Biomedical subjects

F Walker

Publications and source records attributed to F Walker.

At least 55 records · Page 3Linked to original sources

PubMed1998

Hibernoma development in transgenic mice identifies brown adipose tissue as a novel target of aldosterone action.

Aldosterone is a major regulator of salt balance and blood pressure, exerting its effects via the mineralocorticoid receptor (MR). To analyze the regulatory mechanisms controlling tissue-specific expression of the human MR (hMR) in vivo, we have developed transgenic mouse models expressing the SV40 large T antigen (TAg) under the control of each of the two promoters of the hMR gene (P1 or P2). Unexpectedly, all five P1-TAg founder animals died prematurely from voluminous malignant liposarcomas originating from brown adipose tissue, as evidenced by the expression of the mitochondrial uncoupling protein ucp1, indicating that the proximal P1 promoter was transcriptionally active in brown adipocytes. No such hibernoma occurred in P2-TAg transgenic mice. Appropriate tissue-specific usage of P1 promoter sequences was confirmed by demonstrating the presence of endogenous MR in both neoplastic and normal brown adipose tissue. Several cell lines were derived from hibernomas; among them, the T37i cells can undergo terminal differentiation into brown adipocytes, which remain capable of expressing ucp1 upon adrenergic or retinoic acid stimulation. These cells possess endogenous functional MR, thus providing a new model to explore molecular mechanisms of mineralocorticoid action. Our data broaden the known functions of aldosterone and suggest a potential role for MR in adipocyte differentiation and regulation of thermogenesis.

Adipose Tissue, Brown↗

PubMed1998

The Raf-1 protein mediates insulin-like growth factor-induced proliferation of erythroid progenitor cells.

Previous studies from this and other laboratories have shown that insulin-like growth factor-1 (IGF-I) and insulin-like growth factor-2 (IGF-II) support erythroid colony formation in cultures supplemented with serum substitute and recombinant erythropoietin. Subpopulations of IGF-I- and IGF-II-dependent, erythropoietin-independent colony-forming unit-erythroid (CFU-E)-derived colonies and BFU-E-derived colonies were identified under serum-substituted conditions for adult bone-marrow-derived erythroid progenitors which proliferate in the absence and presence of exogenous anti-erythropoietin receptor monoclonal antibody and in serum-substituted medium that was preadsorbed with anti-erythropoietin IgG. To assess whether Raf-1 is required for the formation of IGF-dependent, erythropoietin-independent human erythroid colonies, 5-15 microM sense or antisense oligomer to raf-1 were added to serum-substituted cultures containing either 2 U/ml recombinant human erythropoietin (rHuEpo) alone or 0-1,000 ng/ml IGF-I or IGF-II with/without 2 U/ml rHuEpo. Both erythropoietin-induced and IGF-induced erythroid colony formation were completely blocked by antisense (but not sense) oligomers to raf-1. Purified human CFU-Es were examined for Raf-1 message and protein. Total RNA was extracted, and raf-1 mRNA was detected on Northern blots. Furthermore, a 74 kD protein, corresponding to Raf-1, was also detected in CFU-Es purified from human adult sources. Together, these studies support the hypothesis that the Raf-1 protein mediates both erythropoietin-induced and IGF-induced signal transduction in human erythroid progenitor cells.

Adult↗

PubMed1998

Activation of the Ras/mitogen-activated protein kinase pathway by kinase-defective epidermal growth factor receptors results in cell survival but not proliferation.

Signalling by the epidermal growth factor (EGF) receptor (EGFR) has been studied intensively, but for most cell types the analysis is complicated by the fact that EGFR not only homodimerizes but can also form heterodimers with other EGFR family members. Heterodimerization is a particular problem in the study of EGFR mutants, where the true phenotype of the mutants is confounded by the contribution of the heterodimer partner to signal transduction. We have made use of the murine hemopoietic cell line BaF/3, which does not express EGFR family members, to express wild-type (WT) EGFR, three kinase-defective EGFR mutants (V741G, Y740F, and K721R), or a C-terminally truncated EGFR (CT957) and have measured their responses to EGF. We found that under the appropriate conditions EGF can stimulate cell proliferation of BaF/3 cells expressing WT or CT957 EGFRs but not that of cells expressing the kinase-defective mutants. However, EGF promotes the survival of BaF/3 cells expressing either of the kinase-defective receptors (V741G and Y740F), indicating that these receptors can still transmit a survival signal. Analysis of the early signalling events by the WT, V741G, and Y740F mutant EGF receptors indicated that EGF stimulates comparable levels of Shc phosphorylation, Shc-GRB-2 association, and activation of Ras, B-Raf, and Erk-1. Blocking the mitogen-activated protein kinase (MAPK) signalling pathway with the specific inhibitor PD98059 abrogates completely the EGF-dependent survival of cells expressing the kinase-defective EGFR mutants but has no effect on the EGF-dependent proliferation mediated by WT and CT957 EGFRs. Similarly, the Src family kinase inhibitor PP1 abrogates EGF-dependent survival without affecting proliferation. However blocking phosphatidylinositol-3-kinase or JAK-2 kinase with specific inhibitors does arrest growth factor-dependent cell proliferation. Thus, EGFR-mediated mitogenic signalling in BaF/3 cells requires an intact EGFR tyrosine kinase activity and appears to depend on the activation of both the JAK-2 and PI-3 kinase pathways. Activation of the Src family of kinases or of the Ras/MAPK pathway can, however, be initiated by a kinase-impaired EGFR and is linked to survival.

Adaptor Proteins, Signal Transducing↗

PubMed1998

Biochemical characterization of mutant EGF receptors expressed in the hemopoietic cell line BaF/3.

The Epidermal Growth Factor (EGF) receptor appears to require a fully active tyrosine kinase domain to transmit mitogenic signals. However, waved-2 mice carrying a mutation in the alpha-helix C of their EGF-R, which abolishes tyrosine kinase activity, only display a mild phenotype and are fully viable. This suggests that the mutant EGF-R signals through heterodimerization with endogenous, kinase active members of the EGF-R family such as ErbB-2 or ErbB-4. We have examined the biochemistry of EGF-Rs carrying mutations in the alpha-helix C of the human EGF-R (V741G and Y740F), in the ATP binding site (K721R) and at the C-terminus (CT957), by expression in BaF/3 cells which are devoid of EGF-R family members. The in vitro kinase activity of the alpha-helix C EGF-R mutants was severely impaired as a result of reduced phosphotransfer activity without appreciable changes in the affinity for either ATP or peptide substrate. Surprisingly, EGF stimulation of cells carrying the different mutant or wild type EGF-Rs resulted in tyrosine phosphorylation of EGF-R proteins; this phosphorylation was abolished in crude plasma membrane preparations, and appears to be due to activation of a membrane-associated or a cytosolic kinase. Receptor-mediated internalization of EGF was profoundly suppressed in the V741G, K721R and CT957 receptor mutant, and high affinity EGF binding was undetectable in the V741G and K721R receptors. We conclude that specific residues in the C-helix of the EGF-R kinase are essential for full kinase activity; mutations in this region do not affect ATP binding, but impair the receptors' phosphotransfer ability. High affinity binding of EGF is not dependent on tyrosine kinase activity or sequences in the C-terminus.

Adenosine Triphosphate↗

PubMed1997

A model for the activation of the epidermal growth factor receptor kinase involvement of an asymmetric dimer?

The binding of epidermal growth factor (EGF) to its receptor leads to receptor dimerization, which activates the intracellular kinase domain. Homology models of the inactive and active forms of the EGF-receptor kinase domains have been derived, and these models suggest that the active form can be stabilized by the interaction of helix C and the surrounding area in one receptor monomer with one of two possible complementary surfaces on a second receptor monomer. Both hydrophobic interaction sites are strongly conserved within the EGF-receptor family but not in other tyrosine kinases. Two of the three predicted kinase dimers are symmetric; the other is asymmetric and is predicted to contain only one active kinase. One of the symmetric models and the asymmetric model would account for the effects of two mutations in helix C (Y740F and V741G) on kinase activity. They also provide an explanation for previously reported dominant negative mutants of the EGF receptor and have interesting implications for the signaling through homo- and heterodimers of the family members: EGF receptor, erbB2, erbB3, and erbB4.

Amino Acid Sequence↗

PubMed1997

A highly sensitive detection method for immunohistochemistry using biotinylated tyramine.

A highly sensitive method for light microscopic immunohistochemistry is described. The increased sensitivity compared with current methodologies is based on the horseradish peroxidase-catalysed deposition of biotinylated tyramine at the sites of immunoreactivity, followed by the detection of the biotin with streptavidin biotin horseradish peroxidase complex. This method is of general applicability in immunohistochemistry and has several important advantages over currently used immunohistochemical detection procedures. The most significant advantage is that several antibodies which to data have been non-reactive, even in antigen-retrieval formalin-fixed, was-embedded sections, now show strong and reproducible immunoreactivity using biotinylated tyramine amplification. In addition, many other antibodies can be used at significantly higher dilutions.

Antigens, CD↗

PubMed1997

[Educational strategies and tools in a public health program at the University of Geneva].

In the Swiss context, the newly developed MPH programme at the University of Geneva is experimental in educational matters. Indeed the programme is fully learner-centered and community-oriented. Throughout the curriculum students plan, implement and evaluate intervention programmes or/and research projects related to health problems of the communities they are in charge of. In this article, we describe the educational strategies and tools used in this MPH curriculum (professional profile, mind-mapping procedures, field-work either on research projects or on intervention programmes, group work and evaluation procedures). These strategies and tools might assist some educational experimentation in MPH programmes in search of public health relevance and pedagogic efficacy.

Certification↗

PubMed1997

Developmental expression and functionality of hepatocyte growth factor and c-Met in human fetal digestive tissues.

BACKGROUND & AIMS: Hepatocyte growth factor (HGF) and its receptor c-Met are presumed to play a morphogenic role during embryogenesis. The possible implication of HGF and c-Met during the digestive system development was approached by investigating their ontogeny, distribution, and functionality in human fetal tissues. METHODS: Thirty fetuses, 7-24 weeks old, were obtained. HGF and c-Met messenger RNAs and proteins were detected in liver, pancreas, esophagus, stomach, and small and large intestine. Tyrosine phosphorylation assays were realized on homogenates and membrane preparations from fetal tissues. RESULTS: The temporal appearance of HGF and c-Met was established between 7 and 8 weeks of gestation in digestive tissues. Immunoblot analysis showed the presence of the c-Met beta-subunit 145-kilodalton band and of the HGF alpha-subunit 70-kilodalton band. c-Met was localized in epithelia, especially in fundic parietal cells, pancreatic and gut endocrine cells, and in muscular layers. HGF immunoreactivity was first detected in epithelia and then in mesenchyme and muscular layers. In young fetal stages, the c-Met immunoprecipitated 145-kilodalton band showed tyrosine phosphorylation after HGF stimulation. CONCLUSIONS: This study provides evidence for HGF and c-Met expression early in all human fetal digestive tissues and implicates HGF-c-Met in the digestive system morphogenesis.

Digestive System↗

PubMed1997

Observer variability in the histopathological reporting of needle biopsy specimens of the prostate.

The Scottish Pathology Consistency Group has in previous studies examined the consistency of histopathological reporting of biopsies from the cervix, bladder, bronchus, and rectum. In the current study, consisting of 100 needle biopsy specimens of the prostate, a single hematoxylin-eosin (H&E) slide from each case was circulated in batches of 10 to the 12 pathologists, who filled in a simple proforma. This had two sections: a diagnostic category (benign; suspicious or malignant) along with a standard Gleason score for those regarded as malignant. The majority diagnosis of the 100 cases was benign, 53; suspicious, 1; and malignant, 46. The Kappa value for benign cases versus others was 0.86 and for malignant cases versus others was 0.91. Analysis of the data on Gleason scores showed a value of 0.54 when cases were divided into two categories (2 to 6 v 7 to 10) and 0.41 when three categories were used (2 to 4; 5 to 6; 7 to 10). Although not initially part of the design of the study, the majority diagnosis was compared with the original reported diagnosis. In a small subset, examination of further levels, basal cell antibody staining, along with further clinical information, was obtained. With this added information, it appears that there were probably 52 benign and 48 malignant cases. Of the 48 malignant cases, the group majority diagnosis was malignant, 46; suspicious, 1; and benign, 1. The original reported diagnosis was 56 benign, 1 suspicious, and 43 malignant. The group therefore appeared to perform better than the original reporting pathologists. When compared with the results of our previous studies, this study has shown that the diagnosis of carcinoma of the prostate on a needle biopsy is robust.

Biopsy, Needle↗

PubMed1997

Persistence of hepatitis B virus DNA in serum and liver from patients with chronic hepatitis B after loss of HBsAg.

BACKGROUND/AIMS: The persistence of serum and liver hepatitis B virus (HBV) sequences in patients with chronic hepatitis B after loss of HBsAg has already been described. We have attempted to elucidate the significance of these HBV sequences after loss of HBsAg. METHODS: Fifteen patients were studied. We looked for serum and liver HBV DNA, using polymerase chain reaction (PCR) with different sets of primers and in situ hybridization. The sedimentation velocity of serum HBV DNA was measured on a gradient of sucrose in two patients. RESULTS: Serum HBV DNA was detected by PCR in four of the 14 patients tested at 0 months after loss of HBsAg, two patients remained HBV DNA positive until 12 months, and none was positive at 24 months. The sedimentation velocity of serum HBV DNA in sucrose was relatively similar to that of a chronic HBV carrier with active viral replication. Liver HBV DNA was demonstrated by PCR in all 15 patients and by in situ hybridization in six patients. CONCLUSIONS: Our results show that: 1) HBV DNA may persist in the serum in a minority of patients and may be associated with circulating viral particles; 2) HBV DNA persists in the liver in all patients and its extrachromosomal localization was shown by in situ hybridization technique in some cases. These results suggest the persistence of low-level HBV replication after loss of HBsAg.

Adult↗

PubMed1997

Use of a new mouse model of Acinetobacter baumannii pneumonia to evaluate the postantibiotic effect of imipenem.

Acinetobacter baumannii is responsible for severe nosocomial pneumonia. To evaluate new therapeutic regimens for infections due to multiresistant strains and to study the pharmacodynamic properties of various antibiotics, we developed an experimental mouse model of acute A. baumannii pneumonia. C3H/HeN mice rendered transiently neutropenic were infected intratracheally with 5 x 10(6) CFU of A. baumannii. The mean log10 CFU/g of lung homogenate (+/- the standard deviation) were 9 +/- 0.9, 9.4 +/- 0.8, 8.6 +/- 1.2, and 7.7 +/- 1.4 on days 1, 2, 3, and 4 postinoculation. The lung pathology was characterized by pneumonitis with edema and a patchy distribution of hemorrhages in the peribronchovascular spaces of both lungs. Abscesses formed on days 3 and 4. Four days after inoculation, subacute pneumonitis characterized by alveolar macrophage proliferation and areas of fibrosis was observed. The cumulative mortality on day 4 was 85%. This new model was used to study the effects of 1, 2, or 3 50-mg/kg doses of imipenem. Imipenem concentrations in lungs were above the MIC for 2 h after the last dose. The in vivo postantibiotic effect (PAE) was determined during the 9-h period following the last dose; it decreased in duration with the number of doses: 9.6, 6.4, and 4 h after 1, 2, and 3 50-mg/kg doses, respectively. In contrast, no in vitro PAE was observed. This model offers a reproducible acute course of A. baumannii pneumonia. The presence of a prolonged in vivo PAE supports the currently recommended dosing intervals of imipenem for the treatment of human infections due to A. baumannii, i.e., 15 mg/kg three times a day.

Acinetobacter↗

PubMed1997

Intra-abdominal sepsis: an immunocytochemical study of the small intestine mucosa.

AIM: To investigate immunocytochemical changes in intestinal tissues from patients with intra-abdominal sepsis, and to relate the changes to the possibility of enhanced bacterial adhesion and translocation. METHODS: Tissues from 17 patients suffering from intra-abdominal sepsis and from controls were sectioned and stained immunocytochemically for IgA, IgM, secretory component, J chain, and HLA-DR. Differences in the distribution and characteristics of positively staining cells between the patient groups were assessed. RESULTS: Patients with intra-abdominal sepsis had noticeable reductions in numbers of IgA and IgM plasma cells, reduced J chain staining, and had little immunoglobulin on the surfaces of enterocytes. In contrast, HLA-DR positive cells were increased in the sepsis compared with the control group. The plasma cells present showed cytological changes suggestive of apoptosis. CONCLUSIONS: Stress associated with sepsis and its immediate causes might result in increased plasma glucocorticoid levels that bring about apoptosis of mucosal plasma cells (or their precursors). The consequent reduction in expression of IgA and IgM may favour bacterial adhesion to the enterocytes and facilitate bacterial translocation into the tissues.

Adult↗

PubMed1996

[Eating behavior of 12-16-year olds of French-speaking Valais: cross-sectional survey among 962 secondary students].

Eating habits that are potentially harmful are picked up during the teens. The aim of the study is to identify the main eating habits of 12-16 year-old teenagers of central Valais in order to plan better targeted health promotion interventions. The study design is cross sectional. An anonymous questionnaire was taken by a sample of 962 teenagers. Some results: 25.2% did not have breakfast every day, 37.0% did not eat neither fruit nor vegetable the day before, 58.2% had a soda drink the day before, 5.5% of the 12 of the 16 year-old ones and 51.2% year old ones drink some alcohol over the weekends. Possible health promotion interventions are listed.

Adolescent↗

PubMed1996

Epithelial ovarian carcinoma of low malignant potential.

OBJECTIVE: The aim of this study was to evaluate the interest of sonographic features, serum marker tumors and conservative treatment especially by laparoscopic approach of epithelial ovarian tumors of low malignant potential (or borderline tumors). STUDY DESIGN: Thirty-four patients with 40 epithelial ovarian tumors of low malignant potential treated at Bichat Claude Bernard hospital were reviewed. Of these patients, 29 had preoperative sonographic control (85.2%), 19 (55.8%) had pretreatment serum CA 125 determination, 17 (50%) serum CA 199 determination and 19 CEA serum levels. Twenty patients (58.8%) had serous tumor and 14 (41.2%) had mucinous tumor. Six out of the 20 serous lesions were bilateral. Of the patients 31 (91%) had a stage I disease, one had stage II (3%) disease and two had stage III (6%) disease. Nine patients (26.5%) had laparotomic management, 25 (73.5%) had first laparoscopic management with seven laparoconversions (28%). RESULTS: The majority of borderline tumors (67.7%) exhibited multilocular aspects and seven patients had benign features (unilocular smooth sonolucent). Seven out of the 19 CA 125 serum levels were up to 35 U/ml; 4/17 CA 199 serum levels were above 40 U/ml but the CEA serum levels were always normal. Eighteen patients (52.9%) had radical surgery and 16 patients (47.1%) had conservative surgery including 7 cystectomies. Seven patients (20.6%) had subsequent laparotomic treatment, (five after laparoscopic management, one after laparoconversion management and another one after laparotomic surgery) and no residual disease was found. Fourteen patients (41.1%) had preoperative cyst rupture (13 during laparoscopic management). Four recurrences in stage I disease occurred; one after unilateral salpingo-oophorectomy, three after cystectomy but only one recurrence was in ipsilateral ovary. All four recurrences were diagnosed by sonographic control and one was associated with CA 125 serum level elevation. Ten laparoscopic second look were negative. CONCLUSION: No specific sonographic aspect exists for borderline tumors but it is the best way to detect recurrence. The preoperative CA 125 serum levels was elevated in 36.8% of patients. The conservative treatment including laparoscopic procedure in borderline ovarian tumors appears to have a high risk of peroperative rupture but the rate of recurrence is similar to laparotomic treatment.

Adolescent↗

PubMed1996

Relationship between composition of lymphoid cell infiltrates in the liver and replication status in chronic hepatitis B: an immunohistochemical study.

The purpose of this in situ immunophenotyping study was to assess the nature of lymphoid cells involved in chronic hepatitis B with various levels of replication. Replicative status was studied in 25 patients with chronic hepatitis B. Different subsets of T cells, B cells, natural killer cells, and follicular dendritic cells were counted with a computerized image analyzer. Twelve patients had chronic hepatitis B with a high level of replication, nine without replication, and four with a low level of replication. The number of CD3+ T cells was significantly higher in the liver of patients with replication. In all cases, CD8+ T cells were the most numerous cells in the lobules and CD4+ T cells were the most numerous cells in the portal tract. Intra-portal lymphoid follicles were observed in 33% of patients with a high level of replication. The CD57+ natural killer cells were more numerous in patients with a low level of replication. This study suggests that in immunocompetent patients, the intensity and type of hepatic immune response are directly determined by the HBV replication.

Adult↗

PubMed1996

Improved detection of human papillomavirus infection in genital intraepithelial neoplasia in human immunodeficiency virus positive (HIV +) women by polymerase chain reaction-in situ hybridization.

The prevalence of genital human papillomavirus (HPV) infection was evaluated in 30 consecutive human immunodeficiency virus (HIV) + women by polymerase chain reaction (PCR)-in situ hybridization (ISH) on paraffin-embedded tissue sections and compared with that found with standard ISH. Biopsies were removed from normal or neoplastic areas in the cervix, vagina, and vulva, and ISH was performed with biotinylated or fluorescein isothiocyanate genomic DNA probes. One probe was used for HPV screening and others for HPV typing (types 6, 11, 16, 18, 31, and 33). Sequences were amplified by the "hot-start" PCR method and followed by standard ISH. Among the 30 HIV + women, 90% scored HPV + in one or several locations by PCR-ISH, whereas only 67% were positive by ISH. Oncogenic HPV types were found in 63% by PCR-ISH and in only 43% by ISH. The same HPV types detected by standard ISH were also recognized by PCR-ISH, but with the latter the signal was amplified. Moreover, some HPV types were found with PCR-ISH but not by ISH. We conclude that PCR-ISH is a valuable and sensitive method for specific detection of HPV.

Adult↗

PubMed1996

The heart rate response to carotid artery massage in a sample of healthy elderly people.

Carotid artery massage was carried out in a cross-section of 95 healthy elderly subjects (51 men and 44 women, mean age of 74 years) to assess the heart rate response and associated symptoms. These subjects were drawn from a larger group of randomly selected healthy elderly people living in Southampton. They had no history of myocardial infarction or stroke and were not taking any drugs that might enhance the carotid sinus reflex. A continuous distribution of response to carotid artery massage was seen with a median percent change in RR interval of 11.6% (IQR 22.9%); 66% had a percent change in RR interval of < 20%, 90% had a change of < 50%, and 9.5% a change of > 100%. Carotid sinus hypersensitivity (sinus arrest > 3 s) was found in 4 previously asymptomatic subjects; a prevalence of 4.2% (95% CI 1.2%-10.4%), and this was associated with dizziness in 2 of these 4 subjects. Dizziness also occurred in one other subject who had a change in RR interval of 178%. The definition of an abnormal carotid sinus reflex is arbitrary given the continuous RR interval response to carotid artery massage. In diagnosing carotid sinus syndrome, the RR interval response to carotid artery massage alone is not sufficient, as this maneuver must also reproduce the patient's presenting symptoms.

Aged↗

PubMed1996

Observer variability in histopathological reporting of non-small cell lung carcinoma on bronchial biopsy specimens.

AIMS: To evaluate the ability of histopathologists to sub-classify non-small cell lung carcinomas on bronchial biopsy material using the current World Health Organisation (WHO) classification. METHODS: Twelve histopathologists each reviewed 100 randomly selected bronchial biopsy specimens which had originally been reported as showing non-small cell lung carcinoma. For each case, two sections were circulated, one stained by haematoxylin and eosin and the other by a standard method for mucin (alcian blue/periodic acid Schiff). The participants were allowed to indicate their degree of confidence in their classification of each case. A standard proforma was completed and the results were analysed using kappa statistics. RESULTS: Where the participants were confident in their classification, they were actually quite good at sub-classifying the non-small cell carcinoma sections (kappa = 0.71, standard error = 0.058). Overall, however, the results were only fair (kappa = 0.39, standard error = 0.034). CONCLUSIONS: The majority of non-small cell lung carcinomas can be correctly categorised on adequate bronchial biopsy material. Where a confident diagnosis was made, both squamous carcinoma (kappa = 0.73) and adenocarcinoma (kappa = 0.83) were well recognised.

Adenocarcinoma↗