Search PubMed⌕ Search

Biomedical subjects

E A Roberts

Publications and source records attributed to E A Roberts.

At least 37 records · Page 2Linked to original sources
PubMed2000

Nonalcoholic steatohepatitis in children.

BACKGROUND: Nonalcoholic steatohepatitis occurs commonly in adults with obesity or diabetes mellitus. There are only a few reports of this condition in children. METHODS: Prospective consecutive clinical series. RESULTS: Between December 1985 and April 1995, 36 children (21 boys, 15 girls) were diagnosed with nonalcoholic steatohepatitis at the Hospital for Sick Children, Toronto. The median age at diagnosis was 12 years (range, 4-16 years). Most patients were referred because of elevated serum aminotransferases or abnormal hepatic sonogram. Thirty patients (83%) were obese. Two patients had diabetes mellitus at diagnosis, and it developed later in two. Fifteen patients had palpable hepatomegaly, and one of these had splenomegaly. None had physical signs of chronic liver disease. Thirteen of 36 patients had acanthosis nigricans. Serum aminotransferases were elevated in all but one patient. Tests for Wilson disease and chronic hepatitis B and C were negative. Serum lipid profiles were abnormal in 18 patients: 7 had hypercholesterolemia, and 11 had hypertriglyceridemia. Twenty-four of 31 examined had abnormal liver sonograms suggestive of fatty infiltration. Twenty-four patients underwent percutaneous liver biopsy: all showed large-droplet fat. Inflammation was present in 88% and fibrosis-cirrhosis in 75%. One 10-year-old patient had established cirrhosis at diagnosis. CONCLUSIONS: Nonalcoholic steatohepatitis occurs in children, is clinically diverse, and may not be benign.

Acanthosis Nigricans↗
PubMed2000

A dose ranging study of the pharmacokinetics, safety, and preliminary efficacy of lamivudine in children and adolescents with chronic hepatitis B.

Fifty-three patients with chronic hepatitis B and active viral replication were studied for 4 weeks while on treatment and for 12 weeks after treatment with the oral nucleoside analogue lamivudine. Children aged 2 to 12 years were randomized to receive twice-daily doses of 0.35, 1.5, or 4 mg of lamivudine solution per kg of body weight or once-daily doses of 3 mg of lamivudine solution per kg. Adolescents aged 13 to 17 years received lamivudine at 100 mg (as tablets). Blood samples for pharmacokinetic assay were taken on days 1 and 28. Lamivudine was rapidly absorbed following oral administration, with the maximum concentration in serum being reached 0.5 to 1 h postdosing. Apparent oral clearance (CL/F) was higher in younger children and decreased with age, with CL/F values for adolescents reaching those seen for adults by the age of 12. All doses produced a dramatic fall in serum hepatitis B virus (HBV) DNA levels, with a median reduction of >/=99.5% after 4 weeks of treatment and with the levels returning to the baseline levels posttreatment. The correlation of dose, area under the concentration-time curve (AUC), and changes in HBV DNA levels, as measured by the Chiron Quantiplex assay, showed maximal antiviral effects (99.9% inhibition and a reduction of the amount of HBV DNA of approximately 3 log(10)) at 3 mg/kg/day, with no discernible increase in effect seen whether the drug was given at 4 mg/kg twice daily or whether it was given once daily or twice daily. The limit of detection of the assay (2.5 pg/ml) was reached for some but not all patients across the dose ranges, with the smallest number (n = 2) of those having values negative by the Chiron Quantiplex assay being in the lowest-dose group. The 13- to 17-year-olds showed a similar overall response in terms of the HBV DNA level reduction compared to that for patients younger than age 13. Analysis of the same samples by PCR, which has a lower limit of sensitivity than the Chiron Quantiplex assay, also showed average drops in HBV DNA levels of about 3 log(10) at 4 weeks for patients for which the AUC was >/=4,000 ng. h/ml, confirming the conclusions given above. Lamivudine treatment was well tolerated at all doses, with no significant adverse events or laboratory data changes. On the basis of pharmacokinetic and pharmacodynamic data, a 3-mg/kg/day dose in children (ages 2 to 12 years) with chronic hepatitis B provides levels of exposure and trough concentrations similar to those seen in adults following the receipt of doses of 100 mg. The 100-mg dose is being evaluated in a large phase III study with HBV-infected pediatric patients.

Adolescent↗
PubMed2000

Identification of a Mycobacterium tuberculosis gene that enhances mycobacterial survival in macrophages.

Intracellular survival plays a central role in the pathogenesis of Mycobacterium tuberculosis. To identify M. tuberculosis genes required for intracellular survival within macrophages, an M. tuberculosis H37Rv plasmid library was constructed by using the shuttle vector pOLYG. This plasmid library was electroporated into Mycobacterium smegmatis 1-2c, and the transformants were used to infect the human macrophage-like cell line U-937. Because M. smegmatis does not readily survive within macrophages, any increased intracellular survival is likely due to cloned M. tuberculosis H37Rv DNA. After six sequential passages of M. smegmatis transformants through U-937 cells, one clone (p69) was enriched more than 70% as determined by both restriction enzyme and PCR analyses. p69 demonstrated significantly enhanced survival compared to that of the vector control, ranging from 2.4- to 5.3-fold at both 24 and 48 h after infection. DNA sequence analysis revealed three open reading frames (ORFs) in the insert of p69. ORF2 (1.2 kb) was the only one which contained a putative promoter region and a ribosome-binding site. Deletion analysis of the p69 insert DNA showed that disruption of ORF2 resulted in complete loss of the enhanced intracellular survival phenotype. This gene was named the enhanced intracellular survival (eis) gene. By using an internal region of eis as a probe for Southern analysis, eis was found in the genomic DNA of various M. tuberculosis strains and of Mycobacterium bovis BCG but not in that of M. smegmatis or 10 other nonpathogenic mycobacterial species. Sodium dodecyl sulfate-polyacrylamide gel electrophoretic analysis showed that all M. smegmatis eis-containing constructs expressed a unique protein of 42 kDa, the predicted size of Eis. The expression of this 42-kDa protein directly correlated to the enhanced survival of M. smegmatis p69 in U-937 cells. These results suggest a possible role for eis and its protein product in the intracellular survival of M. tuberculosis.

Amino Acid Sequence↗
PubMed1999

Defective kinesin heavy chain behavior in mouse kinesin light chain mutants.

Conventional kinesin, kinesin-I, is a heterotetramer of two kinesin heavy chain (KHC) subunits (KIF5A, KIF5B, or KIF5C) and two kinesin light chain (KLC) subunits. While KHC contains the motor activity, the role of KLC remains unknown. It has been suggested that KLC is involved in either modulation of KHC activity or in cargo binding. Previously, we characterized KLC genes in mouse (Rahman, A., D.S. Friedman, and L.S. Goldstein. 1998. J. Biol. Chem. 273:15395-15403). Of the two characterized gene products, KLC1 was predominant in neuronal tissues, whereas KLC2 showed a more ubiquitous pattern of expression. To define the in vivo role of KLC, we generated KLC1 gene-targeted mice. Removal of functional KLC1 resulted in significantly smaller mutant mice that also exhibited pronounced motor disabilities. Biochemical analyses demonstrated that KLC1 mutant mice have a pool of KIF5A not associated with any known KLC subunit. Immunofluorescence studies of sensory and motor neuron cell bodies in KLC1 mutants revealed that KIF5A colocalized aberrantly with the peripheral cis-Golgi marker giantin in mutant cells. Striking changes and aberrant colocalization were also observed in the intracellular distribution of KIF5B and beta'-COP, a component of COP1 coatomer. Taken together, these data best support models that suggest that KLC1 is essential for proper KHC activation or targeting.

Animals↗
PubMed1999

Behavior of a cell line derived from normal human hepatocytes on non-physiological and physiological-type substrates: evidence for enhancement of secretion of liver-specific proteins by a three-dimensional growth pattern.

The behavior of a recently described cell line, HH25, derived from normal human hepatocytes, has been investigated on several different substrates--tissue-culture plastic, glass, a thin layer of rat-tail collagen I, and thin layers or thick gels of extracellular matrix derived from the Engelbreth-Holm-Swarm murine sarcoma (EHS matrix). Cellular morphology, proliferation, and secretion of three hepatocyte-specific proteins (albumin, alpha1 acid glycoprotein, and alpha1 antitrypsin) have been examined. There were no differences in morphology, proliferation, or differentiated function in the cells on either plastic, glass, collagen, I, or a thin layer of EHS matrix, but on a thick EHS matrix gel the cells altered their morphology (forming three-dimensional colonies with canalicular-like structures) and their production of albumin and alpha1 acid glycoprotein was enhanced. This suggests that the enhanced differentiated function is associated with the morphological change (occurring only on the thick EHS gel) rather than with receptor-mediated cell-matrix interactions (which can also occur on the thin layer of EHS matrix). This cell line is therefore a good in vitro cellular model for the investigation of the roles of morphological changes and of cell-cell and cell-matrix interactions in the control of human hepatocyte behavior without the need for an extensive source of primary tissue.

Albumins↗
PubMed1999

Primary sclerosing cholangitis in children.

Primary sclerosing cholangitis (PSC), a chronic inflammatory process affecting the extrahepatic and/or medium to large bile ducts, is not rare in children. It has features suggesting an autoimmune pathogenesis, although the mechanism of tissue damage remains unknown. The clinical presentation of childhood primary sclerosing cholangitis is highly variable and frequently without obvious features of cholestasis. Clinical similarity to autoimmune hepatitis is common. Association with chronic colitis is less common than in adults. Cholangiography is essential for the diagnosis and examination of the medium to large intrahepatic ducts is mandatory, as 40% of children lack extrahepatic duct involvement. Histological findings may help to distinguish childhood PSC from autoimmune hepatitis. In children, sclerosing cholangitis may also develop secondary to other disease processes, notably Langerhans histiocytosis, congenital immunodeficiencies and cystic fibrosis. Neonatal sclerosing cholangitis is chronic inflammatory disease of bile ducts which presents initially with neonatal cholestasis; its pathogenesis remains uncertain and may not be the same as for primary sclerosing cholangitis. Effective treatment modalities for childhood PSC remain undetermined. Liver transplantation is required for children who progress to biliary cirrhosis and hepatic decompensation.

Adolescent↗
PubMed1998

Interferon alfa therapy for chronic hepatitis B in children: a multinational randomized controlled trial.

BACKGROUND & AIMS: Treatment of chronic hepatitis B with interferon alfa is not approved in children. The aim of this study was to evaluate the safety and efficacy of interferon alfa (IFN-alpha) in children with chronic hepatitis B and increased transaminase levels. METHODS: Children were given either IFN-alpha2b (6 megaunits/m2 thrice weekly for 24 weeks) or no treatment. Clearance of markers of viral replication was evaluated 24 weeks after therapy and after 48 weeks of observation in controls. RESULTS: Of 149 children enrolled, 144 were evaluable (70 treated and 74 controls). Serum hepatitis B e antigen and viral DNA became negative in 26% of treated children and 11% of controls (P < 0.05). Serum aminotransferase levels normalized and liver histology improved among responders. Hepatitis B surface antigen became undetectable in 10% of treated patients and 1% of controls. Female gender and interferon treatment were the only significant predictors of response. Ethnic origin, baseline aminotransferase level, initial DNA levels, and histology did not correlate with response. Most adverse reactions were mild or moderate, and dose was reduced in 24% of children. CONCLUSIONS: In children with chronic hepatitis B, INF-alpha promotes loss of viral replication markers and surface antigen and improves aminotransferases and histology.

Adolescent↗
PubMed1998

Pemoline hepatotoxicity in children.

Pemoline hepatotoxicity ranges from asymptomatic elevations in levels of serum aminotransferases to fulminant liver failure. We report five cases of pemoline hepatotoxicity in children (four boys, one girl), including the only reported case resulting in orthotopic liver transplantation. We conclude that pemoline causes toxic liver damage in children. The severity of the damage is highly variable, and its onset may be late in the course of treatment. Pemoline and methylphenidate may act synergistically to cause liver damage. The levels of serum aminotransferases should be monitored throughout treatment with these agents.

Adolescent↗
PubMed1998

Wilson disease.

Wilson disease is a recessively inherited disorder of copper transport. Clinical features are highly variable, with any combination of neurological, hepatic or psychiatric illness. The age of onset varies from 3 to 50 years of age. Diagnosis is challenging because no specific combination of clinical or biochemical features is necessarily definitive. The genetic defect is due to a variety of abnormalities in a copper-transporting membrane ATPase. Most of the more than 80 mutations are present at a low frequency, and mutations differ between ethnic groups. At least two mutations are sufficiently common to aid in rapid diagnosis, in European and Asian populations respectively. Molecular analysis can provide a definitive diagnosis for asymptomatic sibs. Treatment, using chelating agents or zinc, is most effective when started before permanent tissue damage occurs.

Adolescent↗
PubMed1997

Distinction of CYP1A1 and CYP1A2 activity by selective inhibition using fluvoxamine and isosafrole.

Ethoxyresorufin O-deethylation (EROD) has been used as a specific probe for CYP1A1 and CYP1A2. Selective inhibition of one of these cytochromes P450 may differentiate their activity in human liver. Four inhibitors were chosen to examine the selective inhibition of EROD activity, using cDNA of CYP1A1 and CYP1A2. The two flavones, alpha-naphthoflavone and apigenin, while differing in potency, inhibited expressed human CYP1A1, CYP1A2, and human liver microsomes to a similar extent. Isosafrole and fluvoxamine were found to inhibit CYP1A2 selectively, with Ki values of 14 and 800 times, respectively, lower than those for CY1A1. A set of equations was developed to estimate both CYP1A1 and CYP1A2 activity. Levels of CYP1A2 in four human liver specimens ranged from 44.4 to 76.7 pmol/mg protein, which significantly correlated with phenacetin O-deethylase activity (r = 0.99; P < 0.001). Low levels of CYP1A1 activity were present in all four investigated livers, ranging from 0.4 to 2.7 pmol/mg protein.

Cell Line↗
PubMed1997

Extended primary culture of human hepatocytes in a collagen gel sandwich system.

To develop a strategy for extended primary culture of human hepatocytes, we placed human hepatocytes between two layers of collagen gel, called a "collagen gel sandwich." Maintenance of hepatocellular functions in this system was compared with that of identical hepatocyte preparations cultured on dry-collagen coated dishes or cocultured with rat liver epithelial cells. Human hepatocytes in a collagen gel sandwich (five separate cultures) survived for more than 4 wk, with the longest period of culture being 78 d. They maintained polygonal morphology with bile canaliculuslike structures and high levels of albumin secretion throughout the period of culture. In contrast, hepatocytes on dry-collagen became feature-less, and albumin secretion could not be detected after 14 d of culture. This loss of albumin secretion was partially recovered by overlaying one layer of collagen gel. Ethoxyresorufin O-deethylase activity, associated with cytochrome P450 1A2, was detected basally up to 29 d in collagen gel sandwich culture. These activities were induced four- to eightfold after induction with dibenz(a,h)anthracene. Cocultures also maintained basal activity up to 29 d. However, their inducibility was lower than that of hepatocytes in collagen gel sandwich. No ethoxyresorufin O-deethylase activity was detected in hepatocytes cultured on dry-collagen at 7 d. Thus, the collagen gel sandwich system preserves differentiated morphology and functions of human hepatocytes in primary culture for a prolonged period of time. This system is a promising model for studying human hepatocellular function, including protein synthesis and drug metabolism in vitro.

Adolescent↗
PubMed1997

Hepatitis B virus replication and viral antigen synthesis in hepatocyte lines derived from normal human liver.

Transient transfection and in vitro infection experiments were performed to characterize replication and antigen synthesis of the hepatitis B virus (HBV) in human hepatocyte lines HH29 and HHY41, derived from normal liver tissue. These liver cell lines are capable of supporting HBV replication and gene expression at levels similar to the human hepatoma cell line HuH-7. Strikingly, a very tight adhesion of HBV to the outer cell membrane of HH29 and HHY41 was observed under conditions that removed HBV to undetectable levels from HuH-7 hepatoma cells. However, no productive HBV infection could be established in these cells as determined by the absence of viral transcripts and de novo antigen synthesis. In conclusion, the human hepatocyte cell lines HH29 and HHY41 may be useful to study important aspects of late steps in the replication of HBV, but appear to lack certain cellular components that play a pivotal role during early steps of the viral life cycle.

Cell Line↗
PubMed1997

Modulation of LDL receptor mRNA stability by phorbol esters in human liver cell culture models.

In the human hepatocarcinoma cell lines HepG2 and Hep3B, low density lipoprotein receptor (LDLR) mRNA levels were rapidly and transiently induced after treatment with phorbol-12-myristate-13-acetate (PMA), increasing by approximately 50-fold and 8-fold, respectively, within 4 h before returning to near basal levels by 24 h. The difference in magnitude of mRNA accumulation between these cell lines is at least partly due to a rapid 2- to 2.5-fold stabilization of LDLR mRNA in HepG2 cells after PMA treatment. Stabilization of LDLR mRNA in response to PMA was also observed in HH01 cells, a human hepatocyte coculture system derived from normal human liver. In both HepG2 and HH01 cells, PMA treatment induced a rapid morphological change with characteristics of cytoskeletal reorganization. The changes in morphology and stabilization of LDLR mRNA by PMA were coincident in the cell lines tested and were independent of de novo gene expression. Subcellular fractionation studies indicated that LDLR polysomes may be associated with the cytoskeleton in HepG2 cells. Disruption of the action cytoskeleton but not microtubules abrogated stabilization of LDLR mRNA by PMA. These data suggest that components of the actin cytoskeleton are involved in the regulated decay of LDLR mRNA in some human liver cell culture systems.

Actins↗
PubMed1997

Outcome of hepatobiliary scanning in neonatal hepatitis syndrome.

UNLABELLED: To evaluate the diagnostic information gained from hepatobiliary scanning in infants, we reviewed 86 consecutive infants who were < or = 4 mo old and were treated for conjugated hyperbilirubinemia at the Hospital for Sick Children in Toronto between 1985 and 1993 and who had technetium iminodiacetic hepatobiliary scanning and a percutaneous liver biopsy performed in close temporal proximity. METHODS: Retrospective reviews of hospital charts and blinded reviews of hepatobiliary scans were performed. RESULTS: There were 58 male and 28 female infants (age range, 2-124 days; mean = 65 days). Hepatobiliary scanning failed to show biliary excretion into the gastrointestinal tract in 53 of 86 patients. Forty of these 53 had extrahepatic biliary atresia. The remaining 33 patients demonstrated biliary excretion into the gastrointestinal tract; 24 of 33 had neonatal hepatitis. Among 13 of the 53 patients who had no evidence of biliary excretion and who also did not have extrahepatic biliary atresia, 8 had idiopathic neonatal hepatitis, 4 had interlobular bile duct paucity and 1 had total parenteral nutrition-associated cholestasis. In this large series, no patient with extrahepatic biliary atresia showed bile drainage on hepatobiliary scanning. Fifty percent of patients with interlobular bile duct paucity but no extrahepatic obstruction failed to show biliary excretion of radionuclide. Twenty-five percent of patients (8 of 32) with idiopathic neonatal hepatitis demonstrated no biliary excretion. Hepatocellular extraction was examined by semiquantitative analysis in the nondraining, nonbiliary atresia patients (12 of 53). Four of these 12 patients demonstrated poor liver extraction. Three patients had idiopathic neonatal hepatitis, and one had bile duct paucity. Therefore, four of eight neonatal hepatitis patients had normal extraction, suggesting that poor versus good liver hepatocyte clearance cannot accurately identify neonatal hepatitis. CONCLUSION: Hepatobiliary scanning requires cautious interpretation. Nondraining scans may indicate severe neonatal hepatitis or the presence of interlobular bile duct paucity.

Alagille Syndrome↗
PubMed1996

Excess zinc associated with severe progressive cholestasis in Cree and Ojibwa-Cree children.

BACKGROUND: High hepatic copper concentrations have been reported in several liver disorders. We report six Native Canadian children with severe chronic cholestatic liver disease, who had excess hepatic copper and zinc. METHODS: The children, aged 22 months to 8 years, came from northern Ontario, Canada. All were referred for possible liver transplantation because of end-stage liver disease. We examined explanted liver samples (or liver biopsy material in one case) by scanning transmission electronmicroscopic (STEM) X-ray elemental microanalysis and atomic absorption spectrophotometry. Samples from four controls (two with no liver pathology, one with biliary atresia, and one with Wilson's disease) were also analysed by atomic absorption spectrophotometry. FINDINGS: The explanted livers showed similar distinctive signs of advanced biliary cirrhosis, and on electronmicroscopy there were dense deposits in enlarged lysosomes and in cytoplasm. Hepatic copper concentrations were many times higher in the five patients with measurements (47.6-56.9 microgram/g dry weight) than in two samples of normal control liver tissue (2.3 and 2.9 microgram/g). Similarly, hepatic zinc concentrations were many times higher in the patients than in controls (104-128 vs 1.9-3.2 microgram/g dry weight). INTERPRETATION: The excess copper may be due to chronic cholestasis but the excess zinc is unexplained. Since three of the patients are related (shared grandparents), a genetic disorder of metal metabolism is possible, but we cannot exclude environmental factors.

Child↗