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Biomedical subjects

W H Peters

Publications and source records attributed to W H Peters.

At least 109 records · Page 6Linked to original sources
PubMed1991

Genetic deficiency of human class mu glutathione S-transferase isoenzymes in relation to the urinary excretion of the mercapturic acids of Z- and E-1,3-dichloropropene.

Mononuclear lymphocytes were isolated from the blood of 12 individuals, who had been exposed to the vapour of the soil fumigant 1,3-dichloropropene (DCP). Western blot experiments were performed on the crude lymphocyte homogenates, using a monoclonal antibody against human hepatic glutathione S-transferase (GST) isoenzyme mu, to determine the presence or absence of mu-class isoenzymes mu and/or psi. Nine of the individuals were found to be positive for mu and/or psi, the remaining three individuals being negative. In addition, all individuals showed a positive staining on immunoblot of a protein of somewhat lower molecular mass than the hepatic standard. This protein was bound by the S-hexylglutathione affinity column, and presumably constitutes a new mu-class isoenzyme, which is not subject to genetic polymorphism. Determination of the specific activities of individual human GST isoenzymes towards Z-(cis-) and E-(trans-)-DCP demonstrated that mu-class isoenzymes show a considerably higher specific activity with Z-DCP than alpha-class or pi-class isoenzymes. In addition, mu-class isoenzymes were found to be 2- to 3-fold more active with Z-DCP than with E-DCP. Their activity towards E-DCP was similar to the specific activity of alpha-class isoenzymes. Genetic polymorphism for mu-class isoenzymes could thus be a determinant in the extent of excretion of mercapturic acids from Z- and E-DCP. The urinary excretion of Z- and E-DCP mercapturic acids and the respiratory exposure to Z- and E-DCP were determined for nine and eight phenotyped individuals, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylcysteine↗
PubMed1991

Biotransformation enzymes in human intestine: critical low levels in the colon?

Biotransformation or drug-metabolising enzymes have an important function in the detoxication of ingested toxic, carcinogenic, or tumour promoting compounds. Enzyme activity and isoenzyme composition of three biotransformation systems: glutathione S-transferase, uridine diphosphate-glucuronosyltransferase, and cytochrome P-450 were studied in normal small and large intestinal mucosa from three kidney donors. The activity of most drug-metabolising enzymes decreases slightly from proximal to distal small intestine, whereas in the mucosa of the large intestine a sharp fall in activity was observed. The isoenzyme composition for each of the three biotransformation systems changed from the small to the large intestine. Class Alpha glutathione S-transferases were not expressed in the colon, in contrast to the small intestine where both Alpha and Pi class isoenzymes are present. In addition, with monoclonal antibodies fewer protein bands for UDP-glucuronosyltransferases and cytochrome P-450 were detected in the colon. In the small intestine both isoforms P-450(4) and P-450(5) were present, whereas in the colon only reduced amounts of cytochrome P-450(4) could be visualised. For UDP-glucuronosyltransferase, 53 and 54 kDa proteins could be detected in the small intestine, but in the colon there was only weak staining of the 54 kDa band. In the normal human colon enzymes are less active and there are fewer isoenzymes present in the mucosa than in the small intestine. This implies a lower level of the detoxifying potential in the colon, which might be important in regard to the high rates of carcinogenesis in the colon.

Adolescent↗
PubMed1990

Quantification of human hepatic glutathione S-transferases.

Human hepatic glutathione S-transferase (GST) subunits were characterized and quantified with the aid of a recently developed h.p.l.c. method. In 20 hepatic tissue specimens the absolute amounts of the basic Class Alpha subunits B1 and B2, the near-neutral Class Mu subunits mu and psi and the acidic subunit pi were determined. The average total amount of GST was 37 micrograms/mg of cytosolic protein, with the Class Alpha GST being the predominant class (84% of total GSTs), and pi as the sole representative of the Class Pi GSTs present in the lowest concentration (4% of total GSTs). Large interindividual differences were observed for all subunits, with variations up to 27-fold, depending on the subunit. For the Class Alpha GST-subunits B1 and B2, a biphasic ratio was observed. The genetic polymorphism of the subunits mu and psi was confirmed by h.p.l.c. analysis, and correlated with the enzymic glutathione conjugation of trans-stilbene oxide and with Western blotting of cytosols, using a monoclonal anti-(Class Mu GST) antibody. Of the 20 livers examined, ten contained only mu, whereas the occurrence of psi alone, and the combination of mu and psi, were found in only one liver each.

Chromatography, High Pressure Liquid↗
PubMed1990

A monoclonal antibody against pig gastric H+/K(+)-ATPase, which binds to the cytosolic E1.K+ form.

Monoclonal antibodies were raised against a purified membrane fraction from hog gastric mucosa containing H+/K(+)-ATPase. The properties of one of these monoclonal antibodies (5B6) were further evaluated. On immunoblot it recognized the 95 kDa peptide of the H+/K(+)-ATPase-rich membrane fraction. The K(+)-ATPase activity was inhibited by 65% under standard assay conditions (pH 7.0). At pH 6.0 and 8.0 this enzyme activity was inhibited by 40% and 100%, respectively. The maximal inhibition in inside-out vesicles was also 65% at pH 7.0. The inhibition was uncompetitive with respect to K+ and noncompetitive with respect to ATP. Mg2(+)-ATPase activity and K(+)-dependent p-nitrophenylphosphatase activity were not influenced. The monoclonal antibody lowered the steady-state phosphorylation level at pH 6.0, 7.0 and 8.0 by 30%, 40% and 60% respectively. The rate of the K(+)-stimulated dephosphorylation step was not inhibited. These findings demonstrate that 5-B6 recognizes the E1.K+ dephosphoenzyme at the cytosolic side.

4-Nitrophenylphosphatase↗
PubMed1990

Immunodetection with a monoclonal antibody of glutathione S-transferase mu in patients with and without carcinomas.

Several monoclonal antibodies against human liver glutathione S-transferase mu were developed. One of these monoclonal antibodies, called GST-3H4 was further characterized and used in this study. In hepatic tissue, after immunoblotting, GST-3H4 strains a 27 kDa protein with a pI value of 6.2. GST-3H4 recognizes other human class-mu glutathione S-transferases, but does not detect acidic or basic glutathione S-transferases. By immunodetection with this monoclonal antibody, glutathione S-transferase mu can be demonstrated in human breast, stomach, liver, small and large intestine, mononuclear blood cells, kidney and placenta. A 100% correlation is found in the distribution of glutathione S-transferase mu when different tissues or mononuclear blood cells from the same individuals are investigated. In 62.5% of the mononuclear blood cells from controls, glutathione S-transferase mu is present. In patients with polyposis coli, breast cancer or colon cancer a similar distribution is found. Therefore no important role for glutathione S-transferase mu deficiencies in the aetiology of these diseases is suggested.

Animals↗
PubMed1990

Expression of glutathione S-transferases in normal gastric mucosa and in gastric tumors.

Glutathione S-transferases from both normal gastric mucosa and its matched gastric tumors from 10 different patients were investigated. The transferases were purified and subsequently the isoenzyme composition was studied. Glutathione S-transferase (GST)-pi was present in all specimens in large amounts. Class alpha GSTs were present in 9 out of 10 normal specimens and in six tumors. In malignant tissue, expression of GST-pi was increased at the expense of class alpha GST. In six patients, the ratio GST-pi/GST-alpha was higher in tumorous versus normal tissue. On a Western blot, using a monoclonal antibody, GST-mu was shown to be present in both normal and malignant tissue from four patients, the other six patients completely missed the enzyme in their gastric tissue. When present, GST-mu amounts to only a few per cent of total GST protein. GST-pi was quantified by densitometric analysis of Western blots, treated with a monoclonal antibody against GST-pi. Both total GST enzyme activity as well as the absolute amounts of GST-pi protein were significantly higher in the tumors, as compared to its matched normal mucosa. The importance of this overexpression of GST-pi was previously unknown. However, the frequent occurrence of this phenomenon in many refractory tumors, and as shown now also in gastric cancers, suggests a role for GST-pi in the mechanism of anti-cancer drug resistance.

Adult↗
PubMed1989

Time-dependent activity and expression of glutathione S-transferases in the human colon adenocarcinoma cell line Caco-2.

The human colon carcinoma cell line Caco-2 was examined for glutathione S-transferase (GST) composition and activity. Freshly seeded cells and cells until 4 days after confluency contain only the placental (Pi) form of glutathione transferase. Cells in culture for longer periods start to express class-Alpha GST isoenzymes. Confluent cells in culture for 20 days or longer contain up to 90% class-Alpha GST. Class-Mu GSTs are not detectable. GST activity gradually increases from 564 +/- 28 to 5381 +/- 165 nmol/min per mg of protein at day 0 and 32 after confluency respectively. With regard to GST composition, Caco-2 cells in culture for longer periods most resemble small-intestinal cells, whereas short-time cultures have characteristics of colonic cells. This cell line is very well suited for the study of both the in vitro properties and the expression of class Alpha and Pi GSTs.

Adenocarcinoma↗
PubMed1989

Cytochromes P-450 in the intestinal mucosa of man.

With monoclonal antibodies against cytochrome P-450(5) and P-450(4,5,6), 52 and 54 kDa bands are visualized in microsomes from proximal as well as distal human small intestine. These bands most probably correspond to cytochrome P-450(5) and P-450(4), respectively. This and several other cytochrome P-450 related proteins are present in hepatic microsomes from the same patient. In both hepatic and intestinal microsomes from this patient cytochrome P-450(8) is hardly detectable. In contrast to the small intestine and liver, large intestinal tissue from several other patients does not contain cytochrome P-450(5). Here the 54 kDa isoenzyme, possibly cytochrome P-450(4), is most prominent. Earlier we found a higher content of total cytochrome P-450 in the proximal as compared to the distal small intestine. A similar distribution is now found with regard to aldrin epoxidase activity.

Cytochrome P-450 Enzyme System↗
PubMed1989

Human intestinal glutathione S-transferases.

Cytosolic glutathione S-transferases were purified from the epithelial cells of human small and large intestine. These preparations were characterized with regard to specific activities, subunit and isoenzyme composition. Isoenzyme composition and specific activity showed little variation from proximal to distal small intestine. Specific activities of hepatic and intestinal enzymes from the same patient were comparable. Hepatic enzymes were mainly composed of 25 kDa subunits. Transferases from small intestine contained 24 and 25 kDa subunits, in variable amounts. Colon enzymes were composed of 24 kDa subunits. In most preparations, however, minor amounts of 27 and 27.5 kDa subunits were detectable. Separation into isoforms by isoelectric focusing revealed striking differences: glutathione S-transferases from liver were mainly basic or neutral, enzymes from small intestine were basic, neutral and acidic, whereas large intestine contained acidic isoforms only. The intestinal acidic transferase most probably was identical with glutathione S-transferase Pi, isolated from human placenta. In the hepatic preparation, this isoform was hardly detectable. The specific activity of glutathione S-transferase showed a sharp fall from small to large intestine. In proximal and distal colon, activity seemed to be about equal. In the ascending colon there might be a relationship between specific activity of glutathione S-transferases and age of the patient, activity decreasing with increasing age.

Adult↗
PubMed1989

Auxiliary liver transplantation in jaundiced rats with UDP-glucuronyltransferase deficiency and defective hepatobiliary transport.

In this study auxiliary liver transplantation (ALT) has been tested as a means of correcting the UDP-glucuronyltransferase deficiency in Gunn rats and the UDP-glucuronyltransferase deficiency and impaired hepatobiliary bilirubin transport in double mutant rats. In both groups serum bilirubin normalized and remained low until the end of the study at 12 weeks after transplantation in 4 out of 6 rats. Excretion of 99mTc-HIDA in non-transplanted double mutants was considerably slower than in Gunn rats (kel 0.9 x 10(-3) versus 4.3 x 10(-3) s-1). HIDA excretion by transplants in double mutants and Gunn rats was about equal (kel 1.6 x 10(-3) and 1.1 x 10(-3) s-1). Experiments with bile duct-cannulated transplants showed that in double mutants bile flow, bile acid and bilirubin excretion was 2-4 times higher than in Gunn rats. This study shows that auxiliary liver transplants can conjugate and excrete bilirubin when one of these or both functions are lacking in the recipient's liver.

Animals↗
PubMed1989

Glutathione S-transferases in normal and cancerous human colon tissue.

In order to evaluate the role of the placental form of glutathione S-transferase (GST-Pi) as a tumour marker, activity and composition of GSTs from human colon were investigated. GSTs were purified from normal colon mucosa and from colonic tumours by affinity chromatography on glutathione-agarose. After SDS-PAGE or isoelectric focusing these purified preparations revealed only one band that comigrated with GST-Pi from human placenta. A monoclonal antibody (mAb) very specific for GST-Pi was developed and characterized. On immunoblot this mAb stains purified GST from normal and diseased colon tissue. GST activity was significantly higher in most cancerous (247 +/- 38 nmol/min/mg protein; n = 7), compared with the corresponding normal tissues (171 +/- 18 nmol/min/mg protein; n = 7). In colon from patients without large bowel malignancies GST-Pi is also by far the most prominent isoform detectable. In conclusion, both normal and tumorous colon tissue predominantly express GST-Pi and therefore GST-Pi is not suitable as a tumour marker for colonic carcinomas. However, the increased GST-Pi levels in colonic tumours could possibly contribute to the relatively high resistance to anti-cancer drugs.

Biomarkers, Tumor↗
PubMed1989

Glutathione S-transferase, cytochrome P450, and uridine 5'-diphosphate-glucuronosyltransferase in human small intestine and liver.

In humans, data on biotransformation enzymes in the intestine, and to a lesser extent in the liver, are rather scarce. Much knowledge about these enzymes is, therefore, obtained from animal studies. We were able to examine both small intestinal and hepatic tissue from a kidney donor and performed a systematic study on enzyme contents and distribution in these organs. In the small intestine the longitudinal distribution of cytochrome P450, glutathione S-transferase, and bilirubin uridine 5'-diphosphate (UDP)-glucuronosyltransferase declined from duodenum to ileum. Activity of 4-nitrophenol- and 4-methylumbelliferone UDP-glucuronosyltransferase increased or remained constant, respectively. Total and specific activity of most enzymes was much higher in the liver, except for bilirubin UDP-glucuronosyltransferase and glutathione S-transferase, where the small intestine contained 28.6% and 7.4% of total hepatic activity, respectively. The relatively great amount of bilirubin UDP-glucuronosyltransferase activity in the small intestinal mucosa of this patient, who probably suffered from Gilbert's syndrome, could indicate that under pathological conditions intestinal metabolism may contribute significantly to the clearance of bilirubin. With a monoclonal antibody, UDP-glucuronosyltransferase isoforms were immunodetectable in microsomes. In the liver, two bands, one of 57 kilodaltons and one in between 53 and 54 kilodaltons, were seen. In the proximal small intestine two isoforms (53 and 54 kilodaltons) were detected. However, in the distal small intestine where bilirubin UDP-glucuronosyltransferase activity was low, only one isoform (54 kilodaltons) was seen. This may indicate that bilirubin UDP-glucuronosyltransferase activity is correlated with the 53-kilodalton isoform. The presence of multiple UDP-glucuronosyltransferase isoforms in humans, similar to that described before in the rat, is further established by this study.

Adolescent↗
PubMed1988

The serum amino acid spectrum of insulin-dependent diabetics and controls from Ethiopia.

The serum amino acid spectrum was examined in healthy men and insulin-dependent diabetics from Ethiopia. Comparison of serum amino acids of controls from Gondar with Ethiopians after adaptation to a free European diet revealed a marginal low protein nutrition, but not the characteristic changes of malnutrition or experimental starvation. There was no apparent nutritional deficiency of sulphur-containing amino acids in Ethiopians. Insulin-dependent diabetics showed significantly elevated serum levels of BCAA indicating an accelerated protein catabolism in recent-onset insulin-deficient patients and known diabetics respectively, most of them in poor metabolic condition. Serum glutamine levels were reduced, suggesting a considerable renal contribution to the hyperglycaemia/glucosuria of diabetics. The data may be best explained by the low residual insulin secretion at diabetes onset or by the poor degree of metabolic control of known Ethiopian diabetics.

Adult↗