„‹Ž‹–›Ǥ ‘—‰‡”•‹„Ž‹‰‘ˆ–ͷǤ”‡ƒ–‡†ˆ”‘‡ƒ” „‹”–Š ™‹–Š ˆ”‡“—‡– ˆ‡‡†‹‰•Ǥ ‘ †‘ —‡–‡††‡”ƒ‰‡‡–•Ǥ ‘—† Ž‡–Šƒ”‰‹  ƒ† Š›’‘–‘‹  ‹ „‡†ǡ ƒ ‘’ƒ‹‡† „› •‡‹œ—”‡• ƒ† Š›’‘‰Ž› ‡‹ƒǤ ‘ƒ–‘•‡ ƒ† Š›’‘‰Ž› ‡‹  ƒˆ–‡” –™‘ †ƒ›•‘ˆ˜‘‹–‹‰ƒ†Š‹‰Šˆ‡˜‡”Ǥ ‘—‰‡”•‹„Ž‹‰‘ˆ–ͺǤ”‡ƒ–‡†ˆ”‘‡ƒ” „‹”–Š ™‹–Š ˆ”‡“—‡– ˆ‡‡†‹‰•ǡ Ǧ ƒ” ƒ† ˜‹–ƒ‹ ʹǤ ‘ †‘ —‡–‡† †‡”ƒ‰‡‡–•Ǥ ‹• ‘˜‡”‡† „› ‡–ƒ„‘Ž‹  ’”‘ˆ‹Ž‡ ƒˆ–‡” ’‘•‹–‹˜‡‘ˆ†ƒ—‰Š–‡”Ǥ‘†‘ —‡–‡† †‡”ƒ‰‡‡–•ƒ• Š‹Ž†Ǥ. ›’–ǣ•›’–‘ƒ–‹ ǡ•›’–ǣ•›’–‘ƒ–‹ Ǥƒ–‡‰‘”‹œƒ–‹‘‘ˆ•›’–‘ƒ–‘Ž‘‰›ƒ ‘”†‹‰–‘ȏͳͲȐ.ǣƒŽ‡ǡ ǣˆ‡ƒŽ‡Ǥ Ǧ ƒ”ǣǦ ƒ”‹–‹‡Ǥ–ǣƒ–‹‡–Ǥ.  Š‡ Ǥͻͺͷε  ACADM —–ƒ–‹‘ ƒ—•‡• ‹’ƒ‹”‡– ‘ˆ –Š‡  ’”‘–‡‹ ˆ‘Ž†‹‰ ȏʹͷȐ ƒ† ’”‘–‡‹‹•–ƒ„‹Ž‹–›ȏʹ͸ǡʹ͹Ȑ”‡•—Ž–‹‰‹†‡‰”ƒ†ƒ–‹‘‘ˆ–Š‡—–ƒ–‡†’”‘–‡‹ƒ†Žƒ ‘ˆƒ –‹˜‹–› ȏʹͺȐǤ‡•’‡ ‹ˆ‹ ƒŽŽ›•–—†‹‡†ˆ‹„”‘„Žƒ•–•–Šƒ–‘”‹‰‹ƒ–‡†ˆ”‘’ƒ–‹‡–•ȋͶ›’–‘ƒ–‹  ƒ†͸ƒ•›’–‘ƒ–‹ ‹†‹˜‹†—ƒŽ•ˆ”‘͹ˆƒ‹Ž‹‡•Ȍ™Š‘™‡”‡ Ž‹‹ ƒŽŽ›ƒ• ‡”–ƒ‹‡†„‡ˆ‘”‡–Š‡. ʹͲͻ. 6.

(15) Chapter 6 ‹–”‘†— –‹‘‘ˆ–Š‡‡™„‘”• ”‡‡‹‰ȋȌ’”‘‰”ƒƒ†‘ˆ™Š‘–Š‡•›’–‘ƒ–‘Ž‘‰›Šƒ† „‡‡ ’”‡˜‹‘—•Ž› †‡• ”‹„‡† ȏͳͲȐǤ

(16) – •Š‘—Ž† „‡ ‘–‡† –Šƒ– •‘‡ ‘ˆ –Š‡ ‹†‹˜‹†—ƒŽ• –Šƒ– ™‡”‡ ƒ–‡‰‘”‹œ‡† ƒ• ƒ•›’–‘ƒ–‹ ǡ ”‡ ‡‹˜‡† –”‡ƒ–‡– „‡ˆ‘”‡ –Š‡ ƒ˜‡”ƒ‰‡ ƒ‰‡ ‘ˆ †‹•‡ƒ•‡ ’”‡•‡–ƒ–‹‘ ȋƒ„Ž‡ ͳȌǤ Š‡•‡ Š‹Ž†”‡ ™‡”‡ †‹ƒ‰‘•‡† ƒˆ–‡” ƒ •‹„Ž‹‰ ’”‡•‡–‡† ™‹–Š •›’–‘•Ǥ

(17) ˆƒ –ǡ–Š‡‡†‹ƒȋ”ƒ‰‡Ȍƒ‰‡‘ˆ‘•‡–‹•›’–‘ƒ–‹ ‰”‘—’™ƒ•ʹȋͳǦʹȌ›‡ƒ”•ǡ ™Š‹Ž‡ –Š‡ ‡†‹ƒ ƒ‰‡ ‘ˆ †‹ƒ‰‘•‹• ‹ –Š‡ ƒ•›’–‘ƒ–‹  ‰”‘—’ ™ƒ• ͳǤʹͷ ȋͲǦ͵ͲȌ ›‡ƒ”•Ǥ • –”‡ƒ–‡– ƒ ‹–‡”ˆ‡”‡ ™‹–Š –Š‡ ƒ–—”ƒŽ ‘—”•‡ ‘ˆ †‹•‡ƒ•‡ †‡˜‡Ž‘’‡–ǡ ‹– ‹• ‘– ‡”–ƒ‹ ™Š‡–Š‡” •‘‡ ‘ˆ –Š‡  ‹†‹˜‹†—ƒŽ• ƒ–‡‰‘”‹œ‡† ƒ• ƒ•›’–‘ƒ–‹  ƒ”‡ –”—Ž› ƒ•›’–‘ƒ–‹ ǤŠ‡”‡ˆ‘”‡ǡ‹–‡”’”‡–ƒ–‹‘‘ˆ”‡•—Ž–•‘„–ƒ‹‡†‹ˆ‹„”‘„Žƒ•–•‘ˆƒ•›’–‘ƒ–‹  ‹†‹˜‹†—ƒŽ••Š‘—Ž†„‡ ‘•‹†‡”‡†™‹–Š ƒ—–‹‘Ǥ

(18) ’‘”–ƒ–Ž›ǡ’ƒ–‹‡–ͳͲȋ–ͳͲȌ™ƒ• –Š‡ ‡Ž†‡•–  ‹†‹˜‹†—ƒŽ ‹ –Š‡ •–—†‹‡† ‘Š‘”–Ǥ –ͳͲ Šƒ• ”‡ƒ‹‡† ƒ•›’–‘ƒ–‹  –Š”‘—‰Š‘—–Ž‹ˆ‡™‹–Š‘—––”‡ƒ–‡–ǤŠ‡™ƒ•‹†‡–‹ˆ‹‡†ƒ–ƒ‰‡͵Ͳ„‡ ƒ—•‡‘ˆƒ’‘•‹–‹˜‡”‡•—Ž–‹ –Š‡ ‡™„‘” • ”‡‡‹‰ ‘ˆ Š‡” ‡™„‘” Š‹Ž†Ǥ –ͳͲ ƒ –Š—• „‡ ‘•‹†‡”‡† –Š‡ ‘•– ƒ• ‡”–ƒ‹‡† ƒ•›’–‘ƒ–‹   ‹†‹˜‹†—ƒŽǤ ‘” –Š‹• ”‡ƒ•‘ǡ –Š‡ ”‡•—Ž–• ‘ˆ –ͳͲ ™‹ŽŽ „‡ ’”‡•‡–‡† ƒ† †‹• —••‡† •‡’ƒ”ƒ–‡Ž›Ǥ Šƒ”ƒ –‡”‹•–‹ • ‘ˆ –Š‡ ‘–”‘Ž ‘Š‘”– ƒ”‡ ‰‹˜‡ ‹ —’’Ž‡‡–ƒ”›ƒ„Ž‡ͳǤ . Acyl-carnitines ‹”•–ǡ ™‡ ’‡”ˆ‘”‡† ƒ ‡š’Ž‘”ƒ–‹˜‡ •–—†› ‘ ƒ •‹‰Ž‡ •ƒ’Ž‡ ‘ˆ ‡ƒ Š ‘ˆ –Š‡ —Ž–—”‡† •‹ ˆ‹„”‘„Žƒ•–• ‘ˆ ƒŽŽ ‹†‹˜‹†—ƒŽ•Ǥ Š‡ ‘–‹˜ƒ–‹‘ ˆ‘” –Š‹• ƒ’’”‘ƒ Š ™ƒ• –Šƒ– ™‡ ‡š’‡ –‡† †‹ˆˆ‡”‡ ‡• „‡–™‡‡ –Š‡ ‰”‘—’• ”ƒ–Š‡” –Šƒ „‡–™‡‡ –Š‡ ‹†‹˜‹†—ƒŽ•Ǥ ‡ ‡ƒ•—”‡† ƒ ›ŽǦ ƒ”‹–‹‡ ’”‘ˆ‹Ž‡• ƒ† –Š‡ –ƒ”‰‡–‡† ’”‘–‡‘‡ ‘ˆ ‹–‘ Š‘†”‹ƒŽ ‡‡”‰› ‡–ƒ„‘Ž‹•Ǥ Š‡ ‘ ‡–”ƒ–‹‘•‘ˆ͸ǦǡͺǦƒ†ͳͲǣͳǦƒ ›ŽǦ ƒ”‹–‹‡ǡ„—–‘–‘ˆͳͲǦƒ ›ŽǦ ƒ”‹–‹‡ǡ™‡”‡Š‹‰Š‡” ‹ –Š‡ —Ž–—”‡ ‡†‹— ‘ˆ  ˆ‹„”‘„Žƒ•–• –Šƒ ‹ –Šƒ– ‘ˆ ‘–”‘Ž ˆ‹„”‘„Žƒ•–• ȋ—’’Ž‡‡–ƒ”› ‹‰—”‡ͳȌǤ•–Š‡ˆ”‡•Š‡†‹—ƒŽ”‡ƒ†› ‘–ƒ‹‡†ƒ ›ŽǦ ƒ”‹–‹‡•ǡ™‡ƒŽ•‘ ƒŽ —Žƒ–‡†–Š‡ Šƒ‰‡‹ƒ ›ŽǦ ƒ”‹–‹‡ ‘ ‡–”ƒ–‹‘•‹–Š‡‡†‹—†—”‹‰ —Ž–‹˜ƒ–‹‘ƒ† ‘”ƒŽ‹œ‡†–Š‡”‡•—Ž––‘ ‡ŽŽ—Žƒ”’”‘–‡‹ ‘–‡–ǡ–‘ƒ ‘—–ˆ‘”†‹ˆˆ‡”‡ ‡•‹ ‘ˆŽ—‡ ‡‘ˆ –Š‡ ‡ŽŽ•Ǥ Š‡  ˆ‹„”‘„Žƒ•–• •‡ ”‡–‡† ‘”‡ ͸Ǧ ƒ† ͺǦƒ ›ŽǦ ƒ”‹–‹‡• ‘’ƒ”‡† –Š‡ ‘–”‘Ž ˆ‹„”‘„Žƒ•–•Ǥ

(19)  ƒ††‹–‹‘ǡ –Š‡ ˆ‹„”‘„Žƒ•–• ˆ”‘ –ͳͲ •‡ ”‡–‡† ‘”‡ ͸ǦͳͲǦƒ ›ŽǦ ƒ”‹–‹‡–Šƒ–Š‡‘–Š‡”ˆ‹„”‘„Žƒ•–•ȋ ‹‰—”‡ͳǦȌǤ‡ƒŽ•‘‡ƒ•—”‡†‹–”ƒ ‡ŽŽ—Žƒ” ƒ ›ŽǦ ƒ”‹–‹‡ Ž‡˜‡Ž•Ǥ Š‹••Š‘™‡†–Šƒ– •›’–‘ƒ–‹  ƒ†ƒ•›’–‘ƒ–‹  ˆ‹„”‘„Žƒ•–• Šƒ† •‹‰‹ˆ‹ ƒ–Ž› Š‹‰Š‡” ‹–”ƒ ‡ŽŽ—Žƒ” ͸Ǧǡ ͺǦ „—– •‹‹Žƒ” ͳͲǣͳǦ ƒ† ͳͲǦƒ ›ŽǦ ƒ”‹–‹‡ ‘ ‡–”ƒ–‹‘• ‘’ƒ”‡† –‘ ‘–”‘Ž ˆ‹„”‘„Žƒ•–• ȋ ‹‰—”‡ ͳǦ ȌǤ —”–Š‡”‘”‡ǡ ™‡ †‹† ‘– ‘„•‡”˜‡ †‹ˆˆ‡”‡ ‡• „‡–™‡‡ –Š‡ ‹–”ƒ ‡ŽŽ—Žƒ” ‘ ‡–”ƒ–‹‘• ‘ˆ ƒ ›ŽǦ ƒ”‹–‹‡• ‹ –Š‡ •‹ ˆ‹„”‘„Žƒ•–‘ˆ–ͳͲƒ†–Š‘•‡‘ˆ–Š‡•›’–‘ƒ–‹ ƒ†‘–Š‡”ƒ•›’–‘ƒ–‹ ˆ‹„”‘„Žƒ•–•ȋ ‹‰—”‡ ͳǦ ȌǤ–Š‡”ƒ ›ŽǦ ƒ”‹–‹‡•™‡”‡‘–†‹ˆˆ‡”‡–„‡–™‡‡ƒ›‘ˆ–Š‡‰”‘—’•ȋ†ƒ–ƒ‘–•Š‘™ȌǤ ‹ ‡–ͳͲ†‹ˆˆ‡”‡†ˆ”‘ƒŽŽ‘–Š‡”‹†‹˜‹†—ƒŽ•ǡ ™‡•‡–‘—––‘˜ƒŽ‹†ƒ–‡–Š‡‘„•‡”˜ƒ–‹‘•™‹–Š —Ž–‹’Ž‡ ”‡’Ž‹ ƒ–‡ ˆ‹„”‘„Žƒ•– —Ž–—”‡• ’‡” ‹†‹˜‹†—ƒŽǤ Š‡ ‘ ‡–”ƒ–‹‘• ‘ˆ ͸Ǧǡ ͺǦǡ ƒ† ͳͲǣͳǦƒ ›ŽǦ ƒ”‹–‹‡‹–Š‡‡†‹—‘ˆƒŽŽ„—–‘‡’ƒ–‹‡– ‡ŽŽŽ‹‡ȋ–͹Ȍ™‡”‡Š‹‰Š‡”–Šƒ–Š‘•‡ ‘ˆ–Š‡ ‘–”‘Žˆ‹„”‘„Žƒ•–•ȋ—’’Ž‡‡–ƒ”› ‹‰—”‡ʹǦȌǤ‡˜‡”ƒŽ’ƒ–‹‡–ˆ‹„”‘„Žƒ•– —Ž–—”‡• Šƒ†ƒŽ•‘Š‹‰Š‡”ͳͲǦƒ ›ŽǦ ƒ”‹–‹‡Ž‡˜‡Ž•‹–Š‡‡†‹—ȋ—’’Ž‡‡–ƒ”› ‹‰—”‡ʹȌǤ. ʹͳͲ.

(20) In vitro metabolic adaptations that explain symptomatology in MCADD individuals A. B 15. ## 1.0. 0.5. §§. ' medium C8 (Pmol/g protein). ' medium C6 (P mol/g protein). 1.5. 0.0 Asympt (ex. Pt10) Pt10. Sympt. Asympt (ex. Pt10) Pt10. Ctrl. D 2.0. ' medium C10 (Pmol/g protein). 2.5. ' medium C10:1 (P mol/g protein). §,¥. Ctrl. C 2.0 1.5 1.0 0.5 0.0. # 1.5. 6. 1.0 0.5 0.0. Sympt. Asympt (ex. Pt10) Pt10. Ctrl. E. Sympt. Asympt (ex. Pt10) Pt10. Ctrl. F 0.25. 1.0. 0.5. §§. intracellular C8 (nmol/g protein). 1.5. intracellular C6 (nmol/g protein). 5. 0 Sympt. 0.0. 0.20 0.15 0.10 0.05. §§. 0.00 Sympt. Asympt (ex. Pt10) Pt10. Ctrl. G. Sympt. Asympt (ex. Pt10) Pt10. Ctrl. Sympt. Asympt (ex. Pt10) Pt10. Ctrl. H 1.0. intracellular C10 (nmol/g protein). 1.0. intracellular C10:1 (nmol/g protein). ##,* 10. 0.8 0.6 0.4 0.2 0.0. 0.8 0.6 0.4 0.2 0.0. Sympt. Asympt (ex. Pt10) Pt10. Ctrl. . Figure 1Ǥ ͸ǦͳͲǦƒ ›ŽǦ ƒ”‹–‹‡ •‡ ”‡–‹‘ ‹–‘ ‹ –Š‡ ‡†‹— ȋǦȌ ƒ† ‹–”ƒ ‡ŽŽ—Žƒ” ͸Ǧ ƒ† ͺǦƒ ›ŽǦ ƒ”‹–‹‡ ‘ ‡–”ƒ–‹‘•ȋǦ ȌǤ‡ ”‡–‹‘‘ˆƒ ›ŽǦ ƒ”‹–‹‡•‹••Š‘™ƒ•–Š‡†‹ˆˆ‡”‡ ‡‹ ‘ ‡–”ƒ–‹‘‘ˆ ƒ ›Ž ƒ”‹–‹‡• ‹ –Š‡ ‡†‹— „‡ˆ‘”‡ ƒ† ƒˆ–‡” —Ž–—”‡ ‘ˆ ˆ‹„”‘„Žƒ•–•ǡ ‘”ƒŽ‹œ‡† ˆ‘” –Š‡ ‡ŽŽ—Žƒ” ’”‘–‡‹ ‘ ‡–”ƒ–‹‘ǤŠ‡”‡•—Ž–’‡”–ƒ‹–‘–Š‡‡š’Ž‘”ƒ–‹˜‡’ƒ”–‘ˆ–Š‡•–—†›ǤαͶǡͷǡƒ†Ͷ†‹ˆˆ‡”‡–‹†‹˜‹†—ƒŽ•ǡˆ‘” •›’–‘ƒ–‹ ǡƒ•›’–‘ƒ–‹ ƒ† ‘–”‘Žˆ‹„”‘„Žƒ•–•ǡ”‡•’‡ –‹˜‡Ž›ǤŠ‡ ‘–”‘Žˆ‹„”‘„Žƒ•–•™‡”‡ͳǡ͵ǡͶǡ ͷƒ†͸Ǥȋ—’’Ž‡‡–ƒ”›ƒ„Ž‡ͳȌǤȚǣ•‹‰‹ˆ‹ ƒ–†‹ˆˆ‡”‡ ‡ ‘’ƒ”‡†–‘ˆ‹„”‘„Žƒ•–•ȋ‹ Ž—†‹‰ –ͳͲȌǡ͓ǣ•‹‰‹ˆ‹ ƒ–†‹ˆˆ‡”‡ ‡ ‘’ƒ”‡†–‘ˆ‹„”‘„Žƒ•–•ȋ‡š Ž—†‹‰–ͳͲȌǡ͎ǣ•‹‰‹ˆ‹ ƒ–†‹ˆˆ‡”‡ ‡ ‘’ƒ”‡† –‘ –ͳͲǡ șǣ •‹‰‹ˆ‹ ƒ– †‹ˆˆ‡”‡ ‡ ‘’ƒ”‡† –‘ •›’–‘ƒ–‹   ˆ‹„”‘„Žƒ•–•ǡ Șǣ •‹‰‹ˆ‹ ƒ– †‹ˆˆ‡”‡ ‡ ‘’ƒ”‡†–‘ƒ•›’–‘ƒ–‹ ˆ‹„”‘„Žƒ•–•ǡƬǣ‹‰‹ˆ‹ ƒ– ‘’ƒ”‡†–‘–Š‡‘–Š‡”ˆ‹„”‘„Žƒ•–•Ǥ ȗǣ•‹‰‹ˆ‹ ƒ–†‹ˆˆ‡”‡ ‡ ‘’ƒ”‡†–‘ ‘–”‘Žˆ‹„”‘„Žƒ•–•Ǥ‡•›„‘Ž†‡‘–‡•’δͲǤͲͷǡ–™‘•›„‘Ž•†‡‘–‡• ’δͲǤͲͳǤ.  ‘•‡“—‡–Ž›ǡ„‘–Š•›’–‘ƒ–‹ ƒ†ƒ•›’–‘ƒ–‹ ‰”‘—’•Šƒ†•‹‰‹ˆ‹ ƒ–Ž›Š‹‰Š‡” ͸ǦͳͲǦƒ ›ŽǦ ƒ”‹–‹‡ ‘ ‡–”ƒ–‹‘•‹–Š‡‡†‹—–Šƒ–Š‡ ‘–”‘Ž‰”‘—’ȋ—’’Ž‡‡–ƒ”› ‹‰—”‡ ʹǦ ȌǤ

(21)  ƒ††‹–‹‘ǡ –Š‡ ‘–”‘Ž ˆ‹„”‘„Žƒ•–• ‘•—‡† ͸ǦͳͲǦƒ ›ŽǦ ƒ”‹–‹‡•ǡ ™Š‹Ž‡ ˆ‹„”‘„Žƒ•–•†‹†‘–ȋ—’’Ž‡‡–ƒ”›ˆ‹‰—”‡͵ƒ† ‹‰—”‡ʹǦȌǤ ʹͳͳ.

(22) Chapter 6. ' medium C6 (P mol/g protein). B. ‚‚. 0.6. 6. 0.4 0.2. ††ÁÁ‚‚–– 0.0 Sympt. Asympt (ex. Pt10) Pt10. Ctrl. -0.2. ' medium C8 (Pmol/g protein). A. C. Sympt. Asympt (ex. Pt10) Pt10. Sympt. Asympt (ex. Pt10) Pt10. Ctrl. 0.3. 0.2 0.1. ††ÁÁ‚–– 0.0 Sympt. Asympt (ex. Pt10) Pt10. Ctrl. -0.1. ' medium C10 (Pmol/g protein). ' medium C10:1 (Pmol/g protein). ††ÁÁ‚–– 0. D 0.3. -0.2. 0.2 0.1. ††Á–– 0.0 Ctrl. -0.1. E. F 0.20. 0.4 0.3 0.2 0.1. ††ÁÁ‚‚––. intracellular C8 (nmol/g protein). 0.5. intracellular C6 (nmol/g protein). 2. -2. -0.4. 0.0. 0.15 0.10. ††ÁÁ‚‚––. 0.05 0.00. Sympt. Asympt (ex. Pt10) Pt10. Ctrl. G. Sympt. Asympt (ex. Pt10) Pt10. Ctrl. Sympt. Asympt (ex. Pt10) Pt10. Ctrl. H 0.04. intracellular C10 (nmol/g protein). 0.08. intracellular C10:1 (nmol/g protein). 4. 0.06 0.04 0.02 0.00. 0.03 0.02 0.01 0.00. Sympt. Asympt (ex. Pt10) Pt10. Ctrl. . Figure 2Ǥ͸ǦͳͲǦƒ ›ŽǦ ƒ”‹–‹‡•‡ ”‡–‹‘‹–‘‹–Š‡‡†‹—ȋǦȌƒ†‹–”ƒ ‡ŽŽ—Žƒ”͸ǦͳͲǦƒ ›ŽǦ ƒ”‹–‹‡ ‘ ‡–”ƒ–‹‘• ȋǦ ȌǤ ‡ ”‡–‹‘ ‘ˆ ƒ ›ŽǦ ƒ”‹–‹‡• ‹• •Š‘™ ƒ• –Š‡ †‹ˆˆ‡”‡ ‡ ‹ ‘ ‡–”ƒ–‹‘ ‘ˆ ƒ ›ŽǦ ƒ”‹–‹‡• ‹ –Š‡ ‡†‹— „‡ˆ‘”‡ ƒ† ƒˆ–‡” —Ž–—”‡ ‘ˆ ˆ‹„”‘„Žƒ•–•ǡ ‘”ƒŽ‹œ‡† ˆ‘” –Š‡ ‡ŽŽ—Žƒ” ’”‘–‡‹ ‘ ‡–”ƒ–‹‘ǤŠ‡”‡•—Ž–•’‡”–ƒ‹–‘–Š‡˜ƒŽ‹†ƒ–‹‘’Šƒ•‡‘ˆ–Š‡•–—†›Ǥαͳˆ‘”ƒ–‹‡–

(23) ͓͹ƒ† ‘–”‘Ž ˆ‹„”‘„Žƒ•–͓ͷǡαʹˆ‘”ƒ–‹‡–

(24) ͓ͳǦ͸ǡͺǦͻƒ† ‘–”‘Žˆ‹„”‘„Žƒ•–•͓͵ǡͶǡ͸ƒ†α͵ˆ‘”ƒ–‹‡–

(25) ͓ͳͲƒ† ‘–”‘Žˆ‹„”‘„Žƒ•–͓͸ǤǤ‹‰‹ˆ‹ ƒ ‡˜‹•—ƒŽ‹œƒ–‹‘‹•ƒ ‘”†‹‰–‘ ‹‰—”‡ͳǤ. 

(26) Ž‹‡™‹–Š–Š‡‡š’Ž‘”ƒ–‹˜‡•–—†›ǡ–Š‡ˆ‹„”‘„Žƒ•–•‘ˆ–ͳͲ•‡ ”‡–‡†•‹‰‹ˆ‹ ƒ–Ž›‘”‡͸ǦͳͲǦ ƒ ›ŽǦ ƒ”‹–‹‡–Šƒ–Š‡ˆ‹„”‘„Žƒ•–•‘ˆƒŽŽ‘–Š‡”‹†‹˜‹†—ƒŽ• ‘„‹‡†ȋ—’’Ž‡‡–ƒ”› ˆ‹‰—”‡͵ƒ† ‹‰—”‡ʹǦȌǤ ‹ƒŽŽ›ǡ–Š‡ˆ‹„”‘„Žƒ•–•‘ˆ•›’–‘ƒ–‹ ƒ†ƒ•›’–‘ƒ–‹  ‹†‹˜‹†—ƒŽ•Šƒ†•‹‰‹ˆ‹ ƒ–Ž›Š‹‰Š‡”‹–”ƒ ‡ŽŽ—Žƒ”͸Ǧƒ†ͺǦǡ„—–•‹‹Žƒ”ͳͲǣͳǦƒ†ͳͲǦƒ ›ŽǦ ƒ”‹–‹‡ ‘ ‡–”ƒ–‹‘• ‘’ƒ”‡† –‘ ‘–”‘Ž ˆ‹„”‘„Žƒ•–• ȋ—’’Ž‡‡–ƒ”› ˆ‹‰—”‡ Ͷ ƒ† ‹‰—”‡ʹǦ ȌǤŠ‡”‡™ƒ•‘†‹ˆˆ‡”‡ ‡„‡–™‡‡–Š‡‹–”ƒ ‡ŽŽ—Žƒ”ƒ ›ŽǦ ƒ”‹–‹‡ ‘ ‡–”ƒ–‹‘• ‘ˆ–Š‡ˆ‹„”‘„Žƒ•–•‘ˆ–ͳͲƒ†–Šƒ–‘ˆ‘–Š‡”•›’–‘ƒ–‹ ‘”ƒ•›’–‘ƒ–‹ ‹†‹˜‹†—ƒŽ•. ʹͳʹ.

(27) In vitro metabolic adaptations that explain symptomatology in MCADD individuals ȋ—’’Ž‡‡–ƒ”›ˆ‹‰—”‡Ͷƒ† ‹‰—”‡ʹǦ ȌǤ˜‡”ƒŽŽǡ–Š‡•‡”‡•—Ž–••—‰‰‡•––Šƒ– ‡ŽŽ•‘ˆ–ͳͲǡ –Š‡‡Ž†‡•–ƒ•›’–‘ƒ–‹ ‹†‹˜‹†—ƒŽǡ•‡ ”‡–‡‘”‡͸ǦͳͲǦƒ ›ŽǦ ƒ”‹–‹‡•Ǥ . Targeted proteomics Š‡‡ƒ•—”‡†‹–‘ Š‘†”‹ƒŽ’”‘–‡‹• ‘’”‹•‡‡œ›‡•‹˜‘Ž˜‡†‹ ƒ† › Ž‡ǡ ‘”‡ •—„—‹–• ‘ˆ ‡ƒ Š ‘ˆ –Š‡ ”‡•’‹”ƒ–‘”›Ǧ Šƒ‹ ‘’Ž‡š‡•ǡ ƒ† –”ƒ•’‘”–‡”• ”‡“—‹”‡† –‘ –”ƒ•’‘”–‡–ƒ„‘Ž‹–‡•‹–‘ƒ†‘—–‘ˆ–Š‡‹–‘ Š‘†”‹ƒȏʹͶȐǤ

(28) ƒŽŽ ‘–”‘Ž•ƒ’Ž‡•™ƒ• †‡–‡ –‡†ǡ ™Š‹Ž‡ ‹– ™ƒ• Ž‘™ ‹ –Š‡ ’ƒ–‹‡– •ƒ’Ž‡• ȋ ‹‰—”‡ ͵ȌǤ –Š‡”™‹•‡ –Š‡”‡ ™‡”‡ ‘ •‹‰‹ˆ‹ ƒ–†‹ˆˆ‡”‡ ‡•„‡–™‡‡ƒ† ‘–”‘Žˆ‹„”‘„Žƒ•–•‹–Š‡‡š’Ž‘”ƒ–‹˜‡’Šƒ•‡‘ˆ–Š‡ •–—†›Ǥ B. MCAD (fmol/Pg). 1.5. §§. 1.0. 0.5 l.o.d. 0.0. 6. &&. 15 10 5 0. Sympt. Asympt (ex.Pt10). Pt10. Ctrl. C. Sympt. Asympt (ex.Pt10). Pt10. Ctrl. D &&. 0.6 0.4 0.2 0.0 Sympt. Asympt (ex.Pt10). Pt10. &&. 0.6. ETFQO (fmol/Pg). 0.8. ANT1 (fmol/P g). 20. Superoxide dismutase (fmol/Pg). A. Ctrl. 0.4 l.o.d. 0.2. 0.0 Sympt. Asympt (ex.Pt10). Pt10. Ctrl. E &&. SCAD (fmol/ P g). 0.5 0.4 l.o.d. 0.3 0.2 0.1 0.0 Sympt. Asympt (ex.Pt10). Pt10. Ctrl. . Figure 3Ǥ‡Ž‡ –‹‘‘ˆ’”‘–‡‹•‘ˆ–Š‡‡š’Ž‘”ƒ–‘”›’Šƒ•‡‘ˆ–Š‡•–—†›Ǥ‡’‹ –‡†ƒ”‡–Š‡’”‘–‡‹•™Š‡”‡–ͳͲ ˆ‹„”‘„Žƒ•–••Š‘™‡†–Š‡Žƒ”‰‡•–†‹ˆˆ‡”‡ ‡‹ ‘ ‡–”ƒ–‹‘ ‘’ƒ”‡†–‘–Š‡ˆ‹„”‘„Žƒ•–•‘ˆ–Š‡‘–Š‡” ƒ† ‘–”‘Ž‹†‹˜‹†—ƒŽ•ƒ†Ǥ‹‰‹ˆ‹ ƒ ‡˜‹•—ƒŽ‹œƒ–‹‘‹•ƒ ‘”†‹‰–‘ ‹‰—”‡ͳǤ. 

(29) –Š‡‡š’Ž‘”ƒ–‘”›’Šƒ•‡‘ˆ–Š‡•–—†›ǡ‹–ƒ’’‡ƒ”‡†–Šƒ––Š‡’”‘–‡‘‡‘ˆ–Š‡ˆ‹„”‘„Žƒ•–•‘ˆ–ͳͲ ™ƒ•“—‹–‡†‹ˆˆ‡”‡–ǡ•Š‘™‹‰Š‹‰Š‡”Ž‡˜‡Ž•‘ˆ•‡˜‡”ƒŽ’”‘–‡‹•–Šƒ–Š‡‘–Š‡”ˆ‹„”‘„Žƒ•–•ȋ–‘’ ͵ƒ†•Š‘™‹ ‹‰—”‡͵ǦȌǤŠ‡’”‘–‡‹ ‘ ‡–”ƒ–‹‘•–Šƒ–™‡”‡‘•–‡Ž‡˜ƒ–‡†‹ –ͳͲ ‡ŽŽ• ‘’ƒ”‡† –‘ –Š‡ ‘–Š‡” ˆ‹„”‘„Žƒ•–•ǡ ™‡”‡ –Š‘•‡ ‘ˆ ‹–‘ Š‘†”‹ƒŽ •—’‡”‘š‹†‡ †‹•—–ƒ•‡ʹȋ ‘†‡†„›ʹȌǡȀ–”ƒ•Ž‘ ƒ•‡ͳȋͳǡ ‘†‡†„›ʹͷͶȌƒ†  †‡Š›†”‘‰‡ƒ•‡ȋ Ǧ—„‹“—‹‘‡‘š‹†‘”‡†— –ƒ•‡ǡ ǡ ‘†‡†„›  Ȍȋ ‹‰—”‡͵ǦȌǤ

(30)  ƒ††‹–‹‘ǡ–ͳͲ•Š‘™‡†Š‹‰Š‡”Ž‡˜‡Ž•‘ˆ ‘’ƒ”‡†–‘–Š‡‘–Š‡”ˆ‹„”‘„Žƒ•–•ȋ ‹‰—”‡͵ȌǤ. ʹͳ͵.

(31) Chapter 6

(32) –Š‡˜ƒŽ‹†ƒ–‹‘’Šƒ•‡‘ˆ–Š‡•–—†›™‡ ‘ˆ‹”‡†–Š‡Ž‘™ ‘ ‡–”ƒ–‹‘•‹ ‘’ƒ”‡†–‘ ‘–”‘Žˆ‹„”‘„Žƒ•–•ȋ ‹‰—”‡Ͷƒ†—’’Ž‡‡–ƒ”› ‹‰—”‡ͷȌǤŠ‡ˆ‹„”‘„Žƒ•–• ‘ˆ –Š‡  ‹†‹˜‹†—ƒŽ• •Š‘™‡† •Ž‹‰Š– „—– •‹‰‹ˆ‹ ƒ–Ž› Š‹‰Š‡” Ž‡˜‡Ž• ‘ˆ –Š‡ ƒ–‹‘š‹†ƒ– ’”‘–‡‹‡”‘š‹”‡†‘š‹Ǧ͸ȋ ‘†‡†„›”†š͸Ȍȋ ‹‰—”‡Ͷƒ†—’’Ž‡‡–ƒ”› ‹‰—”‡ͷȌǤŠ‹•‹• ‹Ž‹‡™‹–Š’”‘–‡‹‘š‹†ƒ–‹˜‡†ƒƒ‰‡ƒ†ƒŽ–‡”‡†ƒ–‹‘š‹†ƒ–†‡ˆ‡•‡‘„•‡”˜‡†‹ ‹†‹˜‹†—ƒŽ• ȏʹͻȐǤ

(33)  ƒ††‹–‹‘ǡ –Š‡ ˆ‹„”‘„Žƒ•–• –ͳͲ †‹† ‘– •Š‘™ •‹‰‹ˆ‹ ƒ– †‹ˆˆ‡”‡ ‡• ‹ ’”‘–‡‘‡ ‘’ƒ”‡†–‘ƒ› ‘ˆ –Š‡ ‰”‘—’•ǡ ‡š ‡’– ˆ‘” •‹‰‹ˆ‹ ƒ–Ž› Š‹‰Š‡” ‘ ‡–”ƒ–‹‘• ‘ˆ ‹ˆ‹„”‘„Žƒ•–‘ˆ–ͳͲ ‘’ƒ”‡†–‘–Š‡‘–Š‡”ƒ•›’–‘ƒ–‹ ˆ‹„”‘„Žƒ•–•ȋ ‹‰—”‡Ͷƒ† —’’Ž‡‡–ƒ”› ‹‰—”‡ͷȌǤ B §§. MCAD (fmol/Pg). 4 3 2 1 l.o.d. 0 Sympt. Asympt (ex.Pt10). Pt10. Ctrl. Peroxiredoxin-6 (fmol/Pg). A. 20 15. §§ 10 5 0 Sympt. Asympt (ex.Pt10). Pt10. Ctrl. C ‚. SCAD (fmol/Pg). 0.5 0.4 0.3 0.2 0.1 0.0 Sympt. Asympt (ex.Pt10). Pt10. Ctrl. Figure 4Ǥ ‡Ž‡ –‹‘ ‘ˆ ’”‘–‡‹• ‘ˆ –Š‡ ˜ƒŽ‹†ƒ–‹‘ ’ƒ”– ‘ˆ –Š‡ •–—†›Ǥ αʹ ˆ‘” ƒ–‹‡–

(34)  ͓ͳǦͻ ƒ† ‘–”‘Ž ˆ‹„”‘„Žƒ•–• ͓͵ǡͶǡ͸Ǥ ƒ† ͸Ǥ ƒ† α͵ ˆ‘” ƒ–‹‡–

(35)  ͓ͳͲ ‹‰‹ˆ‹ ƒ ‡ ˜‹•—ƒŽ‹œƒ–‹‘ ‹• ƒ ‘”†‹‰ –‘ ‹‰—”‡ͳǤ . Discussion

(36)  –Š‹• •–—†› ™‡ ‹˜‡•–‹‰ƒ–‡† ™Š‡–Š‡” –Š‡”‡ ƒ”‡ ‡–ƒ„‘Ž‹  ƒ†ƒ’–ƒ–‹‘• ‹ —Ž–—”‡† •‹ ˆ‹„”‘„Žƒ•–• ‘ˆ  ‹†‹˜‹†—ƒŽ• –Šƒ– •‡’ƒ”ƒ–‡ •›’–‘ƒ–‹  ˆ”‘ ƒ•›’–‘ƒ–‹   ‹†‹˜‹†—ƒŽ•Ǥ ‡ †‹† ‘– ‘„•‡”˜‡ ‘•‹•–‡– †‹ˆˆ‡”‡ ‡• ‹ ƒ ›ŽǦ ƒ”‹–‹‡ ‘ ‡–”ƒ–‹‘• „‡–™‡‡ˆ‹„”‘„Žƒ•–‘ˆ•›’–‘ƒ–‹ ƒ†ƒ•›’–‘ƒ–‹ ‹†‹˜‹†—ƒŽ•ǡ‡š ‡’–ˆ‘”–ͳͲǡ –Š‡ ‡Ž†‡•–  ‹†‹˜‹†—ƒŽ ™‹–Š ƒ Ž‹ˆ‡Ž‘‰ ƒ„•‡ ‡ ‘ˆ •›’–‘•Ǥ

(37) – ƒ’’‡ƒ”•ǡ ˆ”‘ „‘–Š ’Šƒ•‡•‘ˆ–Š‡•–—†›–Šƒ–ˆ‹„”‘„Žƒ•–‘ˆ–ͳͲ•‡ ”‡–‡‹–”ƒ ‡ŽŽ—Žƒ”ƒ ——Žƒ–‡†ƒ ›ŽǦ ƒ”‹–‹‡• ‘”‡ ‡ˆˆ‹ ‹‡–Ž›Ǥ ‡˜‡”–Š‡Ž‡••ǡ –Š‡ ‹–”ƒ ‡ŽŽ—Žƒ” ‘ ‡–”ƒ–‹‘• ‘ˆ ƒ ›ŽǦ ƒ”‹–‹‡• ”‡ƒ‹‡† Š‹‰Š ‘’ƒ”‡†–‘ ‘–”‘Žˆ‹„”‘„Žƒ•–•ǤŠ‡“—‡•–‹‘ƒ”‹•‡•ǡ™Šƒ– ƒ—•‡•–Š‡Š‹‰Š‡””ƒ–‡‘ˆƒ ›ŽǦ ƒ”‹–‹‡ •‡ ”‡–‹‘Ǥ

(38) – ‘—Ž† „‡ ƒ ”‡•—Ž– ‘ˆ Š‹‰Š‡” ”ƒ–‡• ‘ˆ   ™‹–Š ‘ ‘‹–ƒ– ƒ ›ŽǦ ƒ”‹–‹‡’”‘†— –‹‘‹ˆ‹„”‘„Žƒ•–•‘ˆ–ͳͲ ‘’ƒ”‡†–‘–Š‡ˆ‹„”‘„Žƒ•–•‘ˆ–Š‡‘–Š‡” ‹†‹˜‹†—ƒŽ•ǤŽ–‡”ƒ–‹˜‡Ž›ǡ–Š‡’”‘–‡‹•‹˜‘Ž˜‡†‹–Š‡•‡ ”‡–‹‘‘ˆƒ ›ŽǦ ƒ”‹–‹‡• ‘—Ž†„‡ —’”‡‰—Žƒ–‡† ‹ –ͳͲǤ Š‹• ™ƒ• ‘– –‡•–‡†ǡ •‹ ‡ –Š‡ –ƒ”‰‡–‡† ‡–Š‘† †‹† ‘– ‹ Ž—†‡ ƒ› –”ƒ•’‘”–‡”•‘—–•‹†‡–Š‡‹–‘ Š‘†”‹ƒǤ”‘–‡‘‹ ••Š‘™‡†Šƒ”†Ž›ƒ›ƒ†ƒ’–ƒ–‹‘•‹ ‡ŽŽ• ‘’ƒ”‡†–‘ ‘–”‘Ž•ǤŠ‹•‹• ‘•‹•–‡–™‹–Š‡ƒ”Ž‹‡”ˆ‹†‹‰•‹Ǧ‘—•‡Ž‹˜‡” ȋChapter 3ƒ†ȏʹͶȐȌǤ‡‘„•‡”˜‡†ƒ†ƒ’–ƒ–‹‘•‹–Š‡’”‘–‡‘‡‘ˆ–ͳͲǡ„—––Š‡›™‡”‡Ž‡•• ʹͳͶ.

(39) In vitro metabolic adaptations that explain symptomatology in MCADD individuals ’”‘‘— ‡†‹–Š‡˜ƒŽ‹†ƒ–‹‘’Šƒ•‡–Šƒ‹–Š‡‡š’Ž‘”ƒ–‘”›’Šƒ•‡‘ˆ–Š‡•–—†›ǤŠ‡”‡ˆ‘”‡ǡ™‡ ‘—Ž† ‘– —ƒ„‹‰—‘—•Ž› ‡•–ƒ„Ž‹•Š ‹ˆ –Š‡› ‘–”‹„—–‡ –‘ –Š‡ Žƒ  ‘ˆ •›’–‘• ‹ –ͳͲǤ ’”‡‰—Žƒ–‹‘‘ˆǡƒ• ‘•‹•–‡–Ž›‘„•‡”˜‡†‹–Š‡‡š’Ž‘”ƒ–‘”›ƒ†–Š‡˜ƒŽ‹†ƒ–‹‘’Šƒ•‡ǡ ‘—Ž†„‡ƒˆ— –‹‘ƒŽƒ†ƒ’–ƒ–‹‘ǤŠ‹•‡œ›‡ ƒ–ƒŽ›œ‡•–Š‡•ƒ‡”‡ƒ –‹‘ƒ•ǡ™‹–Šƒ ‘˜‡”Žƒ’’‹‰•‡–‘ˆ•—„•–”ƒ–‡•ǡƒ† ƒ–Š‡”‡ˆ‘”‡’ƒ”–Ž› ‘’‡•ƒ–‡ˆ‘”–Š‡Ž‘••‘ˆǤ †‡”–Š‡ —Ž–—”‡ ‘†‹–‹‘•–‡•–‡†‹–Š‹••–—†›ǡ™‹–Š͸Ǥͳ‘ˆ‰Ž— ‘•‡ǡ–Š‡ ‡ŽŽ•ƒ›‘– †‡’‡† •–”‘‰Ž› ‘ ˆƒ––›Ǧƒ ‹† ‘š‹†ƒ–‹‘Ǥ ‘ ‡Ž‹ ‹– Žƒ”‰‡” ƒ† ‘•‹•–‡– ƒ†ƒ’–ƒ–‹‘•ǡ ‘”‡ •–”‹‰‡– ‘†‹–‹‘• ƒ› „‡ ”‡“—‹”‡†ǡ •— Š ƒ• Ž‘™‡” ‰Ž— ‘•‡ Ž‡˜‡Ž• ƒ† Š‹‰Š‡” ˆƒ––›Ǧƒ ‹† ‘ ‡–”ƒ–‹‘•Ǥ . Materials and Methods Patients and controls ”‹ƒ”› Š—ƒ •‹ ˆ‹„”‘„Žƒ•–• ‘”‹‰‹ƒ–‡† ˆ”‘ ’ƒ–‹‡–• Š‘‘œ›‰‘—• ˆ‘” –Š‡ Ǥͻͺͷε  ACADM —–ƒ–‹‘ǡ ™Š‘ ™‡”‡ Ž‹‹ ƒŽŽ› ƒ• ‡”–ƒ‹‡† „‡ˆ‘”‡ ʹͲͲ͹ ™‡”‡ •‡Ž‡ –‡†Ǥ ›’–‘ƒ–‘Ž‘‰› ™ƒ• ƒ–‡‰‘”‹œ‡†ƒ ‘”†‹‰–‘ȏͳͲȐǤ

(40) •Š‘”–ǡ–Š‡•›’–‘ƒ–‹ ’Š‡‘–›’‡™ƒ•†‡ˆ‹‡†„›–Š‡‘ —””‡ ‡‘ˆ ƒ–Ž‡ƒ•–‘‡ƒ —–‡‡–ƒ„‘Ž‹  ”‹•‹•™‹–Š†‘ —‡–‡†Š›’‘‰Ž› ‡‹ƒȀ ‘ƒȀ‡ ‡’ŠƒŽ‘’ƒ–Š›‘”•‡‹œ—”‡•Ǥ ‘–”‘Ž ˆ‹„”‘„Žƒ•–• ™‡”‡ ˆ”‘ ’ƒ–‹‡–• ™‹–Š‘—– ƒ †‘ —‡–‡† ‡–ƒ„‘Ž‹  †‹•‡ƒ•‡ ™‡”‡ —•‡† ȋ—’’Ž‡‡–ƒ”›–ƒ„Ž‡ͳȌǤŠ‡ˆ‹„”‘„Žƒ•–•™‡”‡‘„–ƒ‹‡†ˆ”‘–Š‡‡’ƒ”–‡–‘ˆŽ‹‹ ƒŽ ‡‡–‹ •‘ˆ –Š‡‹˜‡”•‹–›‡†‹ ƒŽ‡–‡” ”‘‹‰‡Ǥ Š‡‡†‹ ƒŽ –Š‹ ƒŽ‘‹––‡‡‘ˆ–Š‡‹˜‡”•‹–›‡†‹ ƒŽ ‡–‡” ”‘‹‰‡ ‘ˆ‹”‡† –Šƒ– –Š‡ ‡†‹ ƒŽ ‡•‡ƒ” Š

(41) ˜‘Ž˜‹‰ —ƒ —„Œ‡ –•  – ™ƒ• ‘– ƒ’’Ž‹ ƒ„Ž‡ƒ†–Šƒ–‘ˆˆ‹ ‹ƒŽ•–—†›ƒ’’”‘˜ƒŽ„›–Š‡‡†‹ ƒŽ–Š‹ ƒŽ‘‹––‡‡™ƒ•‘–”‡“—‹”‡†ȋ  ʹͲͳ͸ȀͷͻͲȌǤ †‡” –Š‡•‡ ‹” —•–ƒ ‡• ƒ† ™Š‡ †ƒ–ƒ ƒ”‡ —•‡† ƒ‘›‘—•Ž›ǡ ‹†‹˜‹†—ƒŽ •—„Œ‡ –̵• ‹ˆ‘”‡† ‘•‡–‹•‘–”‡“—‹”‡†ǡƒ•’‡”‹•–‹–—–‹‘ƒŽƒ†ƒ–‹‘ƒŽ”—Ž‡•Ǥȋ ʹͲͳ͸ȀͷͻͲȌǤ . Cell culture and harvest Š‡ ‡ŽŽ• ™‡”‡ —Ž–—”‡† ƒ– ͵͹ι ‹ ͷΨ ʹ ‹ ƒ̵• ǦͳͲ —–”‹‡– ‹š ȋŠ‡”‘ ‹•Š‡”  ‹‡–‹ˆ‹ Ȍǡ •—’’Ž‡‡–‡† ™‹–Š ͳͲΨ ˆ‡–ƒŽ ƒŽˆ •‡”— ȋ ǡ ‹„ ‘Ȍ ƒ† ͳΨ ’‡‹ ‹ŽŽ‹Ȁ•–”‡’–‘› ‹ ȋ‡–”‡’ǡ. ‹„ ‘ȌǤ ‘” –Š‡ ‡š’‡”‹‡–•ǡ –Š‡ —Ž–—”‡ ‡†‹— ™ƒ• •—’’Ž‡‡–‡† ™‹–Š ͶͲͲ Ɋ Ǧ ƒ”‹–‹‡ ȋ‡š’‡”‹‡–ƒŽ‡†‹—ȌǤ ‘”–ƒ”‰‡–‡†’”‘–‡‘‹ •ǡ ‡ŽŽ•™‡”‡ˆ‹”•–•‡‡†‡†ƒ–̱͹ͷΨ ‘ˆŽ—‡ ›‹͹ͷ —Ž–—”‡ˆŽƒ••ȋ‘”‹‰Ž‹ˆ‡• ‹‡ ‡•ȌǤˆ–‡”ʹͶŠ‘—”•ǡ–Š‡ ‡ŽŽ•™‡”‡‹ —„ƒ–‡†‹‡š’‡”‹‡–ƒŽ‡†‹—Ǥ ˆ–‡”͹ʹŠ‘—”•ǡ–Š‡ ‡ŽŽ•™‡”‡™ƒ•Š‡†™‹–Šȋǡ ‹„ ‘Ȍǡ–”›’•‹‹œ‡†™‹–ŠͲǤʹͷΨ–”›’•‹ ȋ ‹„ ‘Ȍƒ†”‡•—•’‡†‡†‹ˆ”‡•Š‡†‹—ǤŠ‡ ‡ŽŽ•™‡”‡•’—†‘™ȋͷǯƒ–ͳǡͲͲͲ”’Ȍƒ†™ƒ•Š‡† ™‹–ŠͳͲŽ–Š”‡‡–‹‡•Ǥ ‹ƒŽŽ›ǡ–Š‡ ‡ŽŽ•™‡”‡ƒŽ‹“—‘–‡†ȋʹȉͳͲͷǦͳȉͳͲ͸ ‡ŽŽ•’‡”ƒŽ‹“—‘–Ȍ‹ͳǤͷ ”‘–‡‹ ‘‹† –—„‡• ȋ’’‡†‘”ˆȌǡ ’‡ŽŽ‡–‡† ȋͷǯ ƒ– ͳǡͷͲͲ ”’Ȍ ƒ† •–‘”‡† ƒ– ǦͺͲι —–‹Ž ˆ—”–Š‡” ’”‘ ‡••‹‰ƒ†ƒƒŽ›•‹•Ǥ‡ŽŽ’‡ŽŽ‡–•™‡”‡Ž›•‡†‹ʹͷρ‹ ‡Ǧ ‘Ž†ǦͶͲŽ›•‹•„—ˆˆ‡”ȋͲǤͳΨǦͶͲǡͲǤͶ ƒŽǡͳͲ”‹•’ ͺǤͲƒ†ͳ’ ͺǤͲȌˆ‘”ʹͲ‹—–‡•ǡ˜‘”–‡š‡†•Š‘”–Ž›ǡ•’—‘ƒ”‘–ƒ–‘” ƒ–ͶͲ”’ƒ†Ͷιˆ‘”ͳͷ‹—–‡•ƒ†•’—ˆ‘”ͳͲ‹—–‡•ƒ–ͳʹǡͲͲͲ‰ƒ†ͶιǤʹͲρ‘ˆ–Š‡•—’‡”ƒ–ƒ– ™ƒ•–”ƒ•ˆ‡””‡†–‘ƒ•‡ ‘†‘‹†–—„‡ƒ†ͷρ™ƒ•—•‡†ˆ‘”’”‘–‡‹†‡–‡”‹ƒ–‹‘ȋ‹‡” ‡̹ ”‘–‡‹••ƒ›‹–ǡŠ‡”‘ ‹‡–‹ˆ‹ ȌǤ‡”–‹‡’‘‹–͵™‡ŽŽ•‘ˆƒ͸Ǧ™‡ŽŽ’Žƒ–‡ȋ ”‡‹‡”‹‘Ǧ‡ǡͲͷͳ͸ͲȌ ™‡”‡ •‡‡†‡† ƒ– ά͹ͷΨ ‘ˆŽ—‡ ›Ǥ ʹͶ Š‘—”• ƒˆ–‡” •‡‡†‹‰ǡ ‡ŽŽ• ™‡”‡ ‹ —„ƒ–‡† ™‹–Š ͳ Ž ‘ˆ ‡š’‡”‹‡–ƒŽ‡†‹—Ǥ ‘”–Š‡‡–ƒ„‘Ž‘‹ •ǡ•’‡–‡†‹—™ƒ• ‘ŽŽ‡ –‡†ƒ† ‡ŽŽ•™‡”‡Šƒ”˜‡•–‡† ‘‹ ‡Ǥ‡ŽŽ•ˆ”‘–™‘͸Ǧ ™‡ŽŽ•™‡”‡™ƒ•Š‡†–™‹ ‡™‹–Š‹ ‡Ǧ ‘Ž†ǡ• ”ƒ’‡†™‹–ŠͷͲͲρ‡–Šƒ‘Ž ȋǦʹͲιȌǡ ƒŽ‹“—‘–‡† ȋͳͲͲͲ ρŽ ’‡” ƒŽ‹“—‘–Ȍ ƒ† •–‘”‡† ƒ– ǦͺͲι —–‹Ž ˆ—”–Š‡” ƒƒŽ›•‹•Ǥ ‘” ’”‘–‡‹ †‡–‡”‹ƒ–‹‘ǡ ‡ŽŽ•‹‘‡™‡ŽŽ™‡”‡• ”ƒ’‡†™‹–ŠʹͲͲρ‹ ‡Ǧ ‘Ž†ǡƒŽ‹“—‘–‡†ƒ†•–‘”‡†ƒ–ǦͺͲιǤ . Targeted quantitative proteomics of mitochondrial proteins  •‡– ‘ˆ εͷͲ ‹–‘ Š‘†”‹ƒŽ ’”‘–‡‹• ‹˜‘Ž˜‡† ‹ •—„•–”ƒ–‡ –”ƒ•’‘”–ǡ ‘š‹†ƒ–‹˜‡ ’Š‘•’Š‘”›Žƒ–‹‘ ȋ Ȍǡ‹–‘ Š‘†”‹ƒŽˆƒ––›Ǧƒ ‹†ȾǦ‘š‹†ƒ–‹‘ǡ–”‹ ƒ”„‘š›Ž‹ ƒ ‹† › Ž‡ǡƒ†ƒ–‹‘š‹†ƒ–ƒ –‹˜‹–›™‡”‡. ʹͳͷ. 6.

(42) Chapter 6 “—ƒ–‹ˆ‹‡†‹–Š‡’”‡’”‘ ‡••‡† ‡ŽŽ•ƒ’Ž‡•—•‹‰‹•‘–‘’‹ ƒŽŽ›Žƒ„‡ŽŽ‡†•–ƒ†ƒ”†•ȋ ͳ͵ǦŽƒ„‡Ž‡†Ž›•‹‡• ƒ†ƒ”‰‹‹‡•Ȍ†‡”‹˜‡†ˆ”‘•›–Š‡–‹ ’”‘–‡‹ ‘ ƒ–‡‡”•ȋ ‘Ȍȋ‘Ž›—ƒ– „ ǡƒ†„„ƒ Šǡ. ‡”ƒ›Ȍƒ ‘”†‹‰–‘‘Ž–‡”•et al.ȏʹͶȐǤ . Acyl-carnitine levels  ›ŽǦ ƒ”‹–‹‡Ž‡˜‡Ž•™‡”‡‡ƒ•—”‡†ƒ ‘”†‹‰–‘‡”•et al.ȏͳʹȐǤ . Statistical analysis Š‡•‹‰‹ˆ‹ ƒ ‡‘ˆ†‹ˆˆ‡”‡ ‡•ǡ™‡”‡ƒ••‡••‡†—•‹‰’ƒ”ƒ‡–”‹ ȋ‘‡Ǧ™ƒ›ƒ†•—„•‡“—‡– ‘”—‡–ǯ•’‘•–Š‘ –‡•–Ȍ‘”‘’ƒ”ƒ‡–”‹ –‡•–•ȋ”—•ƒŽǦƒŽŽ‹•—ǯ•’‘•–Š‘ –‡•–Ȍ †‡’‡†‹‰‘–Š‡‹”†‹•–”‹„—–‹‘Ǥ–ƒ’˜ƒŽ—‡‘ˆδͲǤͲͷ–Š‡—ŽŽŠ›’‘–Š‡•‹•™ƒ• ‘•‹†‡”‡†–‘„‡ ”‡ˆ—–‡†ǤƮ–ƒ–‹•–‹ ƒŽƒƒŽ›•‡•™‡”‡’‡”ˆ‘”‡†—•‹‰ȋ˜ʹʹȌƒ†–Š‡†ƒ–ƒ™ƒ•˜‹•—ƒŽ‹œ‡†—•‹‰. ”ƒ’Šƒ†”‹••‘ˆ–™ƒ”‡ȋ ”ƒ’Šƒ†‘ˆ–™ƒ”‡

(43)  Ǥǡ˜‡”•‹‘ͷǤͲͲǡʹͲͲ͹ȌǤ. References ͳǤ ʹǤ ͵Ǥ ͶǤ ͷǤ ͸Ǥ ͹Ǥ. ͺǤ ͻǤ. ͳͲǤ ͳͳǤ ͳʹǤ. ʹͳ͸. Žƒƒ” ƒ Ǥƒ•••’‡ –”‘‡–”›‹ Ž‹‹ ƒŽ Š‡‹•–”›ǣŠ‡ ƒ•‡‘ˆ‡™„‘”• ”‡‡‹‰Ǥ Šƒ” ‹‘‡†ƒŽǤʹͲͳͶǢͳͲͳǣͳ͹ͶȂͺʹǤ ƒ’Š‡”ǡ”—‡––‹Ǧ‹‡””‹ǡ‡‡Ǥ

(44) „‘”‡””‘”•‘ˆ‡–ƒ„‘Ž‹•ǣ–Š‡ˆŽ—šˆ”‘‡†‡Ž‹ƒ–‘ ‘’Ž‡š†‹•‡ƒ•‡•Ǥƒ–‡˜ ‡‡–ǤʹͲͲ͸ —Ǣ͹ȋ͸ȌǣͶͶͻȂ͸ͲǤ ƒŽ†‘ƒ†‘ ǡ ƒŠƒ ǡ ƒŠƒ ǡ ƒŠƒ Ǥ ›•–‡• ‹‘Ž‘‰› ’’”‘ƒ Š‡• ‘™ƒ”† †‡”•–ƒ†‹‰”‹ƒ”›‹–‘ Š‘†”‹ƒŽ‹•‡ƒ•‡•Ǥ ”‘– ‡‡–ǤʹͲͳͻǢͳͲȋ ‡„”—ƒ”›ȌǣͳͻǤ ”‰ƒǡ ‘—–‡ǡŠ— ǡ Šƒ†–Ǥ‡š– ‡‡”ƒ–‹‘—Ž–‹• ƒŽ‡‹‡™‘ˆ

(45) „‘”””‘”• ‘ˆ‡–ƒ„‘Ž‹•Ǥ‡ŽŽ‡–ƒ„ǤʹͲͳ͸Ǣʹ͵ȋͳȌǣͳ͵Ȃʹ͸Ǥ ‹Ž ‡ǡ ƒƒ•ǡ ‘›ǡ‹Ž‡›ǡŠƒ’Ž‹ǡŽƒ ǡ Ž‡– Š‡” ǡ ‹ŽŽ ǡ‘‡ŠǤ—– ‘‡‘ˆ ‡‘ƒ–ƒŽ• ”‡‡‹‰ˆ‘”‡†‹—Ǧ Šƒ‹ƒ ›ŽǦ‘†‡Š›†”‘‰‡ƒ•‡†‡ˆ‹ ‹‡ ›‹—•–”ƒŽ‹ƒǣƒ ‘Š‘”– •–—†›Ǥƒ ‡–ǤʹͲͲ͹Ǣ͵͸ͻȋͻͷͷͷȌǣ͵͹ȂͶʹǤ ‘ŠŽ—ŽŽ‡” ǡ ƒ›ƒ–‡’‡ ǡ  Š—Žœ‡ ǡ ‘ˆˆƒ ǡ ‹†‡” ǡ Ž‰‡‘ŽŽ‡” Ǥ š’ƒ†‡† ‡™„‘” ”‡‡‹‰ˆ‘”

(46) „‘”””‘”•‘ˆ‡–ƒ„‘Ž‹•„›Ž‡ –”‘•’”ƒ›

(47) ‘‹œƒ–‹‘Ǧƒ†‡ƒ•• ’‡ –”‘‡–”›ǣ‡•—Ž–•ǡ—– ‘‡ǡƒ†

(48) ’Ž‹ ƒ–‹‘•Ǥ‡†‹ƒ–”‹ •ǤʹͲͲͶǢͳͳͳȋ͸Ȍǣͳ͵ͻͻȂͶͲ͸Ǥ ƒ‰‡”ǡ—‹Œ’‡”•ǡ‘• Šǡ—Ž†‡” ǡ—„‹‘Ǧ ‘œƒŽ„‘ǡ‹••‡” ǡ†‡”‹‡•ǡ‹ŽŽ‹ƒ• ǡ ƒ–‡”Šƒ ǡ ˜ƒ ’”‘•‡ ǡ  Š‹‡Ž‡ 

(49) ǡ ‡”•  Ǥ  ƒ–‹‘™‹†‡ ‡–”‘•’‡ –‹˜‡ „•‡”˜ƒ–‹‘ƒŽ –—†› ˆ ‘’—Žƒ–‹‘ ‡™„‘”  ”‡‡‹‰ ‘” ‡†‹—ǦŠƒ‹  ›ŽǦ‘ ‡Š›†”‘‰‡ƒ•‡ȋȌ‡ˆ‹ ‹‡ ›

(50) Š‡‡–Š‡”Žƒ†•Ǥ 

(51) Š‡”‹–‡–ƒ„‹•ǤʹͲͳͻ’”ʹ͵Ǣ͵ͳȋͲȌǤ  Šƒ–œ ǡ •‡ƒ—‡” Ǥ Š‡ Ž‹‹ ƒŽ ƒ‹ˆ‡•–ƒ–‹‘ ‘ˆ  †‡ˆ‹ ‹‡ ›ǣ ŠƒŽŽ‡‰‡• –‘™ƒ”†• ƒ†—Ž–Š‘‘†‹–Š‡• ”‡‡‡†’‘’—Žƒ–‹‘Ǥ 

(52) Š‡”‹–‡–ƒ„‹•ǤʹͲͳͲǢ͵͵ȋͷȌǣͷͳ͵ȂʹͲǤ ‡–Ž‡”ǡŠƒ‹ǡŽ•„‡ ‡”ǡ”‘Ž† ǡ—”–‘ǡ‘ Ž‡› ǡƒ‡”‘ǡ ‹‡” ǡ†‹   ǡ ‡””› ǡ Š‘ƒ• ǡ ‘†‰‡ ǡ ‹‰Š ǡ ƒ•Šƒ ǡ ‘ƒŽ‡› ǡ –‡„”‹†‰‡ ǡ —••‹ ǡ Š‹ŽŽ‹’•ǡ—”–‘ǡ†ƒ‘ǡŠ”‡•–Šƒǡ ‘‰ƒ•‘ ǡ™›‡”ǡ ƒ‹Ž‹‡ǡ‘‹‡ǡ ƒ‹Ž‹‡ǡ •ƒ‘ƒŠ ǡ ‘‘†‹ ǡ ƒŒƒƒ”—ƒ ǡ ƒ •‘ ǡ ƒ‘•Š ǡ ‡”‘ ǡ ‹–Š ǡ Šƒ† ǡ ‹’‹•‹ǡ ‡Ž†ƒ ǡ‡””›ǡŽ•„‡ ‡”ǡ‡–Ž‡”ǡ ‘–Ǧ‘–‰‘‡”›ǡ‡ ǡ‡ƒǡ ‘‡„‡”Žǡ ‹ƒ‰Š—‹ǡ‹•Šƒ‹ǡ‹œœ‘ǡƒ™•‘ǡ„”‘•‡ǡ‡˜›ǡ”‘ǡ ‘‰ –‘ǡǯ ‘ǡ ƒ”–ǡ”„‡ǡƒ‘•ǡ‡•Ž‹‡ǡ‘™‡”•ǡƒ”–Š‘Ž‘‡™ǡ ‘ˆˆǡ˜ƒƒŽ ƒ”ǡ ƒ•‡ ǡ”‘Ž† ǡ‘ Ž‡› ǡƒŽ•ŠǦ‘ Ž‡›ǡŠ‡ƒ†ǡ‹‘ ǡ‰‡Ž‹‰ǡ‹”†ǡ™ƒ ǡ Š™‘‡”‡” ǡ ‡”›ǡƒ”—ƒ Š‹ǡ —‡Žǡ‹Ž‹• ǡƒ˜‹•Ǧ‡’’‡ǡ–‡‹ǡ‘ƒ ǡ ƒ‡”‘ ǡ †‹   ǡ ‹‡”  ǡ —•–‹ ‡ ǡ Šƒ‹ Ǥ ʹʹͳ ‡™„‘”Ǧ• ”‡‡‡† ‡‘ƒ–‡• ™‹–Š ‡†‹—Ǧ Šƒ‹ƒ ›ŽǦ ‘‡œ›‡†‡Š›†”‘‰‡ƒ•‡†‡ˆ‹ ‹‡ ›ǣ ‹†‹‰•ˆ”‘–Š‡

(53) „‘”””‘”•‘ˆ ‡–ƒ„‘Ž‹•‘ŽŽƒ„‘”ƒ–‹˜‡Ǥ‘Ž ‡‡–‡–ƒ„ǤʹͲͳ͸ǢͳͳͻȋͳȂʹȌǣ͹ͷȂͺʹǤ ‡”• ǡ‡‹Œ‰‘—† ǡƒ–‡”Šƒ ǡ ‡”˜‡” ǡ˜ƒ†‡‡”‰ǡƒ—‡” ǡ‹– ǤŠ‡ ƒ–—”ƒŽ Š‹•–‘”› ‘ˆ ‡†‹—Ǧ Šƒ‹ ƒ ›Ž ‘ †‡Š›†”‘‰‡ƒ•‡ †‡ˆ‹ ‹‡ › ‹ –Š‡ ‡–Š‡”Žƒ†•ǣ Ž‹‹ ƒŽ’”‡•‡–ƒ–‹‘ƒ†‘—– ‘‡Ǥ ‡†‹ƒ–”ǤʹͲͲ͸ǢͳͶͺȋͷȌǤ ‡”• ǡƒ’”‘•‡ ǡƒ‡ ǡƒ‡” ‹Ž•–ǡ’ƒǡ‹– Ǥ ƒˆ‡ƒ†—•ƒˆ‡ †—”ƒ–‹‘‘ˆˆƒ•–‹‰ˆ‘” Š‹Ž†”‡™‹–Š†‡ˆ‹ ‹‡ ›Ǥ—” ‡†‹ƒ–”ǤʹͲͲ͹Ǣͳ͸͸ȋͳȌǣͷȂͳͳǤ ‡”• ǡ‘‡”ǡ••‡ǡ‘•ǡ—‹–‡” ǡƒ–‡”Šƒ ǡ‹‡œ‡Ǧ‘‹‰ǡƒ†‡”• ǡ ‘†‡‡Ž ǡ‘‡„‡” ǡ‡ƒ–‡ǡ‹– ǡ‡‹Œ‰‘—† Ǥ‡‘ƒ–ƒŽ• ”‡‡‹‰ˆ‘”‡†‹—Ǧ.

(54) In vitro metabolic adaptations that explain symptomatology in MCADD individuals. ͳ͵Ǥ ͳͶǤ. ͳͷǤ ͳ͸Ǥ ͳ͹Ǥ ͳͺǤ ͳͻǤ ʹͲǤ ʹͳǤ ʹʹǤ ʹ͵Ǥ ʹͶǤ. ʹͷǤ ʹ͸Ǥ. ʹ͹Ǥ. ʹͺǤ ʹͻǤ. Šƒ‹ ƒ ›ŽǦ‘ †‡Š›†”‘‰‡ƒ•‡ ȋȌ †‡ˆ‹ ‹‡ › ‹ Š‡ ‡–Š‡”Žƒ†•ǣ Š‡ ‹’‘”–ƒ ‡ ‘ˆ ‡œ›‡ƒƒŽ›•‹•–‘ƒ• ‡”–ƒ‹–”—‡†‡ˆ‹ ‹‡ ›Ǥ 

(55) Š‡”‹–‡–ƒ„‹•ǤʹͲͲͺǢ͵ͳȋͳȌǣͺͺȂͻ͸Ǥ ‘—™ǡ‹– ǡ‹‡œ‡Ǧ‘‹‰ǡ‘•‰”ƒƒˆǦ†‡‘‡”ǡ ‡”†‹‰ǡ‡‹Œ‰‘—†Ǧ ǡ‡”•  Ǥ

(56)  ˜‹–”‘ ƒ† ‹ ˜‹˜‘ ‘•‡“—‡ ‡• ‘ˆ ˜ƒ”‹ƒ– ‡†‹—Ǧ Šƒ‹ ƒ ›ŽǦ‘ †‡Š›†”‘‰‡ƒ•‡ ‰‡‘–›’‡•Ǥ”’Šƒ‡– ƒ”‡‹•ǤʹͲͳ͵ǢͺȋͳȌǣͶ͵Ǥ ‘—™ǡ‹– ǡ†‡”‹‡•ǡ†‡Ž‡” ǡ‘• Šǡ‹••‡” ǡ—Ž†‡” ǡ—„‹‘Ǧ ‘œƒŽ„‘ ǡŽ˜‡”•ǡ‹‡œ‡Ǧ‘‹‰ǡƒ†‡”• ǡƒ–‡”Šƒ ǡ‡‹Œ‰‘—†Ǧ ǡ‡”• Ǥ‹• •–”ƒ–‹ˆ‹ ƒ–‹‘„›”‡•‹†—ƒŽ‡œ›‡ƒ –‹˜‹–›ƒˆ–‡”‡™„‘”• ”‡‡‹‰ˆ‘”‡†‹—Ǧ Šƒ‹ƒ ›ŽǦ‘ †‡Š›”‘‰‡ƒ•‡†‡ˆ‹ ‹‡ ›ǣ†ƒ–ƒˆ”‘ƒ ‘Š‘”–•–—†›Ǥ”’Šƒ‡– ƒ”‡‹•ǤʹͲͳʹǢ͹ǣ͵ͲǤ ’‹‡‡”‘‡––‡” ǡ ƒ•–‹ ǡ ‹ŽŽ‹‰Šƒ ǡ ‘””‹• ǡ ‹Œ„—”‰ ǡ ‹Ž ‡ Ǥ —””‡– ‹••—‡• ”‡‰ƒ”†‹‰ –”‡ƒ–‡– ‘ˆ ‹–‘ Š‘†”‹ƒŽ ˆƒ––› ƒ ‹† ‘š‹†ƒ–‹‘ †‹•‘”†‡”•Ǥ 

(57) Š‡”‹– ‡–ƒ„ ‹•Ǥ ʹͲͳͲǢ͵͵ȋͷȌǣͷͷͷȂ͸ͳǤ ”‘‡–›ǡƒ•‘—”‹ƒǡ‘‡ˆ‘– ǡ‘—”–‘‹• ǡ—‹ Šƒǡƒ„‹‡”ǡ‡••ƒ›”‡Ǥ‘•– ƒ•‡• ‘ˆ‡†‹—Ǧ Šƒ‹ƒ ›ŽǦ‘†‡Š›†”‘‰‡ƒ•‡†‡ˆ‹ ‹‡ ›‡• ƒ’‡†‡–‡ –‹‘‹ ”ƒ ‡Ǥ — ‡‡–Ǥ ͳͻͻ͸Ǣͻ͹ȋ͵Ȍǣ͵͸͹ȂͺǤ ‡ŽŽ›ǡŠ‡Žƒ ǡ‰†‡ǡŽ’‡”•ǡŠƒ‰ ǡ‡ŽŽ—• ǡ ”‡‰‡”•‡ǡ‘”Žƒ†ǡ–”ƒ—•• Ǥ‘Ž‡ —Žƒ” Šƒ”ƒ –‡”‹œƒ–‹‘‘ˆ‹Š‡”‹–‡†‡†‹—Ǧ Šƒ‹ƒ ›ŽǦ‘†‡Š›†”‘‰‡ƒ•‡†‡ˆ‹ ‹‡ ›Ǥ ”‘ ƒ–Ž ƒ† ‹ǤͳͻͻͲǢͺ͹ȋʹ͵Ȍǣͻʹ͵͸ȂͶͲǤ —”ƒǡ ‘ˆƒ’ǡŠ‡ƒ† ǡƒ—†—„”ƒ› Ǧǡƒ†ƒǤ—††‡Š‹Ž†‡ƒ–Šƒ†Ʈ ‡ƒŽ–Š›ǯ ˆˆ‡ –‡† ƒ‹Ž› ‡„‡”• ‹–Š ‡†‹—ǦŠƒ‹  ›ŽǦ‘‡œ›‡  ‡Š›†”‘‰‡ƒ•‡ ‡ˆ‹ ‹‡ ›Ǥ ‡†‹ƒ–”‹ •Ǥͳͻͺ͸Ǣ͹ͺȋ͸ȌǣͳͲͷʹȂ͹Ǥ  Šƒ–œƒǤǡ•‡ƒ—‡”ǤŠ‡ Ž‹‹ ƒŽƒ‹ˆ‡•–ƒ–‹‘‘ˆ†‡ˆ‹ ‹‡ ›ǣŠƒŽŽ‡‰‡•–‘™ƒ”†• ƒ†—Ž–Š‘‘†‹–Š‡• ”‡‡‡†’‘’—Žƒ–‹‘Ǥ 

(58) Š‡”‹–‡–ƒ„‹•ǤʹͲͳͲǢ͵͵ȋͷȌǣͷͳ͵ȂʹͲǤ ‘—–‡ ǡ ‹‘Žƒ–‡ ǡ ‡–—”ƒ  Ǥǡ ƒ†‡”•  Ǥ Š‡ ‹‘ Š‡‹•–”› ƒ† Š›•‹‘Ž‘‰› ‘ˆ ‹–‘ Š‘†”‹ƒŽ ƒ––›  ‹† ȾǦš‹†ƒ–‹‘ ƒ†

(59) –• ‡‡–‹  ‹•‘”†‡”•Ǥ — ‡˜ Š›•‹‘ŽǤ ʹͲͳ͸Ǣ͹ͺȋͳȌǣʹ͵ȂͶͶǤ ƒ‰ ǡ ‘Š•‡ ǡ ‡ Š‹ ǡ  Š”‡‹„‡” ǡ ƒŽƒ•—„”ƒƒ‹ ǡ ƒ› ǡ ‹ Šƒ‡Ž ƒ”ƒ†ƒ ǡ ‘ Ž‡› Ǥ ‘’Ž‡š Šƒ‰‡• ‹ –Š‡ Ž‹˜‡” ‹–‘ Š‘†”‹ƒŽ ’”‘–‡‘‡ ‘ˆ •Š‘”– Šƒ‹ ƒ ›ŽǦ‘ †‡Š›†”‘‰‡ƒ•‡†‡ˆ‹ ‹‡–‹ ‡Ǥ‘Ž ‡‡–‡–ƒ„ǤʹͲͳͶǢͳͳʹȋͳȌǣ͵ͲȂͻǤ ‘ Šƒ ǡ ‡””‡‹”ƒǡƒ”˜ƒŽŠ‘ ǡ‹–‘”‹‘ǡƒ–ƒǡ‘’‡•ǡ ”‡‰‡”•‡ǡ‹Žƒ”‹Š‘ǡƒ†‘ Ǥ Šƒ”ƒ –‡”‹œƒ–‹‘ ‘ˆ ‹–‘ Š‘†”‹ƒŽ ’”‘–‡‘‡ ‹ ƒ •‡˜‡”‡ ƒ•‡ ‘ˆ  Ǧ †‡ˆ‹ ‹‡ ›Ǥ  ”‘–‡‘‹ •ǤʹͲͳͳǢ͹ͷȋͳȌǣʹʹͳȂͺǤ †Šƒ‰‡”Ǥǡ–‡„”‘‡ǡ‹‡Ž•‡ǡ”‘••ǡŽ•‡ ǡ ”‡‰‡”•‡ǡƒŽˆ‡Ž†– Ǥ”‘–‡‘‹  ‹˜‡•–‹‰ƒ–‹‘‘ˆ —Ž–‹˜ƒ–‡†ˆ‹„”‘„Žƒ•–•ˆ”‘’ƒ–‹‡–•™‹–Š‹–‘ Š‘†”‹ƒŽ•Š‘”–Ǧ Šƒ‹ƒ ›ŽǦ‘ †‡Š›†”‘‰‡ƒ•‡†‡ˆ‹ ‹‡ ›Ǥ‘Ž ‡‡–‡–ƒ„ǤʹͲͳͶǢͳͳͳȋ͵Ȍǣ͵͸ͲȂͺǤ ‘Ž–‡”• ǡ ‹ƒ’ƒ‹–‡ ǡ ˜ƒ —‡ ǡ ‹‡œ‡Ǧ‘‹‰ ǡ ƒ––‘ ǡ ‘”–‡  ǡ ‘”˜ƒ–‘˜‹ Š ǡ ƒ‡” ǡ ‹• Š‘ˆˆ ǡ ‡”‡–‹‡” Ǥ ”ƒ•Žƒ–‹‘ƒŽ –ƒ”‰‡–‡† ’”‘–‡‘‹ • ’”‘ˆ‹Ž‹‰ ‘ˆ ‹–‘ Š‘†”‹ƒŽ ‡‡”‰› ‡–ƒ„‘Ž‹  ’ƒ–Š™ƒ›• ‹ ‘—•‡ ƒ† Š—ƒ •ƒ’Ž‡•Ǥ —„‹–– –‘ —„ŽǤ ʹͲͳ͸Ǣ ƒ‹Œ‘•ǡ‹ŽŽ‹ƒ ǡƒƒƒŠǤ

(60) –”ƒ‹–‘ Š‘†”‹ƒŽ ‘Ž†‹‰ƒ†••‡„Ž›‘ˆ‡†‹—Ǧ Šƒ‹ ›ŽǦ ‘‡Š›†”‘‰‡ƒ•‡ȋȌǤͳͻͻͶǢʹ͸ͻȋ͸ȌǣͶͶͲͳȂͺǤ ”‘•• ǡ ‡•’‡”•‡ ǡ ‡•‡  ǡ †”‡•‡ ǡ ”‹•–‡•‡  ǡ ‹–‡” ǡ ƒ†› ǡ ”ƒ—–Ž‡ ǡ. Š‹•Žƒǡ‘Ž—†ǡ‹ ǡ ”‡‰‡”•‡Ǥˆˆ‡ –•‘ˆ–™‘—–ƒ–‹‘•†‡–‡ –‡†‹‡†‹— Šƒ‹  ›ŽǦ‘ †‡Š›†”‘‰‡ƒ•‡ ȋȌǦ†‡ˆ‹ ‹‡– ’ƒ–‹‡–• ‘ ˆ‘Ž†‹‰ǡ ‘Ž‹‰‘‡” ƒ••‡„Ž›ǡ ƒ† •–ƒ„‹Ž‹–›‘ˆ‡œ›‡Ǥ‘ŽǤʹ͹Ͳǡ ‘—”ƒŽ‘ˆ‹‘Ž‘‰‹ ƒŽŠ‡‹•–”›ǤͳͻͻͷǤ’ǤͳͲʹͺͶȂͻͲǤ ƒ ǡƒ‹‡”ǡ‡‹„ǡ ƒ•Ž„‡ ǡ‡–‡” ǡ”—‰‡”ǡ‘‡”Š‘ˆˆǡ ‡”†‹ƒ†—••‡ ǡ ƒ†‡”•  ǡ ‡”•–‹‰ ǡ —–ƒ— Ǥ Š‡ †‘ƒ‹Ǧ•’‡ ‹ˆ‹  ƒ† –‡’‡”ƒ–—”‡Ǧ†‡’‡†‡– ’”‘–‡‹ ‹•ˆ‘Ž†‹‰ ’Š‡‘–›’‡ ‘ˆ ˜ƒ”‹ƒ– ‡†‹—Ǧ Šƒ‹ ƒ ›ŽǦ‘ †‡Š›†”‘‰‡ƒ•‡Ǥ ‘ ‡Ǥ ʹͲͳͶǢͻȋͶȌǤ —‹–‡„‡‡ǡ‘‡Ž• ǡ—„—ŽŽǡ ƒ”ƒ˜ƒ‰Ž‹ƒǡŠƒŽ‡”•ǡƒ›Ž‘”Ǥ‘•‡–‡†‹— Šƒ‹ƒ ›ŽǦ‘†‡Š›†”‘‰‡ƒ•‡†‡ˆ‹ ‹‡ ›ǤͳͻͻͷǢʹͲͻȂͳͶǤ ‡”• ǡ‘—™ǡ‹„ƒ• ǡ‹ƒ ‹‹ ǡƒœ‹ǡ‡‰”‡––‘ ǡ‡• ƒǡ‡‹Œ‰‘—† ǡ ‹– ǡ ƒŒ‡” ǡ ƒ”‰ƒ• Ǥ š’‡”‹‡–ƒŽ ‡˜‹†‡ ‡ ˆ‘” ’”‘–‡‹ ‘š‹†ƒ–‹˜‡ †ƒƒ‰‡ ƒ† ƒŽ–‡”‡†ƒ–‹‘š‹†ƒ–†‡ˆ‡•‡‹’ƒ–‹‡–•™‹–Š‡†‹—Ǧ Šƒ‹ƒ ›ŽǦ‘†‡Š›†”‘‰‡ƒ•‡†‡ˆ‹ ‹‡ ›Ǥ 

(61) Š‡”‹–‡–ƒ„‹•ǤʹͲͳͶǢ͵͹ȋͷȌǣ͹ͺ͵ȂͻǤ. . ʹͳ͹. 6.

(62) Chapter 6 . ʹͳͺ. .

(63) In vitro metabolic adaptations that explain symptomatology in MCADD individuals. Supplementary Figures and tables  Supplementary table ST1: Characteristics of the individuals from which the control fibroblasts originated. ID Age (years) Indication Outcome C1 Ͷʹ —††‡†‡ƒ–Š  —–‡ ƒ”†‹ƒ †‡ƒ–Š C2 ͹ʹ —††‡†‡ƒ–Š ƒ”†‹ƒ ƒ””‡•–ǡ‡—”‘†‡‰‡‡”ƒ–‹˜‡†‹•‡ƒ•‡ C3 ͷ͸ ‹‰”ƒ‹‡ ‘‰‡‡–‹ ‘”‹‰‹‘ˆ†‹•‡ƒ•‡ C4 ͳ —††‡†‡ƒ–Š ‡’–‹ •Š‘  C5 Ͳ —††‡†‡ƒ–Š Š‡•—•ƒ–ƒ‰‘‹• C6.K ”‡•—ƒ„Ž›ʹͲǯ•  ‹‡–‹ˆ‹ ‡•‡ƒ” Š Ǧ C6.W Ͳ ‘–ƒ˜ƒ‹Žƒ„Ž‡ ‘–ƒ˜ƒ‹Žƒ„Ž‡ Ǯ‰‡ǯ ”‡ˆ‡”• –‘ –Š‡ ƒ‰‡ ƒ– ™Š‹ Š ˆ‹„”‘„Žƒ•–• ™‡”‡ –ƒ‡Ǥ Ǯ

(64) †‹ ƒ–‹‘ǯ †‡• ”‹„‡• –Š‡ ”‡ƒ•‘ ˆ‘” „‹‘’•›Ǥ Ǯ—– ‘‡ǯ†‡• ”‹„‡•†‹•‡ƒ•‡‘—– ‘‡Ǥ‘‡–ƒ„‘Ž‹ ‘”‹‰‹‘ˆ†‹•‡ƒ•‡™ƒ•‹†‡–‹ˆ‹‡†Ǥ . A. B 0.8. 0.10. 0.05. ††Á‚. medium C8 (PM). medium C6 (PM). 0.15. 0.00. 0.4 0.2. ††ÁÁ‚–. 0.0 Sympt. Asympt (ex. Pt10) Pt10. Ctrl. C. Sympt. Asympt (ex. Pt10) Pt10. Ctrl. D 0.15. #,*. 0.08. 0.10. 0.05. ††Á. 0.00. medium C10 (P M). medium C10:1 (P M). *. 0.6. 0.06 0.04 0.02. ¥ 0.00. Sympt. Asympt (ex. Pt10) Pt10. Ctrl. Sympt. Asympt (ex. Pt10) Pt10. Ctrl. . Supplementary figure S1Ǥ ‘ ‡–”ƒ–‹‘ ‘ˆ ƒ ›ŽǦ ƒ”‹–‹‡• ‹ –Š‡ ‡†‹—Ǥ Š‡ ”‡•—Ž–• ’‡”–ƒ‹ –‘ –Š‡ ‡š’Ž‘”ƒ–‘”›’Šƒ•‡‘ˆ–Š‡•–—†›Ǥ‡–ƒ‹Ž•ƒ•‹ ‹‰—”‡ͳǤ‹‰‹ˆ‹ ƒ ‡˜‹•—ƒŽ‹œƒ–‹‘‹•ƒ ‘”†‹‰–‘ ‹‰—”‡ͳǤ . ʹͳͻ. 6.

(65) Chapter 6. B. 0.15. medium C8 (P M). medium C6 (P M). A. 0.10. 0.05. Pt1 Pt5 Pt7 Pt8 Pt2 Pt3 Pt4 Pt6 Pt9 Pt10 C3. C4. Pt1 Pt5 Pt7 Pt8 Pt2 Pt3 Pt4 Pt6 Pt9 Pt10 C3. C4. C5 C6.KC6.W. Pt1 Pt5 Pt7 Pt8 Pt2 Pt3 Pt4 Pt6 Pt9 Pt10 C3. C4. C5 C6.KC6.W. D 0.08. 0.08. medium C10 (P M). medium C10:1 (P M). 0.5. C5 C6.KC6.W. C 0.06 0.04 0.02 0.00. 0.06 0.04 0.02 0.00. Pt1 Pt5 Pt7 Pt8 Pt2 Pt3 Pt4 Pt6 Pt9 Pt10 C3. C4. C5 C6.KC6.W. E. F 0.15. 1.5. 0.10. 0.05. ††Á‚––. medium C8 (PM). medium C6 (P M). 1.0. 0.0. 0.00. 0.00 Sympt. Asympt (ex. Pt10) Pt10. 1.0. 0.5. ††ÁÁ‚–– 0.0. Ctrl. G. Sympt. Asympt (ex. Pt10) Pt10. Ctrl. H 0.08. 0.06 0.04. ††Á‚–. 0.02 0.00. medium C10 (P M). 0.08. medium C10:1 (P M). 1.5. 0.06 0.04. ¥. 0.02 0.00. Sympt. Asympt (ex. Pt10) Pt10. Ctrl. Sympt. Asympt (ex. Pt10) Pt10. Ctrl. Supplementary figure S2Ǥ ‘ ‡–”ƒ–‹‘ ‘ˆ ƒ ›ŽǦ ƒ”‹–‹‡• ‹ –Š‡ ‡†‹—ǡ †‡’‹ –‡† ˆ‘” –Š‡ ˆ‹„”‘„Žƒ•–• •‡’ƒ”ƒ–‡Ž› ȋǦȌ ƒ† †‡’‹ –‡† ƒ• •›’–‘ƒ–‹ ǡ ƒ•›’–‘ƒ–‹  ƒ† ‘–”‘Ž ˆ‹„”‘„Žƒ•– ‰”‘—’• ȋǦ ȌǤ Š‡ ”‡•—Ž–• ’‡”–ƒ‹ –‘ –Š‡ ˜ƒŽ‹†ƒ–‹‘ ’Šƒ•‡ ‘ˆ –Š‡ •–—†›Ǥ ‡–ƒ‹Ž• ƒ• ‹ ‹‰—”‡ ʹǤ ‹‰‹ˆ‹ ƒ ‡ ˜‹•—ƒŽ‹œƒ–‹‘ ‹• ƒ ‘”†‹‰–‘ ‹‰—”‡ͳǤ . ʹʹͲ.

(66) In vitro metabolic adaptations that explain symptomatology in MCADD individuals. A. B 6. 0.4 0.2 0.0 Pt1 Pt5 Pt7 Pt8 Pt2 Pt3 Pt4 Pt6 Pt9 Pt10 C3. C4. C5 C6.KC6.W. -0.2. ' medium C8 (P mol/g protein). ' medium C6 (P mol/g protein). 0.6. -0.4. 2 0 Pt1 Pt5 Pt7 Pt8 Pt2 Pt3 Pt4 Pt6 Pt9 Pt10 C3. C4. C5 C6.KC6.W. Pt1 Pt5 Pt7 Pt8 Pt2 Pt3 Pt4 Pt6 Pt9 Pt10 C3. C4. C5 C6.KC6.W. -2. C. D 0.3. 0.2 0.1 0.0 Pt1 Pt5 Pt7 Pt8 Pt2 Pt3 Pt4 Pt6 Pt9 Pt10 C3. C4. C5 C6.KC6.W. -0.1. ' medium C10 (P mol/g protein). 0.3. ' medium C10:1 (P mol/g protein). 4. -0.2. 0.2. 6. 0.1 0.0 -0.1. Supplementary figure S3Ǥ ›ŽǦ ƒ”‹–‹‡•‡ ”‡–‹‘‹–‘‹–Š‡‡†‹—ǡ†‡’‹ –‡†’‡”ˆ‹„”‘„Žƒ•– ‡ŽŽŽ‹‡Ǥ Š‡”‡•—Ž–•’‡”–ƒ‹–Š‡˜ƒŽ‹†ƒ–‹‘’Šƒ•‡‘ˆ–Š‡•–—†›Ǥ‡–ƒ‹Ž•ƒ•‹ ‹‰—”‡ʹǤ .  B. 0.5. intracellular C8 (nmol/g. protein). intracellular C6 (nmol/g. protein). A. 0.4 0.3 0.2 0.1 0.0. 0.15 0.10 0.05 0.00. Pt1 Pt5 Pt7 Pt8 Pt2 Pt3 Pt4 Pt6 Pt9 Pt10 C3. C4. C5 C6.KC6.W. C. Pt1 Pt5 Pt7 Pt8 Pt2 Pt3 Pt4 Pt6 Pt9 Pt10 C3. C4. C5 C6.KC6.W. Pt1 Pt5 Pt7 Pt8 Pt2 Pt3 Pt4 Pt6 Pt9 Pt10 C3. C4. C5 C6.KC6.W. D 0.08. intracellular C10 (nmol/g. protein). intracellular C10:1 (nmol/g. protein). 0.20. 0.06 0.04 0.02 0.00 Pt1 Pt5 Pt7 Pt8 Pt2 Pt3 Pt4 Pt6 Pt9 Pt10 C3. C4. C5 C6.KC6.W. 0.04 0.03 0.02 0.01 0.00. . Supplementary figure S4Ǥ

(67) –”ƒ ‡ŽŽ—Žƒ”ƒ ›ŽǦ ƒ”‹–‹‡ ‘ ‡–”ƒ–‹‘•ǡ†‡’‹ –‡†’‡”ˆ‹„”‘„Žƒ•– ‡ŽŽŽ‹‡ǤŠ‡ ”‡•—Ž–•’‡”–ƒ‹–‘–Š‡˜ƒŽ‹†ƒ–‹‘’Šƒ•‡‘ˆ–Š‡•–—†›Ǥ‡–ƒ‹Ž•ƒ•‹ ‹‰—”‡ʹǤ   . . ʹʹͳ.

(68) Chapter 6. B. MCAD (fmol/Pg). 4 3 2 1 l.o.d. 0 Pt1 Pt5 Pt7 Pt8 Pt2 Pt3 Pt4 Pt6 Pt9 Pt10 C3. C4. C5 C6.KC6.W. Peroxiredoxin-6 (fmol/Pg). A. 20 15 10 5 0 Pt1 Pt5 Pt7 Pt8 Pt2 Pt3 Pt4 Pt6 Pt9 Pt10 C3. C4. C5 C6.KC6.W. C SCAD (fmol/Pg). 0.5 0.4 0.3 0.2 0.1 0.0 Pt1 Pt5 Pt7 Pt8 Pt2 Pt3 Pt4 Pt6 Pt9 Pt10 C3. C4. C5 C6.KC6.W. Supplementary figure S5Ǥ‡Ž‡ –‹‘‘ˆ’”‘–‡‘‹ ”‡•—Ž–•‘ˆ–Š‡˜ƒŽ‹†ƒ–‹‘’Šƒ•‡‘ˆ–Š‡•–—†›ǡ†‡’‹ –‡†’‡” ˆ‹„”‘„Žƒ•– ‡ŽŽŽ‹‡Ǥ‡–ƒ‹Ž•ƒ•‹ ‹‰—”‡ͶǤ‹‰‹ˆ‹ ƒ ‡˜‹•—ƒŽ‹œƒ–‹‘‹•ƒ ‘”†‹‰–‘ ‹‰—”‡ͳǤ. . ʹʹʹ. .

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