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R Knowles

Publications and source records attributed to R Knowles.

At least 37 records · Page 2Linked to original sources
PubMed1987

Interleukin-2- and mitogen-activated NK-like killer cells from highly purified human peripheral blood T cell (CD3+ N901-) cultures.

In this study, highly purified (HP) CD3-positive N901-negative T lymphocytes could be induced to become natural killer (NK)-like in culture in the presence of recombinant interleukin-2 (rIL-2) and phytohemagglutinin (PHA). Thus, purified CD3+ N901- T cells from fresh human peripheral blood were obtained by negative selection using an indirect panning technique. To ensure that T lymphocyte fractions were completely devoid of any detectable NK cells, two additional purification procedures were employed: incubation of post-pan T cells with the NK-cytotoxic lysomotropic agent L-leucinemethylester, and complement-mediated lysis using the NK cell specific NKH1a monoclonal antibody. Purity of CD3+ N901- cells could be confirmed by surface marker analysis, whereby two NK-associated antigens, N901 and H-25, were undetectable, while 94 +/- 1% of cells expressed the CD3 (Leu-4) antigen. On functional analysis, fresh HP CD3+ N901- cells exhibited no cytotoxic activity against the standard NK target K562. When HP NK-depleted T lymphocytes were cultured for 7 days in the presence of rIL-2 (100 U/ml), neither surface antigen expression nor cytotoxic activity against K562 changed significantly. However, significant cytotoxicity against K562 [18 +/- 5% specific lysis at 25:1 effector:target (E/T) ratio] could be induced when HP CD3+ N901- cells were grown for 7 days in the presence of rIL-2 and PHA (0.5% v/v). Concomitantly, antigens N901 and H-25 were found to be coexpressed on a minor proportion (22 +/- 16 and 22 +/- 6%, respectively) of CD3+ (88 +/- 2% on day 7) cells. Four-week long-term culture of HP NK-depleted T cells in the presence of rIL-2 and PHA yielded a continuous increase in cytotoxicity against K562 cells (0 up to 46% specific lysis at 25:1 E/T ratio). Of particular interest was the emergence of cytotoxicity against the NK-resistant Daudi cell target (15 +/- 8% specific lysis at 25:1 E/T ratio on day 21). Expression of antigens N901 and H-25 as well as CD3 remained essentially unchanged in long-term culture. In sorting experiments, the H-25+ cell fraction was significantly enriched for cytotoxicity against K562, when compared to both H-25- and unseparated cell fractions. In summary, our results suggest that a proportion of HP CD3+ N901- T lymphocytes may give rise to cells that exhibit NK-like functional and phenotypic properties.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1984

Uptake hydrogenase activity in denitrifying Azospirillum brasilense grown anaerobically with nitrous oxide or nitrate.

zospirillum brasilense Sp7 was grown anaerobically with N2O as the terminal electron acceptor and NH4Cl as the nitrogen source. Hydrogen uptake activity (O2-dependent H3H oxidation) was expressed in the presence and absence of 5% H2; it reached its maximum in late logarithmic phase as the malate became limiting. This activity was very stable in stationary phase, even in the absence of exogenous H2, compared with microaerobically grown cultures; this supports the hypothesis that the exclusion of O2 is critical for maintaining the integrity of the H2 uptake system in this organism. Oxygen, as well as methylene blue and N2O, supported H2 uptake, indicating the presence of electron transport components leading to O2 in anaerobically grown A. brasilense. Nitrite (0.5 mM) inhibited H2 uptake. In cultures grown with NO3- as the terminal electron acceptor and NH4Cl as the nitrogen source, in the presence and absence of exogenous H2, only low H2 uptake activity was observed. Methylene blue, O2, N2O, NO3-, and NO2- were all capable of acting as the electron acceptor for H2 oxidation. Nitrite (0.5 mM) did not inhibit H2 uptake in NO3--grown cells, as it did in N2O-grown cells. A. brasilense appears to be one of the few organisms capable of expressing the H2 uptake system under denitrifying conditions in the absence of molecular H2.

Anaerobiosis↗
PubMed1984

Hydrogenase activity in Azospirillum brasilense is inhibited by nitrite, nitric oxide, carbon monoxide, and acetylene.

Nitrite, NO, CO, and C2H2 inhibited O2-dependent H2 uptake (H3H oxidation) in denitrifying Azospirillum brasilense Sp7 grown anaerobically on N2O or NO3-. The apparent Ki values for inhibition of O2-dependent H2 uptake were 20 microM for NO2-, 0.4 microM for NO, 28 microM for CO, and 88 microM for C2H2. These inhibitors also affected methylene blue-dependent H2 uptake, presumably by acting directly on the hydrogenase. Nitrite and NO inhibited H2 uptake irreversibly, whereas inhibition due to CO was easily reversed by repeatedly evacuating and backfilling with N2. The C2H2 inhibition was not readily reversed, partly due to difficulty in removing the last traces of this gas from solution. The NO2- inhibition of malate-dependent respiration was readily reversed by repeatedly washing the cells, in contrast to the effect of NO2- on H2-dependent respiration. These results suggest that the low hydrogenase activities observed in NO3(-)-grown cultures of A. brasilense may be due to the irreversible inhibition of hydrogenase by NO2- and NO produced by NO3- reduction.

Acetylene↗
PubMed1984

Monoclonal anti thymic cell antibodies detecting epidermal cells.

Two monoclonal antibodies were obtained from a fusion following immunization with human thymic cells. BL6 reacted with an antigen present on 80% of thymocytes found in the cortex, but absent from all other circulating or non circulating T or B lymphocytes. It immunoprecipitated an antigen of 49 Kd heavy chain with a small 12 Kd subunit. BL6 also reacted with dendritic cells of the epidermis identified as Langerhans cells carrying histocompatibility class II antigens and possessing Birbeck granules. The second antibody, BL7, did not stain thymocytes, but reacted with a cytoplasmic antigen of certain thymic epithelial cells. In skin sections, BL7 detected a cytoplasmic antigen of the keratinocytes of the basal cell layer. These cross reactivities between skin and thymic cells are discussed in view of a possible functional similarity between the two tissues.

Adult↗
PubMed1982

Denitrification.

Explore the source record for details and available documents.

Bacteria↗
PubMed1980

Hydrogen metabolism of Azospirillum brasilense in nitrogen-free medium.

Production of H2 by Azospirillum brasilense under N2-fixing conditions was studied in continuous and batch cultures. Net H2 production was consistently observed only when the gas phase contained CO. Nitrogenase activity (C2H2 reduction) and H2 evolution (in the presence of 5% CO) showed a similar response to O2 and were highest at 0.75% dissolved O2. Uptake hydrogenase activity, ranging from 0.3 to 2.5 mumol H2/mg protein per hour was observed in batch cultures under N2. Such rates were more than sufficient to recycle nitrogenase-produced H2. Tritium-exchange assay showed that H2 uptake was higher under Ar than under N2. Uptake hydrogenase was strongly inhibited by CO and C2H2. Cyclic GMP inhibited both nitrogenase and uptake hydrogenase activities.

Acetylene↗
PubMed1978

Lack of specificity of the limulus lysate test in the diagnosis of pyogenic arthritis.

The diagnosis of pyogenic arthritis may be difficult to confirm since culture results are sometimes negative. This study attempted to evaluate the utility of the limulus lysate assay for the early detection of pyogenic arthritis due to gram-negative organisms. Seven-one specimens of synovial fluid from 46 patients were evaluated for reactivity in the limulus test, pyrogenic responses in rabbits, total white blood cell count, total neutrophil count, total red blood cell count, and protein and glucose concentrations. All patients with culture-proven septic arthritis or presumptive septic arthritis had joint fluid specimens that yielded a positive result in the limulus assay (12 patients). However, 52.9% of patients (18 of 34) who had a nonseptic cause for their joint effusion also had a joint fluid specimen that was positive in the limulus test. A positive limulus test result showed a significant correlation with an elevated total white blood cell count (P less than 0.0005), an elevated absolute neutrophil count (P less than 0.0005), and a decreased concentration of glucose (P less than 0.005) in synovial fluid, and the production of fever in rabbits after injection of synovial fluid (P less than 0.05). Thus, this study suggests that a positive result in the limulus test on joint fluid is nonspecific for a septic process, but a negative result would be evidence against it.

Arthritis, Infectious↗
PubMed1978

Effect of oxygen and nitrate on nitrogen fixation and denitrification by Azospirillum brasilense grown in continuous culture.

Azospirillum brasilense was grown continuously at various levels of dissolved oxygen (O2) in a nitrogen-free medium containing malates as the carbon source. Steady-state cultures were established only at O2 concentrations less than 0.0150 atm (1 atm = 101.325 Pa) and rates of acetylene reduction (N2 fixation) and efficiencies of N2 fixation were maximal between 0.0050-0.0075 atm dissolved O2. These cultures appeared to be O2- or N2-limited. There was no evidence of a respiratory protective mechanism in this organism. Anaerobic denitrifying steady-state cultures were established with nitrate (NO3-) as the sole nitrogen source with no detectable N2 fixation. N2 fixation, but no denitrification, was observed when NO3- was decreased to 10 microgram N per millilitre at 0.003 atm dissolved O2. In samples removed from the culture vessel, either activity could be induced with a lag of approximately 120 min by incubation under appropriate conditions.

Acetylene↗
PubMed1977

Potassium homeostasis in chronic diabetes mellitus.

Potassium homeostasis was evaluated in 13 patients with diabetes mellitus. In eight, plasma renin activity was low; plasma aldosterone concentration was decreased in all; seven had a history of spontaneous hyperkalemia. After administration of glucose orally, there were paradoxical increases in serum potassium levels in seven patients. After potassium loading, maximal values and increments of serum potassium were higher and fractional potassium excretion was lower in the diabetic than in the control subjects, although the differences were not statistically significant. Abnormalities of potassium homeostasis in diabetes are probably related to insulin and mineralocorticoid deficiency. Diabetic patients with hypoaldosteronism have the potential for severe hyperkalemia should renal or extrarenal mechanisms for potassium homeostasis be challenged by severe acidosis be challenged by severe acidosis, diminished renal function, marked hyperglycemia, or administration of potassium salts or potassium-sparing diuretics.

Adult↗
PubMed1975

Ultrastructure of the Alnus crispa var. mollis Fern. root nodule endophyte.

Nitrogen-fixing, field-obtained root nodules of the silky green alder were studied by transmission electron microscopy. The nodule endophyte exhibited a prokaryotic cytology and was present in two forms: the hypha(0.3-1.0mum), which was branched and septate, and the vesicle (3-5mum), which was also septate and developed at the parental hypha tip. Bacteria-like cells, previously observed in light microscopy studies, were not seen in the present work. The actinomycete-like endophyte penetrated through the host cell wall and becane enveloped by a capsular material (0.1mum), the whole being enclosed by host membranes. In some host cells, the endophyte appeared to lyse and become a mass of shrunken debris. The fine structure of the Alnus crispa var. mollis root nodule endophyte was found to be similar to that of other nonleguminous root nodule endophytes.

Bacteria↗
PubMed1975

Demonstration of the isolation of non-infective Alnus crispa var. mollis Fern, nodule endophyte by morphological immunolabelling and whole cell composition studies.

Two filamentous, branched, and septate actinomycetes were isolated from field-collected and from axenic in vitro produced root nodules of Alnus crispa var. mollis Fern. host plant. After their transfer to a chemically defined medium, these nodule isolates could not be distinguished from each other on the basis of morphology, cultural reactions, and whole cell composition and were considered to be the same species. They were morphologically similar to the root nodule endophyte, but were incapable of nodulating aseptic host plants growing in a nitrogen-deficient substrate. Whole cells of the nodule isolates were used for the production of rabbit antibodies. The resulting specific antiisolate antibodies were conjugated with fluorescein isothiocyanate and used in staining tests of the nodule endophyte. The immunofluorescence reactions demonstrated the homology of the nodule isolates with the nodule endophyte. After pectinase degradation of the endophyte capsule, the indirect immunoferritin method corroborated the fluorescent anti-body (FA) staining reactions. There was no antigenic relationship between the nodule isolates and 13 known strains of actinomycetes as determined by the FA techique. Fluorescent antibody reactions of adsorbed conjugates suggested that endophytes of both Alnus crispa var. mollis Fern. and Alnus rugosa (DuRoi) Spreng. root nodules belong to a common serotype. The LL and mesoisomers of diaminopimelic acid were present in similar proportions in the nodule endophyte and in the nodule isolates. Glucose, mannose, and an unknown sugar were the predominant whole cell sugars in the nodule isolates, although trace amounts of arabinose and rhamnose were also displayed. The unknown sugar found in the nodule isolates was also present in trace amounts in the endophyte-suspension hydrolysate.

Actinomycetales↗
PubMed1974

Acetylene reduction (nitrogen fixation) by pulp and paper mill effluents and by Klebsiella isolated from effluents and environmental situations.

High rates of acetylene (C(2)H(2)) reduction (nitrogenase activity) were observed in woodroom effluent from a neutral sulfite semi-chemical mill under aerobic (up to 644 nmol of C(2)H(4) produced per ml per h) and under anaerobic (up to 135 nmol of C(2)H(4) produced per ml per h) conditions. Pasteurized effluent developed C(2)H(2) reduction activity when incubated under anaerobic but not under aerobic conditions. Activities were increased by addition of 0.5 to 3.0% glucose or xylose. Enrichment and enumeration studies showed that N(2)-fixing Azotobacter and Klebsiella were abundant, and N(2)-fixing Bacillus was present. Of 129 isolates of Klebsiella from pulp mills, lakes, rivers, and drainage and sewage systems, 32% possessed nitrogen-fixing ability.

Acetylene↗