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Biomedical subjects

A Hall

Publications and source records attributed to A Hall.

At least 253 records · Page 14Linked to original sources
PubMed1993

Cellular responses regulated by rho-related small GTP-binding proteins.

Rho-related proteins are members of the ras superfamily of small GTP-binding proteins. Their function in fibroblasts has been analysed using microinjection of living cells. Rho appears to link plasma membrane receptors to the assembly of focal adhesions and actin stress fibres. The closely related protein rac, on the other hand, links receptors to the polymerization of actin at the plasma membrane to form membrane ruffles and pinocytotic vesicles. In phagocytic cells, rac has been shown to be required for activation of a membrane-bound NADPH oxidase in response to receptor activation. These systems provide the basis for a working model for the mechanism of action of the rho family of small GTPases.

Animals↗
PubMed1993

Importance of the evolutionarily conserved glycine residue in the N-terminal region of human cystatin C (Gly-11) for cysteine endopeptidase inhibition.

Human cystatin C variants in which the evolutionarily conserved Gly-11 residue has been replaced by residues with positively charged (Arg), negatively charged (Glu), bulky hydrophobic (Trp), or small (Ser or Ala) side-chains have been produced by site-directed mutagenesis and expression in Escherichia coli. The five variants were isolated and structurally verified. Their inhibitory properties were compared with those of wild-type recombinant cystatin C by determination of the equilibrium constants for dissociation (Ki) of their complexes with the cysteine endopeptidases papain and human cathepsin B and with the cysteine exopeptidase dipeptidyl peptidase I. The Ser-11 and Ala-11 cystatin C variants displayed Ki values for the two endopeptidases that were approx. 20-fold higher than those of wild-type cystatin C, while the corresponding values for the Trp-11. Arg-11 and Glu-11 variants were increased by a factor of about 2000. In contrast, the Ki values for the interactions of all five variants with the exopeptidase differed from that of wild-type cystatin C by a factor of less than 10. Wild-type cystatin C and the Ser-11, Ala-11 and Glu-11 variants were incubated with neutrophil elastase, which in all cases resulted in the rapid hydrolysis of a single peptide bond, between amino acid residues 10 and 11. The Ki values for the interactions with papain of these three N-terminal-decapeptide-lacking cystatin C variants were 20-50 nM, just one order of magnitude higher than the value for N-terminally truncated wild-type cystatin C, which in turn was similar to the corresponding values for the full-length Glu-11, Arg-11 and Trp-11 variants. These data indicate that the crucial feature of the conserved Gly residue in position 11 of wild-type cystatin C is that this residue, devoid of a side-chain, will allow the N-terminal segment of cystatin C to adopt a conformation suitable for interaction with the substrate-binding pockets of cysteine endopeptidases, resulting in high-affinity binding and efficient inhibition. The functional properties of the remaining part of the proteinase contact area, which is built from more C-terminal inhibitor segments, are not significantly affected even when amino acids with bulky or charged side-chains replace the Gly-11 residue of the N-terminal segment.

Amino Acid Sequence↗
PubMed1993

The BULIT-R: its reliability and clinical validity as a screening tool for DSM-III-R bulimia nervosa in a female tertiary education population.

The Bulimia Test (BULIT) has been updated to accommodate the DSM-III-R criteria for bulimia nervosa. Therefore, in this study, we evaluated the psychometric properties of the BULIT-R using a sample of young women at a tertiary educational institute. The results showed all 28 BULIT-R items correlated highly with the total test score (average = .59) and the internal reliability was high (.92). In terms of its concurrent validity, the BULIT-R correlated highly (.90) with the Bulimia Investigatory Test Edinburgh (BITE), a screening measure argued to detect bulimia nervosa. In terms of criterion-related validity, the optimal cutoff for the BULIT-R as a screening measure was 98 with this sample, using a semistructured DIS-III R interview administered by experienced clinicians who specialize in eating disorders. At this cutoff, the sensitivity was 100%, the specificity 99.0%, the negative predictive value 100%, and the positive predictive value 71.3%.

Adult↗
PubMed1993

A novel role for RhoGDI as an inhibitor of GAP proteins.

RhoGDI inhibits guanine nucleotide dissociation from post-translationally processed Rho and Rac proteins but its biochemical role in vivo is unknown. We show here that N-terminal effector site mutations in the Rac protein do not compromise its interaction with RhoGDI and that, whilst geranylgeranylation and -AAX proteolysis of the C-terminal CAAX motif of Rac1 and RhoA are required for efficient interaction with RhoGDI, methylesterification of the C-terminal cysteine residue is not required. In vitro, RhoGDI can form stable complexes with Rho and Rac proteins in both the GTP and GDP bound states. Furthermore the Rac-GTP--RhoGDI complex is resistent to the action of recombinant RhoGAP and recombinant BCR. Thus GDI, by complexing with Rac-GTP and preventing GAP stimulated GTP hydrolysis, may allow transit of the activated form of the Rac protein between physically separated activator and effector proteins in the cell.

Adenosine Diphosphate Ribose↗
PubMed1993

Ras-related proteins.

A combination of genetic and biochemical techniques has revealed key players in the Ras signal transduction pathway that link receptors to growth and differentiation. Rho and Rac have been shown to couple extracellular signals to the organization of the actin cytoskeleton, while ADP ribosylation factor is required for the formation of non-clathrin coated vesicles.

ADP-Ribosylation Factors↗
PubMed1993

Tumour suppressors and the regulation of GTP-binding protein activity.

GTP-binding proteins regulate a wide variety of intracellular signalling pathways in eukaryotic cells. The Ras GTP-binding proteins have received a great deal of attention since they were found to be modified by amino acid substitutions in a large number of cancers. It is now clear that Ras plays an essential role in regulating normal cell growth and differentiation, although how this is achieved biochemically is not known. The cellular concentration of Ras bound to GTP appears to be the limiting factor for signalling, and, not surprisingly, it is tightly controlled by both positive and negative regulators. There is now convincing evidence that the loss of one of these negative regulators of Ras, neurofibromin, can contribute to the development of malignancy; thus, neurofibromin behaves as a tumour suppressor gene product.

Journal Article↗
PubMed1993

Workshops for consultants on the teaching of clinical communication skills.

A group of senior medical school staff concerned about the short-lived effects of communication training formed the Medical Interview Teaching Association. They felt that communication training needed to be reinforced throughout the curriculum and that this would need active involvement by large numbers of consultants. To achieve this they planned a series of workshops. Seventeen consultants and eight other senior staff agreed to participate in the pilot workshop. This was a 3 1/2-day residential workshop. The structure was adapted from a 'faculty development' model used successfully in the USA. Participants worked mostly in small groups helped by experienced facilitators. The teaching style was learner centred and therefore the details of the problem-based agenda and the choice of working methods were largely determined by the participants themselves. There were also some conventional lectures and demonstrations. Evaluation was by postal questionnaire 2 weeks later. This requested both qualitative comments and Likert scale ratings about every aspect of process and outcome. Most responses were strongly positive. Participants felt they made good progress in developing new skills and new curriculum ideas. They also felt more motivated and self-aware as teachers. The learner-centred approach and the diversity of learning activities were seen as very useful. The unstructured approach to self-awareness training was felt to be less useful. It is concluded that such workshops could well lead to more effective communication training and may also have wider implications for medical education.

Clinical Competence↗
PubMed1993

The oral examination: a study of academic and non-academic factors.

The oral examination in psychiatry for final-year medical students at Wellington and Dunedin School of Medicine, University of Otago, was studied. Between December 1989 and April 1990, 40 medical students were videorecorded during such an examination. The transcripts of the recording of each oral, and at a later date the videorecordings, were individually scored by a panel of six research psychiatrists who were experienced examiners. In addition verbal and non-verbal behaviour was rated using visual analogue scales and the students completed personality and anxiety questionnaires. There was a low level of agreement between research psychiatrists in the allocation of oral marks. The oral score was positively associated with the level of confidence of the student and negatively with anxiety in men.

Body Image↗
PubMed1993

Glutathione-S-transferase pi expression in transitional cell carcinoma of the bladder.

Glutathione-S-transferase pi (GST pi) is a multifunctional protein that acts as an enzyme, involved in the detoxification of drugs and carcinogens. It has been implicated both in drug resistance and malignant transformation of epithelium. Using an indirect immunohistochemical technique, we have evaluated cytoplasmic and nuclear staining in normal urothelium, 23 superficial bladder tumours and 26 invasive tumours. All 26 invasive tumours had been treated with platinum-based chemotherapy. Cytoplasmic staining was seen in normal urothelium and all bladder tumours. Nuclear staining was seen in 1 superficial tumour and in 13 invasive tumours. There was no association between nuclear staining and response to chemotherapy. Nuclear staining was seen in 1 area of dysplasia and in 2 of 3 areas of carcinoma in situ. GST pi expression is not a predictor of response to chemotherapy. Increased intra-nuclear expression of GST pi may be associated with progression of transitional cell carcinoma of the bladder.

Carcinoma, Transitional Cell↗
PubMed1993

Characterization of an apical sodium conductance in rabbit cecum.

Rabbit cecum in vitro exhibits electrogenic Na+ absorption not blocked by amiloride but inhibited by the amiloride analogue phenamil, suggesting transport mediated by modified Na+ channels in the apical membrane. To further characterize the mechanism(s) of Na+ absorption, microelectrode impalements of single epithelial cells were performed to measure intracellular potential difference (psi mc) and fractional resistance of the apical membrane, to characterize ionic conductances of the apical and basolateral membranes, and to determine the response to phenamil. The electrical potential profile of cecum (psi mc = -31 +/- 2 mV, fractional resistance = 0.71 +/- 0.03) was qualitatively similar to distal colon. The apical membrane exhibited responses suggesting both Na+ and K+ conductances, whereas the basolateral membrane appeared to have a predominant K+ conductance. Phenamil elicited a depolarization of psi mc and a decrease in fractional resistance; neither response is consistent with inhibition of an apical Na+ conductance. Studies were performed in apical membrane vesicles to characterize ionic conductances by a second independent methodology. These additional studies confirmed the presence of an apical Na+ conductance not inhibited by either amiloride or phenamil. Thus both microelectrode impalement and vesicle studies demonstrated an apical membrane Na+ conductance in rabbit cecum; this is the likely mechanism of electrogenic Na+ absorption in this epithelium. However, the anomalous response to phenamil suggests that the inhibitory effect of this agent is not directly on the conductance. The cecal transporter may be one of a family of cation channels related to, but significantly different from, the classic Na+ channel found in distal colon and other tight epithelia.

Amiloride↗
PubMed1993

The structured oral self-directed learning evaluation: one method of evaluating the clinical reasoning skills of occupational therapy and physiotherapy students.

The Structured Oral Self-directed Learning Examination (SOSLE) is used to evaluate the clinical reasoning skills of occupational therapy (OT) and physiotherapy (PT) students. It is an oral examination which evaluates a student's problem-solving ability, self-directed learning skills, knowledge level and self-assessment ability. The three parts of the examination are conducted over a 24-hour period. Validation of this instrument was carried out in two groups of OT and PT undergraduate students over two consecutive years (Year 1--n = 20) (Year 2--n = 18). Inter-rater reliability correlations varied from 0.61 to 0.78 the first year to 0.85 to 0.99 in the second year. The results obtained from the SOSLE were also compared to written and tutorial marks obtained in the same course. Pearson Correlation Coefficients (PCC) among mean SOSLE and two written paper scores ranged from 0.0-0.05 (Year 1) to 0.0-0.1 (Year 2). The PCC among the mean SOSLE and tutorial performance scores were 0.57 (Year 1) and 0.0 (Year 2). The results show that good agreement between raters can be reached using this evaluation method. However, the poor correlations between the SOSLE and the other methods of evaluation may show that different skills are being evaluated. Further validity testing needs to be carried out to confirm that this tool is measuring process oriented skills.

Clinical Competence↗
PubMed1993

Different structural organization of Ras and Rho effector domains.

Ras regulates proliferation and differentiation signals in cells, and activation of the protein can lead to malignant transformation. Activation of the related protein, Rho, affects cell morphology, and it has been suggested that it may also have some oncogenic potential. We show here that Rho does not induce a malignant phenotype in NIH3T3 cells but instead is a potent activator of actin stress fibre formation. The limited homology between Ras and Rho has enabled us to determine the amino acids specifying their different biological activities and GTPase-activating protein (GAP) protein sensitivities using chimeras. Rho substituted with amino acids 23-46 of Ras induces transformed foci in NIH3T3 monolayers, and we conclude that Ras has a single effector domain required for downstream signalling. Although mutational analysis of Rho has revealed that residues 32-42 are also essential for its biological activity, Ras substituted with amino acids 25-48 of Rho does not induce actin stress fibre formation. On the basis of these experiments, we propose that Rho may have two effector domains: one at amino acids 32-42 and corresponding to the effector region of Ras and the second located elsewhere in the carboxy-terminal two-thirds of the molecule.

3T3 Cells↗
PubMed1992

Post-translational modifications of p21rho proteins.

Post-translational modifications of the ras proteins, which are required for plasma membrane localization and biological function of the proteins, have been shown to include prenylation and carboxymethylation at the carboxyl terminal cysteine residue of the cysteine-aliphatic amino acid-aliphatic amino acid-any amino acid (CAAX) box. In addition, p21Ha-ras and p21N-ras, but not p21K-ras (B), are palmitoylated. The three mammalian rho proteins (A, B, and C) are also members of the ras superfamily but have distinct biological activities and different intracellular distributions from p21ras. Analysis showed all three rho proteins are modified by a COOH-terminal carboxymethylation similar to p21ras, whereas p21rhoC labeled with [3H]mevalonic acid in vivo revealed the presence of a C20 prenoid, similar to that already described for p21rhoA. However, in vivo and in vitro studies of p21rhoB showed this protein to be modified by both C15 and C20 prenoids. Mutation of C193 in the CAAX box abolished prenylation, whereas mutation of the adjacent C192 resulted in a significant reduction in the amount of the C20, but not C15 prenoid, recovered from p21rhoB. In vivo labeling studies with [3H]palmitic acid and mutational analysis showed that both cysteine residues at 189 and 192 upstream of the CAAX box in p21rhoB are sites for palmitoylation. We conclude that there are different populations of post-translationally modified p21rhoB in the cell and that the sequence specificity for geranylgeranyl- and farnesyltransferases may be more complicated than previously proposed.

Amino Acid Sequence↗
PubMed1992

The small GTP-binding protein rho regulates the assembly of focal adhesions and actin stress fibers in response to growth factors.

Actin stress fibers are one of the major cytoskeletal structures in fibroblasts and are linked to the plasma membrane at focal adhesions. rho, a ras-related GTP-binding protein, rapidly stimulated stress fiber and focal adhesion formation when microinjected into serum-starved Swiss 3T3 cells. Readdition of serum produced a similar response, detectable within 2 min. This activity was due to a lysophospholipid, most likely lysophosphatidic acid, bound to serum albumin. Other growth factors including PDGF induced actin reorganization initially to form membrane ruffles, and later, after 5 to 10 min, stress fibers. For all growth factors tested the stimulation of focal adhesion and stress fiber assembly was inhibited when endogenous rho function was blocked, whereas membrane ruffling was unaffected. These data imply that rho is essential specifically for the coordinated assembly of focal adhesions and stress fibers induced by growth factors.

3T3 Cells↗
PubMed1992

The small GTP-binding protein rac regulates growth factor-induced membrane ruffling.

The function of rac, a ras-related GTP-binding protein, was investigated in fibroblasts by microinjection. In confluent serum-starved Swiss 3T3 cells, rac1 rapidly stimulated actin filament accumulation at the plasma membrane, forming membrane ruffles. Several growth factors and activated H-ras also induced membrane ruffling, and this response was prevented by a dominant inhibitory mutant rac protein, N17rac1. This suggests that endogenous rac proteins are required for growth factor-induced membrane ruffling. In addition to membrane ruffling, a later response to both rac1 microinjection and some growth factors was the formation of actin stress fibers, a process requiring endogenous rho proteins. Using N17rac1 we have shown that these growth factors act through rac to stimulate this rho-dependent response. We propose that rac and rho are essential components of signal transduction pathways linking growth factors to the organization of polymerized actin.

3T3 Cells↗
PubMed1992

Intensity of reinfection with Ascaris lumbricoides and its implications for parasite control.

Intestinal helminths are among the most common and widespread of human infections. Because it is typical to find that most worms are aggregated in a few potential hosts it has been suggested that some individuals are predisposed to heavy infections and that morbidity could be controlled by the treatment of heavily infected individuals only. We have studied the prevalence and intensity of reinfection with the intestinal nematode Ascaris lumbricoides among people living in Dhaka, Bangladesh. 880 people were treated with pyrantel pamoate three times at six month intervals, and on each occasion they collected all their stools for 48 h after treatment. Worms expelled by each subject were counted and weighed. The prevalence of infection at round 1 of treatment was 89% and the mean burden was 18.5 worms. Reinfection was rapid and at rounds 2 and 3 the prevalence was 82% and 80%, respectively, with mean burdens of 14.0 and 11.5 worms. The intensity of reinfection was not random: more subjects than expected became heavily reinfected (greater than or equal to 15 worms) and more subjects than expected remained lightly infected (less than or equal to 14 worms) (p less than 0.001). Worms were highly aggregated at each round of treatment but although just over 10% of all subjects were heavily infected at each and every round of treatment, over 60% of all subjects were heavily infected at least once. The findings show that some individuals seem to be susceptible to heavy infection whereas others are not, that deworming has a greater effect on the intensity of infection than on the prevalence, and that mass chemotherapy is likely to be a more effective means to control morbidity than is selective treatment of heavily infected individuals only.

Adolescent↗