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dc.contributor.advisorSchwartz, Ida Vanessa Doederleinpt_BR
dc.contributor.authorStarosta, Rodrigo Tzovenospt_BR
dc.date.accessioned2020-07-18T03:48:30Zpt_BR
dc.date.issued2020pt_BR
dc.identifier.urihttp://hdl.handle.net/10183/212099pt_BR
dc.description.abstractPacientes com DG tiveram diferença significativa de parâmetros nucleares hepatocelulares (área nuclear 28,16 ± 9,45 vs 26,88 ± 9,42 μm², p=0,006; dimensão fractal média 1,11 ± 0,06 vs 1,09 ± 0,09, p=0,005; e absorbância 4,88 ± 2,52 vs 7,68 ± 4,60, p=0,014) e de parâmetros canaliculares (absorbância 88,24 ± 59,80 vs 173,52 ± 76,78, p=0.001; relação perímetro-Feret 2,70 ± 0,56 vs 2,79 ± 0,66, p=0,006). Etapa 3) 33 pacientes tiveram 19 genes sequenciados, com identificação de 95 variantes (sendo 4 novas). Foi encontrada segregação de variante com fenótipo em 3 genes (A2M, CYBRD1 e TF), e diferenças no teste de carga na comparação com duas bases de dados populacionais em 5 genes (CP, CYBRD1, PSAP, TF e TFR2). A variante CYBRD1 rs10455 foi associada a uma diminuição de chance de osteonecrose (razão de chances de 0,08, intervalo de confiança 0,01 – 0,64, valor p de 0,004) no teste de segregação e com uma menor gravidade geral de doença no teste de carga. Etapa 4) Doze pacientes foram incluídos no estudo com ETH, com elastografia média de 5,25 kPa; aproximadamente metade dos pacientes com DG apresentaram algum grau de fibrose; o escore APRI teve uma área abaixo da curva de 0,701, sendo o mais acurado dos 3 escores testados. Esse teste mostrou sensibilidade 100% e especificidade 100% para detecção de pacientes com fibrose F2 ou superior utilizando-se os pontos de corte 0,201 e 0,604. Objetivo 2) foram incluídos 42 pacientes no estudo de coorte de pacientes com DCGs. Foram encontrados: grande aumento dos marcadores de dano hepatocelular, mas não biliar, com tendência à normalização após os 5 anos de idade nos pacientes com DCGs tipo I, mas não naqueles com tipo II; todos os tipos de esteatose e todos os graus de fibrose, sendo que 3 pacientes apresentaram cirrose; à pesquisa histológica em pacientes com cirrose, houve completa prevalência de acúmulo hepatocelular de glicogênio e completa ausência de hemossiderose. Conclusão: o fenótipo hepático da DG e dos DCGs é amplo e variado, com diversos pontos convergentes e divergentes entre essas doenças. Os DCGs de tipos I e II têm evoluções similares internamente, mas distintas entre si. A histomorfometria foi capaz de identificar alterações morfológicas canaliculares e hepatocelulares previamente não-descritas em pacientes com DG e que podem ajudar a explicar o fenótipo desses pacientes. A variante CYBRD1 rs10455 mostrou-se promissora como modificadora da DG. O escore APRI possui acurácia adequada ao seu uso clínico como preditor de fibrose na DG.pt_BR
dc.description.abstractIntroduction: Gaucher disease (GD) and the congenital disorders of glycosylation (CDGs) are inborn errors of metabolism with a broad and highly variable phenotype. Still understudied in GD and the CDGs is the set of pathophysiological manifestations that arise from the liver tissue – the hepatic phenotype. Objectives: 1) to describe and analyze the hepatic phenotype of GD; 2) to describe and analyze the clinical hepatic phenotype of the CDGs. Methods: for objectives 1), the following steps were performed: 1) describing and analyzing the hepatic phenotype of GD at a clinical level in a cohort study with acquisition of clinical, laboratorial, imaging, histological, and molecular genetic data; 2) describing and analyzing quantitatively, at the histological level, the hepatic phenotype of GD with the development of a new computerized analysis method (histomorphometry) of hepatocellular nuclei and biliary canaliculi in tissue in liver biopsy samples; 3) investigation of modifier factors of GD – candidate genes were sequenced by next-generation sequencing, with correlation of molecular findings with clinical (phenotypic segregation analysis) and populational (burden test) data using the populational databases gnomAD and ABraOM; step 4), transient hepatic elastography (THE) was used and the APRI, FIB-4, and NFS scores calculated for patients with GD. For objective 2), a cohort of patients with CDGs was used, with acquisition of clinical, laboratorial, imaging, histological, and molecular genetic data. Results: Objective 1), step 1) forty-two patients were included in the GD cohort study. Findings included: a high prevalence of abnormalities in hepatocellular and biliary markers both before and during treatment; a significant increase in the prevalence of steatosis after initiation of treatment with enzyme replacement therapy (ERT); evidence for hemosiderosis, canalicular cholestasis, and liver fibrosis in a high proportion of investigated patients; and the presence of steatohepatitis in 2 patients (a previously undescribed feature of GD). Step 2) At the histomorphometric study of patients with GD 4 liver biopsy samples of GD patients and 4 of healthy controls were included. GD patients had significant differences in hepatocellular nuclear parameters (nuclear area 28.16 ± 9.45 vs 26.88 ± 9.42 μm², p=0.006; average fractal dimension 1.11 ± 0.06 vs 1.09 ± 0.09, p=0.005; and corrected absorbance 11 4.88 ± 2.52 vs 7.68 ± 4.60, p=0.014) and in canalicular parameters (absorbance 88.24 ± 59.80 vs 173.52 ± 76.78, p=0.001; perimeter-to-Feret ratio 2.70 ± 0.56 vs 2.79 ± 0.66, p=0.006). Step 3) 33 patients were sequenced for 19 genes, identifying 95 variants (4 of which were novel). Variants in 3 genes were positive for phenotypic segregation (A2M, CYBRD1, and TF), and burden test was positive for both populational databases in 5 genes (CP, CYBRD1, PSAP, TF, and TFR2). The CYBRD1 rs10455 variant was associated with a decreased chance of osteonecrosis in the segregation analysis (odds ratio = 0.08, confidence interval 0.01-0.64, p-value = 0.004) and with a decreased overall disease severity in the burden test, being a strong candidate for a modifier variant. Step 4) Twelve patients were included in the study, with a mean elastography results of 5.25 kPa; approximately half of the DG patients had some degree of liver fibrosis; the APRI score had an area under the curve of 0.701, being the most accurate of the tested scores. This score was 100% sensitive and 100% specific for the detection of patients with an F2 or higher degree of fibrosis at the cut-off respectively of 0.201 and 0.640. Objective 2) Forty-two patients were included the CDG cohort study. Findings included: a stark increase in hepatocellular, but not biliary, markers with a trend towards normalization after 5 years of age in type I CDG patients but not in type II patients; all types of steatosis and all degrees of fibrosis were present, with cirrhosis in 3 patients; at histology of patients with cirrhosis, all patients had hepatocellular glycogen build-up, and none had hemosiderosis. Conclusion: the hepatic phenotype of GD and the CDGs is broad and varied, with many converging and diverging features between those pathologies. CDGs types I and II are internally similar but externally different regarding the hepatic natural history. Histomorphometry was able to identify canalicular and hepatocellular features which had not been described previously in GD, and which may help to explain some aspects of their clinical phenotype. The CYBRD1 rs10455 variant is promising as a putative modifier of GD. The APRI score has adequate accuracy for a clinical use as a fibrosis predictor in GD.en
dc.format.mimetypeapplication/pdfpt_BR
dc.language.isoporpt_BR
dc.rightsOpen Accessen
dc.subjectDoença de Gaucherpt_BR
dc.subjectGlicosilaçãopt_BR
dc.subjectErros inatos do metabolismopt_BR
dc.subjectFígadopt_BR
dc.titleO fenótipo hepático da doença de Gaucher e dos distúrbios congênitos da glicosilaçãopt_BR
dc.typeTesept_BR
dc.contributor.advisor-coSiebert, Marinapt_BR
dc.identifier.nrb001116140pt_BR
dc.degree.grantorUniversidade Federal do Rio Grande do Sulpt_BR
dc.degree.departmentInstituto de Biociênciaspt_BR
dc.degree.programPrograma de Pós-Graduação em Genética e Biologia Molecularpt_BR
dc.degree.localPorto Alegre, BR-RSpt_BR
dc.degree.date2020pt_BR
dc.degree.leveldoutoradopt_BR


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