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Biomedical subjects

C J Chandler

Publications and source records attributed to C J Chandler.

36 records · Page 2Linked to original sources
PubMed1990

Haemodynamic alterations in anaesthetized and acutely intoxicated newborn piglets.

1. We investigated the haemodynamic alterations in 2-day old anaesthetized piglets after acute ethanol intoxication using the microsphere technique. 2. After ethanol infusion, mean arterial blood pressure (MABP) decreased by 6%, cardiac output (CO) decreased by 26%, heart rate (HR) increased by 20% and systemic vascular resistance (SVR) increased by 36%. 3. Arterial perfusion to the kidneys, gastrointestinal (GI) organs and carcass decreased by 39%, 34% and 26%, respectively. 4. In piglets pretreated with 4-methylpyrazole (4-MP), an alcohol dehydrogenase inhibitor, the decrease in MABP and increase in HR were not observed after ethanol infusion, but the reduction in CO and increase in SVR were maintained. 5. Arterial blood flow to the GI organs and carcass, but not the kidneys, remained at significantly reduced levels. 6. These observations indicate that ethanol can adversely affect CO and arterial perfusion to GI and musculoskeletal structures while metabolites of ethanol, such as acetaldehyde, affect MABP and HR. Therefore, the clinical effects observed after acute ethanol intoxication in neonates may vary with their rate of ethanol metabolism.

Alcoholic Intoxication↗
PubMed1990

The synthesis, stability and biological activity of bis-intercalating bis-daunomycin hydrazones.

The synthesis of a series of bis-daunomycin hydrazones (5a-g)--all moderately stable at 37 degrees C, pH 6.8, with a half-life of approximately 30 h--is reported. Under a pulse exposure of 2 h they exhibited growth inhibition of mouse L1210 cells, and were 2-3 fold more active than daunomycin. Under continuous exposure growth inhibition conditions with human colon cell lines (HT-29 and HCT-8) they hydrolysed to daunomycin and a partially hydrolysed mono-derivative of daunomycin, and there was no apparent increase in activity over that of the parent anthracycline. Their rate of hydrolysis was observed to increase rapidly with decreasing pH.

Animals↗
PubMed1989

Kinetic disposition of ethanol in the neonatal piglet and hemodynamic effects in the presence and absence of 4-methylpyrazole.

The hemodynamic effects of acute ethanol intoxication and the kinetic disposition of ethanol are reported for the first time in neonatal piglets under nitrous oxide anesthesia. Two hours after a single dose of ethanol (1.4 g/kg), blood pressure decreased from 76 +/- 4 to 71 +/- 4 mm Hg (p less than 0.05) and heart rate increased from 194 +/- 10 to 227 +/- 8 beats/min (p less than 0.05; means +/- SE). By 5 hr, blood pressure dropped to 67.5 +/- 4 mm Hg and heart rate increased to 239 +/- 8 beats/min. In piglets pretreated with 4-methylpyrazole, an alcohol dehydrogenase inhibitor, there was a transient increase in blood pressure (p less than 0.05) and a decrease in heart rate (p less than 0.05) immediately after the end of the ethanol infusion. However, the hemodynamic alterations observed 2 hr after ethanol treatment alone were prevented with 4-methylpyrazole. These findings indicate that ethanol metabolites play a significant role in hemodynamic alterations observed after acute ethanol intoxication. The mean ethanol metabolic rate derived from plasma data was 94 +/- 9 mg/liter/hr. This corresponded to an apparent Km of 68 +/- 3 mg/liter and a Vm of 123 +/- 11 mg/liter/hr. The Vd was 0.966 +/- 0.031 liter/kg. The metabolic rate for ethanol, derived from plasma data, correlated with in vitro alcohol dehydrogenase activity at pH 7.4 and 25 and 37 degrees C. The optimum pH for hepatic alcohol dehydrogenase activity was 9.9.

Animals↗
PubMed1989

Folate absorption in alcoholic pigs: in vitro hydrolysis and transport at the intestinal brush border membrane.

We used the miniature pig to evaluate the effect of ethanol ingestion on the hydrolysis of pteroylpolyglutamate and on the uptake of pteroylmonoglutamate (PteGlu) by the intestinal brush border membrane, processes that are required for folate absorption. After feeding ethanol or sucrose at 60% of calories for 11 mo, the uptake of PteGlu by jejunal brush-border-membrane vesicles was similar in both groups of animals. Jejunal brush border pteroylpolyglutamate hydrolase was decreased by one-half in the ethanol-fed group. Jejunal brush-border-membrane fluidity, measured by fluorescence polarization, was similar in both groups. Acute exposure of the jejunal vesicles to ethanol increased membrane fluidity and decreased hydrolase activity but had no effect on PteGlu transport. Inhibition of jejunal folate hydrolase by chronic exposure to ethanol may be an early effect in the pathogenesis of folate malabsorption and deficiency in chronic alcoholism.

Alcoholism↗
PubMed1988

Fast atom bombardment mass spectrometry of some anthracycline and bisanthracycline derivatives.

Fast atom bombardment (FAB) mass spectra of daunomycin, four of its derivatives, seven bisanthracyclines and three mixed-functional daunomycin-acridine derivatives are reported. These anthracyclines all exhibited their expected [MH]+ ions and peaks corresponding to the fragmentations which are characteristic of the anthracycline moiety, and in addition the spectra showed enhanced [MH + n]+ (n = 1-4) ions which were attributed to reductive processes occurring in the liquid matrix under FAB conditions. Daunomycin was also observed to form a dimeric cluster ion [M2H]+ together with associated reduced ions under FAB conditions. We have found that FAB mass spectrometry is an ideal method for the qualitative analysis of large, non-volatile derivatives of anthracyclines.

Antibiotics, Antineoplastic↗
PubMed1988

The DNA-association and biological activity of a new bis(14-thiadaunomycin).

A new bis-daunomycin has been synthesized and characterized by 13C-n.m.r. and reversed-phase h.p.l.c. The compound was found to be highly self-associated in aqueous solution and to bis-intercalate into DNA with a residence time about 200-fold greater than the parent compound daunomycin. Although the bis-daunomycin and its parent congener were taken up by V79 Chinese hamster ovarian leukaemia cells to similar extents, the cytotoxicity of the former was considerably lower. The possible in vivo relationship between DNA binding affinity and cytotoxicity is discussed.

Animals↗
PubMed1986

Intracellular pteroylpolyglutamate hydrolase from human jejunal mucosa. Isolation and characterization.

Human jejunal intracellular pteroylpolyglutamate hydrolase was purified 30-fold from intestinal mucosa. The apparent molecular weight of the enzyme was 75,000 by Sephadex G-200 gel filtration, and the isoelectric point was at pH 8.0. The enzyme was maximally active at pH 4.5 and was unstable at increasing temperatures. Intracellular pteroylpolyglutamate hydrolase cleaved both terminal and internal gamma-glutamate linkages. In contrast, brush-border pteroylpolyglutamate hydrolase catalyzed the hydrolysis of only terminal gamma-glutamate linkages. The intracellular enzyme showed greatest affinity for the complete folic acid molecule with longer glutamate chains. Subcellular fractionation studies showed the intracellular enzyme was localized in lysosomes. These data show that the properties of human jejunal intracellular pteroylpolyglutamate hydrolase are distinct from those of the brush-border enzyme but are similar to the properties of intracellular pteroylpolyglutamate hydrolase described in other tissues.

Carboxypeptidases↗
PubMed1986

Pteroylpolyglutamate hydrolase from human jejunal brush borders. Purification and characterization.

Pteroylpolyglutamate hydrolase was solubilized with Triton X-100 from human jejunal mucosal brush borders and purified approximately 5,000-fold using organomercurial affinity chromatography, DEAE-cellulose chromatography, and gel filtration. The apparent molecular weight of the purified enzyme in the Triton micelle was estimated as 700,000 using Bio-Gel A-1.5m gel filtration. Sodium dodecyl sulfate/urea-polyacrylamide gel electrophoresis followed by Coomassie stain demonstrated two polypeptide bands at 145,000 and 115,000 daltons. The purified enzyme had an isoelectric point of 7.2, was maximally active at pH 5.5, and was stable above pH 6.5 and at temperatures up to 65 degrees C for at least 90 min. Human jejunal brush-border pteroylpolyglutamate hydrolase is an exopeptidase which liberated [14C]Glu as the sole labeled product of PteGlu2[14C]Glue (where PteGlun represents pteroylpolyglutamate), failed to liberate a radioactive product from PteGlu2[14C]GluLeu2, and released all possible labeled PteGlun products during incubation with Pte[14C]GluGlu6 with the accumulation of Pte[14C]Glu. PteGlu2, PteGlu3, and PteGlu7 were substrates, each with Km = 0.6 microM, whereas PteGlu was a weak inhibitor of the hydrolysis of PteGlu3 with Ki = 20 microM. Components of the pteroyl moiety, Glu, and short chain Glun in alpha or gamma linkages were not inhibitory. The enzyme was activated by Zn2+ or Co2+. The properties of brush-border pteroylpolyglutamate hydrolase are different from those described for the soluble intracellular pteroylpolyglutamate hydrolase in other species and in human mucosa, yet are consistent with previous data on the process of hydrolysis of PteGlun in the intact human intestine.

Carboxypeptidases↗
PubMed1986

Folate binding and hydrolysis by pig intestinal brush-border membranes.

The intestinal absorption of dietary polyglutamyl folate involves hydrolysis, binding, and transport at the brush-border surface of the enterocyte. We studied the binding of [3H]folic acid ([3H]PteGlu) and the hydrolysis of [14C]pteroyltriglutamate ([14C]PteGlu3) by use of brush-border vesicles prepared from pig jejunal mucosa. [3H] PteGlu associated with the vesicles was not affected by increasing the osmolarity of the incubation solution, verifying that the experiments describe binding and not transport of PteGlu. The binding of [3H]PteGlu was saturable (Kd = 0.08 microM) and pH dependent with maximal binding at pH 5.2. Binding was competitively inhibited by PteGlu3 (Ki = 0.25 microM) and 5-methyltetrahydrofolate (Ki = 0.8 microM) but was not affected by components of the PteGlu molecule. ZnCl2, MgCl2, and MnCl2 enhanced the binding capacity but not the affinity of the binding component for PteGlu. We distinguished the processes of folate binding and hydrolysis by demonstrating differences in metal ion requirements and susceptibilities to various inhibitors. In addition, the binding component and the hydrolytic enzyme had distinct affinities for PteGlu (Ki = 45 microM for PteGlu3 hydrolysis). These data demonstrate pH-dependent, specific, and saturable binding of PteGlu to the intestinal brush-border membrane and suggest that the binding component is separate from the hydrolytic enzyme.

Animals↗
PubMed1986

Clinical studies of intestinal folate conjugases.

Clinical differences between the two human intestinal mucosal folate conjugases were assessed by measurement of their activities in normal individuals and in patients with chronic diarrhea of differing causes. Intracellular folate conjugase (ICFC) was 15-fold more active than brush border folate conjugase (BBFC) in jejunal mucosa from seven obese patients undergoing elective gastric bypass surgery. The activity of ICFC was similar among normal volunteers and patients with diarrhea of unknown origin (DUO), gluten-sensitive enteropathy (GSE), inflammatory bowel disease (IBD), and the short bowel syndrome (IBD-SBS). By contrast, BBFC, sucrase, and lactase were decreased significantly in GSE, and BBFC was increased in IBD-SBS. The activity of BBFC correlated with lactase and with sucrase in the normal subjects and in patients with DUO, whereas no correlations were found with the activity of ICFC in any group. Our clinical studies confirm that ICFC and BBFC are different enzymes. ICFC is not affected by intestinal disease, whereas the activity of jejunal BBFC, like that of other brush border enzymes, is decreased by mucosal injury and is also capable of adapting to distal small intestinal disease or surgical resection.

Adult↗
PubMed1985

Comparative stability and catalytic and chemical properties of the sulfate-activating enzymes from Penicillium chrysogenum (mesophile) and Penicillium duponti (thermophile).

ATP sulfurylases from Penicillium chrysogenum (a mesophile) and from Penicillium duponti (a thermophile) had a native molecular weight of about 440,000 and a subunit molecular weight of about 69,000. (The P. duponti subunit appeared to be a little smaller than the P. chrysogenum subunit.) The P. duponti enzyme was about 100 times more heat stable than the P. chrysogenum enzyme; k inact (the first-order rate constant for inactivation) at 65 degrees C = 3.3 X 10(-4) s-1 for P. duponti and 3.0 X 10(-2) s-1 for P. chrysogenum. The P. duponti enzyme was also more stable to low pH and urea at 30 degrees C. Rabbit serum antibodies to each enzyme showed heterologous cross-reaction. Amino acid analyses disclosed no major compositional differences between the two enzymes. The analogous Km and Ki values of the forward and reverse reactions were also essentially identical at 30 degrees C. At 30 degrees C, the physiologically important adenosine 5'-phosphosulfate (APS) synthesis activity of the P. duponti enzyme was 4 U mg of protein-1, which is about half that of the P. chrysogenum enzyme. The molybdolysis and ATP synthesis activities of the P. duponti enzyme at 30 degrees C were similar to those of the P. chrysogenum enzyme. At 50 degrees C, the APS synthesis activity of the P. duponti enzyme was 12 to 19 U mg of protein-1, which was higher than that of the P. chrysogenum enzyme at 30 degrees C (8 +/- 1 U mg of protein-1). Treatment of the P. chrysogenum enzyme with 5,5'-dithiobis(2-nitrobenzoate) (DTNB) at 30 degrees C under nondenaturing conditions modified one free sulfhydryl group per subunit. Vmax was not significantly altered, but the catalytic activity at low magnesium-ATP or SO4(2-) (or MoO4(2-)) was markedly reduced. Chemical modification with tetranitromethane had the same results on the kinetics. The native P. duponti enzyme was relatively unreactive toward DTNB or tetranitromethane at 30 degrees C and pH 8.0 or pH 9.0, but at 50 degrees C and pH 8.0, DTNB rapidly modified one SH group per subunit. APS kinase (the second sulfate-activating enzyme) of P. chrysogenum dissociated into inactive subunits at 42 degrees C. The P. duponti enzyme remained intact and active at 42 degrees C.

Amino Acids↗
PubMed1985

Evidence that adrenal oestrogens are not involved in benign breast disease.

We have studied 30 women with symptomatic benign breast disease in order to test the hypothesis that in some it might be caused by an enzyme variant leading to excessive adrenal production of oestrone. Synthetic 1-24 ACTH (Synacthen) was given intramuscularly after overnight suppression with dexamethasone, during the early follicular phase of the menstrual cycle. In no subjects did plasma oestrone levels show a consistent and significant response to ACTH despite the expected consistent rises in androstenedione and cortisol. We conclude that it is unlikely that adrenal oestrogens play a significant role in the pathophysiology of even a minority of patients with benign breast disease.

Adult↗
PubMed1984

Effect of age on poly- and monoglutamyl folacin absorption in human subjects.

Intestinal folacin absorption and mucosal conjugase activities were assessed in healthy male elderly (65-83 years) and young (20-29 years) human subjects. Luminal disappearance as measured simultaneously from the perfused jejunum of 3 mumol of both [3H]pteroylmonoglutamate [( 3H]PG-1) and pteroyl-[14C]glutamyl-hexaglutamate [( 14C]PG-7) as well as the urinary recovery of each isotope and mucosal conjugase activities were compared. Luminal disappearance of either [3H]PG-1 or [14C]PG-7 was not different for the two age groups (P greater than 0.05). The average (mean +/- SD) disappearance of [3H]PG-1 was 63 +/- 12% in the elderly and 67 +/- 6% in the young groups. In contrast, the disappearance of [14C]PG-7 averaged 51 +/- 6% in the elderly and 48 +/- 3% in the young group. Urinary recovery (percent in 48 hours) was not different for either isotope when the two age groups were compared (P greater than 0.05). The urinary recovery of 3H averaged 60 +/- 21% in the elderly and 59 +/- 13% in the young in contrast to 35 +/- 22% and 35 +/- 7% for 14C recovery in the elderly and young groups, respectively. Folacin conjugase activities measured at pH 6.5 and at pH 4.5 were not different between the groups (P greater than 0.05). The findings indicate that intestinal absorption of folacin and activity of folacin conjugases are not affected by the aging process.

Adult↗
PubMed1979

ATP sulfurylase from Penicillium chrysogenum: is the internal level of the enzyme sufficient to account for the rate of sulfate utilization?

The in vivo rate of sulfate activation in Penicillium chrysogenum (wild-type strain ATCC 24791) was determined to be 0.19 +/- 0.09 mumol g(-1) (dry weight) min(-1) by the following methods. (i) The maximum growth of the organism in synthetic medium was a linear function of the initial Na(2)SO(4) concentration between 0 and 8 x 10(-4) Na(2)SO(4). The growth yield was 1.64 x 10(-2) g (dry weight) of mycelium per mumol of added sulfate, corresponding to a minimum sulfur requirement of 61 mumol/g (dry weight). Under these conditions (limiting sulfate) the minimum doubling time of P. chrysogenum in submerged culture was about 3.8 h, corresponding to a maximum exponential growth rate constant of 3.0 x 10(-3) min(-1). If all the sulfur in this mycelium passed through adenosine-5'-phosphosulfate, the rate of sulfate activation in vivo must have been 0.183 mumol min(-1) g(-1) (dry weight). (ii) In the presence of excess (35)SO(4) (2-), the total organic (35)S produced varied with the mycelial growth rate. However, until the culture approached maximum density, the product of [(growth rate constant) x (organic (35)S content)] was nearly constant at 0.24 to 0.28 mumol min(-1) g(-1) (dry weight). (iii) A sulfur-starved mycelium pulsed with 10(-4) M (35)SO(4) (2-) produced organic (35)S at a rate of about 0.10 mumol min(-1) g(-1) (dry weight) under conditions where the internal concentrations of ATP and sulfate would permit ATP sulfurylase to operate at about 70% of its V(max). Cell-free extracts of P. chrysogenum growing rapidly on excess sulfate contained 0.22 U of ATP sulfurylase per g (dry weight). Thus, in spite of the relatively low specific activity of homogeneous ATP sulfurylase (0.13 U/mg of protein, corresponding to an active site turnover of 7.15 min(-1)), the mycelial content of the enzyme was sufficient to account for the observed growth rate of the organism on inorganic sulfate as the sole sulfur source.

Biological Transport, Active↗
PubMed1978

Mechanism of the antimicrobial action of pyrithione: effects on membrane transport, ATP levels, and protein synthesis.

Pyrithione is a general inhibitor of membrane transport processes in fungi. A brief preincubation of Penicillium mycelia with pyrithione resulted in a marked decrease in the activities of a variety of independently regulated transport systems, including those for inorganic sulfate, inorganic phosphate, methylamine (actually, the NH(4) (+) permease), choline-O-sulfate, glucose, l-methionine (a specific system), and several hydrophobic l-alpha-amino acids (the general amino acid permease). The degree of inhibition at any fixed pyrithione concentration and exposure time increased as the pH of the incubation medium was decreased. This result strongly suggests that the active species is the un-ionized molecule and that pyrithione acts by collapsing a transmembrane DeltapH driving force. The degree of transport inhibition caused by a given concentration of pyrithione increased with increasing time of exposure to the inhibitor. However, exposure time and pyrithione concentration were not reciprocally related. At "low" pyrithione concentrations, transport inhibition plateaued at some finite value. This observation suggests that the fungi can detoxify low levels of the inhibitor. The concentration of pyrithione required for a given degree of growth inhibition increased as the experimental mycelial density increased. This phenomenon was consistent with the suggestion that the fungi are capable of inactivating pyrithione.

Adenosine Triphosphate↗
PubMedDate not supplied

Gamete intrafallopian transfer.

Gamete intrafallopian transfer (GIFT) is a form of assisted reproduction which may be used as an alternative to in vitro fertilization (IVF) in certain circumstances. It is undertaken as a single day-case episode and requires less embryological expertise than IVF. GIFT has been successfully carried out in a number of district general hospitals. The details of the procedure as presently undertaken in Wigan are described.

Adult↗