RESUMO
OBJECTIVES: To determine how advanced genetic analysis methods may help in clinical diagnosis. STUDY DESIGN: We report a combined genetic diagnosis approach for patients with clinical suspicion of genetic liver diseases in a tertiary referral center, using tools either tier 1: Sanger sequencing on SLC2SA13, ATP8B1, ABCB11, ABCB4, and JAG1 genes, tier 2: panel-based next generation sequencing (NGS), or tier 3: whole-exome sequencing (WES) analysis. RESULTS: In a total of 374 patients undergoing genetic analysis, 175 patients received tier 1 Sanger sequencing based on phenotypic suspicion, and pathogenic variants were identified in 38 patients (21.7%). Tier 2 included 216 patients (39 of tier 1-negative patients) who received panel-based NGS, and pathogenic variants were identified in 60 (27.8%). In tier 3, 41 patients received WES analysis, and 20 (48.8%) obtained genetic diagnosis. Pathogenic variants were detected in 6 of 19 (31.6%) who tested negative in tier 2, and a greater detection rate in 14 of 22 (63.6%) patients with deteriorating/multiorgan disease receiving one-step WES (P = .041). The overall disease spectrum is comprised of 35 genetic defects; 90% of genes belong to the functional categories of small molecule metabolism, ciliopathy, bile duct development, and membrane transport. Only 13 (37%) genetic diseases were detected in more than 2 families. A hypothetical approach using a small panel-based NGS can serve as the first tier with diagnostic yield of 27.8% (98/352). CONCLUSIONS: NGS based genetic test using a combined panel-WES approach is efficient for the diagnosis of the highly diverse genetic liver diseases.
Assuntos
Testes Genéticos , Hepatopatias , Humanos , Sequenciamento do Exoma , Hepatopatias/diagnóstico , Hepatopatias/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , MutaçãoRESUMO
OBJECTIVE: To assess the diagnostic and prognostic usefulness of the serum matrix metallopeptidase-7 (MMP-7) level for biliary atresia in infants with cholestasis after hepatoportoenterostomy. STUDY DESIGN: We enrolled 100 infants with cholestasis (age, 43.56 ± 1.97 days; 62 males) with a direct bilirubin level of >1 mg/dL, of whom 36 (36%) were diagnosed with biliary atresisa. The MMP-7 levels in serum samples collected during the cholestasis workup and 6 months after hepatoportoenterostomy were assessed by enzyme-linked immunosorbent assay. We quantified liver fibrosis by Picro Sirius red staining of collagen in specimens from the 81 infants with cholestasis. RESULTS: Infants with biliary atresisa had a significantly higher serum MMP-7 level than that of non-biliary atresisa infants with cholestasis of equivalent age (P < .0001). Receiver operating characteristic analysis showed that a serum MMP-7 level of >1.43 ng/mL was predictive of biliary atresisa in infants with cholestasis (diagnostic accuracy, 88%). There was a positive correlation between the serum MMP-7 level and the severity of liver fibrosis (P = .0002). Survival analysis showed that the frequency of liver transplantation was significantly higher in infants with biliary atresisa with a serum MMP-7 level of >10.30 ng/mL compared with a serum MMP-7 level of ≤10.30 ng/mL after hepatoportoenterostomy (hazard ratio, 4.22; P = .02). CONCLUSIONS: The serum MMP-7 level, which reflects the severity of liver fibrosis and can be determined noninvasively, may facilitate the diagnosis of biliary atresisa among infants with cholestasis. Moreover, the serum MMP-7 level after hepatoportoenterostomy is associated with a need for liver transplantation in infants with biliary atresisa.
Assuntos
Atresia Biliar/diagnóstico , Atresia Biliar/enzimologia , Colestase/complicações , Cirrose Hepática/etiologia , Metaloproteinase 7 da Matriz/sangue , Portoenterostomia Hepática/efeitos adversos , Atresia Biliar/cirurgia , Colestase/enzimologia , Estudos de Coortes , Feminino , Humanos , Lactente , Transplante de Fígado , Masculino , Valor Preditivo dos Testes , Curva ROCRESUMO
OBJECTIVE: To test the application of a target enrichment next-generation sequencing (NGS) jaundice panel in genetic diagnosis of pediatric liver diseases. STUDY DESIGN: We developed a capture-based target enrichment NGS jaundice panel containing 42 known disease-causing genes associated with jaundice or cholestasis and 10 pathway-related genes. During 2015-2017, 102 pediatric patients with various forms of cholestasis or idiopathic liver diseases were tested, including patients with initial diagnosis of cholestasis in infancy, progressive familial intrahepatic cholestasis, syndromic cholestasis, Wilson disease, and others. RESULTS: Of the 102 patients, 137 mutations/variants in 44 different genes were identified in 84 patients. The genetic disease diagnosis rate was 33 of 102 (32.4%). A total of 79 of 102 (77.5%) of patients had at least 1 heterozygous genetic variation. Those with progressive intrahepatic cholestasis or syndromic cholestasis in infancy had a diagnostic rate of 62.5%. Disease-causing mutations, including ATP8B1, ABCB11, ABCB4, ABCC2, TJP2, NR1H4 (FXR), JAG1, AKR1D1, CYP7B1, PKHD1, ATP7B, and SLC25A13, were identified. Nine patients had unpredicted genetic diagnosis with atypical phenotype or novel mutations in the investigational genes. We propose an NGS diagnosis classification categorizing patients into high (n = 24), moderate (n = 9), or weak (n = 25) levels of genotype-phenotype correlations to facilitate patient management. CONCLUSIONS: This panel enabled high-throughput detection of genetic variants and disease diagnosis in patients with a long list of candidate causative genes. A NGS report with diagnosis classification may aid clinicians in data interpretation and patient management.
Assuntos
Membro 11 da Subfamília B de Transportadores de Cassetes de Ligação de ATP/genética , Colestase Intra-Hepática/diagnóstico , DNA/genética , Mutação , Receptores Citoplasmáticos e Nucleares/genética , Membro 11 da Subfamília B de Transportadores de Cassetes de Ligação de ATP/metabolismo , Pré-Escolar , Colestase Intra-Hepática/genética , Colestase Intra-Hepática/metabolismo , Análise Mutacional de DNA , Feminino , Estudos de Associação Genética , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Lactente , Recém-Nascido , Masculino , Proteína 2 Associada à Farmacorresistência Múltipla , Receptores Citoplasmáticos e Nucleares/metabolismo , Estudos RetrospectivosRESUMO
OBJECTIVE: To investigate the factors predicting spontaneous clearance of hepatitis B surface antigen (HBsAg) in a long-term, prospectively followed cohort from childhood into adult life. STUDY DESIGN: Children with chronic hepatitis B virus (HBV) infection without treatment were followed longitudinally every 6 months. At each visit, liver profiles and HBV markers were assessed. Hepatitis B vaccination history and the maternal HBV markers also were studied. RESULTS: A total of 349 children (205 male) were followed for 20.6 ± 4.4 years with initial ages of 8.4 ± 3.9 years; 42 (12.0%) cleared HBsAg spontaneously. The HBsAg titers decayed with age, with an average annual clearance rate of 0.58%. Children had a lower annual HBsAg decay rate if their mothers are HBsAg carriers (P < .001). Hepatitis B e antigen-seroconversion is a favorable predictor for spontaneous HBsAg clearance (P = .04). Those with HBsAg titer ≤1000 IU/mL at enrollment during childhood have a higher rate of HBsAg clearance (hazard ratio = 5.23; P < .001). Using HBsAg titer ≤1000 IU/mL to predict HBsAg clearance, the sensitivity is 38.1%, specificity is 90.6%, positive predictive value is 35.6%, and negative predictive value is 91.4%. CONCLUSIONS: During long-term follow-up, spontaneous HBsAg clearance is most likely to occur in a patient born to a non-HBsAg-carrier mother, is a hepatitis B e antigen-seroconverter, and had an initial HBsAg level ≤1000 IU/mL.
Assuntos
Antígenos de Superfície da Hepatite B/sangue , Hepatite B Crônica/sangue , Hepatite B Crônica/imunologia , Adolescente , Adulto , Criança , DNA Viral/sangue , Feminino , Seguimentos , Genótipo , Vírus da Hepatite B/imunologia , Humanos , Masculino , Valor Preditivo dos Testes , Modelos de Riscos Proporcionais , Estudos Prospectivos , Sensibilidade e Especificidade , Fatores de Tempo , Transaminases/sangue , Carga Viral , Adulto JovemRESUMO
OBJECTIVE: To investigate the impact of menarche on the natural course of chronic hepatitis B virus (HBV) infection in women. STUDY DESIGN: Young women who are positive for hepatitis B e antigen (HBeAg; n = 101) chronically infected with genotypes B and C HBV were recruited at a mean age of 4.57 ± 3.08 years, and a mean follow-up duration of 23.98 ± 3.77 years. Clinical data, including age at menarche, HBV genotypes, serum HBV viral loads, hepatitis B surface antigen (HBsAg) titers, and serial liver functional profiles were analyzed. RESULTS: Women with earlier onset of menarche had earlier spontaneous HBeAg seroconversion than others (hazard ratio, 2.0; P = .02) adjusting for HBV genotype and peak alanine aminotransferase levels before HBeAg seroconversion. The annual decrease in HBsAg titer from 15 to 20 years of age also was greater in the early menarche group compared with the late menarche group (0.11 ± 0.11 vs 0.05 ± 0.11 log10 IU/mL, P = .04). The baseline HBV viral load was also borderline low in female subjects with earlier menarche as compared with others (P = .06). Earlier menarche onset was associated with higher spontaneous HBeAg seroconversion, HBsAg seroclearance, and HBsAg seroconversion rate before 15 years of age in females with chronic HBV infection. CONCLUSIONS: Earlier puberty-onset, indicated by menarche-onset, was associated with earlier spontaneous HBeAg seroconversion and greater rate of HBV clearance before 15 years of age in female subjects with chronic HBV infection.
Assuntos
Hepatite B Crônica/imunologia , Menarca , Adolescente , Adulto , Criança , Pré-Escolar , Feminino , Seguimentos , Antígenos de Superfície da Hepatite B/sangue , Antígenos E da Hepatite B/sangue , Hepatite B Crônica/sangue , Humanos , Estudos Retrospectivos , Adulto JovemRESUMO
OBJECTIVE: To assess the diagnosis of neonatal intrahepatic cholestasis caused by citrin deficiency (NICCD) by using high-resolution melting (HRM) analysis and a clinical scoring system. STUDY DESIGN: Genetic variations in the 18 coding exons were prescreened using HRM analysis and then confirmed by direct sequencing. To establish a scoring system, clinical features of 20 patients with NICCD diagnosed in Taiwan between the years 2000 and 2008 were compared with those of 47 patients with biliary atresia and 35 with infantile cholestasis. RESULTS: Eight types of mutations/polymorphisms were identified in patients with NICCD, including 5 mutations in the coding region or splice site (c.851del4, c.1638ins23, R553Q, IVS6+5G > A, IVS11+1G > A), and 3 single-nucleotide polymorphisms (IVS11+17C > G, IVS4+6A > G/rs6957975, and c.1194A > G/rs2301629). The 3 hotspot mutations (c.851del4, c.1638ins23, and IVS6+5G > A) comprised 33/35 (94.3%) mutated alleles. The patients with NICCD had a higher frequency of the rs6957975 polymorphism compared with 103 healthy controls (P < .0001). A 6-point scoring system was proposed according to clinical parameters. The patients with NICCD tended to score ≥ 4 points, whereas biliary atresia and other infantile cholestasis tended to score <4 points (P < .0001). CONCLUSIONS: HRM analysis was efficient and effective in detecting mutations. Three common mutations comprised the majority of mutations found in our patients. The IVS4+6A > G polymorphism was associated with NICCD. A scoring system may help to differentiate patients with NICCD from those with biliary atresia.
Assuntos
Citrulinemia/diagnóstico , Desnaturação de Ácido Nucleico , Citrulinemia/genética , Frequência do Gene , Humanos , Recém-Nascido , Proteínas de Transporte da Membrana Mitocondrial/genética , Mutação , Mutação de Sentido Incorreto , Polimorfismo de Nucleotídeo ÚnicoRESUMO
OBJECTIVE: To investigate the risk factors for hyperbilirubinemia in infants who are exclusively breast-fed. STUDY DESIGN: A prospective study was conducted to investigate the effects of birth body weight, sex, mode of delivery, glucose-6-phosphate dehydrogenase (G6PD) deficiency, variant UDP-glucuronosyltransferase 1A1 (UGT1A1) gene, and hepatic solute carrier organic anion transporter 1B1 (SLCO1B1) gene on hyperbilirubinemia in neonates who were breast-fed. Hyperbilirubinemia was diagnosed when a full term neonate had a bilirubin level â§15.0 mg/dL (256.5 µM) in serum at 3 days old. The polymerase chain reaction-restriction fragment length polymorphism method was used as a means of detecting the known variant sites in the UGT1A1 and SLCO1B1 gene. RESULTS: Of 252 infants born at term who were exclusively breast-fed, 59 (23.4%) had hyperbilirubinemia. The significant risk factors were a variant nucleotide 211 in UGT1A1 (2.48; 95% CI, 1.29 to 4.76; P = .006), G6PD deficiency (12.24; 95% CI, 1.08 to 138.62; P < .05), and vaginal delivery (3.55; 95% CI, 1.64 to 7.66; P < .001). CONCLUSION: Breast-fed neonates who are 211 variants in the UGT1A1, G6PD deficiency, and vaginal delivery are at high-risk for hyperbilirubinemia.
Assuntos
Aleitamento Materno , Hiperbilirrubinemia/epidemiologia , Hiperbilirrubinemia/etiologia , Estudos de Casos e Controles , Parto Obstétrico , Feminino , Deficiência de Glucosefosfato Desidrogenase/epidemiologia , Glucuronosiltransferase/genética , Humanos , Lactente , Recém-Nascido , Transportador 1 de Ânion Orgânico Específico do Fígado , Masculino , Transportadores de Ânions Orgânicos/genética , Mutação Puntual , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Estudos Prospectivos , Fatores de RiscoRESUMO
OBJECTIVE: To elucidate the association between human interleukin-10 (IL-10) genotypes and hepatitis B virus (HBV) precore/core gene mutation in children with chronic HBV infection. STUDY DESIGN: The study group comprised of 21 children with chronic HBV infection with spontaneous hepatitis B e antigen (HBeAg) seroconversion who were followed for more than 10 years. Another nine children without HBeAg seroconversion served as the control subjects. Sera at the immune tolerance and inflammatory phase (alanine aminotransferase, >80 IU/L) were subjected to HBV precore/core sequence analysis. IL-10 -1082 polymorphism was also determined. RESULTS: HBV precore/core gene mutation increased significantly more in the inflammatory phase than in the tolerance phase (G1896A, 76.2% versus 4.8%; C1913A, 33.3% versus 0%; C2189A, 28.6% versus 4.8%; G2304A, 52.4% versus 14.3%) in study group (n = 21) but not the control group (n = 9). Subjects with the G/G genotype at the IL-10-1082 polymorphism site had higher C2189A mutation rate than the A allele carriers (P = .02). C2189A mutation carriers are associated with more viral load decrement from tolerance to inflammatory phase (P = .01) and earlier spontaneous HBeAg seroconversion (P = .01). CONCLUSIONS: The G/G genotype at the IL-10 -1082 polymorphism is associated with higher C2189A mutations, lower HBV viral load at immune inflammatory phase, and earlier spontaneous HBeAg seroconversion than A allele carriers.
Assuntos
DNA/genética , Hepatite B Crônica/genética , Interleucina-10/genética , Mutação , Pré-Escolar , Progressão da Doença , Feminino , Seguimentos , Genótipo , Antígenos E da Hepatite B/análise , Vírus da Hepatite B/imunologia , Hepatite B Crônica/sangue , Hepatite B Crônica/virologia , Humanos , Interleucina-10/sangue , Masculino , Reação em Cadeia da Polimerase , Regiões Promotoras Genéticas , Fatores de Tempo , Carga ViralRESUMO
OBJECTIVE: To test the hypothesis that the presence of the PNPLA3 rs738409 G allele would increase the susceptibility of non-alcoholic fatty liver disease (NAFLD) in obese Taiwanese children. STUDY DESIGN: A total of 520 obese children aged 6-18 years were recruited. Their PNPLA3 rs738409 genotypes-CC, CG, or GG-were detected by the 5'-nuclease assay. The effects of the PNPLA3 rs738409 G allele on pediatric NAFLD were evaluated based on liver ultrasonography findings and mean serum alanine aminotransferase levels in these children. RESULTS: NAFLD was present in 19.6% of the obese children. In comparison to the subjects with CC alleles, the risk of NAFLD was increased by 2.96-fold (95% CI, 1.57 to 5.59, P = .0008) in the subjects with CG alleles and by 5.84-fold (95% CI, 2.59 to 13.16; P < .0001) in those with GG alleles. Variant PNPLA3 rs738409 genotypes were associated with increases in mean serum alanine aminotransferase level of 4.77 IU/L (P = .0435) in subjects with CG alleles and of 10.86 IU/L (P < .0001) in those with GG alleles compared with subjects with CC alleles. CONCLUSIONS: The variant PNPLA3 rs738409 genotypes increased the risk of NAFLD in our population of obese Taiwanese children. The effect of the G allele on pediatric NAFLD followed a dominant genetic model.
Assuntos
DNA/genética , Predisposição Genética para Doença , Lipase/genética , Fígado/metabolismo , Proteínas de Membrana/genética , Obesidade/genética , Polimorfismo de Nucleotídeo Único , Adolescente , Alelos , Criança , Fígado Gorduroso/complicações , Fígado Gorduroso/epidemiologia , Fígado Gorduroso/genética , Feminino , Genótipo , Humanos , Lipase/metabolismo , Fígado/diagnóstico por imagem , Masculino , Proteínas de Membrana/metabolismo , Hepatopatia Gordurosa não Alcoólica , Obesidade/complicações , Obesidade/epidemiologia , Prevalência , Estudos Retrospectivos , Fatores de Risco , Taiwan/epidemiologia , UltrassonografiaRESUMO
OBJECTIVE: To test the hypothesis that a mutation in uridine diphosphate-glucuronosyl transferase 1A1 (UGT1A1) gene of breast-fed infants is a contributory factor to prolonged unconjugated hyperbilirubinemia. STUDY DESIGN: Of 125 breast-fed term infants, 35 infants had prolonged unconjugated hyperbilirubinemia; another 90 breast-fed neonates without prolonged jaundice were control infants. The polymerase chain reaction-restriction fragment length polymorphism method was used to detect the known variant sites (promoter area, nucleotides 211, 686, 1091, and 1456) of the UGT1A1 gene. RESULTS: Of 35 breast-fed infants with prolonged unconjugated hyperbilirubinemia, 29 had at least 1 mutation of the UGT1A1 gene. Variation at nucleotide 211 was most common. The percentages of the neonates carrying the variant nucleotide 211 were significantly different between the prolonged hyperbilirubinemia group and control neonates. Male breast-fed infants had a higher risk than female infants for prolonged hyperbilirubinemia. CONCLUSIONS: Male breast-fed neonates with a variant nucleotide 211 in UGT1A1 have a high risk for developing prolonged hyperbilirubinemia.