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Biomedical subjects

P Stephens

Publications and source records attributed to P Stephens.

53 records · Page 3Linked to original sources
PubMed1993

Effect of mutations in the V3 loop of HIV-1 gp120 on infectivity and susceptibility to proteolytic cleavage.

It has been suggested that the V3 domain of human immunodeficiency virus type 1 (HIV-1) isolates has to interact with a cell-surface-associated or endosomal proteinase during virus entry into susceptible cells. To investigate this hypothesis, we examined the effect of several mutations in the V3 loop on its susceptibility to proteolytic cleavage by thrombin and cathepsin E and compared it with the effect of these mutations on viral infectivity. The data obtained indicate that, if an interaction between the V3 loop and a proteinase is indeed crucial for viral entry, the substrate requirements for such a proteinase(s) would have to be very complex. In particular, it seems unlikely that a single enzyme with a unique specificity would be able to interact with all of the different HIV-1 and HIV-2/SIV strains isolated so far. Therefore, one would have to postulate the involvement of several cellular proteinases, or proteases with multiple specificities, in V3-based viral tropism.

Amino Acid Sequence↗
PubMed1992

Antibody against the Leu-CAM beta-chain (CD18) promotes both LFA-1- and CR3-dependent adhesion events.

The Leukocytic cell-adhesion molecule (beta 2 integrin) family of adhesion molecules play a key role in the intercellular adhesive interactions necessary for normal immune cell function. In this study, we report an antibody that recognizes an epitope on the Leukocytic cell-adhesion molecule common beta-chain (CD18) and promotes both lymphocyte function-associated Ag-1- and CR3-dependent adhesion events. The antibody recognizes a temperature-sensitive epitope that is not dependent on the presence of divalent cations. It is proposed that antibody binding promotes a conformational change in both lymphocyte function-associated Ag-1 and CR3, which may mimic a natural activation mechanism, resulting in increased cellular adhesion.

Antibodies, Monoclonal↗
PubMed1990

The expression of a synthetic rainbow trout metallothionein gene in E. coli.

A synthetic gene for rainbow trout metallothionein was constructed and inserted into a dual origin plasmid where expression was induced by a temperature shift in a proteinase-deficient strain of Escherichia coli. The recombinant protein was purified to homogeneity, and a partial amino acid sequence was determined to confirm its identity. Its immunochemical characteristics were similar to those of native metallothionein from rainbow trout. The amounts of recombinant metallothionein produced were quantified in soluble cell extracts by ELISA. Low concentrations were detected when growth was performed either in L-broth or defined (GMM-II) medium. Supplementation of the medium with zinc or copper had no effect on the amount of metallothionein produced. By contrast, when cadmium was included in either L-broth or GMM-II medium, much higher concentrations of the protein within the cells (approx. 13 micrograms/mg soluble cell protein) were detected. This stabilisation of the protein by metal reconstitution in vivo is considered in relation to the selective uptake/exclusion of metals by the cells and its significance for the scavenging of certain precious or toxic heavy metals is discussed.

Amino Acid Sequence↗
PubMed1990

Effect of maternal-fetal movement on concentration of cells obtained at genetic amniocentesis.

The number of viable amniotic fluid cells in the aliquot obtained at amniocentesis is an important determinant of the success of subsequent cell culture. However, cell concentration and viability are highly variable, even in samples of equivalent gestational age. We studied the effect of prescribed maternal position change or spontaneous fetal movement on the concentration, viability, and subsequent harvest time of amniotic fluid cells removed at genetic amniocentesis. Samples from 108 patients at 15 to 22 weeks' gestation were evaluated. We did not find a relationship between this type of maternal or fetal movement and the retrieval of amniotic fluid cells.

Adult↗
PubMed1984

Codon usage can affect efficiency of translation of genes in Escherichia coli.

By inserting synthetic oligonucleotides into a highly expressed gene in E. coli it has been shown that unfavourable codon usage can reduce the maximum translation rate of a protein. However, in the case of the codon used (AGG), a significant effect on translation was only seen at very high transcription rates from a gene containing multiple copies of the unfavourable codon.

Bacterial Proteins↗
PubMedDate not supplied

A review of clozapine: an antipsychotic for treatment-resistant schizophrenia.

This report reviews over 25 years of literature of the development, pharmacology, proposed mechanism of action, efficacy, adverse effects, and recommendations for use of clozapine. Clozapine, synthesized in 1960, is an efficacious antipsychotic that rarely causes extrapyramidal side effects. However, in the mid-1970s, it was associated with an increased incidence of agranulocytosis resulting in restrictions of use. Recent trials with treatment-resistant schizophrenic patients found clozapine to be superior to chlorpromazine and haloperidol, fortifying the potential contribution of this drug. This has generated optimism that clozapine will obtain Food and Drug Administration approval. Generally well tolerated, the 1% to 2% risk of agranulocytosis can be minimized with careful patient selection, white blood cell (WBC) count monitoring, and weighing of risks versus benefits for use beyond the relatively safe initial 4-week period.

Clozapine↗
PubMedDate not supplied

Development of a multilayered in vitro model for studying events associated with wound healing.

Simplified in vitro models such as cellular monolayer cultures have only limited usefulness in the study of cutaneous wound repair processes. This has stimulated the investigation of three-dimensional tissue equivalent systems such as the dermal and skin equivalent models. With the use of a wound system constructed of rat tail type I collagen and human dermal fibroblasts, experimental wounding was accompanied by problems with mechanical scoring of the plastic substratum which prevented cell migration. These problems were overcome with the use of a multilayered model in which a punch biopsy-wounded dermal equivalent (bilayered model) or skin equivalent (tri-layered model) was placed onto an acellular collagen lattice and fixed in place with polymerizing collagen. This model permitted observation of the process of cellular repopulation of the "wound space," into which both fibroblast and keratinocyte migration commenced within 1 day. The number of fibroblasts in this space increased dramatically over a period of 9 days, the cells appearing to migrate both over and through the acellular lower collagen layer. Keratinocyte reepithelialization of the "wound space" was completed after 5 days. With the model it was shown that platelet-derived growth factor--AB and epidermal growth factor had positive effects in increasing fibroblast number within the wound space. In conclusion, the model described here should facilitate the study of fibroblast and keratinocyte responses to a wound stimulus in vitro and be a plausible in vitro system for evaluating agents which may have a potential stimulatory or inhibitory effect on numerous cellular responses associated with wound healing.

Journal Article↗
PubMedDate not supplied

Randomized clinical trial of the effect of semi-occlusive dressings on the microflora and clinical outcome of acute facial wounds.

This study investigated whether treatment with occlusive, hydroactive polyurethane dressings affects the microflora and clinical outcome of acute wounds. A randomized, controlled trial was performed on 60 patients with acute facial lacerations. Following primary closure, patients were provided with either a dry-gauze dressing or a hydroactive polyurethane dressing (Cutinova Thin). The wounds were assessed clinically and microbiologically prior to closure, then after 5, 28 and 56 days. The dressing, which was removed at day 5, was also sent for microbiological culture. Wounds treated with the polyurethane dressing showed improved comfort and contour (p < 0.04), less erythema (p < 0.03) and less potential for scarring (p < 0.01) at day 5. At day 28 and day 56, there were no significant differences in the clinical assessment between the test and control groups (p > 0.05). A total of 518 isolates were recovered from the patients during the study. Mean number of isolates and bacterial growth density were calculated in both groups. A similar range of microorganisms were obtained from both treatment groups with no clear difference in organism colonization. This study shows the potential usefulness of facial wounds as a human model for studying acute wound healing responses and anti-scarring therapy. While short-term, clinical benefits of occlusive dressings were evident in the management of acute wounds, these data clearly show that all studies of scarring following acute wounding should extend over at least 3 months post-injury to allow for spontaneous improvement to occur.

Adolescent↗
PubMedDate not supplied

Phenotypic variation in the production of bioactive hepatocyte growth factor/scatter factor by oral mucosal and skin fibroblasts.

Hepatocyte growth factor/scatter factor (HGF/SF) is a pleiotropic growth factor produced principally by cells of mesenchymal origin. HGF/SF is an important mitogen, morphogen, and motogen and plays an important role in wound healing, tumorigenesis and particularly fetal development. Oral mucosal fibroblasts exhibit a fetal phenotype, including an increased extracellular matrix reorganizational ability, cellular migration and experimental wound repopulation in comparison to skin fibroblasts. In this study the expression, production, and bioactivity of HGF/SF by oral mucosal and skin fibroblasts was investigated. Although both oral mucosal and skin fibroblasts expressed HGF/SF, the oral mucosal fibroblasts produced significantly increased amounts of total HGF/SF (p < 0.01) as measured by enzyme-linked immunosorbent assay and bioactive HGF/SF as measured by cell scatter and cell-dissociation techniques (p < 0.01). The possible effect of increased HGF/SF in production mediating the previously described preferential responses of oral mucosal fibroblasts was studied in vitro. Reverse transcriptase-polymerase chain reaction-Western blotting and immunocytochemistry methods all showed that both oral mucosal and skin fibroblasts expressed and produced the c-Met receptor. Recombinant HGF (20-40 ng/mL) however, failed to affect fibroblast repopulation of monolayer wounds or cellular proliferation. In contrast, recombinant HGF significantly increased ECV304 wound repopulation. These studies provide direct evidence of another mechanism by which site-specific variations in fibroblast phenotype may contribute in a paracrine fashion to the rapid reepithelialization and revascularization of oral wounds.

Adolescent↗
PubMedDate not supplied

Use of molecular techniques to study microbial diversity in the skin: chronic wounds reevaluated.

The skin is colonized by an array of microorganisms which form its natural microflora. Disruption to the normal barrier function of the skin (due to trauma or disease) may result in invasion of the dermis by opportunistic bacteria. To date, these organisms, which may contribute to the chronicity of skin wounds, have been analyzed solely by culture methods. It is increasingly realized that standard culture methods of analysis do not accurately reflect the full diversity of complex microflora. This review discusses the limitations of traditional culture approaches and reviews recent advances in molecular microbiological techniques which facilitate a more comprehensive characterization of the microflora within clinical samples. The currently available technologies and techniques are described, as is their use in clinical practice and their potential for diagnostic screening. Chronic venous ulceration of the lower limbs is an important skin disorder in which the microflora invading the dermal tissues contribute to the observed delayed healing. Using chronic leg ulcers as a working example, we show how strict culture and molecular microbiological techniques may be employed, for the first time in combination, to definitively characterize the invading microbial community of the dermis.

Chronic Disease↗
PubMedDate not supplied

Ultrastructural study on experimental infection of rotavirus in a murine heterologous model.

Viral replication, histopathological and ultrastructural changes were observed for a period of nine days in the small intestine of suckling mice infected with a simian rotavirus (SA11). Samples taken from duodenum, jejunum and ileum were prepared for light microscopy, transmission and scanning electron microscopy analysis. Histopathologic effect could be detected within 8 hr post-infection, when only a few altered cells were observed. Damage was extensive after 16 hr post-infection, showing swollen enterocytes and reduced and irregularly oriented microvilli at intestinal villi tips. Virus particles were detected at 16 and 48 hr post-infection, budding from the viroplasm into the rough endoplasmic reticulum cisternae in ileum enterocytes. Clear evidence of viral replication, observed by electron microscopy was not described before in heterologous murine models. Regeneration of the intestinal villi began at the third day post-infection. Despite some differences observed in clinical symptoms and microscopic analysis of homologous and heterologous rotavirus infections, we concluded that mechanisms of heterologous rotavirus infection in mice follow similar patterns to those observed in the homologous models.

Animals↗