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Biomedical subjects

G Harris

Publications and source records attributed to G Harris.

At least 145 records · Page 8Linked to original sources
PubMed1985

Typing of Pseudomonas cepacia by bacteriocin susceptibility and production.

The significance of Pseudomonas cepacia as an opportunistic pathogen in immunocompromised patients has become increasingly recognized. Particularly disturbing is its increased incidence, reported by several North American centers, in respiratory tract cultures from patients with cystic fibrosis. Epidemiological studies of P. cepacia have been hampered by a lack of typing methods. In this paper we report the development of a typing scheme based on bacteriocin production and susceptibility. For bacteriocin production, test isolates of P. cepacia were rapidly applied to the surfaces of agar plates with a multiple inoculator. After incubation of these test isolates for 5.5 h and their exposure to chloroform, indicator strains were applied in agar overlays without prior removal of the test strain growth. After 18 h of incubation, inhibition zones caused by bacteriocin activity were recognized. A similar procedure was used to examine the bacteriocin susceptibility of the test strain. The bacteriocin type of the test strain was defined based on its bacteriocin production as judged by zones of inhibition against a set of eight indicator strains and by susceptibility or resistance of the test strain to bacteriocin produced by six producer strains. Of 373 strains of P. cepacia, 95.2% were typed into a total of 44 type combinations. Bacteriocin typing provided a suitable procedure for epidemiological studies of colonization or infection by P. cepacia. The technique described in this paper was simple to perform, gave a result within 24 h, provided good strain discrimination, and was suitable for clinical, environmental, and phytopathogenic strains.

Bacteriocins↗
PubMed1985

Prolongation of life in female NZB/NZW (F1) hybrid mice by cyclosporin A.

Oral administration of cyclosporin A (Cy A), an immunosuppressive agent, prolonged the life span of female NZB/NZW (F1) hybrid mice (NZB/W). The most prominent feature of this study was the reduction of glomerular proliferation, proteinurea, and delay of renal failure. Protection from renal damage, in treated mice, occurred despite similar degrees of perivascular cellular infiltration in the kidney and other organs, and glomerular deposition of immunoglobulin and complement. Treatment did not influence the hypergammaglobulinaemia, high levels of circulating immune complexes and later development of autoimmune haemolytic anaemia typically associated with this murine disease. The levels of antibodies to double stranded DNA (dsDNA), an important disease marker, and the response to sheep red blood cells were reduced in the Cy A treated mice, in contrast with the untreated mice. Levels of rheumatoid factors were increased in Cy A treated mice, when compared with untreated mice, and this could play a role in prolongation of life associated with protection from glomerular damage, although it was considered that inhibitory effects on T cell function were a more likely mechanism.

Animals↗
PubMed1984

Relationship between intracellular oxygenation and neuromuscular conduction during hypoxic hypoxia.

Previous studies have shown that high-altitude hypoxic hypoxia is associated with reduced ventilatory capacity that may be related to skeletal muscle weakness. In the present investigation, ascent to high altitude (4,000 m) was simulated experimentally by exposure of male rats (Sprague-Dawley, 250-350 g), anesthetized with thiopental sodium (25 mg/kg, i.p.), to a breathing gas mixture of 12% oxygen diluted in 88% nitrogen (FiO2 = 0.12). Determinations of oxygen saturation on microsamples (250 ul) of arterial and central venous blood were made spectrophotometrically. Neuromuscular conduction latency was measured following electrostimulation of the sciatic nerve (1-5 V, 0.5 msec duration, 1-40 Hz) and recording of the electromyogram from the gastrocnemius muscle. Experimental hypoxia (FiO2 = 0.12) produced a highly significant increase in conduction latency from a control value (mean +/- SEM) of 3.06 +/- 0.16 msec to 4.02 +/- 0.31 msec (n = 10, P less than 0.001). Conduction latency increased with decreasing arterial oxygen saturation from a control value of 92.9% +/- 0.18% to 83.2% +/- 0.76% (P less than 0.001) in the absence of statistically significant changes in central venous oxygen saturation, central venous pressure, arterial and central venous pH, and heart rate. A significant decrement in the mean arterial blood pressure from a control value of 85 +/- 1.5 mm Hg to 69 +/- 1.5 mm Hg suggests that local ischemia may be a component of this model. These responses were accompanied by marked reduction in uptake of 3,3'-diaminobenzidine (DAB) by gastrocnemius muscle mitochondria, suggesting decreased intracellular activity of cytochrome oxidase. It was concluded that exposure of rodents to hypoxic gas mixtures may provide a suitable model for studying the mechanism of skeletal muscle weakness associated with ascent to high altitude and of other conditions wherein the supply of oxygen to tissues is limited.

3,3'-Diaminobenzidine↗
PubMed1984

Mechanism of inactivation of Escherichia coli and Lactobacillus leichmannii ribonucleotide reductases by 2'-chloro-2'-deoxynucleotides: evidence for generation of 2-methylene-3(2H)-furanone.

Incubation of 2'-chloro-2'-deoxy[3'-3H]uridine 5'-diphosphate ([3'-3H]ClUDP) with Escherichia coli ribonucleotide reductase (RDPR) and use of thioredoxin-thioredoxin reductase as reductants result in release of 4.7 equiv of 3H2O/equiv of B1 protomer, concomitant with enzyme inactivation. Inactivation is accompanied by the production of 6 equiv of inorganic pyrophosphate [Stubbe, J. A., & Kozarich, J.W. (1980) J. Am. Chem. Soc. 102, 2505-2507] and by the release of uracil as previously shown [Thelander, L., Larsson, A., Hobbs, J., & Eckstein, F. (1976) J. Biol. Chem. 251, 1398-1405]. Reisolation of RDPR by Sephadex chromatography and analysis by scintillation counting indicate that 0.96 equiv of 3H is bound per protomer of the B1 subunit of the inactivated enzyme. Incubation of [5'-3H]ClUDP with RDPR followed by similar analysis indicates that 4.6 mol of 3H is bound per protomer of the B1 subunit of the inactivated enzyme. No 3H2O is released, and 6 equiv of inorganic pyrophosphate is produced during the inactivation. RDPR is protected against inactivation when dithiothreitol (DTT) is used as a reductant in place of thioredoxin-thioredoxin reductase. Incubation of [5'-3H]ClUDP with RDPR and DTT results in the isolation of CHCl3-extractable material that exhibits infrared absorptions at 1710 and 1762 cm-1. The infrared spectrum and the NMR spectrum of the CHCl3-extracted material are very similar to model compounds prepared by the interaction of 2-methylene-3(2H)-furanone with ethanethiol. Incubation of ribonucleoside-triphosphate reductase (RTPR) from Lactobacillus leichmannii with [3'-3H]ClUTP and 3 mM DTT also results in time-dependent 3H2O release concomitant with enzyme inactivation. Reisolation of the inactive protein by Sephadex chromatography followed by radiochemical analysis indicates that 0.4 equiv of 3H is bound covalently per mol of inactivated enzyme. Similar studies with [5'-3H]ClUTP indicate that 2.9 equiv of 3H is bound covalently per mol of inactivated enzyme. No 3H2O is released. High concentrations of DTT protect the enzyme against inactivation. Extraction of the enzymatic reaction mixture with CHCl3 and analysis of the isolated products result in an infrared spectrum and an NMR spectrum remarkably similar to those observed with the E. coli RDPR. Data presented are consistent with the proposal that both the E. coli and L. leichmannii enzymes are able to catalyze the breakdown of the appropriate 2'-chloro-2'-deoxynucleotide to a 3'-keto-2'-deoxynucleotide that can collapse to form the reactive sugar intermediate 2-methylene-3(2H)-furanone.(ABSTRACT TRUNCATED AT 400 WORDS)

Deoxyuracil Nucleotides↗
PubMed1984

Revised pyocin typing method for Pseudomonas aeruginosa.

In the Gillies and Govan method of pyocin typing for Pseudomonas aeruginosa a cross-streaking technique was used, and 105 main types and 25 subtypes were identified by the patterns of inhibition observed on 13 indicator strains. Disadvantages of the technique included the need to remove test strain growth before application of the indicator strains, the 48-h period needed to obtain a result, and the inability to reliably type mucoid P. aeruginosa. Recent studies have enabled us to overcome these disadvantages and significantly improve the speed and application of pyocin typing. Our revised technique utilizes the same 13 indicator strains which are already used internationally. Test strains were rapidly applied to the surface of agar plates with a multiple inoculator. After incubation for 6 h and exposure to chloroform, the indicator strains were applied in agar overlays without prior removal of the test strain growth. After 18 h of incubation, the pyocin type was recognized by inhibition of particular indicator strains. Additionally, the activity of particulate (R and F) and nonparticulate (S) pyocins could be distinguished on the basis of inhibition zone size, which thus allowed further discrimination. The revised technique allows typing within 24 h, increases the number of identifiable types, and can be used to type mucoid strains.

Bacteriocins↗
PubMed1983

Adjuvant chemotherapy with doxorubicin (Adriamycin) and 5-fluorouracil in T3, NX, MO bladder cancer treated with radiotherapy.

Radical radiotherapy alone has been compared with radical radiotherapy followed by chemotherapy using doxorubicin (Adriamycin) and 5-fluorouracil in a randomised prospective study on 129 patients presenting with T3, NX, MO transitional cell carcinoma of the bladder. One hundred and ten patients were evaluable with a minimum follow-up of 2 years. The addition of this form of chemotherapy did not appear to influence the survival rate or the proportion of patients free from tumour. It cannot be recommended for routine use in the primary treatment of infiltrating bladder cancer.

Carcinoma, Transitional Cell↗
PubMed1983

Autoimmune haemolytic disease in mice after exposure to a methylating carcinogen.

N-Methyl-N-nitrosourea (MNU) but not methyl methanesulphonate (MMS) induced both autoimmune haemolytic anaemia and thymic lymphoma in susceptible strains of mice, particularly the C57BL/6. These effects could be positively correlated with the formation of O6-methylguanine in target DNA. All murine lymphoid cells showed lack of ability to remove O6-methylguanine from their DNA, therefore the variation of responses between different mouse strains indicated that other host factors, probably genetic, must be involved. The results do indicate however that a potent pre-mutagenic DNA base modification can initiate the events leading to autoimmune disease in susceptible mice.

Anemia, Hemolytic, Autoimmune↗
PubMed1982

Defective repair of 0(6)-methylguanine in autoimmune diseases.

On the premise that somatic mutations may be involved in the pathogenesis of diseases such as systemic lupus erythematosus and rheumatoid arthritis, the proficiency of repair of 0(6)-methylguanine, a powerful, premutagenic, directly miscoding base lesion, was examined in the DNA of peripheral-blood mononuclear cells (mainly lymphocytes) from patients with such diseases. The capacity of lymphocytes to repair this lesion was impaired in many patients with autoimmune disease. In some healthy controls repair was also defective whereas in other, nonautoimmune diseases, it was not. These findings support the hypothesis that this defect of repair was not simply a result of the disease state. It is therefore postulated that defective repair of 0(6)-methylguanine could be one of the factors determining the susceptibility of autoimmune diseases in genetically predisposed individuals.

Arthritis, Rheumatoid↗
PubMed1982

Direct cloning of human malignant melanoma.

An in vitro soft agar technique was used to culture human malignant melanoma cells from 61 solid tumors, 17 lymph nodes, 11 effusions, and four bone marrow specimens from 93 patients with malignant melanoma. Colonies grew in soft agar from 64 (69%) of the 93 specimens. Fifty-five percent of the specimens cultured formed greater than or equal to 30 colonies per 500,000 nucleated cells plated. Light microscopy, electron microscopy, tumor marker, and athymic nude mouse studies provided evidence the colonies were composed of malignant melanoma cells. Drug sensitivity studies utilizing the cloning technique showed similarities between in vitro results and the general clinical experience noted with the same drugs. The human tumor cloning system represents a new model for future basic biology and clinical studies of human malignant melanoma.

Agar↗
PubMed1982

DNA chain elongation rates in marrow cells from vitamin B12-deficient patients and methotrexate-treated mice.

The DNA synthesized by marrow cells from patients with vitamin B12 deficiency and mice previously given methotrexate (MTX), has been investigated. Suspensions of bone marrow cells were pulse-labelled with [methyl-3H]thymidine or deoxy [5-3H]cytidine for 30 s and the radioactivity in the DNA was chased thereafter in the presence of 10 microM non-radioactive nucleoside for periods up to 60 min. The rates of elongation of new daughter strands were then assessed by hydroxyapatite chromatography of alkali-denatured DNA samples. No significant differences were found between the average rates of elongation of daughter strands from control marrow cells on the one hand and the vitamin B12-deficient or the methotrexate-affected cells on the other. This is to be contrasted with the results of previous studies which have shown a retardation in the rates of movement of replication forks in stimulated, cultured lymphocytes obtained from vitamin B12- or folate-deficient patients.

Anemia, Megaloblastic↗
PubMed1982

Acquisition of beta-glucuronidase activity by deficient fibroblasts during direct contact with lymphoid cells.

Fibroblasts deficient in beta-glucuronidase acquired high levels of this enzyme when they were co-cultured with concanavalin A-stimulated lymphocytes. Acquired enzyme activity, determined using a single-cell cytochemical assay, was directly proportional to the number of lymphocytes added and persisted for several days in fibroblasts maintained at high density. Lymphocytes did not secret significant levels of beta-glucuronidase into their culture medium, and did not release other substances able to induce synthesis of the enzyme by the deficient fibroblasts. Nor did beta-glucuronidase acquisition result from concanavalin A-mediated uptake of enzyme, since alpha-methylmannoside did not reduce acquired activity. Moreover, lymphocytes from various sources, whether unstimulated or activated by a different mitogen, bacterial lipopolysaccharide, were equally effective in promoting the appearance of beta-glucuronidase. Deficient fibroblasts did not acquire beta-glucuronidase by active endocytosis when co-cultured with lymphocytes, since enzyme extracted from lymphocytes was not itself effective in this respect. Furthermore, mannose 6-phosphate, which did inhibit, endocytosis by deficient fibroblasts of exogenous beta-glucuronidase prepared from 3T3 cells, had no effect on enzyme acquisition by fibroblasts during their co-culture with lymphocytes. Conversely, inhibitors of protein synthesis and energy metabolism, which did not interfere with endocytosis of exogenous enzyme, abolished the acquisition of beta-glucuronidase during co-culture. Deficient fibroblasts did not acquire beta-glucuronidase when they were cultured together with lymphocytes but separated from them by Millipore membranes permeable to exogenous enzyme. Thus, although the mechanism of acquisition is still unclear, the present results suggest that beta-glucuronidase is transferred from lymphocytes to deficient fibroblasts by a process in which direct cell-to-cell contact is obligatory.

Animals↗