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Biomedical subjects

J Buxton

Publications and source records attributed to J Buxton.

At least 19 recordsLinked to original sources
PubMed2006

Effects of light level, time of harvest and position within field on the variability of tissue nitrate concentration in commercial crops of lettuce (Lactuca sativa) and endive (Cichorium endiva).

Seven commercial crops of lettuce and one crop of endive were sampled in order to study the variability in plant tissue nitrate concentration (TNC). Assuming that an appropriate sampling pattern was employed, ten plants were sufficient to give an acceptable estimate of the mean TNC. Short-term shading (24-48 h) had no significant effects on mean TNC, unlike the increase in TNC known to occur following dull periods 10-14 days before harvest. The effect on TNC of time of day harvested was significant, but there was no obvious pattern of diurnal variation. Averaged over all experiments, the coefficient of variation for TNC was in the order of 35%. Increasing the sample size from ten to 40 plants would only be expected to decrease the standard error of measurement of TNC from 16 to 12% of the mean because of the underlying analytical error, which would remain constant.

Agriculture↗
PubMed2004

Risk factors for Salmonella typhimurium DT104 and non-DT104 infection: a Canadian multi-provincial case-control study.

To identify risk factors for sporadic Salmonella Typhimurium definitive phage-type 104 (DT104) and non-DT104 diarrhoeal illness in Canada, we conducted a matched case-control study between 1999 and 2000. Cases were matched 1:1 on age and province of residence. Multivariate analysis suggested that recent antibiotic use [odds ratio (OR) 5.2, 95% confidence interval (CI) 1.8-15.3], living on a livestock farm (OR 4.9, 95% CI 1.9-18.9), and recent travel outside Canada (OR 4.1, 95% CI 1.2-13.8) are independent risk factors for DT104 illness. Similar analyses suggested that recent travel outside North America is a sizable risk factor for non-DT104 illness (OR 66.8, 95% CI 6.7-665.3). No food exposure was a risk factor in either analysis. Educating health-care providers and the public about appropriate antibiotic use and antimicrobial resistance is important. Appropriate administration of antibiotics to livestock, particularly cattle, and hygienic measures such as handwashing after contact with farm animals may reduce risk. Travel represents an important and probably underestimated risk factor for sporadic illness with S. Typhimurium. Improved national surveillance and detailed investigation of travel-related illness are required.

Adolescent↗
PubMed2001

Signaling complexes of the FERM domain-containing protein GRSP1 bound to ARF exchange factor GRP1.

GRP1 is a member of a family of proteins that contain a coiled-coil region, a Sec7 homology domain with guanosine nucleotide exchange activity for the ARF GTP-binding proteins, and a pleckstrin homology domain at the C terminus. The pleckstrin homology domain of GRP1 binds phosphatidylinositol (3,4,5) trisphosphate and mediates the translocation of GRP1 to the plasma membrane upon agonist stimulation of PI 3-kinase activity. Using a (32)P-labeled GRP1 probe to screen a mouse brain cDNA expression library, we isolated a cDNA clone encoding a GRP1-binding partner (GRSP1) that exists as two different splice variants in brain and lung. The GRSP1 protein contains a FERM protein interaction domain as well as two coiled coil domains and may therefore function as a scaffolding protein. Mapping experiments revealed that the interaction of GRP1 and GRSP1 occurs through the coiled coil domains in the two proteins. Immunodepletion experiments indicate that virtually all of the endogenous GRSP1 protein exists as a complex with GRP1 in lung. When co-expressed in Chinese hamster ovary cells expressing the human insulin receptor, both proteins display a diffuse, cytoplasmic localization. Acute translocation and co-localization of GRSP1 and GRP1 to ruffles in the plasma membrane was evident after insulin stimulation. These results identify GRSP1 as a novel member of GRP1 signaling complexes that are acutely recruited to plasma membrane ruffles in response to insulin receptor signaling.

ADP-Ribosylation Factors↗
PubMed2001

Insulin action on GLUT4 traffic visualized in single 3T3-l1 adipocytes by using ultra-fast microscopy.

A novel imaging technology, high-speed microscopy, has been used to visualize the process of GLUT4 translocation in response to insulin in single 3T3-L1 adipocytes. A key advantage of this technology is that it requires extremely low light exposure times, allowing the quasi-continuous capture of information over 20-30 min without photobleaching or photodamage. The half-time for the accumulation of GLUT4-eGFP (enhanced green fluorescent protein) at the plasma membrane in a single cell was found to be of 5-7 min at 37 degrees C. This half-time is substantially longer than that of exocytic vesicle fusion in neuroendocrine cells, suggesting that additional regulatory mechanisms are involved in the stimulation of GLUT4 translocation by insulin. Analysis of four-dimensional images (3-D over time) revealed that, in response to insulin, GLUT4-eGFP-enriched vesicles rapidly travel from the juxtanuclear region to the plasma membrane. In nontransfected adipocytes, impairment of microtubule and actin filament function inhibited insulin-stimulated glucose transport by 70 and 50%, respectively. When both filament systems were impaired insulin-stimulated glucose transport was completely inhibited. Taken together, the data suggest that the regulation of long-range motility of GLUT4-containing vesicles through the interaction with microtubule- and actin-based cytoskeletal networks plays an important role in the overall effect of insulin on GLUT4 translocation.

3T3 Cells↗
PubMed2000

A randomised, prospective, phase III clinical trial of primary bleomycin, ifosfamide and cisplatin (BIP) chemotherapy followed by radiotherapy versus radiotherapy alone in inoperable cancer of the cervix.

BACKGROUND: Phase II studies have shown primary (neo-adjuvant) chemotherapy with bleomycin, ifosfamide and cisplatin (BIP) is active against inoperable cervical cancer. We present here results of a randomised phase III multicentre trial comparing radical radiotherapy with neo-adjuvant BIP chemotherapy followed by radical radiotherapy in patients with inoperable cervical cancer, designed to discover whether this combination might improve survival. PATIENTS AND METHODS: Patients with inoperable cervical carcinoma were randomised to pelvic radiotherapy alone [RT] or two to three cycles of bleomycin 30 units/24-hour infusion, ifosfamide 5 g/m2/24 hours, and cisplatin 50 mg/m2) chemotherapy followed by pelvic radiotherapy (BIP + RT). Randomisation was stratified by stage and radiotherapy centre. RESULTS: One hundred seventy-two eligible women were randomised into this trial; eighty-six to RT and eighty-six to BIP + RT. A total of 190 cycles of chemotherapy were given. Median follow-up for the 47 patients still alive is 9 years with a minimum follow-up of 3 years. Complete or partial response occurred in 51 of 86 (59%) of those randomised to RT and 60 of 86 (69%) of those randomised to BIP + RT. The difference between response rates does not reach statistical significance (chi2 = 2.06, P = 0.15). Median survival is two years with an actuarial survival at five years of 32% (95% confidence interval (95% CI): 25%-39%). There is no significant difference between the treatment groups (chi2log-rank = 0.11, P = 0.74). CONCLUSIONS: This study does not show any survival benefit from the use of neo-adjuvant BIP chemotherapy in advanced cervical cancer.

Adult↗
PubMed2000

Falling through the cracks. Women's experiences of ineligibility for genetic testing for risk of breast cancer.

OBJECTIVE: To describe experiences of women seeking information about their risk of hereditary breast cancer who fail to meet strict eligibility criteria for genetic counseling and testing. DESIGN: Qualitative descriptive study. SETTING: Hereditary cancer program in western Canada. PARTICIPANTS: Women who had received notification of their ineligibility for referral for hereditary breast cancer risk assessment (n = 20) and some of their referring physicians (n = 10). Of 28 attempted contacts, five women had moved, one declined the invitation to participate, and two could not be interviewed because of scheduling conflicts. Ten of 20 physicians declined the invitation to participate. METHOD: In-depth, open-ended telephone interviews were conducted. Transcribed interviews were systematically analyzed to identify salient themes. MAIN FINDINGS: Three themes emerged. The first theme, "It's always on your mind," points to the profound concern about breast cancer that underlies women's experiences in seeking genetic testing. The second theme, "A test is a test," reflects women's beliefs that the test was relatively simple and similar to other medical tests in that it would provide a definitive answer. The third theme, "Falling through the cracks," captures the experience of ineligibility. Women reacted with a range of emotional responses and were left frustrated in their search for more specific information about their personal risk for breast cancer. Although women were encouraged to contact their physicians, few did. CONCLUSION: These findings point to the psychological consequences in women who seek genetic testing for risk of breast cancer when they are told they are ineligible and they are not given adequate information and support.

Adult↗
PubMed1998

Patients' experiences using a computerized program with a touch-sensitive video monitor for the assessment of health-related quality of life.

This study assessed patients' experiences using a computerized program with a touch-sensitive video monitor (TSVM) for the assessment of health-related quality of life (HRQoL). A software program was developed for a computerized form of the European Organization for Research and Treatment of Cancer Quality of Life Questionnaire, the QLQ-C30. One hundred and seventy-eight patients completed the QLQ-C30, followed by a structured interview designed to assess perceived difficulties with the use of the TSVM. Patients were asked to evaluate the ease of use of different aspects of the TSVM system (using the touch-sensitive screen, entering the patient identification number, reading the screen and following the on-screen instructions). The majority of patients found all aspects of the TSVM system 'very easy' to use. A few patients (1-2%) admitted finding any aspect of the TSVM use 'somewhat difficult' and none ranked any aspect as 'very difficult'. There were no unanswered items in the QLQ-C30. All patients found the amount of time spent on answering the questionnaire acceptable, (the mean time to complete was 6.8 min with a median of 5 min) and 96% stated they were willing to complete a similar questionnaire on a future occasion. From the patients' perspective the TSVM system appears to be a highly acceptable approach for the collection of HRQoL data in clinical practice.

Aged↗
PubMed1997

Interactions of Drosophila Cbl with epidermal growth factor receptors and role of Cbl in R7 photoreceptor cell development.

The human proto-oncogene product c-Cbl and a similar protein in Caenorhabditis elegans (Sli-1) contain a proline-rich COOH-terminal region that binds Src homology 3 (SH3) domains of proteins such as the adapter Grb2. Cb1-Grb2 complexes can be recruited to tyrosine-phosphorylated epidermal growth factor (EGF) receptors through the SH2 domain of Grb2. Here we identify by molecular cloning a Drosophila cDNA encoding a protein (Drosophila Cbl [D-Cbl]) that shows high sequence similarity to the N-terminal region of human c-Cbl but lacks proline-rich sequences and fails to bind Grb2. Nonetheless, in COS-1 cells, expression of hemagglutinin epitope-tagged D-Cbl results in its coimmunoprecipitation with EGF receptors in response to EGF. EGF also caused tyrosine phosphorylation of D-Cbl in such cells, but no association of phosphatidylinositol 3-kinase was detected in assays using anti-p85 antibody. A point mutation in D-Cbl (G305E) that suppresses the negative regulation of LET-23 by the Cbl homolog Sli-1 in C. elegans prevented tyrosine phosphorylation of D-Cbl as well as binding to the liganded EGF receptor in COS-1 cells. Colocalization of EGF receptors with both endogenous c-Cbl or expressed D-Cbl in endosomes of EGF-treated COS-1 cells is also demonstrated by immunofluorescence microscopy. In lysates of adult transgenic Drosophila melanogaster, GST-DCbl binds to the tyrosine-phosphorylated 150-kDa torso-DER chimeric receptor. Expression of D-Cbl directed by the sevenless enhancer in intact Drosophila compromises severely the development of the R7 photoreceptor neuron. These data suggest that despite the lack of Grb2 binding sites, D-Cbl functions as a negative regulator of receptor tyrosine kinase signaling in the Drosophila eye by a mechanism that involves its association with EGF receptors or other tyrosine kinases.

Adaptor Proteins, Signal Transducing↗
PubMed1997

Clinical-neurologic, cytogenetic and molecular aspects of the Prader-Willi and Angelman syndromes.

The Prader-Willi syndrome (PWS) and the Angelman syndrome (AS) are human neurogenetic disorders involving the imprinting mechanism, at the 15q11-13 chromosome region. The predominant genetic defects in PW are 15q11-13 deletions of paternal origin and maternal chromosome 15 uniparental disomy. In contrast, maternal deletions and paternal chromosome 15 uniparental disomy are associated with a different neurogenetic disorder, the AS. In both disorders, these mutations are associated with parent-of-origin specific methylation at several 15q11-13 loci. We studied 5 patients suspect of PWS and 4 patients suspect of AS who were referred to the Medical Genetics Unit at the University Hospital of Medical School from Ribeirão Preto. Our objective was to establish the correct clinical and etiological diagnosis in these cases. We used conventional cytogenetics, methylation analysis with the probe KB17 (CpG island of the SNRPN gene) by Southern blotting after digestion with the Xba I and Not I restriction enzymes. We studied in patients and their parents the segregation of the (CA)*** repeats polymorphisms by PCR, using the primers 196 and IR4-3R. All the patients had normal conventional cytogenetical analysis. We confirmed 3 cases of PWS: one by de novo deletion, one by maternal chromosome 15 uniparental disomy and one case with no defined cause determined by the used primers. We confirmed 2 cases of AS, caused by de novo deletion at the 15q11-13 region, and one case with normal molecular analysis but with strong clinical characteristics.

Adolescent↗
PubMed1996

Imprint switching on human chromosome 15 may involve alternative transcripts of the SNRPN gene.

Imprinting on human chromosome 15 is regulated by an imprinting centre, which has been mapped to a 100-kb region including exon 1 of SNRPN. From this region we have identified novel transcripts, which represent alternative transcripts of the SNRPN gene. The novel exons lack protein coding potential and are expressed from the paternal chromosome only. We have also identified intragenic deletions and a point mutation in patients who have Angelman or Prader-Willi syndrome due to a parental imprint switch failure. This suggests that imprint switching on human chromosome 15 may involve alternative SNRPN transcripts.

Alternative Splicing↗
PubMed1994

Differential intracellular localizations of GDP dissociation inhibitor isoforms. Insulin-dependent redistribution of GDP dissociation inhibitor-2 in 3T3-L1 adipocytes.

Insulin action on fat cell/skeletal muscle glucose transporter (GLUT4) redistribution to cell surface membranes appears to involve small GTP-binding proteins. It has been recently recognized that at least two GDP dissociation inhibitor (GDI) isoforms, GDI-1 and GDI-2, can bind and release GDP-bound Rab proteins from membranes (Shisheva, A., Südhof, T.C., and Czech, M. P. (1994) Mol. Cell. Biol. 14, 3459-3468). The present studies show that a strikingly higher level of GDI-2 fractionates with total membranes of COS-1 cells, Chinese hamster ovary cells, and 3T3-L1 adipocytes compared to GDI-1, which is virtually totally cytosolic. In 3T3-L1 adipocytes, most of the membrane-bound GDI-2 was present in a low density, intracellular membrane fraction. Immunodepletion of GLUT4-enriched vesicles from this membrane fraction also depleted significant amounts of GDI-2 proteins. Localization of both GDI-2 and GLUT4 in the same perinuclear regions of these cells was established by immunofluorescence microscopy, whereas GDI-1 displayed a diffuse, cytoplasmic distribution. Insulin acutely decreased both GLUT4 and GDI-2 protein levels in the low density microsomes by about 50%. Concomitantly, GLUT4 but not GDI-2 protein content of plasma membranes increased, suggesting release of GDI-2 into the cytoplasm in response to insulin. Taken together, these data suggest functional differences for the GDI-1 and GDI-2 protein isoforms, as well as a potential role of GDI-2 in the action of insulin on membrane movements.

3T3 Cells↗
PubMed1994

Insulin-sensitive association of GLUT-4 with endocytic clathrin-coated vesicles revealed with the use of brefeldin A.

The interaction of the adipocyte/skeletal muscle glucose transporter (GLUT-4) with clathrin lattices may be important in maintaining its intracellular distribution. To better understand the role of clathrin lattices in the sorting of GLUT-4, we have attempted to determine the cellular origin of clathrin-coated vesicles (CCVs) that contain this transporter. The fungal toxin brefeldin A (BFA) causes the selective disassembly of clathrin lattices at the trans-Golgi network (TGN), but not at the plasma membrane (PM), thus providing a way of estimating the proportion of GLUT-4 in PM- versus TGN-derived clathrin lattices. Exposure of 3T3-L1 adipocytes to BFA resulted in a rapid disassembly of clathrin lattices at the TGN, observed by optical sectioning microscopy, and to a pronounced decrease in the yield of CCVs purified from these cells. Thus, CCVs isolated from BFA-treated cells are likely to be derived from the PM. Immunoblotting experiments revealed the presence of GLUT-4 in such CCVs, suggesting that under basal conditions the transporter is continually retrieved from the PM through the CCV pathway. Exposure of both BFA-treated or non-treated cells to insulin resulted in a 4-6-fold increase in the concentration of GLUT-4 at the PM. In parallel, the concentration of GLUT-4 in PM-derived CCVs decreased by 60%. These results suggest (a) that the effect of insulin to increase the cell surface concentration of GLUT-4 is not inhibited by BFA, and (b) that a decreased association of GLUT-4 with endocytic CCVs may be important in facilitating its increased cell surface concentration in response to the hormone.

3T3 Cells↗