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E A Roberts

Publications and source records attributed to E A Roberts.

At least 55 records · Page 3Linked to original sources
PubMed1996

Modulation of the expression of liver-specific functions in novel human hepatocyte lines cultured in a collagen gel sandwich configuration.

Liver-specific functions were examined in two novel human hepatocyte lines established from normal liver tissue (HH29, HHY41) and one human hepatoma line (Hep G2), cultured either in a collagen gel sandwich configuration or on plain plastic dishes. At confluence, levels of albumin and alpha-fetoprotein mRNA were markedly increased in all lines cultured in the collagen gel sandwich over those on plastic, while levels of mRNA for mitochondrial elongation factor (EFTU), a ubiquitous enzyme, were equivalent. Secretion of albumin by cells cultured in the collagen gel sandwich was significantly increased in parallel with the abundance of albumin mRNA. Hepatocytes in the collagen gel sandwich configuration secreted alpha-fetoprotein of a higher molecular weight in addition to alpha-fetoprotein of standard molecular weight. The three-dimensional configuration of the collagen gel sandwich enhances the expression of liver-specific functions of human hepatocytes from continuous cell lines, probably by preserving proper cellular polarity in vitro.

Carcinoma, Hepatocellular↗
PubMed1996

Cavitation at radiolucent areas on proximal surfaces of posterior teeth.

To investigate the factors that influence the probability of clinical cavitation at radiolucent areas of proximal surfaces of posterior teeth, 108 molars and premolars with varying depths of proximal radiolucency were examined clinically, after cavity preparation on the carious contiguous tooth surfaces. The data obtained were subjected to logistic regression analysis with cavitation as the dependent variable, while age, tooth type and past caries experience (DMFT and DFS) were independent variables. When proximal radiolucency was confined to the outer half of enamel, there was no cavitation, but when it extended to the amelodentinal junction and the outer and inner half of dentine, there was cavitation in 19.3, 79.1 and 100% of cases, respectively. Moreover, there was a statistically significant relationship between the probability of cavitation, depth of radiolucency and age, suggesting that these should be among the main factors considered when restorative management of a radiolucent proximal surface of a posterior tooth is contemplated.

Adolescent↗
PubMed1995

Different mechanisms of hydroxylation site selection by liver and kidney cytochrome P450 species (CYP27 and CYP24) involved in vitamin D metabolism.

A series of homologated 1 alpha-hydroxyvitamin D3 and 1,25-dihydroxyvitamin D3 molecules with one to three extra carbons in the side chain were used to examine the substrate preferences and hydroxylation site selection mechanisms of the liver vitamin D3-25-hydroxylase (CYP27) and the target cell 25-hydroxyvitamin D3-24-hydroxylase (CYP24). Cultured and transfected cell models, used as sources of these hydroxylases, gave 23-, 24-, 25-, and 27-hydroxylated metabolites which were identified by their high performance liquid chromatography and GC-MS characteristics. Lengthening the side chain is tolerated by each cytochrome P450 isoform such that 25-hydroxylation or 24-hydroxylation continues to occur at the same rate as in the native side chain, while the site of hydroxylation remains the same for the liver enzyme in that CYP27 continues to hydroxylate at C-25 and C-27 (minor) despite the two-carbon-atom extension. Somewhat surprising is the finding that C-24 and C-23 (minor) hydroxylations also do not change as the side chain is extended by as much as three carbons. We conclude that CYP24 must be directed to its hydroxylation site(s) by the distance of carbon 24 from the vitamin D ring structure and not as in CYP27 by the distance of the hydroxylation site from the end of the side chain.

Calcitriol↗
PubMed1995

Establishment of a human hepatocyte line derived from primary culture in a collagen gel sandwich culture system.

A human hepatocyte line (HHY41) was established from normal human liver tissue. This cell line was derived from a primary culture of human hepatocytes maintained between two layers of collagen gel for 4 weeks. It differs from other human hepatocyte lines in that transfection with the simian virus 40 gene was not used for cellular transformation and nonhepatocellular coculture cells were not present. HHY41 cells have proliferated freely in serum and hormone-supplemented medium after more than 1 year in continuous culture, exhibiting typical morphological characteristics of hepatocytes. HHY41 cells retain glucose-6-phosphatase activity. They also retain the ability to secrete liver-specific proteins such as albumin, transferrin, and alpha-fetoprotein. Northern blot analysis confirmed the presence of albumin mRNA. Cytochromes P450 induced by polycyclic aromatic hydrocarbons are maintained in these cells. Detection of cell surface antigens revealed that HHY41 cells express alpha 1 beta 1-integrin, which is expressed by normal hepatocytes and not by bile duct epithelial cells. High-molecular-weight cytokeratin, a marker for bile duct cells, is also absent in HHY41. Cytogenetic analysis showed hyperdiploid karyotype with a consistent deletion in the short arm of chromosome 1. HHY41 can be considered a new human hepatocyte line which retains liver-specific functions of differentiated hepatocytes. Derived from normal liver tissue, not a hepatocellular carcinoma, it provides a new model system for studying the regulation of cell growth and differentiated functions in human hepatocytes.

Adult↗
PubMed1995

Glycine conjugation of para-aminobenzoic acid (PABA): a quantitative test of liver function.

OBJECTIVE: To evaluate glycine conjugation of para-aminobenzoic acid (PABA) to the hippurated metabolites, para-aminohippuric acid (PAHA), and para-acetamidohippuric acid (PAAHA) as a quantitative liver function test in patients with liver disease. DESIGN AND METHODS: Serum concentrations of PABA and metabolites were measured by high pressure liquid chromatography in 24 controls and 50 patients with hepatobiliary disease. RESULTS: Hippurate formation was significantly decreased in all patient groups with chronic liver disease versus controls. The hippurate ratio (% hippurated metabolites formed) correlated with severity of disease, serum albumin, and factor VII concentrations. PAHA concentration was a better prognostic indicator than factor VII concentrations in patients with acute liver disease; concentrations of zero correctly predicted a poor outcome in patients with fulminant liver failure. CONCLUSIONS: Glycine conjugation of PABA may be useful as a quantitative liver function test in patients with hepatobiliary disease and as a prognostic index in patients with fulminant liver failure.

4-Aminobenzoic Acid↗
PubMed1995

The Wilson disease gene: spectrum of mutations and their consequences.

We have previously reported the cloning of a gene that encodes a copper transporting P-type ATPase (ATP7B) which is defective in Wilson disease. We have now identified in 58 WND patients, 20 new mutations as well as three of five previously published mutations: 11 small insertions and deletions, seven missense, two nonsense and three splice site mutations. Two of the mutations are relatively frequent, representing 38% of the mutations in patients of European origin. Our findings suggest a wider spectrum of age of onset than is considered typical of Wilson disease: mutations that completely disrupt the gene can produce liver disease in early childhood when Wilson disease may not typically considered in the differential diagnosis. The mutations identified provide an explanation for at least part of the wide phenotypic variation observed in Wilson disease.

Adenosine Triphosphatases↗
PubMed1995

Safety and immunogenicity of two acellular pertussis vaccines with different pertussis toxoid and filamentous hemagglutinin content in infants 2-6 months old.

The optimal composition and antigen content of acellular pertussis vaccines is not known. Two vaccines with different quantities of pertussis toxoid (10 and 20 micrograms) and filamentous hemagglutinin (5 and 20 micrograms) and identical 69 kD protein (3 micrograms) and fimbriae 2 and 3 (5 micrograms) combined with diphtheria and tetanus toxoids were compared in a randomized, double-blind study in 2,050 infants undergoing their primary immunization series at 8 centers in the US and Canada. A 6:1 increased antigen to lower antigen allocation was used; 96% of infants received 3 doses and completed the study. A 'clinically significant' local reaction was reported in 3-6% of participants after each dose. Erythema was the most common reaction occurring in 3-5% of infants after the second or third dose. A clinically significant systemic adverse reaction was reported in 28-34% of vaccinees (or vaccinated children) after each dose; fever (7-18%) and fussiness (12-17%) were most common. There were no differences in adverse events between the 2 vaccine formulations. Antibody responses were measured in 292 infants at 1 center. At 7 months, geometric mean anti-filamentous hemagglutinin antibody titers were higher in recipients of the higher antigen content vaccine (p < 0.001) whereas recipients of the lower antigen content formulation had higher anti-fimbriae antibody (p < 0.001) and agglutinin titers (p < 0.05). No differences were detected in anti-pertussis toxin or other antibody responses between the formulations. We conclude that increasing the antigen content of the acellular pertussis vaccine had a variable effect on antibody response but was not associated with increased adverse reactions.

Adhesins, Bacterial↗
PubMed1995

Haplotypes and mutations in Wilson disease.

Wilson disease is a disorder of copper transport, resulting in neurological and hepatic damage due to copper toxicity. We have recently identified > 20 mutations in the copper-transporting ATPase defective in this disease. Given the difficulties of searching for mutations in a gene spanning > 80 kb of genomic DNA, haplotype data are important as a guide to mutation detection. Here we examine the haplotypes associated with specific mutations. We have extended previous studies of DNA haplotypes of dinucleotide-repeat polymorphisms (CA repeats) in the Wilson disease region to include an additional marker, in 58 families. These haplotypes, combining three markers (D13S314, D13S316, and D13S301), are usually specific for each different mutation, even though highly polymorphic CA repeat markers have been used. Haplotypes, as well as their accompanying mutations, differ between populations. In the patients whom we have studied, the haplotype data indicate that as many as 20 mutations may still be unidentified. The use of the haplotypes that we have identified provides an important guide for the identification of known mutations and can facilitate future mutation searches.

Adenosine Triphosphatases↗
PubMed1995

Timely referral of infants with jaundice: case report.

Family physicians should see all infants between 2 and 4 weeks of age to exclude persistent jaundice. Early identification of infants with jaundice due to liver disease might lead to better outcomes not only for infants with extrahepatic biliary atresia but for all infants with neonatal liver diseases. The case illustrates the consequences of late identification.

Biliary Atresia↗
PubMed1994

Joubert syndrome with congenital hepatic fibrosis: an entity in the spectrum of oculo-encephalo-hepato-renal disorders.

Joubert syndrome is an autosomal recessive inherited condition characterized by agenesis or hypoplasia of the cerebellar vermis, retinal dystrophy, chorioretinal colobomata, oculomotor abnormalities, episodic hyperpnea, ataxia, and mental retardation. Congenital hepatic fibrosis has not previously been described in Joubert syndrome. We report two unrelated children with Joubert syndrome and hepatosplenomegaly. On histopathological examination, both had congenital hepatic fibrosis. Both were also found to have congenital medullary cystic disease of the kidneys. Joubert syndrome appears to be one of a spectrum of congenital malformation syndromes involving the central nervous system, eye, liver and kidneys.

Abnormalities, Multiple↗
PubMed1994

Caffeine biotransformation in human hepatocyte lines derived from normal liver tissue.

Caffeine biotransformation was demonstrated in three novel human hepatocyte cell lines established from normal liver tissue and cultured continuously for 19 to 30 months. Caffeine and its metabolites were identified and quantified by high performance liquid chromatography. Without induction, caffeine was metabolized to the four primary metabolites [theobromine (37X), paraxanthine (17X), theophylline (13X), 1,3,7-trimethylurate (137U)]. Under these basal conditions 137U was the predominant metabolite. The actual pattern of metabolite production was a reproducible characteristic of each line. After induction with dibenz(a,h)anthracene, the formation of 17X was increased 4-17 fold. Induction with phenobarbital did not change the metabolic profile. These human hepatocyte lines can reproduce in vitro metabolism of caffeine observed in man in vivo.

Adult↗
PubMed1994

Characterization of human hepatocyte lines derived from normal liver tissue.

Four separate continuous lines of human hepatocytes (HH01, HH02, HH09, HH25) were developed from normal liver tissue by subjecting cocultures of human hepatocytes with rat liver epithelial cells in a highly enriched medium to frequent subculturing. The addition of conditioned medium from either the human hepatoma line Hep G2 or one of these stable human hepatocyte lines (HH09) appeared to facilitate establishment of line HH25. These human hepatocyte lines have been in continuous culture for 2 to 5 yr and consist of approximately 95% human cells by analysis of cell surface antigens. Cytogenetic analysis also confirmed the human origin of these cells and showed clonal origin with abnormal ploidy. Cells in these human hepatocyte lines retain morphological features of hepatocytes by both light and electron microscopy. They also retain glucose-6-phosphatase activity and secrete proteins characteristic of hepatocytes, such as albumin, alpha-fetoprotein and transferrin. After incubation with 13 mumol/L dibenz(a,h) anthracene for 24 hr, each line had detectable activity of aryl hydrocarbon hydroxylase, ethoxyresorufin O-deethylase and methoxyresorufin O-demethylase. Thus, these human hepatocyte lines retain important differentiated characteristics of hepatocytes. Derived from normal liver tissue, they appear to be immortalized. They provide a new model system for studying human hepatocellular drug metabolism. These lines may also be useful for studying the regulation of synthesis of albumin, alpha-fetoprotein and other proteins in human hepatocytes, determining the effects of cytokines and growth factors and designing systems to effect gene transfer into human hepatocytes for the purpose of gene therapy.

Albumins↗
PubMed1994

Haplotype studies in Wilson disease.

In 51 families with Wilson disease, we have studied DNA haplotypes of dinucleotide repeat polymorphisms (CA repeats) in the 13q14.3 region, to examine these markers for association with the Wilson disease gene (WND). In addition to a marker (D13S133) described elsewhere, we have developed three new highly polymorphic markers (D13S314, D13S315, and D13S316) close to the WND locus. We have examined the distribution of marker alleles at the loci studied and have found that D13S314, D13S133, and D13S316 each show nonrandom distribution on chromosomes carrying the WND mutation. We have studied haplotypes of these three markers and have found that there are highly significant differences between WND and normal haplotypes in northern European families. These findings have important implications for mutation detection and molecular diagnosis in families with Wilson disease.

Base Sequence↗
PubMed1993

DNA markers for the diagnosis of Wilson disease.

Wilson disease is an autosomal recessive disorder of copper transport for which the basic defect is unknown. Laboratory diagnosis of Wilson disease is usually made by measuring serum ceruloplasmin concentration, urinary copper excretion, and liver copper concentration. However, discrimination between heterozygotes and patients is sometimes difficult. The gene for Wilson disease has been assigned to chromosome-13 at q14-q21. In this study, 10 markers from the 13q14-13q21 region were investigated in 12 families with a well-established diagnosis, to confirm reported linkage results. Markers from the same region were tested in two additional families, in which a sib of each index case had unclear results with conventional biochemical assays. The linkage results in this study are similar to those of Middle Eastern families, and support the hypothesis of a single disease locus. In the two families studied for diagnostic purposes, the status of 2 presymptomatic sibs was established as affected and 1 as unaffected. This study therefore shows that DNA markers can be used to discriminate between presymptomatic patients and non-affected individuals when biochemical results are equivocal, as long as an index case with Wilson disease of known status is available and markers are informative.

Adolescent↗
PubMed1993

Allelic association and linkage studies in Wilson disease.

We have studied 21 families with Wilson disease (WND), using restriction fragment length polymorphisms (RFLPs) in the 13q14.3 region, to measure linkage of these markers to the disease locus. In addition to previously described markers, we include linkage data for a newly isolated marker (D13S86) and an established marker (D13S56), which were previously not placed on the genetic map in the region of the WND locus. Our data, including those from two recombinant families, support the location of WND between the markers D13S31 and D13S59. We have examined the distribution of marker alleles at the loci studied and have found that D13S31 and D13S228, and associated microsatellite marker, show a non-random distribution on chromosomes carrying the WND mutation. The significant linkage disequilibrium indicates that these two markers must be close to the WND locus.

Alleles↗