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Biomedical subjects

J L Mitchell

Publications and source records attributed to J L Mitchell.

At least 19 recordsLinked to original sources
PubMed1992

Recovery of ornithine decarboxylase activity after inhibition with alpha-difluoromethylornithine.

alpha-Difluoromethylornithine is an effective inhibitor of polyamine biosynthesis because of its specificity for ornithine decarboxylase and the fact that its attachment to this enzyme is considered to be irreversible. We have found, however, that ornithine decarboxylase inactivated with this inhibitor in intact cells, as well as purified enzyme inactivated in vitro, both are capable of releasing this inhibitor and recovering enzyme activity. This reactivation can be initiated by freezing of inactivated enzyme samples in the presence of reducing agents at -7 or -20 degrees C and can be partially induced at 37 degrees C. These results reveal an unexpected lability of this enzyme-inhibitor complex that needs to be considered in future experimental designs.

Animals↗
PubMed1992

Abnormal accumulation and toxicity of polyamines in a difluoromethylornithine-resistant HTC cell variant.

Mammalian cells possess an inducible, active polyamine transport system that is stringently regulated by feedback controls. This study provides evidence that DH23b cells, which were initially selected from the rat hepatoma HTC line for overproduction of ornithine decarboxylase, demonstrate an abnormality in the regulation of polyamine transport. Exposure of these cells to micromolar levels of spermidine or spermine resulted in inhibition of protein synthesis and eventual cell lysis. These effects were not due to by-products of polyamine oxidation by serum oxidases as neither inhibition of protein synthesis nor cell lysis was mitigated by aminoguanidine, reduced glutathione, dithiothreitol, or catalase. Although the polyamine transport system in the DH23b cells has the same Km and Vmax as that in the parental HTC line, the variant cells accumulated abnormally high levels of both spermidine (8-times normal) and spermine (4-times normal). In the HTC line, however, transport of both polyamines as well as putrescine was feedback inhibited within approx. 3 h, while in the variant cells uptake was not diminished by 12 h and terminated only with cell lysis. The DH23b cells appear to lack the normal mechanism responsible for feedback control of active polyamine incorporation. This defect provided the opportunity to manipulate intracellular levels of spermidine from 30 to approx. 800% of normal, allowing the demonstration that cellular protein synthesis is as sensitive to spermidine levels as previous in-vitro studies had suggested.

Animals↗
PubMed1992

Feedback repression of polyamine uptake into mammalian cells requires active protein synthesis.

Two mammalian cell lines, rat hepatoma (HTC) and Chinese hamster ovary (CHO), were fed 10 to 50 microM spermidine while changes were monitored in intracellular polyamine levels and spermidine uptake activity. Normal feedback control preventing excessive polyamine uptake was found to be completely blocked by the addition of inhibitors of protein synthesis at the time of polyamine exposure. Under these conditions the cells accumulated abnormally high, toxic concentrations of spermidine. Further, continuous protein synthesis was needed to maintain repression of polyamine transporter proteins that had been inhibited previously by normal or elevated intracellular polyamines. These results suggest that a major factor in the regulation of polyamine uptake is the rapid, reversible inactivation of existing polyamine carrier molecules by an unstable protein whose synthesis is stimulated by intracellular polyamines.

Animals↗
PubMed1992

Measuring care and dependency of elderly people in residential homes: an unreliable exercise.

OBJECTIVES: to determine the correlation between a scale measuring the amount of care required by people in rest homes and two scales, the Booth scale and the composite scale which measure dependency. To examine the scales for retest reliability. SUBJECTS: one hundred and twenty-five people aged 47 to 98 years, median age 84 years, living in seven rest homes in Dunedin. METHOD: subjects were assessed using the care, Booth and composite scales and the assessment repeated after four weeks. RESULTS: the care scale test-retest reliability based on rank order correlates was 0.86. For the physical care items retest reliability was high but supervision and the management of behavioural problems showed poor retest reliability. Correlation between the care scale and the two dependency scales was only moderate (composite scale 0.64, Booth scale 0.55). Behavioural problems and the time for supervision were assessed on the care scale only and it was these components which accounted for most of the poor correlation between the care and dependency scales. CONCLUSION: the time required for the management of behavioural problems and supervision for some people in rest homes was considerable. Measurement of this was difficult and made assessment of individual residents in homes unreliable.

Activities of Daily Living↗
PubMed1991

Stable ornithine decarboxylase in a rat hepatoma cell line selected for resistance to alpha-difluoromethylornithine.

Ornithine decarboxylase (ODC) is extremely unstable in mammalian cells. This unusual characteristic facilitates rapid fluctuations in the activity of this enzyme in response to variations in its biosynthesis. Unfortunately, very little is known about the mechanism or regulation of this ODC-specific proteolytic pathway. This study describes the production and characterization of a variant of the rat hepatoma HTC cell line that is strikingly deficient in this pathway. This cell variant was induced by selection for growth in stepwise increasing concentrations (up to 10 mM) of the irreversible ODC inhibitor, alpha-difluoromethylornithine (DFMO). Resistance to this inhibitor appears to result from a combination of elevated (10X) ODC biosynthesis and inhibited degradation, producing greater than a 2000-fold increase in the level of ODC protein. In these variant cells (DH23b) inhibition of protein synthesis by cycloheximide did not result in rapid loss of enzyme activity or ODC protein determined by radioimmunoassay. Pulse-chase studies with [35S]methionine confirmed that this enzyme was not preferentially degraded, even when spermidine was added to the media. ODC purified from the variant cells was found to be identical to the control cell enzyme in size, isoelectric point, substrate binding kinetics, and sensitivity to the inhibitor DFMO. Also, as in the control cells, a major fraction of the ODC molecules extracted from DH23b cells was shown to be phosphorylated on a serine residue. The inability to detect physical or kinetic differences between the parent and the variant cell ODC suggests that the unusual stability of ODC in this cell is associated with a defect in a cellular mechanism for ODC-specific degradation.

Animals↗
PubMed1991

Ornithine decarboxylase: a biochemical marker of repair in damaged tissue.

The purpose of this study was to: (a) determine the temporal pattern of expression of ornithine decarboxylase (ODC), an established marker of cells engaged in proliferation and differentiation, during repair of traumatized skeletal muscle, and (b) evaluate ODC as a biochemical marker for indexing the extent and rate of repair of traumatized skeletal muscle in response to therapeutic agents. Adult female Wistar rats weighing 240 to 280 grams were anesthetized and injected with 100 ul of 2% lidocaine into the right or left anterior tibialis muscle to induce a localized injury. The animals were treated 1 hour post-injection and every 12 hours for five minutes up to the time of sacrifice using ultrasonic irradiation at 1.5 watts/cm2, sham-irradiation, or no treatment. An analysis of variance for repeated measures and a Tukey's post hoc test were used to determine the significance of treatment effects. Animals irradiated with ultrasound demonstrated an accelerated pattern of change in ODC activity at 24 hours (P less than 0.001), 30 hours (P less than 0.003), and at 48 hours (P less than 0.002) compared to control and sham-irradiated animals at these time intervals. Irradiated animals also demonstrated less edema at 48 hours compared to sham-irradiated and control animals (P less than 0.003). These findings suggest that ODC is a useful biochemical marker for determining the extent and rate of tissue repair in traumatized skeletal muscle, and it provides a temporal and quantifiable parameter for evaluating the efficacy of therapeutic agents used to treat damaged skeletal muscle.

Animals↗
PubMed1991

So you want to go freelance?

Freelance working may seem an attractive alternative when faced with redundancy. Jack Lamport Mitchell discusses the factors which could affect the success or failure of any professional worker contemplating such a move.

Humans↗
PubMed1991

The quest for stress management.

Although much has been written about the debilitating effects of stress at work and courses designed to help staff understand and cope with the problem proliferate, many managers still have to be convinced that stress needs to be tackled--if only because it eats into profits. Occupational psychologist Jack Lamport Mitchell examines two new developments designed to raise stress awareness.

Humans↗
PubMed1990

Conformational changes in ornithine decarboxylase enable recognition by antizyme.

Rapid, polyamine-induced degradation of mammalian ornithine decarboxylase (L-ornithine carboxy-lyase, EC 4.1.1.17) (ODC) is though to be controlled by the availability of a small, ODC-binding protein termed antizyme. In this study we have investigated the ability of antizyme to bind ODC protein in various altered physiological states. In particular, cold, NaCl, spermidine and deprivation of coenzyme and substrate enhance enzyme-antizyme complex formation and are all found to promote ODC homodimer dissociation. Conversely, conditions that maintain the active ODC homodimer state prevent antizyme binding and inactivation of ODC. Further, covalent modification of ODC near its active site by difluoromethylornithine or phosphate also increases its sensitivity to antizyme. These results suggest that the initial signal in ODC degradation may actually be a subtle conformational change in the enzyme that enables antizyme to bind to the enzyme and may subsequently facilitate its degradation.

Animals↗