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D J Thomas

Publications and source records attributed to D J Thomas.

At least 145 records · Page 8Linked to original sources
PubMed1991

Measurement of in vivo urological tumour cell kinetics using multiparameter flow cytometry. Preliminary study.

The in vivo labelling of urological tumour cells using the S phase marker bromodeoxyuridine (BRdU) for histochemical studies is reported. The use of multiparameter flow cytometry (FCM) with BRdU labelling to study tumour proliferation offers significant advantages. It provides simultaneous measurements of the DNA ploidy (DI), the duration of the S phase (Ts), the potential doubling time (Tpot) and the total and aneuploid tumour labelling indices (LI) from a single specimen. Heterogenous tumour cell populations can be measured with high sensitivity. We report a preliminary study to evaluate the method in the measurement of the kinetics of transitional cell carcinoma of the bladder (TCCB). Nineteen patients with TCCB, 1 with leukoplakia of the bladder, 2 with renal carcinoma, 1 with prostatic carcinoma and 1 with a squamous carcinoma of the penis were studied. Of the bladder tumours, 3 were aneuploid, DI = 1.32, 1.58 and 1.89. BRdU uptake was detected in all tumours. The median LI was 1.5% (range 0.5-10.0). In 15/19 tumours the labelling profile was satisfactory for calculation of the Ts and Tpot. The median Ts was 6.2 h and the median Tpot was 17.1 days. This study demonstrates that measurement of multiple parameters of urological tumour proliferation in vivo is possible. These parameters require further assessment as indices of biological aggressiveness and clinical prognosis.

Aneuploidy↗
PubMed1991

gp160, the envelope glycoprotein of human immunodeficiency virus type 1, is a dimer of 125-kilodalton subunits stabilized through interactions between their gp41 domains.

The molecular masses, carbohydrate contents, oligomeric status, and overall molecular structure of the env glycoproteins of human immunodeficiency virus type 1--gp120, gp160, and gp41--have been determined by quantitative electron microscopy. Using purified gp160s, a water-soluble form of env purified from a recombinant vaccinia virus expression system, we have measured the masses of several hundred individual molecules by dark-field scanning transmission electron microscopy. When combined with sequence-based information, these mass measurements establish that gp160s is a dimer of subunits with an average monomer mass of 123 kDa, of which approximately 32 kDa is carbohydrate and 91 kDa is protein. Similarly, gp120 was found to be a monomer of 89 kDa and to contain virtually all of env's glycosylation. gp41 is glycosylated only slightly, if at all, and is responsible for the interactions that stabilize the gp160s dimer. A molecular mass map of gp160s derived by image processing depicts an asymmetric dumbbell whose two domains have masses of approximately 173 and approximately 73 kDa, corresponding to a gp120 dimer and a gp41 dimer, respectively. We infer that the average monomer mass of native gp160 is 125 kDa and that in situ, env is either a dimer or a tetramer but is most unlikely to be a trimer.

Gene Products, env↗
PubMed1991

Altered organ growth and zinc and copper distribution in endotoxin-treated neonatal mice.

Organ weights and the distribution of zinc and copper were compared in HLA/ICR mice that received intraperitoneal injections of 10 micrograms of Serratia marcescens lipopolysaccharide W or of sterile physiologic saline at 2 d of age. Between 5 and 28 d of age, body weight gains were similar in both groups. At 5 and 7 d of age, lipopolysaccharide W-treated mice had significantly lower thymus weights (p less than 0.01). At 7 d of age, liver weight was significantly increased (p less than 0.01) in lipopolysaccharide W-treated mice. Compared with tissue copper concentration in coeval saline-treated mice, lipopolysaccharide W treatment significantly increased copper concentration in thymus at 5 d of age (p less than 0.05) and significantly decreased concentration of this metal in liver at 7 d of age (p less than 0.05) and in spleen at 14 d of age (p less than 0.05). Liver zinc concentration was significantly lower (p less than 0.05) in 28-d-old mice that had received lipopolysaccharide W. When expressed on the basis of total organ burdens of zinc or copper, only the liver burden of zinc in 5-d-old lipopolysaccharide W-treated mice was significantly increased (p less than 0.05). Lipopolysaccharide W treatment consistently decreased copper concentration in liver cytosol and the amounts of zinc and copper bound to metallothionein, a transition metal-binding protein, in liver cytosol. These effects of lipopolysaccharide W on organ size and metal distribution may contribute to the adverse effects that persist after endotoxin exposure in early life.

Animals↗
PubMed1990

Effect of 5-azacytidine on metallothionein inducibility and sensitivity to lethality of cadmium in rat osteosarcoma (ROS 17/2.8) cells.

ROS 17/2.8 cells, a cloned rat osteoblastic osteosarcoma cell line, were found to be extremely sensitive to the lethal effects of cadmium and to synthesize little, if any, metallothionein in response to cadmium exposure. Culture of cells for 24 h in the presence of 1 microM 5-azacytidine, a cytidine analog, increased the inducibility of metallothionein by cadmium and significantly reduced (P less than 0.001) cytotoxicity. Anion exchange chromatographic analysis of cadmium binding to low molecular mass cytotoxicity. Anion exchange chromatographic analysis of cadmium binding to low molecular mass cytosolic proteins showed that cells treated with cadmium and 5-azacytidine expressed at least 2 isoforms of metallothionein. One isoform of metallothionein with a low affinity for cadmium was constitutively expressed by these cells. The association of poor inducibility of metallothionein by cadmium with extreme sensitivity of cells to cadmium emphasizes the role of this protein in the cellular response to this toxic metal. The modulation of metallothionein inducibility and sensitivity to cadmium by 5-azacytidine treatment suggest that metallothionein gene structure and regulation are altered in ROS 17/2.8 cells.

Animals↗
PubMed1990

The entropic tension of protein loops.

A large survey of reliably known structures supports the hypothesis that protein loops can behave like elastic strings, and has enabled a parameter describing the entropically generated tensions to be determined. Such tensions may be of major importance in the folding process; here they are discussed mainly in the context of the final collapse of a molten globule into the native structure, though they must also play a dominant rôle in the earlier development of the tertiary fold.

Mathematics↗
PubMed1990

Management of "brittle" diabetes with a preprogrammable implanted insulin pump delivering intraperitoneal insulin.

OBJECTIVE: Glycaemic control in a young woman with "brittle" diabetes. DESIGN: Use of a preprogrammable fully implanted pump (Infusaid) to deliver insulin intraperitoneally at variable rates, giving a total dose of about 60 units/24 h. SETTING: Endocrinology department in a teaching hospital. PATIENT: Thirty year old woman with 15 years' history of "brittle" diabetes. MAIN OUTCOME MEASURES: Glycated haemoglobin concentration; plasma glucose concentration. RESULTS: After implantation of the pump there was an immediate and sustained improvement in diabetic control. The patient's glycated haemoglobin concentration decreased from 15.2% to 9.2% over seven months. Her daily glucose concentrations were in the range 3.5-12 mmol/l. She has not been admitted to hospital since implantation of the pump, which was eight months before the time of writing. CONCLUSION: The implanted programmable intraperitoneal insulin pump may be of value in the management of patients with "brittle" diabetes in whom other attempts at glycaemic control have failed.

Adult↗
PubMed1990

Toxicity of cadmium to rat osteosarcoma cells (ROS 17/2.8): protective effect of 1 alpha,25-dihydroxyvitamin D3.

Inadequate vitamin D intake is an important cofactor in clinical and experimental bone disease induced by chronic cadmium exposure. The interaction was investigated by culture of rat osteoblastic osteosarcoma cells (ROS 17/2.8) in a serum-free medium with equimolar concentrations of cadmium chloride and 1 alpha,25-(OH)2 vitamin D3. After addition of cadmium alone to culture medium, the unstimulated secretion of osteocalcin and cellular alkaline phosphatase activity were inhibited at 10 pM, and of DNA synthesis and proline incorporation into collagen at 500 nM. In the presence of equimolar amounts of cadmium and 1 alpha,25-(OH)2 vitamin D3, all four responses paralleled those of 1 alpha,25-(OH)2 vitamin D3 alone up to the inhibitory concentration of 500 nM cadmium. Neither 10 nM 1 alpha,25-(OH)2 vitamin D3 nor 1 microM cadmium induced synthesis of metallothionein in these cells indicating that the protective effect of D3 was not related to the induction of a metallothionein-like protein in ROS 17/2.8 cells. In the presence or absence of D3, cadmium inhibited osteoblastic function at concentrations below the whole-organ concentration of cadmium in bone as reported in experimental and clinical cadmium-induced osteotoxicity. The extreme sensitivity of ROS 17/2.8 cells to cadmium may relate to the absence of metallothionein synthesis.

Animals↗
PubMed1990

Lead inhibits the basal and stimulated responses of a rat osteoblast-like cell line ROS 17/2.8 to 1 alpha,25-dihydroxyvitamin D3 and IGF-I.

Low-level exposure to lead impairs longitudinal growth in children and in experimental animals. The proposed mechanisms include decreased osteocalcin secretion in response to 1 alpha,25-(OH)2 vitamin D3 and decreased response to insulin-like growth factor-I. The interaction of lead, 1 alpha,25-(OH)2 vitamin D3, and insulin-like growth factor-I was investigated in an osteoblast-like cell line from rat osteosarcoma, ROS 17/2.8. Cells were cultured 24 hr in a serum-free medium with lead, 1 alpha,25-(OH)2 vitamin D3, and insulin-like growth factor-I. 1 alpha,25-(OH)2 vitamin D3 (10 nM) evoked a 4-5 X increase in osteocalcin secretion and a 100% increase in cellular alkaline phosphatase activity but no increase in DNA/cell layer. Insulin-like growth factor-I (92.5 ng/ml) evoked a 100% increase of osteocalcin secretion and a 20% increase in cellular DNA contents but no change in cellular alkaline phosphatase activity. Basal and stimulated cellular osteocalcin secretion, cellular alkaline phosphatase activity, and DNA contents were significantly inhibited by addition of 1-10 microM lead. The data are consistent with a toxic effect of lead on osteoblastic function and the cellular responses to 1 alpha,25-(OH)2 vitamin D3 and insulin-like growth factor-I. This interaction may be relevant to impaired childhood growth at low levels of lead exposure.

Alkaline Phosphatase↗
PubMed1990

Quantitative analysis of one-dimensional gel electrophoresis profiles.

A general method for the one-dimensional quantitative analysis of band profiles from an autoradiographed or stained gel is described. Curved lanes may be tracked, noise across the width of the band reduced by local averaging of densities within a selected window, and background-corrected density profiles produced. Sections of each profile can be subjected to a choice of three methods of peak-area determination, depending on the type of gel and the requirements of the analysis being carried out. In the first method, simple integration is used to measure the area under the profile between any two positions. The second method integrates a Gaussian distribution fitted to a single peak. Where bands overlap, the third method resolves peaks into their component areas, taking into account the observed increase in the width of the band in the direction of electrophoretic migration. This is incorporated into a least-squares profile-fitting method, which is shown to be both accurate and efficient.

Electrophoresis↗
PubMed1990

The functional nature of cerebellar diaschisis.

We report a patient who presented with transient clumsiness of his right hand due to a small hemorrhage in the left globus pallidus. Ten days later, positron emission tomography performed at rest showed decreased oxygen metabolism and blood flow at the site of the anatomic lesion and in remote areas such as the ipsilateral frontotemporoparietal cortex and the contralateral cerebellar hemisphere. Cerebellar hypometabolism has been ascribed to functional disconnection of the contralateral hemisphere from the cerebral cortex and has been termed crossed cerebellar diaschisis. One month later, positron emission tomography performed during unilateral motor activation (finger opposition) showed increased blood flow in the sensorimotor and supplementary motor areas contralateral to the hand engaged in the motor task. An at-rest study at this time showed resolution of the crossed cerebellar diaschisis observed acutely, but cerebellar asymmetry was demonstrated during performance of the motor task with the normal as well as with the previously paretic hand. Our activation study demonstrated cerebellar asymmetry in the chronic phase during a motor task, even though resting cerebellar blood flow was symmetrical. This observation reveals the dynamic, functional nature of crossed cerebellar diaschisis and may partially explain the lack of any clinical counterpart in functional studies of the cerebellum performed with the patient at rest.

Brain↗
PubMed1990

Histological and environmental changes in longstanding ileal conduits.

Seventeen patients with urinary diversions present for between 10 and 17 years were examined endoscopically. The region of the anastomosis was successfully biopsied in 13 cases. Biopsy specimens were examined both histologically and cytologically. Urine was aspirated from the conduits under sterile conditions and sent for qualification of bacterial flora. Histology revealed a variable degree of villous atrophy, mucosal metaplasia, crypt hyperplasia and in one case, antral type gland metaplasia. The finding of hyperplasia was corroborated by flow cytometry which demonstrated a high percentage of cells in G2 and metaphase. The majority of cases showed a colonic type of bacterial colonization and mucin histochemistry demonstrated a colonic type of mucin.

Colon↗
PubMed1990

Activation and deactivation of quinones catalyzed by DT-diaphorase. Evidence for bioreductive activation of diaziquone (AZQ) in human tumor cells and detoxification of benzene metabolites in bone marrow stroma.

Bioactivation of diaziquone (AZQ) in HT-29 human colon carcinoma cells and detoxification of benzene metabolites in bone marrow stromal cells were used as examples of the potential role of DT-diaphorase in both activation and deactivation processes. HT-29 cell cytosol contained high levels of DT-diaphorase activity and removed AZQ in the presence of either NADH or NADPH. Prior boiling of cytosol, omission of NADH or NADPH or inclusion of dicoumarol, an inhibitor of DT-diaphorase, inhibited removal of AZQ. AZQ-induced cytotoxicity in HT-29 cells was also inhibited by dicoumarol. Chemical reduction of AZQ in a cell free system enhanced formation of a GSH conjugate of AZQ. Two of the major cell types in bone marrow stroma are macrophages and fibroblastoid stromal cells. A fibroblastoid cell line derived from stromal cells contained approximately fourfold higher levels of DT-diaphorase than macrophages. Inclusion of dicoumarol in incubations containing 14C-hydroquinone and the respective stromal cell type, significantly increased covalent binding of radiolabel to macromolecules in stromal fibroblasts but not in macrophages.

Animals↗
PubMed1990

Bone marrow stromal cell bioactivation and detoxification of the benzene metabolite hydroquinone: comparison of macrophages and fibroblastoid cells.

Bone marrow stroma consists predominately of two cell types, macrophages and fibroblastoid stromal cells, which regulate the growth and differentiation of myelopoietic cells via the production of growth factors. We have previously shown that macrophages are more sensitive than fibroblastoid stromal cells (LTF cells) to the toxic effects of the benzene metabolite hydroquinone. In this study, the role of selective bioactivation and/or deactivation in the macrophage-selective effects of hydroquinone was examined. LTF and macrophage cultures were incubated with 10 microM [14C]hydroquinone to examine differential bioactivation. After 24 hr, the amount of 14C covalently bound to acid-insoluble macromolecules was determined. Macrophages had 16-fold higher levels of macromolecule-associated 14C than did LTF cells. Additional experiments revealed that hydroquinone bioactivation to covalent-binding species was hydrogen peroxide dependent in macrophage homogenates. Covalent binding in companion LTF homogenates was minimal, even in the presence of excess hydrogen peroxide. These data suggest that a peroxidative event was responsible for bioactivation in macrophages and, in agreement with this, macrophages contained detectable peroxidase activity whereas LTF cells did not. Bioactivation of [14C]hydroquinone to protein-binding species by peroxidase was confirmed utilizing purified human myeloperoxidase in the presence of hydrogen peroxide and ovalbumin as a protein source. High performance liquid chromatographic analysis of incubations containing purified myeloperoxidase, hydroquinone, and hydrogen peroxide showed that greater than 90% of hydroquinone was removed and could be detected stoichometrically as 1,4-benzoquinone. 1,4-Benzoquinone was confirmed as a reactive metabolite formed from hydroquinone in macrophage incubations using excess GSH and trapping the reactive quinone as its GSH conjugate, which was measured by high performance liquid chromatography with electrochemical detection. The activity of DT-diaphorase, a quinone reductase that has been invoked as a protective mechanism in quinone-induced toxicity, was 4-fold higher in LTF cells than macrophages. These data suggest that the macrophage-selective toxicity of hydroquinone results from higher levels of peroxidase-mediated bioactivation and/or lower levels of DT-diaphorase-mediated detoxification.

Animals↗
PubMed1990

Parental hypertension as a predictor of hypertension in black physicians: the Meharry Cohort Study.

Parental histories were obtained for a cohort of black medical students in a longitudinal study of hypertension precursors. At follow-up, 25 to 30 years later, initial and current parental histories for hypertension were compared with other precursor characteristics as well as resulting cohort hypertension. The number of participants having no positive parental history for hypertension as parents aged declined from 55% to 24%. Hypertension among black physicians varied according to parental history: 38.9% for both parents negative, 41.4% for mother only positive, 60.5% for father only positive, and 73.7% for both parents positive. Parental history of hypertension was an independent predictor of subject hypertension. Positive parental history in combination with weight gain and high normal baseline systolic and diastolic blood pressure produced a gradient of risk corresponding to the number of risk factors present. Having all four risk factors increased the hypertension risk by 15 times. Parental history together with weight gain, blood pressure, and smoking provide a strong predictor of hypertension. Weight and blood pressure control, along with cessation of smoking, should be considered important factors in the clinical management of such patients.

Adult↗
PubMed1989

Inhibition of aspartic proteinases by alpha 2-macroglobulin.

The effect of alpha 2-macroglobulin, one of the major antiproteinases in the plasma of vertebrates, on the action of the aspartic proteinases chymosin, cathepsin D and cathepsin E towards peptide and protein substrates at pH 6.2 was examined. Activities towards protein substrates were blocked, thus demonstrating that alpha 2-macroglobulin can inhibit aspartic proteinases, in addition to serine proteinases, cysteine proteinases and metalloproteinases.

Cathepsin D↗
PubMed1989

Macrophage regulation of myelopoiesis is altered by exposure to the benzene metabolite hydroquinone.

Hydroquinone, a myelotoxic metabolite of benzene, decreases the ability of murine bone marrow stromal cells to support myelopoiesis in vitro. Bone marrow stroma consists of macrophages and fibroblastoid stromal cells that participate coordinately in regulating myelopoiesis. The goal of this study was to determine if macrophage or fibroblastoid cell function is more sensitive to the myelotoxic actions of hydroquinone. To address this question, we developed purified populations of macrophages and fibroblastoid stromal cells and treated each population with hydroquinone. These cells were reconstituted together with nontreated cells of the opposite type and assayed for their ability to support the formation of granulocyte and macrophage colonies in an agar overlay. Reconstituted cultures containing hydroquinone-treated macrophages supported fewer colonies than did corresponding cultures containing untreated macrophages. Reconstituted cultures containing hydroquinone-treated fibroblastoid stromal cells were not affected. Moreover, hydroquinone reduced detectable interleukin-1 activity in purified macrophage cultures stimulated with lipopolysaccharide. These results indicate that hydroquinone selectively interferes with macrophage function possibly, in part, via alteration of macrophage interleukin-1 secretion.

Animals↗