Polimorfizam gena endotelne sintaze dušikova monoksida i perinatalno hipoksično-ishemično oštećenje mozga : doktorska disertacija

Cilj: Hipoksično-ishemična encefalopatija (HIE) je oštećenje mozga, koja se razvija zbog manjka kisika i poremećaja cirkulacije. Kod hipoksije i ishemije u mozgu dolazi do niza biokemijskih događaja, između ostalog i pojačana aktivnost sintaze dušikova monoksida (NOS). Aktivacija endotelne NOS (eNOS) ima neuroprotektivu ulogu. Cilj istraživanja je utvrditi povezanost mutacija polimorfizama gena za NOS3 s HIE. Ispitanici i metode: Ova studija uključila je 110 djece (69 dječaka i 41 djevojčica) s hipoksično-ishemičnim (HI) oštećenjem mozga i kontrolnu skupinu od 128 djece (60 dječaka i 68 djevojčica) rođenih u 16-godišnjem periodu na području Dalmacije – Hrvatska. Djeca s HI oštećenjem mozga ispunjavala su dijagnostičke kriterije za perinatalnu asfiksiju. Analizirali smo 6 tag SNP-ova NOS3 gena (rs3918186, rs3918188, rs1800783, rs1808593, rs3918227, rs1799983), i dva funkcionalna polimorfizma istog gena SNP rs1800779 i rs2070744. Polimorfizmi su analizirani metodom real-time PCR. Asocijacijska analiza napravljena je na razini alelske i genotipske distribucije. Rezultati: Genotipiziranjem je utvrđena je povezanost rs1808593 tag SNP-a s hipoksično-ishemičnim oštećenjem mozga (p=0.03). Analizom omjera izgleda (engl. odd ratio – OR) genotipova osam polimorfizama uočena je veća učestalost TT genotipa polimorfizma rs1808593 (-10G/T u intronu 23) u djece s hipoksično-ishemičnim oštećenjem mozga [OR 1.06 (95% CI 1.0-1.1) p=0.025]. Utvrđena je povezanost TGT haplotipa polimorfizama rs1800783 (-1474T/A), rs1800779 (-922 A/G) i rs2070744 (-786T/C) s hipoksično-ishemičnim oštećenjem mozga (p=0.001). Distribucijom genotipova ispitivanih polimorfizma s obzirom na kliničke parametre utvrđena je povezanost TT genotipa polimorfizma rs1808593 s patološkim nalazom magnetske rezonancije. Uočena je veća učestalost AA genotipa polimorfizma rs3918186 u ispitanika s urednim Apgar indeksom. Analizom povezanosti alelske distribucije svih osam ispitivanih polimorfizama nije dobivena statistički značajna povezanost s HIE. Analizom distribucije alela ispitivanih polimorfizama s obzirom na kliničke parametre utvrdili smo da je T alel SNP-a rs1808593 učestaliji u djece s niskim AS (p=0.015). Drugi polimorfizmi nisu pokazali povezanost distribucije alela i genotipova s kliničkim parametrima. Zaključak: Usprkos ograničenom broju ispitanika s HIE što umanjuje statističku snagu studije, utvrdili smo genotipsku i haplotipsku povezanost polimorfizma NOS3 gena s hipoksično-ishemičnim oštećenjem mozga.Aim: Perinatal hypoxic-ischemic encephalopathy (HIE) is characterized with impaired cerebral circulation and increased activity of nitric oxide synthase (NOS). Activation of the eNOS in endothelial cells has a neuroprotective role. Aim of this study was to test the association of NOS3 gene with HIE. Patients and Methods: The study included 110 unrelated term or preterm born children (69 boys and 41girls) with HIE and 128 term and preterm born children (60 boys and 68 girls) without any neurological problems after two years. Children with perinatal HIE fulfilled the diagnostic criteria for perinatal asphyxia. All children were admitted to the Clinical Hospital Split between 1992 and 2008. We analyzed 6 tagging SNPs within NOS3 gene (rs3918186, rs3918188, rs1800783, rs1808593, rs3918227, rs1799983), in addition to previously confirmed NOS3-associated SNP rs1800779 and rs2070744. Genotyping was conducted using real-time polymerase chain reaction (PCR). Association analyses were performed according to allelic and genotypic distribution. Results: Genotypic test detected association of rs1808593 tag SNP with HIE (p=0.03). We found that TT genotype SNP rs1808593 (-10G/T intron 23) was more common in HIE [OR 1.06 (95% CI 1.0-1.1) p=0.025]. We found that the TT genotype of SNP rs 1808593 is more frequent in children with brain damage confirmed by MRI (p=0,015). We also found that the genotype AA of SNP rs 3918186 is more frequent in children with normal Apgar score (p=0,025). We also observed rs1800783-rs1800779-rs2070744 TGT haplotype association with HIE (p=0.001). Allelic test did not observe any SNP association with HIE. We found that the T allele of SNP rs1808593 is more frequent in children with low Apgar score (p=0.015). Conclusion: Despite the limited number of HIE patients that reduced the statistical power of this study, we observed genotypic and haplotype associations of NOS3 polymorphisms with HIE

Polimorfizam gena endotelne sintaze dušikova monoksida i perinatalno hipoksično-ishemično oštećenje mozga : doktorska disertacija

Cilj: Hipoksično-ishemična encefalopatija (HIE) je oštećenje mozga, koja se razvija zbog manjka kisika i poremećaja cirkulacije. Kod hipoksije i ishemije u mozgu dolazi do niza biokemijskih događaja, između ostalog i pojačana aktivnost sintaze dušikova monoksida (NOS). Aktivacija endotelne NOS (eNOS) ima neuroprotektivu ulogu. Cilj istraživanja je utvrditi povezanost mutacija polimorfizama gena za NOS3 s HIE. Ispitanici i metode: Ova studija uključila je 110 djece (69 dječaka i 41 djevojčica) s hipoksično-ishemičnim (HI) oštećenjem mozga i kontrolnu skupinu od 128 djece (60 dječaka i 68 djevojčica) rođenih u 16-godišnjem periodu na području Dalmacije – Hrvatska. Djeca s HI oštećenjem mozga ispunjavala su dijagnostičke kriterije za perinatalnu asfiksiju. Analizirali smo 6 tag SNP-ova NOS3 gena (rs3918186, rs3918188, rs1800783, rs1808593, rs3918227, rs1799983), i dva funkcionalna polimorfizma istog gena SNP rs1800779 i rs2070744. Polimorfizmi su analizirani metodom real-time PCR. Asocijacijska analiza napravljena je na razini alelske i genotipske distribucije. Rezultati: Genotipiziranjem je utvrđena je povezanost rs1808593 tag SNP-a s hipoksično-ishemičnim oštećenjem mozga (p=0.03). Analizom omjera izgleda (engl. odd ratio – OR) genotipova osam polimorfizama uočena je veća učestalost TT genotipa polimorfizma rs1808593 (-10G/T u intronu 23) u djece s hipoksično-ishemičnim oštećenjem mozga [OR 1.06 (95% CI 1.0-1.1) p=0.025]. Utvrđena je povezanost TGT haplotipa polimorfizama rs1800783 (-1474T/A), rs1800779 (-922 A/G) i rs2070744 (-786T/C) s hipoksično-ishemičnim oštećenjem mozga (p=0.001). Distribucijom genotipova ispitivanih polimorfizma s obzirom na kliničke parametre utvrđena je povezanost TT genotipa polimorfizma rs1808593 s patološkim nalazom magnetske rezonancije. Uočena je veća učestalost AA genotipa polimorfizma rs3918186 u ispitanika s urednim Apgar indeksom. Analizom povezanosti alelske distribucije svih osam ispitivanih polimorfizama nije dobivena statistički značajna povezanost s HIE. Analizom distribucije alela ispitivanih polimorfizama s obzirom na kliničke parametre utvrdili smo da je T alel SNP-a rs1808593 učestaliji u djece s niskim AS (p=0.015). Drugi polimorfizmi nisu pokazali povezanost distribucije alela i genotipova s kliničkim parametrima. Zaključak: Usprkos ograničenom broju ispitanika s HIE što umanjuje statističku snagu studije, utvrdili smo genotipsku i haplotipsku povezanost polimorfizma NOS3 gena s hipoksično-ishemičnim oštećenjem mozga.Aim: Perinatal hypoxic-ischemic encephalopathy (HIE) is characterized with impaired cerebral circulation and increased activity of nitric oxide synthase (NOS). Activation of the eNOS in endothelial cells has a neuroprotective role. Aim of this study was to test the association of NOS3 gene with HIE. Patients and Methods: The study included 110 unrelated term or preterm born children (69 boys and 41girls) with HIE and 128 term and preterm born children (60 boys and 68 girls) without any neurological problems after two years. Children with perinatal HIE fulfilled the diagnostic criteria for perinatal asphyxia. All children were admitted to the Clinical Hospital Split between 1992 and 2008. We analyzed 6 tagging SNPs within NOS3 gene (rs3918186, rs3918188, rs1800783, rs1808593, rs3918227, rs1799983), in addition to previously confirmed NOS3-associated SNP rs1800779 and rs2070744. Genotyping was conducted using real-time polymerase chain reaction (PCR). Association analyses were performed according to allelic and genotypic distribution. Results: Genotypic test detected association of rs1808593 tag SNP with HIE (p=0.03). We found that TT genotype SNP rs1808593 (-10G/T intron 23) was more common in HIE [OR 1.06 (95% CI 1.0-1.1) p=0.025]. We found that the TT genotype of SNP rs 1808593 is more frequent in children with brain damage confirmed by MRI (p=0,015). We also found that the genotype AA of SNP rs 3918186 is more frequent in children with normal Apgar score (p=0,025). We also observed rs1800783-rs1800779-rs2070744 TGT haplotype association with HIE (p=0.001). Allelic test did not observe any SNP association with HIE. We found that the T allele of SNP rs1808593 is more frequent in children with low Apgar score (p=0.015). Conclusion: Despite the limited number of HIE patients that reduced the statistical power of this study, we observed genotypic and haplotype associations of NOS3 polymorphisms with HIE

Diagnostic, clinical and therapeutic challenges in patients with Duchenne muscular dystrophy – patient series reports

Duchenneova mišićna distrofija najčešća je mišićna distrofija koja se javlja u djece. Cilj istraživanja: Procijeniti pravodobno prepoznavanje kliničke slike, provođenje multidisciplinskog pristupa prema najnovijim smjernicama te analizirati klinički tijek u oboljelih na suvremenoj terapiji. Materijali i metode: Retrospektivnom analizom medicinske dokumentacije izdvojeni su bolesnici s distrofinopatijama. Izdvojeni su podatci o kliničkom tijeku bolesti, kliničkoj slici u trenutku postavljanja dijagnoze, metodama njene potvrde, tipovima mutacija i terapiji koju primaju. Rezultati: U istraživanje je uključeno ukupno devet ispitanika, osam s dijagnozom Duchenneove i jedan s dijagnozom Beckerove mišićne distrofije. Dob pri postavljanju dijagnoze Duchenneove mišićne distrofije bila je u rasponu između dvije i šest i pol godina, dok je kod ispitanika s Beckerovom mišićnom distrofijom postavljena u dobi od jedanaest godina. Od tipova mutacija četvorica ispitanika ima delecije, trojica duplikacije i dvojica točkaste mutacije. Samostalno pokretna su šestorica ispitanika, dok su trojica ovisna o uporabi kolica. Plućna funkcija uredna je kod sedmorice ispitanika, dok su dvojica razvila kroničnu plućnu insuficijenciju i kod njih se primjenjuju metode neinvazivne ventilacije. Na glukokortikoidnoj terapiji ukupno je šestorica ispitanika i svi primjenjuju deflazacort. Zaključak: Pacijenti s Duchenneovom mišićnom distrofijom uz osnovnu progresivnu slabost mišića, razvijaju i komplikacije drugih organskih sustava, što zahtijeva multidisciplinsku skrb. Pojava genske terapije kao uzročne terapije Duchenneove mišićne distrofije stavila je dodatni naglasak na što ranijem postavljanju dijagnoze. Time se postiže dodatno produljenje životnog vijeka te podizanje kakvoće života u bolesnika s Duchenneovom mišićnom distrofijom.The aim of the study was to assess timely recognition of clinical characteristics and implementation of a multidisciplinary approach according to the latest guidelines, and to analyse clinical course in patients on gene therapy. The research was designed as a crosssectional and retrospective study. Patients with dystrophinopathies and their contacts were singled out by retrospective analysis of medical documentation. Data on current age, age at diagnosis, type of mutations, methods of confi rming the diagnosis, clinical picture and therapy administered were extracted. Nine subjects were included in the study, eight with the diagnosis of Duchenne muscular dystrophy (DMD) and one with the diagnosis of Becker muscular dystrophy (BDM). The age at diagnosis of DMD ranged between two and six and a half years, while in the subject with BMD it was set at the age of eleven. Considering types of mutations, four patients had deletions, three of them duplications and two of them point mutations. Six subjects were mobile independently, while three subjects were dependent on wheelchair use. Pulmonary function was normal in seven subjects, while two developed chronic pulmonary insuffi ciency and used non-invasive ventilation methods. Six subjects were on glucocorticoid therapy and all were using defl azacort. In conclusion, patients with DMD, in addition to basic progressive muscle weakness, also develop complications of other organ systems, which requires multidisciplinary care. The advent of gene therapy as a causal therapy for DMD has placed additional emphasis on diagnosing the disease as early as possible. This achieves an additional prolongation of life expectancy and increase in the quality of life in patients with DMD

Duchenne Muscular Dystrophy: Clinical and Therapeutic Approach

Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD) are dystrophinopathies, a group of muscular dystrophies caused by mutations in the dystrophin gene. Duchenne muscular dystrophy is the most common muscular dystrophy that occurs in children. A mutation in the DMD gene leads to a loss of expression of the dystrophin protein, a subsarcolemmal protein that provides strength, stability, and functionality to the myofibrils. Patients with dystrophinopathies with basic progressive weakness of the musculoskeletal system develop complications of many organ systems that significantly contribute to the deterioration of the clinical condition and shorter life expectancy. Multidisciplinary care has extended the patients’ life expectancy and the development of subspecialist branches has enabled the improvement of diagnostic methods and treatment. Recently, therapeutic options in the treatment of DMD have advanced significantly, and new genetic and molecular therapies are emerging. The advent of gene therapy as a causal therapy for DMD has placed additional emphasis on diagnosing and treating the disease as early as possible. This achieves an additional prolongation of life expectancy, increases the quality of life in patients with DMD, and provides hope for patients and their families

Neurological manifestations and complications of autoimmune gastrointestinal diseases in children

Upalna bolest crijeva i celijakija autoimune su bolesti kojima su crijeva zajednički specifični ciljni organ, ali obje se smatraju sistemskim bolestima koje se mogu manifestirati i izvanintestinalnim simptomima. Cilj istraživanja: Istražiti učestalost neuroloških poremećaja sa celijakijom i upalnom bolesti crijeva u dječjoj i adolescentnoj dobi. Materijali i metode: Istraživanje je provedeno retrospektivno prikupljanjem podataka u Klinici za dječje bolesti KBC-a Split u razdoblju od 1. siječnja 2015. do 31. prosinca 2019. U istraživanje je uključeno 122-je bolesnika s dijagnozom upalne bolesti crijeva ili celijakije. Ispitanicima su analizirani podatci: spol i dob prilikom dijagnoze autoimune bolesti crijeva, prisutnost i tip neuroloških simptoma te dob u kojoj je došlo do manifestacije. Rezultati: Najviše bolesnika je bolovalo od Crohnove bolesti (35,2%), potom od ulceroznog kolitisa (30,3%) te celijakije (27,9%). Udio djece s neurološkim simptomima u skupini oboljelih od celijakije je 32,4%, ulceroznog kolitisa 16,2% i Crohnove bolesti 4,7%. Udio neuroloških simptoma u istraživanoj skupini je veći kod oboljelih od celijakije nego kod oboljelih od upalnih bolesti crijeva i postoji statistički značajna povezanost neuroloških simptoma s vrstom bolesti (P=0,005). Najčešći neurološki simptom u istraživanoj skupini je glavobolja, a ostale neurološke manifestacije su: epilepsija, cerebrovaskularna bolest, pervazivni razvojni poremećaj, zastoj u psihomotornom razvoju i tikovi. Zaključak: Upalne bolesti crijeva i celijakija kronične su bolesti kojima je potrebno dugoročno praćenje liječnika gastroenterologa, ali iako neurološke manifestacije nisu učestale, određeni dio bolesnika ih ima ili će ih razviti, stoga je važno pravodobno prepoznavanje, a može biti potrebno i dugoročno praćenje neurologa i u odrasloj dobi.The aim was to investigate the association of neurological disorders with celiac disease and infl ammatory bowel disease (Crohn’s disease, ulcerative colitis and indeterminate colitis) in children and adolescents. The research was conducted retrospectively by collecting data from available hospital documentation at Department of Paediatrics, Split University Hospital Centre, during the period from January 1, 2015 to December 31, 2019. The study included 122 patients diagnosed with infl ammatory bowel disease or celiac disease. The following data were analysed: sex and age at the time of autoimmune bowel disease diagnosis, presence and type of neurological symptoms, and age at the onset of disease manifestation. The greatest proportion of patients suff ered from Crohn’s disease (35.2%), followed by ulcerative colitis (30.3%) and celiac disease (27.9%). The proportion of children with neurological symptoms was 32.4% in patients with celiac disease, 16.2% in those with ulcerative colitis and 4.7% in those with Crohn’s disease. The proportion of children with neurological symptoms was higher in patients with celiac disease than in those with infl ammatory bowel disease, and there was a statistically signifi cant association of neurological symptoms with the type of disease (p=0.005). The most common neurological symptom in the study group was headache; other neurological manifestations were epilepsy, cerebrovascular disease, pervasive developmental disorder, psychomotor developmental delay and tics. In conclusion, infl ammatory bowel diseases and celiac disease are chronic diseases that require long-term follow-up by a gastroenterologist; however, although neurological manifestations are uncommon, some patients develop them, so timely recognition is important, and long-term follow-up by neurologists may be necessary in adulthood

Inovativne terapije redefiniraju terapijske ciljeve u multiploj sklerozi

The treatment of multiple sclerosis (MS) is becoming more complex, especially with the expanding number of available therapies for relapsing forms of MS. Greater understanding of the degenerative aspects of MS pathogenesis is redefining treatment goals and creating new treatment strategies. The existing immunomodulation drugs (disease-modifying therapies, DMTs) used in MS treatment have shown only partial benefits in controlling disease progression, primarily by reducing the inflammation component. However, new therapies for MS have been shown to be effective in delaying disease progression by protecting against brain atrophy, which is considered the most important preindicator of future patient disability. The favorable effect on reducing brain atrophy suggests the potential neuroprotective or even neuroregenerative effects of new treatments, marking progress in the treatment of MS.Liječenje multiple skleroze (MS) postaje sve složenije, naročito zbog rastućeg broja dostupnih terapija za relapsni oblik MS. Sve bolje razumijevanje degenerativnog aspekta patogeneze MS redefinira terapijske ciljeve i stvara nove terapijske strategije. Dosadašnji imunomodulacijski lijekovi (disease-modifying therapies, DMTs) u terapiji MS pokazuju samo djelomičnu korist u kontroli progresije bolesti, prvenstveno smanjujući upalnu komponentu. Međutim, nove terapije u MS pokazuju djelotvoran učinak na odgađanje progresije bolesti štiteći od atrofije mozga koja se smatra najvažnijim predskazateljem budućeg invaliditeta bolesnika. Pozitivan utjecaj na smanjenje atrofije mozga ukazuje na potencijalno neuroprotektivno ili čak neuroregenerativno djelovanje novih terapija, što predstavlja korak naprijed u liječenju MS

The Floppy Infant : Evaluation of Hypotonia

Hipotonija u novorođenčadi i dojenčadi predstavlja dijagnostički izazov za neonatologe i pedijatre, budući da je to klinički simptom koji upućuje na dobroćudna, ali i ozbiljna stanja. Diferencijalna dijagnoza neonatalne i dojenačke hipotonije jest opsežna, a metodičan pristup pomaže u lokalizaciji problema na određeni dio živčanog sustava i formuliranju diferencijalne dijagnoze. Postavljanje dijagnoze pomaže u planiranju liječenja i informiranju roditelja o prognozi. Ovaj pregledni članak predstavlja strukturirani pristup koji naglašava početnu procjenu, pregled i liječenje novorođenčeta i dojenčeta s generaliziranom hipotonijom.Hypotonia in newborns and infants represents a diagnostic challenge for neonatologists and pediatricians, since it is a clinical symptom that points to benign as well as serious conditions. The differential diagnosis of neonatal and infant hypotonia is extensive, and a methodical approach helps in localizing the problem to a specific part of the nervous system and formulating a differential diagnosis. Establishing a diagnosis helps in planning treatment and informing parents about the prognosis. This review article presents a structured approach that emphasizes the initial assessment, examination, and management of the neonate and infant with generalized hypotonia

Analysis of the C609T Polymorphism of NQO1 Gene in South Croatian Patients with Hematological Malignancies

In this study we analyzed the effect of polymorphic variation of NAD(P)H: quinone oxidoreductase1 (NQO1) gene that encode enzyme which detoxifies harmful quinines and protect hematopoietic stem cells against oxidative stress. C609T polymorphism of NQO1 gene leads to loss of enzyme activity, which may be a risk factor in the etiology of specific types of hematopoietic malignancies.We analyzed C609T polymorphism in NQO1 gene in the group of 82 patients (56 adult and 26 children) with different type of hematopoietic malignancies and 99 healthy participants (61 adult and 38 children) using PCR and the RFLP method. We confirmed that the polymorphism C609T in NQO1 gene was more frequent in the adult patients’ group with myeloid disorders, (p=0.0267) compared with adult controls.We could not confirm the association C609T polymorphism with recurrent chromosome translocations (clonal karyotype changes) neither in the adult nor in pediatric group of patients

West Syndrome with Periventricular Leukomalacia: Ten-year Clinical Study

The aim of the study was to evaluate magnetic resonance imaging (MRI) findings in infants with periventricular leukomalacia (PVL) andWest syndrome (WS) and determine the neurodevelopmental outcome in children withWest syndrome and PVL. Ultrasound and brain MRI were performed in 37 infants with recognized PVL. PVL was categorized according to De Vries, whereas West syndrome was categorized according to International League Against Epilepsy 1989. West syndrome in our patients developed during the first 2 years of life. The most common interictal abnormality was hypsarrhythmia. All, except two patients had delayed development and various degrees of mental retardation.The most characteristic neuroimaging findings were major reduction in cerebral cortical gray matter volume, reduction in the volume of brain myelin, and delayed myelination. These findings may explain the anatomical association between the West syndrome onset and PVL and intellectual and cognitive deficit in premature infants with PVL

Neurodevelopmental Outcome in Children with Periventricular Leukomalacia

The purpose of this study was to question the correlation of different grades of periventricular leukomalacia (PVL) and subsequent neurodevelopmental outcome. In a prospective study we followed 52 preterm infants. Infants were divided into three groups according to their cranial ultrasound findings of PVL (De Vries classification). Seventeen children had PVL 1, 20 children had PVL 2, and 15 children had PVL 3. All 15 (100%) children with PVL 3 developed cerebral palsy with additional visual perceptual dysfunctions and epilepsy. Children with PVL 1 had high frequency of mild neuromotoric delay and visual impairment. PVL 2 and 3 have great predictive value for subsequent severe neurodevelopmental disorder which refers to cerebral palsy, different cognitive deficits, vision impairment and epilepsy. We have determined that due to high frequency of visual impairment and epilepsy we need to include neurophysiologic examinations very early in children with PVL lesions