Biomedical subjects
P Cook
Publications and source records attributed to P Cook.
Phenotypic difference between Bcg(r) and Bcg(s) macrophages is related to differences in protein-kinase-C-dependent signalling.
Mice of diverse genetic backgrounds may be classified as being either resistant or susceptible to infection with Mycobacteria. These phenotypes appear to be determined by a single gene on chromosome 1, the Bcg gene, and are expressed at the level of the macrophage in vitro. When compared to macrophages from mice of the susceptible phenotype (Bcg[s]), macrophages from mice of the resistant phenotype (Bcg[r]) show enhanced functional properties including increased expression of MHC class II molecules, increased nitric oxide production, and greater capacity to inhibit the growth of several intracellular pathogens. The bacteriostatic activity of B10R and B10S macrophages correlated with the amount of nitric oxide produced by the macrophages. Since protein kinase C (PKC) has been shown to be involved in the induction of a range of macrophage functional activities, experiments were conducted to examine the possibility that phenotypic differences between Bcg(r) and Bcg(s) macrophages may be related to differences in PKC-dependent signalling. Macrophage cell lines were derived from mice congenic at the Bcg locus that are either resistant (B10R) or susceptible (B10S) to infection with Mycobacteria. In the basal state, PKC-specific activity was significantly increased in the cytosolic fractions of B10R cells when compared to B10S cells. Following phorbol myristate acetate (PMA) treatment and following the stimulation with Mycobacteria bovis BCG, PKC-specific activity increased significantly in membrane fractions of both B10R and B10S cells, but the absolute level was significantly greater in particulate fractions from B10R macrophages. Furthermore, B10R cells had a superior ability to phosphorylate endogenous substrates compared to B10S macrophages. Scatchard analysis of phorbol ester receptors revealed no differences between B10R and B10S cells. In contrast, the sensitivity of partially purified PKC from B10S cells to activation in vitro by diacylglycerol was decreased by approximately 50% when compared to enzyme from B10R cells. Western-blotting analysis using antibodies specific for PKC isoforms (alpha, beta, delta, epsilon, zeta and eta) showed similar levels of PKC isoforms present in B10R and B10S cells. To examine whether differences in PKC activity of B10R and B10S cells had functional consequences, the induction of c-fos gene expression was compared in the two cell lines. In response either to infection with M. bovis BCG or to stimulation with PMA, c-fos mRNA levels in B10R macrophages were increased 2-4-fold in comparison to B10S macrophages. Since we have previously found that the bacteriostatic activity of B10R and B10S macrophages correlated with the amount of nitric oxide produced by the macrophages, we have tested if the enhancement of PKC activity in these macrophages affects their ability to produce nitric oxide. We have found that interferon-gamma-(IFNgamma)-induced secretion of nitric oxide by B10R macrophages could be augmented a few fold by the activation of PKC whereas, in B10S macrophages stimulated with IFNgamma, nitric oxide release could be augmented by only about 10-20%. These results indicate that the differences in PKC activity between B10R and B10S macrophages may contribute to altered responsiveness to IFNgamma that results in different production of effector molecules crucial for bacteriostatic activity against M. bovis BCG.
Philadelphia-negative chronic myelogenous leukemia in a patient with a unique complex translocation: 46,XY,t(9;12;15)(q34;12;q21).
Chronic myelogenous leukemia (CML) is associated with an acquired karyotypic abnormality, the Philadelphia (Ph) chromosome, in 95% of cases. The Ph chromosome is the product of a balanced translocation that results in a hybrid gene that is considered essential for the pathogenesis of this disease. We have found a complex translocation involving chromosomes 9, 12, and 15 in a 42-year-old Haitian male with the clinical findings of CML. Complex translocations have been shown to result in the masking of the Ph chromosome. We used a mixture of two BCR-specific DNA probes for Southern blot analysis in order to test this hypothesis in our patient. High-molecular weight DNA was digested with the restriction enzymes BglII, BamHI and HindIII. The BglII digestion revealed the presence of two abnormal fragments of 3.9 and 3.0 kb and the BamHI digestion an abnormal 15-kb fragment. These data suggest there is a breakpoint in region 2 of M-bcr. The identification of this breakpoint confirms our hypothesis that a rearrangement involving 22q11 has occurred in the leukemic cells of our patient. A secondary translocation involving chromosomes 12 and 15 has hidden the effects of this translocation. Combined cytogenetic and molecular analysis establishes the karyotype of our patient as 46,XY,t(9;12;15;22)(q34;q12;q21;q11).
The ectopic maxillary canine: a case report.
A case is described of a severely ectopic maxillary canine, which underwent initial spontaneous improvement, and following extraction of the deciduous canine erupted satisfactorily. It provides a cautionary tale when considering surgical removal of ectopic maxillary canines.
Toxic equivalency factors (TEFs) for PCBs, PCDDs, PCDFs for humans and wildlife.
An expert meeting was organized by the World Health Organization (WHO) and held in Stockholm on 15-18 June 1997. The objective of this meeting was to derive consensus toxic equivalency factors (TEFs) for polychlorinated dibenzo-p-dioxins (PCDDs) and dibenzofurans (PCDFs) and dioxinlike polychlorinated biphenyls (PCBs) for both human, fish, and wildlife risk assessment. Based on existing literature data, TEFs were (re)evaluated and either revised (mammals) or established (fish and birds). A few mammalian WHO-TEFs were revised, including 1,2,3,7,8-pentachlorinated DD, octachlorinated DD, octachlorinated DF, and PCB 77. These mammalian TEFs are also considered applicable for humans and wild mammalian species. Furthermore, it was concluded that there was insufficient in vivo evidence to continue the use of TEFs for some di-ortho PCBs, as suggested earlier by Ahlborg et al. [Chemosphere 28:1049-1067 (1994)]. In addition, TEFs for fish and birds were determined. The WHO working group attempted to harmonize TEFs across different taxa to the extent possible. However, total synchronization of TEFs was not feasible, as there were orders of a magnitude difference in TEFs between taxa for some compounds. In this respect, the absent or very low response of fish to mono-ortho PCBs is most noticeable compared to mammals and birds. Uncertainties that could compromise the TEF concept were also reviewed, including nonadditive interactions, differences in shape of the dose-response curve, and species responsiveness. In spite of these uncertainties, it was concluded that the TEF concept is still the most plausible and feasible approach for risk assessment of halogenated aromatic hydrocarbons with dioxinlike properties.
Effect of time between farm loading and processing on carcase quality of broiler chickens.
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A Bayesian analysis of regression models with continuous errors with application to longitudinal studies.
We employ a regression model with errors that follow a continuous autoregressive process to analyse longitudinal studies. In this way, unequally spaced observations do not present a problem in the analysis. We employ a Bayesian approach, where our inferences are based on a direct resampling process that generates values from the posterior distribution of the parameters of the model. We illustrate these Bayesian inferences with an analysis of a longitudinal study that involves the regression of foetal head circumference on menstrual age. Using these same data, we contrast the Bayesian approach with a maximum likelihood technique.
Prognostic factors for patients with advanced seminoma treated with platinum-based chemotherapy.
Prognostic factors for 3-year progression-free survival (PFS) were defined in 286 patients with advanced seminoma treated with cisplatin-based chemotherapy at 10 European oncology units (no prior treatment: 236; prior radiotherapy: 50). Previously irradiated patients displayed a 69% PFS as compared to 87% in those presenting with advanced seminoma at the time of diagnosis (P = 0.009). In the univariate analysis, the extent and site of disease before chemotherapy and the level of serum LDH (< 2.0 versus > or = 2.0 x upper limit of normal) correlated with PFS in previously non-irradiated patients, but not in patients with prior radiotherapy. The multivariate analysis was, therefore, restricted to previously non-irradiated patients. The presence of non-pulmonary visceral metastases and a serum LDH level of > or = 2 x normal (N) proved to be independent prognostic factors. Based on these variables, two prognostic models were constructed and validated in an external data set of 166 comparable patients. For clinical use, Model 2 is recommended. The good-prognosis group comprises non-irradiated patients with stage II seminoma and any LDH level at presentation, or stage III and IV patients (with lung metastases only) whose serum LDH level is < 2 x N. These patients display a 94% 3-year PFS. The poor prognosis group includes all other patients with a 56% PFS. With this prognostic model, individualisation of the therapeutic approach may be considered in patients with advanced seminoma and a high risk of chemotherapy-related toxicity.
Treatment of recurrent malignant glioma with BCNU-fluosol and oxygen inhalation. A phase I-II study.
OBJECTIVES: To evaluate the toxicity and response rate following BCNU with oxygen inhalation and escalating dosages of fluosol administered to patients with radiographic progression of malignant glioma after definitive surgery and radiotherapy. METHOD: This single arm, phase I-II multicenter trial, enrolled 99 patients with malignant gliomas recurrent after definitive surgery and radiotherapy. All patients received a fixed dose (200 mg/m2) of BCNU along with 100% oxygen and fluosol, a perfluorochemical. Fluosol doses were escalated between patients (150, 275, 400 and 600 ml/m2). Treatment was repeated every 6 weeks for a maximum of 6 cycles. Patients were assessed for toxicity at the time of infusion and sequentially thereafter. Response was evaluated clinically and radiologically at least every 6 weeks. RESULTS: Treatment was well tolerated. Dose reductions were required at least once in 18 patients, treatment delays were necessary at least once in 33 patients. Grade 3-4 leukopenia occurred in 6 patients (12 events), grade 3-4 thrombocytopenia in 10 patients (25 events) and grade 3-4 liver enzymes elevations in 18 patients (31 events). Higher fluosol dosages did not produce increases in toxicity or responses. Response or stabilization was seen in 57% (38% were stabilizations) of the patients who entered the trial with progressive disease. The median time to progression was 45 weeks, and median survival was 66 weeks for patients who had response or stabilization. For patients with glioblastoma response/stabilization was seen in 45% with a mean duration of 24 weeks, for patients with anaplastic astrocytoma response/stabilization was seen in 68% with a mean duration of 50 weeks. CONCLUSION: This treatment regimen is well tolerated. Our results suggest fluosol may enhance the effectiveness of BCNU for the treatment of recurrent malignant gliomas. Future studies will be performed using fluosol at the dose of 400 ml/m2.
Single and double autotransplants for relapsing/refractory Hodgkin's disease: results of two consecutive trials.
To evaluate a strategy of one cycle of dose-intensive chemotherapy for patients with Hodgkin's disease in sensitive relapse and two cycles for those with refractory disease, 122 patients received dose-intensive chemotherapy followed by autotransplant in two consecutive studies. Patients with refractory disease were offered a second transplant with different conditioning in the absence of progression or excessive toxicity. CR was present after treatment in 46% while 16% died in the peritransplant period. Of 41 patients with primary refractory disease and 42 with refractory relapse, 24 and 21 respectively received a second cycle. Of these 45 refractory patients, 12 were in CR and 11 in PR after the first cycle and 10 of these 11 in PR achieved a durable CR with the second transplant. The CR rate is 37% in patients with refractory relapse and 19% in those with primary refractory disease. At a median follow-up of 4 years, median survival is 45 months. Progression-free survival of the refractory patients who could receive a second cycle was similar to that of patients with sensitive disease. A sequential transplant strategy is feasible. A subgroup of patients with refractory disease can achieve long-term survival after sequential BMT.
Sheet plastination as a clinically based teaching aid at the University of Auckland.
As a concerted move toward closer integration of the clinical and preclinical aspects of the undergraduate medical curriculum at the University of Auckland, the Department of Anatomy with Radiology has implemented a number of clinical procedures, pathological observations and diagnostic methods into the course resulting in a structured program of clinically based teaching of gross anatomy to second- and third-year medical students. The recently developed method of E12 epoxy resin sheet plastination of sectioned cadaver specimens has expanded the learning process twofold, firstly as a developmental microscopy teaching aid enabling us to link histological slides directly to gross anatomical specimens and secondly as a radiographic training tool in the correlation of clinical imaging techniques of magnetic resonance (MRI) and computed tomography (CT) with the teaching of gross anatomy. The direct comparison between E12 serial sectioned cadaver specimens and the equivalent MRI and CT images provides the student with a much clearer understanding of anatomical structures in relation to clinical diagnosis.
Current implant biomechanical concepts and controversies.
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Postoperative morbidity and mortality after D1 and D2 resections for gastric cancer: preliminary results of the MRC randomised controlled surgical trial. The Surgical Cooperative Group.
BACKGROUND: In Japan the surgical approach to treatment of potentially curable gastric cancer, including extended lymphadenectomy, seems in retrospective surveys to give better results than the less radical procedures favoured in Western countries. There has, however, been no evidence from randomised trials that extended lymphadenectomy (D2 gastric resection) confers a survival advantage. This question was addressed in a trial involving thirty-two surgeons in Europe. METHODS: In a prospective randomised controlled trial, D1 resection (level 1 lymphadenectomy) was compared with D2 resection (levels 1 and 2 lymphadenectomy). Central randomisation (200 patients in each arm) followed a staging laparotomy. FINDINGS: The D2 group had greater postoperative hospital mortality (13% vs 6.5%; p=0.04 [95% Cl 9-18% for D2, 4-11% for D1] and higher overall postoperative morbidity (46% vs 28%; p<0.001); their postoperative stay was also longer. The excess postoperative morbidity and mortality in the D2 group was accounted for by distal pancreaticosplenectomy and splenectomy. In the whole group (400 patients), survival beyond three years was 30% in patients whose gastrectomy included en-bloc pancreatico-splenic resection versus 50% in the remainder. INTERPRETATION: D2 gastric resections are followed by higher morbidity and mortality than D1 resections. These disadvantages are consequent upon additional pancreatectomies and distal splenectomies, and in long-term follow-up the higher mortality when the pancreas and spleen are resected may prove to nullify any survival benefit from D2 procedures.
Anterograde but not retrograde memory loss following combined mammillary body and medial thalamic lesions.
We report the first human case of combined mammillary body and medial thalamic lesions due to focal pathology. A patient presented with a multi-lobular lesion that affected the mammillary bodies, the medial thalamus and the brain stem. On neuropsychological testing, he showed significant anterograde memory impairment, with marked impairment on delayed story recall, but normal or only mildly impaired performance on retrograde memory tasks. Our findings corroborate the results of recent non-human lesion studies and indicate that some of the well-established features of the amnesic syndrome, such as severe retrograde amnesia, may not be due to primary diencephalic pathology. Significant retrograde amnesia may result from cortical pathology or from an interaction between cortical and subcortical pathology.
Cranial computed tomographic findings in tuberous sclerosis are not affected by sex.
Males with the tuberous sclerosis complex (TSC) have been shown to be more at risk of learning disorder than females. A retrospective population based study was undertaken to see if males with TSC have more cerebral lesions than females. 47 of 131 individuals with TSC identified in a prevalence study had a cranial CT scan available for review. No significant difference between the sexes was found in the number of subependymal nodules or the number, site or size of parenchymal lesions. However, males with five or fewer parenchymal lesions were at greater risk of learning disorder than females. The increased risk of learning disorder in males with tuberous sclerosis is not accounted for by an increase in number, site or size of cranial lesions as seen on CT scans. Therefore there must be an alternative mechanism to account for this difference between the sexes.
Stable and specific binding of heat shock protein 90 by geldanamycin disrupts glucocorticoid receptor function in intact cells.
When isolated from cells grown under hormone-free conditions, the glucocorticoid receptor (GR) is known to exist as a large heteroprotein complex that contains, among its multiple components, the stress protein hsp90 (heat shock protein 90). To explore hsp90's role in mediating glucocorticoid hormone action, we have examined the effects of a selective hsp90-binding agent, geldanamycin (GA), or GR structure and function. Using a steroid-responsive reporter construct, we found that GA inhibited the dexamethasone-dependent transactivating activity of GR in transfected cells. At the molecular level, GA bound hsp90, but not GR, in a stable and specific manner in intact cells. GA treatment of cells did not inhibit coprecipitation of hsp90 or hsp70 with the GR but did result in a complete loss of the recently described p23 protein from GR immunoprecipitates. This drug-induced alteration in GR heteroprotein complex composition was associated with a rapid (15-30 min), noncompetitive loss of dexamethasone-binding activity. Longer exposures of cells to GA (2-8 h), resulted in a marked decline in the cellular level of GR protein. Pulse-chase data revealed that this decline resulted from a decrease in GR protein stability, not rate of synthesis. GA-induced declines in GR protein level were blocked by cotreatment of cells with lactacystin, a selective inhibitor of 20S proteasome activity, suggesting the possible involvement of the ubiquitin-proteasome pathway in mediating GA-induced decreases in GR protein abundance. Overall, these findings provide direct pharmacological evidence that hsp90 function is required to maintain both the hormone-binding activity and stability of the GR protein in intact cells and suggest that hsp90 function may provide a novel target for the modulation of steroid hormone signaling.
Response of human multiple myeloma-derived cell lines to alkyl-lysophospholipid.
Clinical studies using high-dose chemotherapy and autologous bone marrow transplantation in multiple myeloma report improvement in tumor regression. The potential success of this approach may be compromised because of the myeloma cell burden. A promising approach is chemopurging of bone marrow with alkyl-lysophospholipids (ALP), a new class of antitumor agents. Using a limiting dilution assay, we tested ALP against several drug-resistant human myeloma cell lines: RPMI 8226/S, 8226/DOX40, 8226/LR-5, 8226/MR-20, 8226/DOX1V, and 8226/MDR10V. IC50 values were approximately 6 microg/mL, for all cell lines evaluated. ALP was effective against all cells, with mean log kills ranging from 2.48 to 4.95. Cytotoxicity was temperature-dependent. Survival curves of cell lines exposed to ALP showed a steep slope during the first 4 hours of exposure, with little additional cell kill after 24 hours. Dose-response curves after drug exposure for 4 and 24 hours reached plateaus at 75 and 25 microg/mL, respectively. Increasing concentrations of ALP caused a sustained increase in resting [Ca2+]i of sufficient magnitude to induce cell death. These results suggest that ALP is an effective cytotoxic agent even in cells known to be resistant to alkylating agents and other classes of cytotoxic drugs.
Replicative advantage in tissue culture of egg-adapted influenza virus over tissue-culture derived virus: implications for vaccine manufacture.
Influenza virus derived from clinical material on MDCK cells has been shown to possess haemagglutinin (HA) indistinguishable from that of the natural, uncultivated virus. In contrast, viurs derived in embryonated hens' eggs are variants with substitutions in their HA in the vicinity of the receptor binding site. We report here the superior growth of egg-adapted virus over cell-derived virus on MDCK cells in studies in which MDCK-derived virus was spiked with small amounts of egg-adapted virus and the mixture sequentially passaged on MDCK cells. Such egg-derived variants bind to and are internalized by MDCK cells with a much higher efficiency than cell-derived virus. These data imply that the natural virus, whilst able to replicate on MDCK cells, is by no means the best fit for the MDCK receptor and variants with appropriate substitutions around the receptor binding site can readily displace the natural virus. Vaccine manufacturers who are investigating the use of tissue culture for vaccine production should minimize passage levels of cell-derived virus and beware of the displacement of the original virus with variants similar to those derived in eggs, which are often antigenically distinct.