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Biomedical subjects

M D Burke

Publications and source records attributed to M D Burke.

At least 55 records · Page 3Linked to original sources
PubMed1994

A comparative study of constitutive and induced alkoxyresorufin O-dealkylation and individual cytochrome P450 forms in cynomolgus monkey (Macaca fascicularis), human, mouse, rat and hamster liver microsomes.

The expression of constitutive and inducible cytochrome P450 forms was measured in cynomolgus monkey liver and compared with man, rat, mouse and hamster. Four alkoxyresorufin O-dealkylation (AROD) activities widely used as indicators of P450 induction were measured: methoxyresorufin O-demethylation (MROD), ethoxyresorufin O-deethylation (EROD), pentoxyresorufin O-dealkylation (PROD) and benzyloxyresorufin O-dealkylation (BROD). In monkeys there were no sex-differences in untreated, phenobarbitone (PB)- or beta-naphthoflavone (BNF)-treated animals in AROD activities, or in individual P450 proteins detected by immunoblotting. Basal MROD and EROD activities varied by less than 7-fold between the five species, but the comparative pattern of basal MROD, EROD, PROD and BROD activities (the "MEPB profile") was very species-specific, with monkeys being similar to rats but different from man, mouse and hamster. The induction of AROD activities by PB and BNF was also highly species-specific. Monkeys expressed constitutive proteins immunorelated to the CYP1A, CYP2A, CYP2B, CYP2C and CYP3A sub-families (human CYP2A6 cross-reacted with the anti-rat CYP2B1 antibodies used, and so CYP2A and CYP2B forms could not be separately identified in the monkey). Single constitutive immunoblot bands were identified in monkey for CYP1A (54 kDa), CYP2A/CYP2B (51 kDa) and CYP3A (51 kDa), respectively, but two strong (51 and 52 kDa) plus two weak (49 and 49.5 kDa) bands were shown for CYP2C. Human liver expressed CYP1A2 (54 kDa), CYP2A6 (51 kDa), CYP3A4 (50.5 kDa) and three CYP2C9-immunorelated protein bands (48, 50 and 54 kDa). In monkeys BNF induced the 54 kDa CYP1A protein and CYP1A-dependent MROD, EROD and PROD activities (18-, 15- and 6-fold increases in activity, respectively), whereas PB strongly induced the 51 kDa CYP2A/CYP2B protein but did not induce PROD activity. PB also induced non-constitutive CYP2A/CYP2B protein bands at 49 and 52 kDa in some monkeys. BROD activity was induced less that four-fold by either PB or BNF in monkeys. In conclusion, cynomolgus monkeys expressed a range of constitutive CYP1A, CYP2A or CYP2B, CYP2C and CYP3A proteins similar to man, and a range of AROD monooxygenase reaction rates similar to both man and rat, but the basal MEPB profile of AROD activities in monkeys was more similar to rat than to man. MROD and EROD were good measures of CYP1A induction by polycyclic aromatic hydrocarbons in cynomolgus monkeys, but neither PROD nor BROD were indices of CYP2B induction by PB.

Adult↗
PubMed1994

Immunohistochemical localization of glutathione S-transferases in sarcomas.

The glutathione S-transferases (GSTs) are a multi-gene family of enzymes involved in detoxifying electrophilic compounds and the expression of these enzymes in tumours has been proposed as one important mechanism of anti-cancer drug resistance. In this study, the localization of the major cytoplasmic forms of GST has been studied in soft tissue sarcomas by immunohistochemistry. The alpha, mu, and pi forms of GST were identified in 59, 68, and 51 per cent of tumours, respectively. In addition, GST pi immunoreactivity was consistently identified in fibroblasts in adjacent non-neoplastic tissue. The presence of specific forms of GST in soft tissue sarcomas may contribute to the drug resistance frequently observed in these tumours.

Adult↗
PubMed1994

Cytochrome P450 expression in oesophageal cancer.

The cytochrome P450 superfamily of enzymes play a central part in the metabolism of carcinogens and anti-cancer drugs. The expression, cellular localisation, and distribution of different forms of P450 and the functionally associated enzymes epoxide hydrolase and glutathione S-transferases have been investigated in oesophageal cancer and non-neoplastic oesophageal tissue using immunohistochemistry. Expression of the different enzymes was confined to epithelial cells in both non-neoplastic samples and tumour samples except the CYP3A was also identified in mast cells and glutathione S-transferase pi was present in chronic inflammatory cells. CYP1A was present in a small percentage of non-neoplastic samples but both CYP2C and CYP3A were absent. Epoxide hydrolase was present in half of the non-neoplastic samples and the different classes of glutathione S-transferase were present in a low number of samples. In carcinomas CYP1A, CYP3A, epoxide hydrolase, and glutathione S-transferase pi were expressed in at least 60% of samples. The expression of glutathione S-transferases alpha and mu were significantly less in adenocarcinoma compared with squamous carcinoma.

Adenocarcinoma↗
PubMed1994

Bromobenzene detoxification in the human liver-derived HepG2 cell line.

1. The applicability of the human hepatoma cell line, HepG2, as a cell culture model for studying xenobiotic liver toxicity has been investigated using the well-characterized hepatotoxic chemical, bromobenzene. 2. Bromobenzene caused a concentration- (0-10 mM) and time-dependent (0-180 min) decrease in HepG2 cell viability. The degree of toxicity was dependent upon the culture medium composition and the state of cell growth. Toxicity in Modified Earle's and Williams' E Media was maximal at 7 days growth compared with 3 and 10 days, and was greater in Williams' than in Earle's medium. Toxicity in Dulbecco's medium was apparent only at 10 days growth and was less than the maximum toxicity in the other media. 3. Bromobenzene was detoxified by epoxide hydrase. The question of metabolic activation by P450 remained unresolved, but any involvement of P450 was by forms not inhibited by ketoconazole. 4. The mechanism of bromobenzene toxicity did not appear to involve lipid peroxidation, depletion of reduced glutathione, calcium-mediated proteolysis or metabolic activation by prostaglandin synthetase, but may have involved direct solvent-induced cell damage. 5. This study demonstrates the potential usefulness of HepG2 cells in toxicity testing and highlights the importance of standardizing culture conditions.

Antipain↗
PubMed1993

Cytochrome P450 2C9 is responsible for hydroxylation of the naphthoquinone antimalarial drug 58C80 in human liver.

2-(4-t-Butylcyclohexyl)-3-hydroxy-1,4-naphthoquinone (58C80) is an experimental naphthoquinone antimalarial drug which undergoes extensive alky hydroxylation in man. By means of purification, N-terminal amino acid sequencing and inhibition by antibodies and sulfaphenazole, we have identified the form of cytochrome P450 primarily responsible for 58C80 hydroxylation in human liver, P450hB20-27, to be a member of the P450 2C9 subfamily. P450hB20-27 is a low-spin haemoprotein with molecular mass 54 kDa. 58C80 hydroxylation in human liver microsomes was dependent on either NADPH or NADH, with the activity supported by NADH being 35% of that supported by NADPH. With purified P450hB20-27 cytochrome b5 stimulated the NADH-dependent activity 8-fold but inhibited the NADPH-dependent activity by 30%. 58C80 is a novel substrate structure for human P450 2C and these results significantly broaden the range of drugs which have been directly shown (i.e. using a purified enzyme as opposed to expressed cDNA) to be metabolized by human P450 2C forms that are incontrovertibly expressed in human liver in vivo.

Amino Acid Sequence↗
PubMed1993

The effects of inducing agents on cytochrome P450 and UDP-glucuronyltransferase activities in human HEPG2 hepatoma cells.

Selective induction in vitro of cytochrome P450-dependent mixed-function oxidase (MFO) and UDP-glucuronyltransferase (GT) activities was observed in the human HepG2 hepatoma cell line. 1,2-Benzanthracene (BA) induced MFO O-dealkylation activities for ethoxyresorufin, methoxyresorufin and benzyloxyresorufin, whereas phenobarbitone (PB) selectively induced pentoxyresorufin O-dealkylation and rifampicin (RIF) selectively induced benzyloxyresorufin O-dealkylation. Antibody inhibition experiments indicated that ethoxyresorufin and methoxyresorufin O-dealkylations were catalysed mainly by the P450 1A subfamily in untreated and BA-induced HepG2 cells, that additional unidentified P450 forms were considerably involved in methoxyresorufin and benzyloxyresorufin O-dealkylations and that the P450 2B subfamily was partially responsible for pentoxyresorufin O-dealkylation in PB-induced cells. Bilirubin GT activity was induced by PB, BA, RIF and dexamethasone, but 1-naphthol, morphine and testosterone GT activities were not induced by any of these treatments.

Benz(a)Anthracenes↗
PubMed1993

The expression of cytochrome P-450, epoxide hydrolase, and glutathione S-transferase in hepatocellular carcinoma.

BACKGROUND: Cytochrome P-450, epoxide hydrolase, and glutathione S-transferase (GST) all play a key role in the metabolism of chemical carcinogens, mutagens, and various anti-cancer drugs. All these functionally associated enzymes might be involved in both the development of hepatocellular carcinoma and in determining the anti-cancer drug sensitivity of such tumors. METHODS: The expression of two forms of cytochrome P-450 (P-450 IA and P-450 IIIA), microsomal epoxide hydrolase, and three classes of cytosolic GST (alpha, mu, and pi) have been studied immunohistochemically in human hepatocellular carcinoma. RESULTS: The hepatocellular carcinomas were characterized by a consistently high expression of epoxide hydrolase and variable expression of the cytochromes P-450 and GST. Cytochrome P-450 IA and IIIA stained in 64.5% and 41.9% of the 31 hepatocellular carcinomas studied, respectively. Epoxide hydrolase was present in all tumors, and GST types alpha, pi, and mu were identified in 48.4%, 38.7%, and 74.2% of the hepatocellular carcinomas, respectively. CONCLUSIONS: This study showed that the expression of xenobiotic-metabolizing enzymes in hepatocellular carcinoma is complex and the presence of different xenobiotic enzymes in hepatocellular carcinoma may contribute to the intrinsic drug resistance of these tumors.

Adult↗
PubMed1993

Expression of xenobiotic metabolizing enzymes in breast cancer.

We have studied the expression of different xenobiotic metabolizing enzymes in primary operable breast cancer of no special type. The expression of two forms of cytochrome P450, microsomal epoxide hydrolase, and three classes of glutathione S-transferase was investigated using immunohistochemistry. The tumours were characterized by consistent expression of microsomal epoxide hydrolase and by variable expression of the two forms of cytochrome P450 and the three types of glutathione S-transferase. Cytochrome P450 1A and cytochrome P450 3A were identified in 39 and 22 per cent of tumours, respectively. In each case, immunostaining was present only in areas of invasive carcinoma. Epoxide hydrolase was identified in 89 per cent of tumours and glutathione S-transferases pi, mu, and alpha were identified in 56, 65, and 44 per cent of tumours, respectively. Immunoreactivity for epoxide hydrolase and glutathione S-transferases was identified in both tumours and non-neoplastic breast tissue. The presence of different xenobiotic metabolizing enzymes may have a role in determining the intrinsic drug resistance of breast cancer to a variety of anti-cancer drugs, and the expression of these enzymes can readily be assessed using immunohistochemistry.

Adult↗
PubMed1993

Cytochrome P450 expression is a common molecular event in soft tissue sarcomas.

Two major xenobiotic metabolizing sub-families of cytochrome P450 (cytochrome P450 1A and cytochrome P450 3A) have been identified in soft tissue sarcomas. Cytochrome P450 1A was present in 70 per cent and cytochrome P450 3A was present in 78 per cent of tumours, respectively. A high proportion (86 per cent) of those tumours which contained cytochrome P450 1A or cytochrome P450 3A demonstrated co-expression of both sub-families. In each tumour, cytochrome P450 immunoreactivity was identified in all tumour cells and there was no intra-tumour heterogeneity. These results indicate that expression of cytochrome P450 is a common molecular event in soft tissue sarcomas and that the presence of different sub-families of cytochrome P450 has implications both for the pathogenesis and for the treatment of these tumours. Cytochrome P450 expression may influence the intrinsic drug resistance of these tumours and also provide a molecular target for anti-cancer drugs which can be activated by cytochrome P450.

Aged↗
PubMed1993

Laboratory medicine education in United States medical schools.

The Academy of Clinical Laboratory Physicians and Scientists conducted a survey of US medical schools to examine the current status of laboratory medicine education, estimate the amount of teaching time available in the curriculum, and identify the most effective teaching practices. Questionnaires were sent to department heads and course directors at 126 US medical schools. Replies were received from 120 schools (95%), 83 of which offered a total of 132 courses in laboratory medicine. Only 68 schools (57%) had required courses. Most of the elective courses (35 of 50; 70%) were in general clinical pathology. Lectures remained the most common teaching format, with or without laboratory sessions and workshops. Computer-assisted instruction was used in only 10 schools. Laboratory medicine courses were offered in all 4 years of medical school, with the majority (70 of 132; 53%) in the second year, often integrated with general pathology. Opinion was divided over the relative importance of laboratory medicine instruction in the preclinical versus clinical years.

Computer-Assisted Instruction↗
PubMed1993

Implementing the Patient Self-Determination Act.

An interdisciplinary process action team developed policies and procedures for implementation of the Patient Self-Determination Act. Specific roles were defined for all involved in the process: Admissions Office, Unit Nurse, Social Worker, Attending Physician, Medical Records, Public Affairs, Patient Administration, Legal Office, Medical Library, Medical Law and Chaplain. An educational effort provides orientation for the hospital staff and the community on development of advance directives by patients.

Advance Directives↗
PubMed1993

The effects of retinoids and terbinafine on the human hepatic microsomal metabolism of cyclosporin.

Following the observation of increased trough whole blood cyclosporin A (CyA) concentrations and reduced renal function in a patient with recalcitrant generalized pustular psoriasis treated with a combination of CyA and etretinate, the effect of vitamin A analogues on human microsomal cytochrome P450-dependent CyA metabolism was investigated in vitro. In addition, the effect of terbinafine, a new allylamine antifungal agent, was also tested. Etretinate, its major metabolite acitretin, and isotretinoin, each at a single concentration of 100 microM, inhibited total hepatic microsomal CyA metabolism to a similar extent (33-45%, compared with control values). The generation of total primary and total secondary CyA metabolites was also inhibited to a similar extent by each of the retinoids. Conversely, terbinafine was without significant effect on CyA metabolism in vitro. The results, which suggest that inhibition of hepatic CyA metabolism by retinoids may contribute to increased circulating CyA concentrations, are discussed in relation to other potential drug interactions, and to the use of etretinate in reducing the CyA administered dose.

Acitretin↗
PubMed1993

Xenobiotic metabolising enzyme expression in colonic neoplasia.

The cytochrome P450, epoxide hydrolase, and glutathione S-transferase enzyme families play an important part in the metabolism of many carcinogens and anti-cancer drugs. The expression of two forms of cytochrome P450 (P450 1A and P450 3A), epoxide hydrolase and of the alpha, mu, and pi forms of glutathione S-transferase in normal colon, colonic adenomas, and adenocarcinoma of the colon were studied by immunohistochemistry. This allowed the precise cellular site and distribution of each enzyme to be determined. Expression of all the xenobiotic metabolising enzymes studied was almost wholly confined to the epithelial cells, whether in normal, adenoma or carcinoma samples, except that cytochrome P450 3A was also identified in mast cells and glutathione S-transferase pi was also present in chronic inflammatory cells. Cytochrome P450 was present in only a small proportion of normal colon samples, whereas epoxide hydrolase and glutathione S-transferase mu were identified in about half, and glutathione S-transferase alpha and pi in most normal samples. By contrast all the enzyme forms studied were expressed in virtually all adenomas and in over half the carcinomas. These results suggest that cytochrome P450 1A and cytochrome P450 3A are more specific markers of colonic neoplasia than epoxide hydrolase or glutathione S-transferases alpha, mu, and pi.

Adenocarcinoma↗
PubMed1992

Cytochrome P450IA expression in adult and fetal human liver.

A monoclonal antibody has been produced that recognizes the cytochrome P450 form, cytochrome P450IA1, but not cytochrome P450IA2 in rats and recognizes a single protein band of similar mol. wt on immunoblots of human liver microsomes. Immunohistochemical studies have been carried out with this antibody to investigate the localization and distribution of cytochrome(s) P450 of the P450IA family in human liver. Cytochrome P450IA was identified in both adult and fetal liver and in each case it was localized predominantly to hepatocytes. In adult liver there was a heterogeneous distribution of cytochrome P450IA immunoreactivity with cytochrome P450IA mainly present in zone 3 hepatocytes of the liver acinus. Within fetal liver there was a uniform distribution of cytochrome P450IA immunoreactivity with no apparent zonal distribution. Bile duct epithelium did not show definite immunostaining for cytochrome P450IA in either adult or fetal liver.

Adult↗