o:31606 Ispitivanje povezanosti promena u mitohondrijskom genomu sa fenotipskim karakteristikama pacijenata sa Leberovom hereditarnom optičkom neuropatijom doktorska disertacija Correlation of changes in the mitochondrial genome with phenotypic characteristics in patients with Leber's hereditary optic neuropathy : doctoral dissertation sr Cilj ove doktorske disertacije bila je detaljna genotipska i fenotipska karakterizacija grupepacijenata sa kliničkom slikom Leberove hereditarne optičke neuropatije (LHON) u akutnoj ilihroničnoj fazi bolesti i funkcionalna analiza mitohondrijske funkcije kod nosilaca mogućihnovih patogenih mutacija.Metodologija: U grupi od 82 pacijenata sa kliničkom slikom LHON-a urađena je genetska analiza(MLPA, sekvenciranje nove generacije mitohondrijske DNK [mtDNK], celog egzoma i genoma),kao i detaljna fenotipska karakterizacija (vidna oštrina, kolorni vid, optička koherentnatomografija [OCT] i elektrofiziologija). Karakteristike genetski različitih grupa međusobno supoređene. Kod nosilaca novih mogućih patogenih mutacija izvedeni su testovi mitohondrijskefunkcije (respirometrija visoke rezolucije Oroboros i protočna citometrija).Metodom spekularne mikroskopije, utvrđivali smo i broj endotelnih čelija rožnjače.Rezultati: Kod 17 pacijenata je identifikovan genetski uzrok LHON-a, a kod 6 pacijenatapromene u mitohondrijskoj DNK (retke mutacije) koje su mogući uzročnici pojave LHON-a.Testovi mitohondrijske funkcije su kod ovih pacijenata pokazali smanjenu aktivnost kompleksaI lanca prenosilaca elektrona, povećanu produkciju slobodnih kiseoničnih radikala (ROS) idepolarizaciju unutrašnje membrane miohondrija, što ukazuje na patogenu priroduidentifikovanih promena. Kod 2 pacijenta, čije su fenotipske karakteristike prikazane zasebno,potvrđen je autozomno recesivni LHON. Kod nekoliko pacijenata sa genetski potvrđenim LHONomdošlo je do poboljšanja vidne oštrine. Poređenjem fenotipskih karakteristika ovih pacijenatasa pacijentima bez poboljšanja utvrđena je bolja očuvanost pojedinih slojeva retine u srednjemETDRS prstenu, kao i bolje elektrofiziološke karakteristike u akutnoj i u hroničnoj fazi bolesti.Kod izvesnog broja pacijenata i pored jasnog fenotipa LHON-a nije identifikovana genetskaosnova, ali su upoređivane fenotipske karakteristike ovih pacijenata sa grupom genetskipotvrđenih LHON pacijenata radi identifikovanja kliničkih biomarkera bolesti. Utvrđeno jepostojanje bolje očuvanosti debljine unutrašnje retine kod LHON pacijenata što se stoga možesmatrati potencijalnim biomarkerom bolesti, kao i smanjen broj endotelnih ćelija rožnice.Zaključak: Rezultati prikazani u okviru ove teze pomogli su boljoj karakterizaciji pacijenata saLHON-om, kao i identifikaciji potencijalnih biomarkera bolesti. Takođe su identifikovane novepromene u mtDNK, i potvrđena je njihova potencijalna patogenost. The goal of this doctoral dissertation was to genotypically and phenotypically characterize agroup of patients with a clinical presentation of Leber’s Hereditary Optic Neuropathy (LHON)in the acute and chronic phase of the disease and perform functional analysis of themitochondrial function in carriers of the possible pathogenic novel changes in mitochondrialDNA.Methodology: Genetic analysis (MLPA, next-generation sequencing of the mtDNK, whole exome,and genome) and detailed clinical work-up (visual acuity testing, color vision, optical coherenttomography (OCT), and electrophysiology) were performed in a group of 82 patients withphenotypic characteristics of LHON. Patients were divided into groups based on the genetictesting results and their phenotypic characteristics were compared. In patients with identifiednovel mtDNK changes testing of mitochondrial function was performed. We have alsodetermined the count of endothelial cells in patients with confirmed LHON.Results: We have identified the genetic cause of LHON in 17 patients, and possible causativepathogenic mtDNK changes (rare mutations) in 6 patients. Mitochondrial function testing inthese patients showed decreased complex I-based respirometry, membrane depolarizationandincreased ROS production. These results indicate the pathogenic nature of the novel changes.Autosomal recessive LHON as a distinct entity has been identified in two patients and presentedseparately. The improvement of the visual function has been noticed in 4 LHON patients. Bycomparing the phenotypic characteristics of LHON patients with and without visual functionimprovement we have concluded that there is better preservation of the retinal layers in themiddle ETDRS ring and that these patients have better electrophysiology results both in theacute and the chronic phase in patients with improvement. A certain number of patientsremained unclarified regardless of the typical LHON phenotype. We have compared thephenotypic characteristics of these patients with a group of patients with genetically confirmedLHON in order to identify clinical biomarkers of the disease and concluded that in geneticallyconfirmed LHON patients there is a better preservation of the ETDRS center which could be anovel biomarker of the disease. We have also found a small decrease in the endothelial cell countin the cornea of LHON patients.Conclusions: The results presented in this doctoral dissertation have attributed to a bettercharacterization of LHON patients and the identification of novel biomarkers of the disease. Wehave also identified novel changes in the mtDNK whose pathogenicity has been proven by theserial of the mitochondrial function tests. Medicina - Molekularna medicina / Medicine- Molecular medicine Datum odbrane: 29.06.2023. Leberova hereditarna optička neuropatija (LHON), sekvenciranje nove generacije, mtDNK, optička koherentna tomografija (OCT), segmentacija, elektrofiziologija, oscilatorni potencijali Leber hereditary optic neuropathy (LHON), next-generation sequencing, mtDNK, optical coherent tomography (OCT), segmentation, electrophysiology, oscillatory potentials 616.8:577.2(043.3) 91552100 127246089 91552101 9319 2023-11-06T09:38:44.053Z 45 no 46 Sanja, 1982- Petrović-Pajić 28273511 2023 63 mentor Ivanka, 1968- Marković 12765287 2023 63 mentor Marko Hawlina 106597641 2023 63 član komisije Dragan, 1959 Vuković 12539239 2023 63 član komisije Sonja, 1975- Misirlić Denčić 25077607 2023 63 član komisije Tanja, 1977- Kalezić 36199271 2023 63 član komisije Jelena, 1979- Vasilijević 35344487 2023 63 član komisije Borut Peterlin 106600201 2023 str. 164-185 10436379 http://phaidrabg.bg.ac.rs/o:31606 no yes 12 2023-11-06T09:38:44.320Z 70 11 1066950 1066958 Leberova hereditarna optička neuropatija (LHON), sekvenciranje nove generacije,mtDNK, optička koherentna tomografija (OCT), segmentacija, elektrofiziologija, oscilatornipotencijali Leber hereditary optic neuropathy (LHON), next-generation sequencing, mtDNK,optical coherent tomography (OCT), segmentation, electrophysiology, oscillatory potentials 616.8:577.2(043.3) 1738 11A32 2023