Biomedical subjects
Publications and source records attributed to M Johnston.
The yeast Saccharomyces cerevisiae is the pre-eminent organism for the study of basic functions of eukaryotic cells. All of the genes of this simple eukaryotic cell have recently been revealed by an international collaborative effort to determine the complete DNA sequence of its nuclear genome. Here we describe some of the features of chromosome XII.
BACKGROUND: Loading doses of warfarin that are larger than those used for maintenance therapy are widely used in clinical practice, but they have never been prospectively evaluated. OBJECTIVE: To compare the effect of 5- and 10-mg loading doses of warfarin on laboratory markers of warfarin's anticoagulant effect. DESIGN: Randomized clinical trial. SETTING: Tertiary care teaching hospital. PATIENTS: 49 patients seen over a 5-month period with a target international normalized ratio (INR) of 2.0 to 3.0. INTERVENTION: Patients were randomly assigned to receive an initial dose of 5 or 10 mg of warfarin. Subsequent doses of warfarin were administered on the basis of dosing nomograms. MEASUREMENTS: INRs and levels of factors II, VII, IX, and X and protein C were measured daily for 5 days. RESULTS: 11 of 25 patients in the 10-mg group (44% [95% CI, 34% to 54%]) and 2 of 24 patients in the 5-mg group (8% [CI, 3% to 14%]) had INRs greater than 2.0 at 36 hours (P = 0.005), at which time the factor VII levels were 27% (CI, 18% to 36%) in the 10-mg group and 54% (CI, 43% to 65%) in the 5-mg group (P < 0.001). In contrast, factor II levels were 74% (CI, 67% to 81%) in the 10-mg group and 82% (CI, 73% to 93%) in the 5-mg group (P > 0.2). At 60 hours, 9 of 25 patients in the 10-mg group (36% [CI, 17% to 54%]) and no patients in the 5-mg group had INRs greater than 3.0. At 84 hours, 15 of 24 patients in the 10-mg group (63% [CI, 43% to 81%]) and 19 of 24 patients in the 5-mg group (79% [CI, 62% to 95%]) had INRs between 2.0 and 3.0. Four patients in the 10-mg group and 1 patient in the 5-mg group received vitamin K for excessive prolongation of the INR. CONCLUSIONS: A 5-mg loading dose of warfarin produces less excess anticoagulation than does a 10-mg loading dose; the smaller dose also avoids the development of a potential hypercoagulable state caused by precipitous decreases in levels of protein C during the first 36 hours of warfarin therapy.
High levels of glucose repress expression of the SUC2 gene in the yeast Saccharomyces cerevisiae. We have discovered that low levels of glucose are required for maximal transcription of SUC2: SUC2 expression is induced about five- to ten-fold in cells growing on low levels of glucose (0.1%) compared to cells growing on galactose or glycerol. Two pieces of evidence suggest that this low-glucose-induced expression is mediated by a repression mechanism that involves an upstream repression site in the SUC2 promoter (URS(SUC2)). First, deletion of the URS(SUC2) results in expression of the SUC2 gene in the absence of glucose, and second the URS(SUC2) mediates a six-fold repression of a reporter gene when inserted into a heterologous promoter. However, this URS(SUC2) mediated repression occurs on all tested carbon sources, suggesting that this URS element acts in concert with all other promoter elements to respond to low concentrations of glucose. This repression requires the general repressor SSn6p. SNF3, which encodes a glucose transporter that appears to be a sensor of low levels of glucose, is also required for low-glucose-induced expression of SUC2.
OBJECTIVES: The International Registry of Lung Metastases was established in 1991 to assess the long-term results of pulmonary metastasectomy. METHODS: The Registry has accrued 5206 cases of lung metastasectomy, from 18 departments of thoracic surgery in Europe (n = 13), the United States (n = 4) and Canada (n = 1). Of these patients, 4572 (88%) underwent complete surgical resection. The primary tumor was epithelial in 2260 cases, sarcoma in 2173, germ cell in 363, and melanoma in 328. The disease-free interval was 0 to 11 months in 2199 cases, 12 to 35 months in 1857, and more than 36 months in 1620. Single metastases accounted for 2383 cases and multiple lesions for 2726. Mean follow-up was 46 months. Analysis was performed by Kaplan-Meier estimates of survival, relative risks of death, and multivariate Cox model. RESULTS: The actuarial survival after complete metastasectomy was 36% at 5 years, 26% at 10 years, and 22% at 15 years (median 35 months); the corresponding values for incomplete resection were 13% at 5 years and 7% at 10 years (median 15 months). Among complete resections, the 5-year survival was 33% for patients with a disease-free interval of 0 to 11 months and 45% for those with a disease-free interval of more than 36 months; 43% for single lesions and 27% for four or more lesions. Multivariate analysis showed a better prognosis for patients with germ cell tumors, disease-free intervals of 36 months or more, and single metastases. CONCLUSIONS: These results confirm that lung metastasectomy is a safe and potentially curative procedure. Resectability, disease-free interval, and number of metastases enabled us to design a simple system of classification valid for different tumor types.
G-CSF administration to normal donors results in granulocyte apheresis yields generally greater than those observed with other neutrophil mobilizing agents. In vitro, neutrophils cultured with G-CSF exhibit prolonged survival; however, the random migration of neutrophils exposed to this agent is inhibited. Although transfused neutrophils mobilized with agents other than G-CSF migrate to sites of inflammation or infection in vivo, this has yet to be demonstrated with infusion of G-CSF-mobilized neutrophils into neutropenic human subjects. Five neutropenic allogeneic bone marrow transplant (BMT) patients each received a fresh infusion of G-CSF-mobilized indium-labeled irradiated white blood cells (WBC) apheresed from HLA-matched normal donors on day +5 post-transplant. Localization of activity on delayed scintigraphic images of indium-labeled WBC scans to sites of tissue damage (oral/nasopharynx in two patients with mucositis and terminal ileum/cecum in one with diarrhea) occurred, and supports the hypothesis that G-CSF-mobilized HLA-matched donor neutrophils which have been irradiated are functional after infusion into neutropenic recipients.
A range of sensitivities exhibited by Escherichia coli O157 to cefixime and potassium tellurite are demonstrated. The sensitivity was shown by growth on cefixime tellurite sorbitol MacConkey agar and by the effect on the metabolic activity in glucuronate trimethylamine-oxide conductance broth. These antimicrobials are regularly used in the isolation of this pathogen from food and clinical specimens, and such sensitivity may lead to the reporting of false negative samples.
BACKGROUND: Granulocyte-colony-stimulating factor (G-CSF) is a safe and effective agent for mobilization of neutrophils in normal donors, consistently resulting in cell yields per leukapheresis (LA) procedure that are superior to those with other agents. LA components also contain platelets, whose clinical relevance is unknown. STUDY DESIGN AND METHODS: This study describes the kinetics of and analyzes the factors determining the ANC and platelet count increments seen with each of three transfusions of granulocytes collected from HLA-matched sibling donors receiving G-CSF (n = 10; maximum of 3 LA procedures/donor). The transfusions were given to recipients (n = 10) on alternate days beginning. Day 1 after allogeneic bone marrow transplant (BMT). RESULTS: Significant, sustained increments in the recipient ANCs were observed after the transfusion of G-CSF-mobilized LA components. The mean peak posttransfusion increments in the ANCs were 1195, 729, and 631 per microL with transfusion of donor LA components on Days 1, 3, and 5, respectively. The length of time that the mean posttransfusion ANC was at or above the baseline (pretransfusion) value was 25 to 37 hours, depending on the post-BMT day when the component was administered. No consistent relationship was observed between LA component granulocyte dose, baseline recipient ANC, or temperature elevation and post-transfusion ANC increments. Large numbers of platelets (mean, 2.55 x 10(11)) were present in LA components, and this resulted in significant increments from baseline in the mean platelet count 1 hour after LA component transfusions. Between Days 1 and 7, the duration of severe neutropenia was shorter and the percentage of patients requiring nondonor platelet transfusions was less in study patients who received LA component transfusions than in a similar historical control group who did not. CONCLUSION: The transfusion of G-CSF-mobilized, HLA-matched LA components to allogeneic BMT recipients resulted in significant and sustained increments in the ANC and the platelet count. Within the range examined, a relationship between neutrophil dose and an increment in the ANC was not demonstrated.
Glucose represses the transcription of many genes in bakers yeast (Saccharomyces cerevisiae). Mig1 is a Cys2-His2 zinc finger protein that mediates glucose repression of several genes by binding to their promoters and recruiting the general repression complex Ssn6-Tup1. We have found that the subcellular localization of Mig1 is regulated by glucose. Mig1 is imported into the nucleus within minutes after the addition of glucose and is just as rapidly transported back to the cytoplasm when glucose is removed. This regulated nuclear localization requires components of the glucose repression signal transduction pathway. An internal region of the protein separate from the DNA binding and repression domains is necessary and sufficient for glucose-regulated nuclear import and export. Changes in the phosphorylation status of Mig1 are coincident with the changes in its localization, suggesting a possible regulatory role for phosphorylation. Our results suggest that a glucose-regulated nuclear import and/or export mechanism controls the activity of Mig1.
The plasma recovery of an intraventricularly administered protein was compared before and after lymph diversion/ligation in the same conscious sheep to determine the relative roles of arachnoid villi and lymphatics in the clearance of a cerebral spinal fluid tracer. 125I-human serum albumin was injected into both lateral ventricles, and venous blood was sampled. One day later, multiple cervical vessels and the thoracic duct were cannulated for lymph collection. Uncannulated vessels were ligated. The experiment was repeated with 131I-human serum albumin as the tracer. Before lymph diversion/ligation, the time-averaged tracer transport into the plasma was 6.4 +/- 1.0%/h, with an average 6-h plasma recovery of 38.2 +/- 5.7% (percentage of injected dose). After lymph diversion/ligation, the values dropped to 2.9 +/- 0.5%/h and 17.7 +/- 2.7%, respectively. The collected lymph contained 8.7 +/- 2.6% of the tracer. No significant differences were observed in sham-operated animals. In conclusion, extracranial lymphatic vessels in sheep transport approximately one-half of the protein tracer from the cerebral spinal fluid compartment into plasma.
The success of cardiovascular disease (CVD) risk factor screening programmes depends heavily on good uptake rates and the extent of risk factors modification following screening. It is thus important to try to understand what affects people's decision to attend a screening appointment and their motivation to change their lifestyle, if necessary, following screening. This paper summarises three studies undertaken to describe uptake of screening and risk factor modification and to identify the psychological factors involved. The studies took place in two settings: a worksite mobile screening service and a city health centre screening programme. The results identified a number of sociodemographic, personal and organizational factors involved in screening behaviour which offer suggestions for effective planning of screening programmes.
This study demonstrated the in vitro effect of diazepam on human ciliary beat frequency (CBF) from fifteen normal subjects using brush cytology. Tube A contained culture medium, tube B the diluent that diazepam intravenous injectate was carried in and culture medium (controls). Tube C, D and E contained, 0.4 mg/l, 4.0 mg/l and 40.0 mg/l of diazepam in culture medium respectively. The mean effective diazepam concentration in plasma is 0.4 mg/l. CBF was measured photometrically. The most vigorous cilia were measured in 5 areas taking 10 readings on each sample, 30 minutes and 1 hour after mixing. Standard deviation (SD) and confidence limits were calculated along with significance testing (p < 0.05) using the paired t-test and ANOVA. The mean of the CBF of tubes A and B were 13.44 (SD 2.65) and 13.67 (SD 2.48). There was a reduction of the CBF with increasing concentrations of diazepam at 30 minutes, 11.32 (SD 2.14), 10.29 (SD 1.58) and 4.14 (SD 1.57) tubes C. D and E respectively. There was a significant lowering in CBF of 17% (p < 0.01) of diazepam at the mean effective plasma level (tube C) when compared against the controls. CBF decreased over time and at 1 hour was 10.57 (SD, 1.36), 9.02 (SD, 1.39) and 3.58 (SD, 1.31) tubes C, D and E respectively. A proposed mechanism of altered intracellular calcium flux via the action diazepam on GABA receptors is described.
Laparoscopic assisted vaginal hysterectomy (LAVH) is a procedure gaining in popularity over abdominal hysterectomy, for a number of different indications. According to a recent study by Wilcox, et al., over 1.7 million women underwent a hysterectomy procedure in the U.S. from 1988 through 1990, and two-thirds of the procedures for benign disease were performed for leiomyomata or endometriosis, indications typically afflicting women of child-bearing age. Seventy-five percent of the procedures included in the Wilcox study were abdominal hysterectomies, the remainder being vaginal hysterectomies. National Hospital Discharge Summary data reveal that 562,000 hysterectomies were performed in 1993, rendering hysterectomy second only to cesarean section in volume of annual Ob/Gyn surgical procedures performed.
Glucose is the preferred carbon source for most eukaryotic cells and has profound effects on many cellular functions. How cells sense glucose and transduce a signal into the cell is a fundamental, unanswered question. Here we describe evidence that two unusual glucose transporters in the yeast Saccharomyces cerevisiae serve as glucose sensors that generate an intracellular glucose signal. The Snf3p high-affinity glucose transporter appears to function as a low glucose sensor, since it is required for induction of expression of several hexose transporter (HXT) genes, encoding glucose transporters, by low levels of glucose. We have identified another apparent glucose transporter, Rgt2p, that is strikingly similar to Snf3p and is required for maximal induction of gene expression in response to high levels of glucose. This suggests that Rgt2p is a high glucose-sensing counterpart to Snf3p. We identified a dominant mutation in RGT2 that causes constitutive expression of several HXT genes, even in the absence of the inducer glucose. This same mutation introduced into SNF3 also causes glucose-independent expression of HXT genes. Thus, the Rgt2p and Snf3p glucose transporters appear to act as glucose receptors that generate an intracellular glucose signal, suggesting that glucose signaling in yeast is a receptor-mediated process.
The genome of the yeast Saccharomyces cerevisiae has been completely sequenced through a worldwide collaboration. The sequence of 12,068 kilobases defines 5885 potential protein-encoding genes, approximately 140 genes specifying ribosomal RNA, 40 genes for small nuclear RNA molecules, and 275 transfer RNA genes. In addition, the complete sequence provides information about the higher order organization of yeast's 16 chromosomes and allows some insight into their evolutionary history. The genome shows a considerable amount of apparent genetic redundancy, and one of the major problems to be tackled during the next stage of the yeast genome project is to elucidate the biological functions of all of these genes.
The green fluorescent protein (GFP) from the jellyfish Aequorea victoria has attracted much attention as a tool to study a number of biological processes. This study describes the use of GFP as a vital reporter molecule for localization and expression studies in Saccharomyces cerevisiae. Construction of GFP expression vectors which allow N- or C-terminal fusion of the gfp gene to a gene of interest allowed the generation of fusion proteins whose subcellular localization was followed by fluorescence microscopy in living yeast cells. Analysis of three unknown open reading frames obtained from the budding yeast chromosome XIV resulted in distinct staining patterns, allowing prediction of the cellular localization of these unknown proteins. Furthermore, GFP was used to construct a gene replacement cassette which, after homologous integration into the genomic locus, placed the gfp gene behind a promoter of interest. The amount of GFP produced from this promoter was then quantified in living yeast cells by flow cytometry. With this novel replacement cassette a gene of interest can be deleted and at the same time its expression level studied under various growth conditions. The experiments presented here suggest that GFP represents a convenient fluorescent marker for localization studies as well as gene expression studies in budding yeast. Systematic studies of a large number of genes should benefit from such assays.
The complete sequencing of the genome of a simple eukaryotic organism - the budding yeast Saccharomyces cerevisiae - is a milestone for biology, and sets the stage for a complete understanding of how a eukaryotic cell functions.
The background to this work was the expected increase in the proportion of elderly people in the population, giving rise to a greater demand for services at a time of scarcity of healthcare resources. The aims of the study were to investigate disability and health in a sample of elderly people living at home in the community. We wished to obtain detailed information about physical disability and health then to explore individual perceptions of health and disability and, finally, to examine relationships between the objective assessments and individual, subjective perceptions. The results reported here represent only part of the work of a larger study which also investigated the use of health and social services in this sample of elderly people. In-depth interviews were undertaken with a random stratified population sample of 200 elderly people living at home. Analysis of interview data showed that, by use of a multidimensional scale of disability, just over one-third of the sample could be classified as 'disabled'. The mobility status of the sample was, in general, good with only small numbers spending most of their time in a wheelchair. The total sample scored highly on activities of daily living, though those classified as disabled were significantly less independent. The mean number of reported health conditions was less than might have been expected in an elderly population. One-third of the subjects in the sample showed discrepancies between their own assessment of their disability and objective assessments. These discrepancies could partly be explained by a combination of individuals' assessment of their own abilities and their beliefs about factors influencing disability. Measures of disability, or activity associated with daily living, were related to perceptions of health, whereas the number of current health problems was not. The factors which determined perceptions of health or disability were: walking unaided and general levels of activity. Health was perceived in terms of what one could do rather than the presence of current diseases or conditions. The results suggested that when planning treatment for elderly people, physiotherapists and others should consider individual variability in perceptions of disability and health, as well as objective assessments. The importance of locomotion in the value systems of elderly people emphasises the high profile of physiotherapy in the care of elderly people.
We wish to stress the importance of remaining alert to the possibility of both depression and suicidality in the geriatric population. There is also a need for flexibility in assessing the elderly who may present with atypical symptoms of depression and who may not express their distress directly. The interaction of risk factors may be instrumental in determining the risk for suicide in a particular patient. In our case, we have identified an impaired ability to communicate, intractable tinnitus, and feelings of helplessness.