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P Ritchie

Publications and source records attributed to P Ritchie.

17 recordsLinked to original sources
PubMed2001

Retrospective collection of exposure data from industry: results from a feasibility study in the United Kingdom.

In the United Kingdom the Health and Safety Executive for some years has stored chemical exposure data in their National Exposure Database. However, it has been difficult to persuade industry and other organizations to contribute to this resource. The aim of this project was to devise a cost-effective method of obtaining occupational exposure data on chemicals from U.K. industry and other sources. Five strategies were used to identify data for three different substances: toluene, acrylonitrile, and ethylene oxide. In total, 810 organizations were contacted and over 45 percent responded. However, only 40 had relevant exposure data. Almost equal numbers of acceptable measurements were identified for toluene and acrylonitrile (2,770 and 2,000 respectively) with lesser ethylene oxide data (800). These measurements were drawn from a wide range of industries and are probably representative of measurements made by U.K. industry, although most of the data were from companies employing more than 100 people. During the second phase of the project, more than 3,000 measurements and associated contextual information were collected (499 for toluene, 1,516 for acrylonitrile, and 17 for ethylene oxide, with a further 1,004 measurements for 1 of 27 substances collected simultaneously with one of the above). The costs of identifying and collecting exposure data ranged from ł7 to ł380 per valid measurement, depending on the source of the data. We suggest that, rather than trying to retrospectively collect data, it is likely to be more cost-effective to enlist a number of occupational hygiene consults and industrial organizations to prospectively provide anonymized exposure measurements for inclusion in the Health and Safety Executives National Exposure Database.

Data Collection↗
PubMed2000

Xenopus NK cells identified by novel monoclonal antibodies.

Early-thymectomized (Tx) Xenopus frogs, which are permanently deficient in T cells, are used as a model sytem for the characterization of novel monoclonal antibodies (mAb) which identify candidate NK cells at the amphibian level of evolution. Hybridomas, generated from mice immunized with splenocytes from Tx Xenopus following B cell and thrombocyte depletion, were screened by flow cytometry. Three mAb (1F8, 4D4 and 1G5) were identified that stained increased proportions of splenocytes from Tx compared with control frogs. These mAb identified lymphoid populations from Xenopus spleen, liver and gut which, after 48 h culture in growth factor-rich medium, exhibited spontanous killing of MHC-deficient allotumor targets. mAb-defined splenocytes also rapidly induced apoptosis of such tumor targets. Dual color analysis confirmed that NK cells are neither T nor B cells. Cytospins of splenocytes isolated with anti-NK mAb revealed large lymphoid cells with distinct pseudopodia. Immunohistology indicated each anti-NK mAb routinely labeled cells within the gut epithelium but NK cells were difficult to visualize in spleen sections. Western blotting of spleen, liver and intestinal lysates subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis showed that 1G5 reacted strongly with protein bands of approximately 70 - 85 kDa, whereas mAb 1F8 and 4D4 stained less intensely, but identified similar protein bands.

Animals↗
PubMed1998

Role of the carboxyterminal residue in peptide binding to protein farnesyltransferase and protein geranylgeranyltransferase.

Protein farnesyltransferase and protein geranylgeranyltransferase-I catalyze the prenylation of a cysteinyl group located four residues upstream of the carboxyl terminus. The identity of the carboxyterminal residue plays a significant role in determining the ability of compounds to bind to each enzyme and to serve as substrate. We compared the binding and substrate specificities of peptides with carboxyterminal substitutions to determine which residues promote selectivity and which residues promote recognition by both enzymes. Using tetrapeptide inhibitors with the general structure l-penicillamine-valine-isoleucine-X and substrates with the structure Lys-Lys-Ser-Ser-Cys-Val-Ile-X, we measured their respective Ki, Km, and kcat values for both recombinant rat protein farnesyltransferase and recombinant rat protein geranylgeranyltransferase-I. We studied the roles of carboxyterminal branched residues (leucine, isoleucine, valine, and penicillamine) and linear residues (methionine, cysteine, homocysteine, alanine, aminobutyrate, and aminohexanoate) in promoting interaction with the enzymes. For protein geranylgeranyltransferase-I, peptide substrates with carboxyterminal branched or linear residues had Km values that were 5- to 15-fold greater than the Ki values of the corresponding peptide inhibitors. For protein farnesyltransferase, peptide substrates with carboxyterminal branched residues, proline, or homoserine had Km values that were 7- to 200-fold greater than the Ki values of the corresponding peptide inhibitors. For protein farnesyltransferase the Km and Ki values for peptides ending with linear residues were in general agreement. Our studies indicate that the substrate and inhibitor binding specificities of protein geranylgeranyltransferase was much more restricted than those of protein farnesyltransferase.

Alkyl and Aryl Transferases↗
PubMed1998

T-cell and natural killer cell development in thymectomized Xenopus.

The Xenopus early-thymectomy model system is used to investigate the extent to which the thymus controls T-cell development and to probe the evolution of natural killer (NK) cells. Loss of T-cell function following thymectomy, together with the paucity of cells expressing monoclonal antibody-defined T-cell surface markers, and greatly reduced expression of T-cell receptor beta transcripts in spleen, liver and intestine, indicate that T-cell development in minimal in the absence of the thymus. Our findings therefore mitigate against the idea that a substantial extrathymic pathway of T-cell development exists in early vertebrate evolution. Rather, they suggest that in this amphibian representative T cells are predominately thymus dependent. In vitro studies with control and thymectomized Xenopus splenocytes reveal that a non-T/non-B population and also two T-cell subsets all display natural cytotoxicity towards allogeneic thymus lymphoid tumour cells (which are deficient in MHC antigen expression). Since Xenopus thymectomized early in larval development are permanently deficient in T cells, they may provide a useful phylogenetic model for the study of NK cells.

Animals↗
PubMed1998

Apoptosis-specific protein (ASP) identified in apoptotic Xenopus thymus tumor cells.

A novel apoptosis-specific protein (ASP) has recently been identified in the cytoplasm of apoptotic mammalian cells. This paper investigates whether ASP is found in Xenopus thymus tumor-derived lymphoid cell lines undergoing apoptosis and also in apoptotic, nontransformed splenocytes. Cultured Xenopus tumor lymphoid cells induced to undergo apoptosis by serum deprivation or treatment with the calcium ionophore, ionomycin, displayed altered morphology typical of apoptotic cells, as judged by flow cytometric light-scatter characteristics and by fluorescence microscopy of acridine-orange-stained cells. Flow cytometry of permeabilized cells and fluorescence microscopy of acetone-fixed cytospins revealed that apoptotic Xenopus tumor cells, especially those displaying loss or condensation of DNA, displayed increased expression of epitopes recognized by a rabbit polyclonal antibody against ASP. Flow cytometry confirmed that ASP is also expressed in splenocytes induced to apoptose by culture in ionomycin or following concanavalin A stimulation. No increased expression of ASP was seen when lymphoid tumor cells or splenocytes were induced into necrosis by overdose with the antifungal agent amphotericin B. Western blotting with antibody against ASP identified the emergence of several protein bands in cell lysates from apoptotic, but not necrotic, Xenopus tumor cells. The new and simple methodology for identifying apoptotic cells described here is likely to be of value to those studying immune system development and associated programmed cell death in Xenopus.

Amphotericin B↗
PubMed1996

NK-like activity against allogeneic tumour cells demonstrated in the spleen of control and thymectomized Xenopus.

This paper addresses the issue of natural killer (NK) cell evolution by searching for NK-like activity in an amphibian representative, the immunologically well-characterized clawed frog, Xenopus laevis. Using in vitro 6 h 51chromium release assays, we have shown that splenocyte effectors from early thymectomized (Tx) year-old frogs, but not from control siblings, are able to spontaneously lyse allogeneic thymus tumour cell lines that lack MHC antigen expression. Such lytic capacity can be readily induced in control Xenopus and elevated in Tx frogs by a single injection of tumour cells, with maximal splenocyte cytotoxicity occurring 3 days postinjection, the amount of 51Cr-release correlating directly with effector: target ratios. Splenocytes, even those from tumour-injected frogs, are unable to lyse allogeneic splenic lymphoblasts or erythrocyte targets, even when the latter are coated with IgY (the Xenopus IgG equivalent); moreover, we were unable to demonstrate any splenocyte-induced lysis of the human NK cell target K562. Lymphokine-activated killing (LAK) in Xenopus is suggested, since Tx splenocytes cultured in cytokine-rich medium (from concanavalin A-stimulated control splenocytes) display significantly elevated killing of allogeneic tumour targets. Flow cytometric analysis highlights the loss of T cell markers from the spleen of Tx frogs and reveals a variable staining pattern of both control and Tx splenocytes when treated with a mAb that binds to both fish non-specific cytotoxic cells and human NK cells. Prospects for identifying the cellular basis of NK-like activity in Xenopus are discussed in the light of these experiments.

Animals↗
PubMed1996

Psychology in Canada.

This chapter reviews how psychology in Canada evolved over half a century to become the most popular discipline in universities and a respected health-care and helping profession. The organization, journals, and funding of the scientific discipline are described. The importance of DO Hebb's research as the stimulus and foundation for discipline growth and significant research contributions to basic processes is identified. The multicultural mosaic of Canadian society and early research on second-language learning are shown to have influenced cross-cultural and social research. Canadian research contributions to basic processes and to the social and health sciences are reviewed. Although late to begin in Canada, clinical research and the profession of psychology are shown to have substantially developed over the past two decades. With large numbers of quality researchers and practitioners, psychology has a bright future in Canada.

Journal Article↗
PubMed1994

Farnesyl-protein transferase and geranylgeranyl-protein transferase assays using phosphocellulose paper absorption.

Farnesyl-protein transferase catalyzes the reaction of farnesyl pyrophosphate and its acceptors to yield farnesyl protein and pyrophosphate. Geranylgeranyl-protein transferases are distinct enzymes that catalyze the reaction of geranylgeranyl pyrophosphate and their acceptors. We used tritiated isoprenoid pyrophosphate donors and synthetic peptide acceptors to measure enzyme activities. The peptide acceptors contained basic amino acid residues on the amino terminus of tetrapeptide substrate determinants specific for each enzyme. Following the incubation, portions of the reaction mixture were applied to numbered phosphocellulose paper strips that were immersed in 95% ethanol/75 mM phosphoric acid (1/1; v/v). Acid promoted binding of positively charged peptide substrates and products to the negatively charged paper, and alcohol eluted the radioactive prenyl groups. Paper strips were processed in the same container in batches for 40 min, and radioactivity adsorbed to the strips was then measured by liquid scintillation spectrometry. The use of peptides makes the expression and purification of recombinant substrates in bacteria unnecessary. However, most proteins bind quantitatively to phosphocellulose at acidic pH, and the washing procedure developed for peptide substrates is applicable for measuring prenyltransferase activities with recombinant Ras proteins as acceptor.

Absorption↗
PubMed1992

Skin xenograft rejection in Xenopus--immunohistology and effect of thymectomy.

Immunohistology, using the T-lineage-specific monoclonal antibody XT-1 and an anti-IgM mAb, illustrates differences in the cellular basis of skin allograft and xenograft destruction displayed by control froglets (X. laevis). Thus T cells predominate within allografts, whereas B-lineage cells accumulate under xenografts (from X. tropicalis). The possibility that T cells do not play a central role in mediating xenograft rejection is consistent with the finding that early thymectomy (at 7 days) has minimal effect on rejection end points of X. tropicalis transplants. However, rejection of skin from a "phylogenetically less distant" xenogeneic species (X. borealis) is shown here to be impaired in early thymectomized X. laevis. Differences in the extent to which the thymus has been shown to influence skin xenograft rejection in Xenopus are discussed.

Animals↗
PubMed1991

Splenocyte response to T cell-derived cytokines in thymectomized Xenopus.

A miniaturized, "hanging-drop" bioassay reveals that splenocytes from early-thymectomized (Tx) Xenopus can respond (by enhanced thymidine incorporation) to thymic-dependent "cytokines" generated in PHA- or alloantigen-stimulated cultures. Preliminary evidence, using fluorescence activated cell sorting, indicates that surface IgM- splenocytes, rather than sIgM+ cells, from Tx toads are sensitive to the crude, splenocyte-derived, active supernatants. Although these responsive cells display residual, but low, reactivity to PHA, their thymus independence is suggested by flow cytometric observations using the anti-T cell monoclonal antibody XT-1. The development of "T-like" cells in Tx Xenopus is discussed.

Animals↗
PubMed1991

Phosphorylation of rat tyrosine hydroxylase and its model peptides in vitro by cyclic AMP-dependent protein kinase.

The enzyme tyrosine hydroxylase catalyzes the first step in the biosynthesis of dopamine, norepinephrine, and epinephrine. Tyrosine hydroxylase is a substrate for cyclic AMP-dependent protein kinase as well as other protein kinases. We determined the Km and Vmax of rat pheochromocytoma tyrosine hydroxylase for cyclic AMP-dependent protein kinase and obtained values of 136 microM and 7.1 mumol/min/mg of catalytic subunit, respectively. These values were not appreciably affected by the substrates for tyrosine hydroxylase (tyrosine and tetrahydrobiopterin) or by feedback inhibitors (dopamine and norepinephrine). The high Km of tyrosine hydroxylase correlates with the high content of tyrosine hydroxylase in catecholaminergic cells. We also determined the kinetic constants for peptides modeled after actual or potential tyrosine hydroxylase phosphorylation sites. We found that the best substrates for cyclic AMP-dependent protein kinase were those peptides corresponding to serine 40. Tyrosine hydroxylase (36-46), for example, exhibited a Km of 108 microM and a Vmax of 6.93 mumol/min/mg of catalytic subunit. The next best substrate was the peptide corresponding to serine 153. The peptide containing the sequence conforming to serine 19 was a very poor substrate, and that conforming to serine 172 was not phosphorylated to any significant extent. The primary structure of the actual or potential phosphorylation sites is sufficient to explain the substrate behavior of the native enzyme.

Animals↗
PubMed1988

Comparative effects of verapamil and isradipine on steady-state digoxin kinetics.

The effects on the steady-state digoxin pharmacokinetics of verapamil (240 mg/day) and a new dihydropyridine calcium channel blocker, isradipine (15 mg/day), were compared. Nineteen healthy white men, aged 23 to 40 years, ingested 0.25 mg digoxin tablets every 12 hours for two consecutive periods of 2 weeks. Each subject also received one of the calcium channel blockers during one of these periods, with agent and sequence randomized. Analyst-blind RIA serum digoxin determinations demonstrated that the nine subjects who received isradipine, 5 mg t.i.d., had a small increment in peak digoxin level from 2.3 +/- 0.6 to 2.9 +/- 0.7 ng/ml (p less than 0.05) but no significant change in steady-state level or AUC over 12 hours. By contrast, the 10 subjects who received verapamil, 80 mg t.i.d., showed significant increases in steady-state (0.9 +/- 0.1 to 1.3 +/- 0.2 ng/ml; p less than 0.001) and peak serum digoxin concentrations (2.5 +/- 0.7 to 3.6 +/- 0.8 ng/ml; p less than 0.001) and in AUC (15.7 +/- 1.7 to 23.6 +/- 2.9 ng . hr/ml; p less than 0.001). Neither calcium channel blocker reduced renal digoxin clearance. Verapamil increases digoxin levels without affecting renal clearance. Isradipine has no clinically important interaction with digoxin.

Adult↗
PubMed1985

A comparison of nalbuphine and pethidine for postoperative pain relief after orthopaedic surgery.

Nalbuphine hydrochloride (Nubain; Du Pont Pharmaceuticals), a synthetic agonist-antagonist analgesic, in a dose of 20 mg was compared with pethidine 100 mg in 60 patients after elective surgery in a random double-blind study. Both drugs were given intramuscularly on the first day after surgery. The pain intensity and visual analogue scales would seem to indicate that nalbuphine has a longer duration of action than pethidine (P less than 0,05). The respiration rates in the pethidine group were significantly more depressed 30 minutes after the injection than in the nalbuphine group (P less than 0,05). Nalbuphine caused less depression of both systolic and diastolic blood pressure at both 30 and 60 minutes (P less than 0,001). The results of the study show that nalbuphine, in the dose used here, may prove to be a useful substitute for pethidine.

Adolescent↗
PubMedDate not supplied

Incomplete tolerance induced in Xenopus by larval tissue allografting: evidence from immunohistology and mixed leucocyte culture.

Application of adult skin allografts to Xenopus larvae has been a favoured protocol for probing the development of self-tolerance. A more physiologic approach is presented here that examines the immunologic outcome of grafting semi- or fully allogeneic larval skin or spleen to age-matched, larval Xenopus (X. laevis/X. gilli clonal hybrids). Following such grafting at 2 or 4 weeks-of-age, young froglets (4-5-months-old) are generally unable to reject second-set skin transplants, but destroy third-party skin vigorously, the MHC class II-rich spleen proving especially effective at inducing this tolerance. In contrast, following larval grafting of semiallogeneic tissues, mixed leucocyte culture performed at the end of metamorphosis (6 weeks) and again at 6 months reveals splenocyte reactivity toward donor-strain stimulators. Immunohistological findings extend this observation of anti-donor reactivity (suggesting incomplete tolerance) to the graft site. Thus despite excellent health when viewed externally, apparently tolerated second-set skin transplants display localised infiltration (especially into the epidermis) by CD8+ T cells and increased numbers of MHC class I and II-expressing cells by 3 weeks post-grafting. The immunologic implications of these findings are discussed.

Animals↗
PubMedDate not supplied

Ontogeny and thymus-dependence of T cell surface antigens in Xenopus: flow cytometric studies on monoclonal antibody-stained thymus and spleen.

Recently generated anti-Xenopus T cell monoclonal antibodies (mAbs) to the 120 kDa XTLA-1 determinant and against the putative CD5 and CD8 homologues, together with anti-IgM and anti-MHC class II mAbs, are used in dual colour flow cytometric experiments to characterize cell surface antigenic expression on lymphocytes in thymus and spleen of Xenopus laevis during larval and early adult life and also in metamorphosis-inhibited animals. Histological confirmation of T cell emergence early in larval ontogeny is supplied by cryostat sections stained for CD8. Five-day thymectomy, i.e. prior to T-lineage cell differentiation in the thymus, abolishes T cell marker expression in the spleen for up to 1 year. Moreover, late larval (20 days) or early adult (3 months) thymectomy (i.e. removal after peripheralization of T cells has occurred) also leads to severe depletion of mAb-defined T cells in the spleen.

Animals↗
PubMedDate not supplied

Natural cytotoxicity towards allogeneic tumour targets in Xenopus mediated by diverse splenocyte populations.

We have recently demonstrated NK-like activity in the spleen of the clawed frog, Xenopus laevis. This paper investigates the cellular basis of this natural cytotoxicity. Significant levels of cytotoxicity towards B3B7 allogeneic thymus tumour targets, that express neither class Ia nor class II MHC proteins, occurred after splenocytes from either control or early-thymectomized (Tx) year-old Xenopus were cultured for 48 hours. Killing by Tx cells required their culture in growth factor-rich medium (GFM) obtained from concanavalin A-stimulated cells. Immunomagnetic cell sorting revealed that cytotoxic effectors in both control and Tx frogs were found in the B cell-depleted population, but never in the B cell-enriched fraction. Splenocytes from control Xenopus, depleted of T cells by magnetic sorting and following culture in GFM, also developed natural cytotoxicity towards allotumour cells. Magnetic cell sorting also revealed that purified (CD5+) T cells cultured for 48 hours in GFM also became able to lyse the allogeneic tumour targets. Cytotoxicity mediated by T cells resided not only in the CD5+, CD8+ population, but also in the CD5+, CD8- (putative CD4+) T cell subset. Ontogenetic studies revealed that splenocytes from 6-7 week-old (stage 56-57) control larvae, even after 48 hr culture in GFM, were unable to spontaneously lyse the allotumour targets, whereas cultured splenocytes from 6 month old froglets were effective killers. Thymocytes from larvae or adults routinely failed to kill tumour cells. The work highlights the need to use Tx Xenopus to further explore non-T-cell-mediated, NK-like cytotoxicity at the amphibian level of evolution.

Animals↗