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Biomedical subjects

J F White

Publications and source records attributed to J F White.

At least 37 records · Page 2Linked to original sources
PubMed1994

Phage surface presentation and secretion of antibody fragments using an adaptable phagemid vector.

Phagemid vectors have been developed which promise to supersede hybridoma technology for the selection and production of human antibodies. We have modified an existing phagemid vector to improve the stability of synthesized soluble antibody fragments. The vector allows the antibody fragment to be produced: i) as a soluble protein incorporating a stable carboxyl terminal octapeptide (FLAG) or, ii) on the surface of a bacteriophage fused to a minor coat protein (the gene III protein). The antibody gene encoding the well characterized monoclonal antibody NC10 (an antibody that recognizes the neuraminidase of the influenza strain N9) was inserted as a single chain Fv construct into the phagemid vectors pHFA and pHFA/SacII. Western blotting, ELISA and electron microscopy studies showed that recombinant clones could be manipulated to either synthesize soluble protein into the periplasm or present the protein on the surface of bacteriophage. Cosynthesis of GroEL and GroES chaperonins resulted in complete proteolysis of the scFvNC10-FLAG-gIIIp fusion product and did not improve total phage production. Coexpression of chaperonins should be used with caution for library construction due to the expected selection pressure for protease resistant gene III fusions.

Amino Acid Sequence↗
PubMed1994

Evaluation of the Omniflow collagen-polymer vascular prosthesis.

The Omniflow Vascular Prosthesis (OVP) is a truly integrated bio-synthetic composite of polyester mesh and ovine connective tissue components. In animal and human studies, the prosthesis has demonstrated long-term patency rates comparable and often better than alternate surgical replacements, with minimal aneurysmal or thrombotic problems. Immunohistological and TEM analysis of explants of OVP I provide good evidence for the excellent in vivo performance and are reviewed below. In addition, new data from explants of the OVP II, a later version of the Omniflow, are also described. These explant analyses have been enhanced by use of a library of highly specific monoclonal antibodies which can discriminate between collagens from different species.

Animals↗
PubMed1993

Structural analysis of a collagen--polyester composite vascular prosthesis.

The Omniflow Vascular Prosthesis is a collagen--polyester composite which has been used successfully for peripheral vascular replacement. In this study, we have examined the distribution of the various connective tissue components and the ultrastructural organisation of these in order to understand and allow improvement of its functional properties. Using immunohistology with specific monoclonal antibodies, types I and III collagens were found to be the major components throughout the prosthesis. Type VI collagen was also present but was mainly associated with cells, particularly around the polyester mesh and silicone interfaces. While elastin was absent, two elastic tissue microfibrillar proteins were present uniformly throughout the structure. Ultrastructurally, clear differences existed between the local environments of the inner surface, which had formed around the silicone mandrel, the polyester mesh within the prosthesis, and the outer collagenous tissue which formed the central wall. At the inner surface, the amount of collagen was less and the orientation of these fibres was not well defined. The collagen fibrils in the polyester region were smaller than those of the main wall, which were well ordered and orientated along the axis of the device.

Antibodies, Monoclonal↗
PubMed1993

Monoclonal antibodies to type VI collagen demonstrate new tissue augmentation of a collagen-based biomaterial implant.

We developed a panel of highly specific monoclonal antibodies (MAb) to either human or dog collagen Type VI. Various degrees of species crossreactivities were found with ELISA and immunohistology. Because of these differentiating species specificities, which allowed distinction between the original donor collagen and newly formed host collagen, the MAb proved to be valuable tools in examination of explanted samples of an ovine composite vascular prosthesis retrieved from a canine model. With an MAb that reacts with dog but not sheep collagen Type VI, newly synthesized pockets of collagen Type VI could readily be detected within the prosthesis as early as 3 months after implantation. These areas were associated with regions of cell infiltration, presumably derived from the host. This association was also apparent in the newly formed intimal region of the prosthesis where only host cells were found. Another of the MAb, which was positive against human but not sheep collagen, was also used to demonstrate marked deposition of host collagen Type VI in a retrieved human sample of the prosthesis. In this case the antibody was able to detect collagen on a formalin-fixed tissue, which would broaden the scope of its use in clinical and pathological situations. Use of these novel antibody probes provides a rapid marker for new tissue augmentation of implanted biological devices which would be an indicator of the long-term performance of a prosthesis.

Animals↗
PubMed1992

Expression of influenza neuraminidase in baculovirus-infected cells.

Recombinant influenza neuraminidase (NA, subtype 2, A/NT/60/68) was produced by recombinant baculovirus-infected insect cells. The recombinant NA retained enzyme activity and was located on the cell surface. Enzyme activity was both cell-associated and in the cell free supernatant; maximal NA activity was found in the supernatant. Recombinant NA was recognised by polyclonal antisera and by three monoclonal antibodies specific for NA (subtype 2). Enzyme activity was also neutralised by polyclonal antisera. Recombinant NA thus retains most of the immunological and activity properties of authentic influenza NA. Immunoprecipitation of [35S]Methionine-labelled cells and supernatant and partial purification of NA indicated that a approximately 50-kDa form of NA was present in the supernatant, whilst the expected size (approximately 67-kDa) was cell-associated. Purified recombinant extracellular virus was also enzymatically active, and contained the 67-kDa NA which was located on the membrane capsule of the virus. This suggests that the virus had acquired the cell-associated form of recombinant NA during the budding process from the cell.

Animals↗
PubMed1992

Stromal Development and Mating System of Balansia epichloë, a Leaf-Colonizing Endophyte of Warm-Season Grasses.

Studies of the stromal development and mating system of Balansia epichloë were conducted. Early development of the stroma consists of both endophytic and epiphytic phases of growth. As development progresses, the epiphytic stromal subiculum on the upper surface of leaves is linked with endophytic mycelium within leaves by hyphal bridges, which may provide carbohydrates for stromal development. Sugar utilization studies suggest that Balansia epichloë is excluded from growth within inflorescence primordia of grasses by the presence of high levels of sugars that are inhibitory to growth of the endophyte. Studies of the mating system of B. epichloë were conducted, and the fungus was shown to be heterothallic, with ephelidial conidia functioning as spermatia. Insect vectoring of ephelidial spermatia is suggested to account for the irregular pattern of perithecial development on stromata.

Journal Article↗
PubMed1991

Immunoelectron microscopic localization of elastic tissue components in archival tissue samples.

Tissue samples that have been stored for many years, in different media and under a variety of conditions, have been examined by modern techniques of immunoelectron microscopy, using antibodies against elastic tissue components. A range of postembedding restorative procedures has been identified, which will allow reliable immunolocalization of antibodies against the elastic tissue component of such specimens. These methods have been applied successfully to autopsy-derived material, fixed in buffered formaldehyde, to archival material stored frozen at -70 or -20 degrees C, to specimens fixed for electron microscopy and stored for many years in buffer, and even to archival material from formaldehyde-fixed, paraffin-embedded blocks, reprocessed for electron microscopic examination. The successful restorative methods included pre-treatment of the sections with 6 M guanidine hydrochloride, or 1 M Tris/saline, each containing 100 mM dithiothreitol (a reducing agent) followed by alkylation with 220 mM iodoacetamide. The application of these techniques allowed reliable study of elastic tissue antibody distributions in archival tissues that could not be obtained again, as well as comparative studies with tissues processed many years previously.

Animals↗
PubMed1991

Link between cell apical morphology and H+ secretion in salamander small intestine.

To determine the morphological basis for the cellular secretion of H+ by in vitro salamander jejunum the transepithelial proton flux (JH) was measured under different conditions and compared with the ultrastructure of the villus absorptive cells using transmission electron microscopy in conjunction with morphometric analysis. Norepinephrine (NE) stimulated JH and significantly increased the density of small cytoplasmic vesicular and tubular profiles in the region immediately below the apical plasma membrane of the absorptive cells. Cytoplasmic tubular elements were often observed in direct contact with the plasma membrane at the base of the microvilli, suggesting that expansion of the plasma membrane had occurred. The alpha 2-antagonist, yohimbine, blocked NE-stimulated H+ secretion and reduced the number of tubulovesicles. Theophylline reduced the rate of spontaneous acid secretion and the density of apical tubules and vesicles. Exposure to horseradish peroxidase revealed a physical connection between the extracellular space and the apical tubulovesicular compartment. The direct relationship between the number of apical vesicular and tubular profiles and JH suggests that the luminal membrane plays a fundamental role in the cellular events in H+ secretion. The membranes of the tubules and vesicles may be elements of a dynamic pool of transporting membranes under the control of exogenous effectors such as catecholamines.

Animals↗
PubMed1990

Multitubular bodies in intestinal cells of Amphiuma means/tridactylum (Urodela): ultrastructural characterization.

The jejunal absorptive cells of the salamander Amphiuma, when examined using transmission electron microscopy, were found to possess a unique type of intracellular vacuole containing membranous tubules. These vacuoles, tentatively named multitubular bodies, were located in the cytoplasm between the nucleus and the brush-border membrane, and were seen with greatest frequency in the summer and fall. The vacuoles containing multitubular bodies had an average diameter of 0.6 microns, and the membranous tubules within had an average diameter of 30 nm. The tubules differed morphologically from the vesicles in the multivesicular bodies, and from the primary lysosomes in the polylysosomal vacuoles. The tubules did not exhibit acid phosphatase activity, and were of similar diameter and membrane thickness as the Golgi saccules. In contrast to the multivesicular bodies, the multitubular bodies did not take up exogenous horseradish peroxidase. Early forms of autophagosomes resembling these vacuoles were often seen in the para-Golgi region of the cell. The multitubular bodies may represent a distinct type of autophagosome. Although the exact origin of the tubules as well as their role in cellular activity is unclear, their seasonal appearance within the multitubular bodies of the absorptive cells suggests a unique means of selective down-regulation of Golgi-like organelles.

Acid Phosphatase↗
PubMed1989

Basolateral impalement of intestinal villus cells: electrophysiology of Cl- transport.

A method of dissecting the serosal muscle layers is described that transforms the villus of isolated Amphiuma small intestine into a flat sheet one cell layer thick, allowing rapid equilibration of the serosal medium with the basolateral membrane of the villus tip cells and direct access of the basal membrane to microelectrodes. The "villus sheet" preparation was used to examine the luminal and basolateral mechanisms of Cl- transport. The serosal membrane potential (Vs), measured with conventional microelectrodes, averaged -79.7 mV in tissues bathed in Cl- -free medium; the mucosal membrane potential (Vm) averaged -80.9 mV. Fractional resistance measured directly was 0.82 and 0.14 for the mucosal and serosal membranes, respectively. Elevation of bath [K] reduced Vm and Vs by 30.3 and 44.5 mV, respectively. Cl- (20 mM) added to the luminal medium reduced Vm by 23.9 mV and stimulated Cl- transport; luminal addition of furosemide then increased Vm by 5.6 mV and reduced Cl- transport. Addition of Cl- (20 mM) to the Cl- -free serosal fluid increased Vs 2.0 +/- 1.9 mV. On reducing the serosal [Cl] 10-fold Vs decreased 2.0 +/- 2.2 mV. These and other results indicate that basolateral Cl- exit is not over a conductive pathway. The villus sheet affords new opportunities for studying enterocyte function in the intact mucosa.

4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfo↗
PubMed1989

Conductive pathways for HCO3- in basolateral membrane of salamander intestinal cells.

To understand the route of HCO3- exit across the basolateral membrane of Amphiuma small intestinal cells during active H+ secretion the influence of bath HCO3- on serosal membrane potential (Vs) was measured using conventional microelectrodes. The villus sheet preparation was used, which allowed direct access to the basal membrane for microelectrode impalement. When tissues were incubated in Cl(-)-free (SO2-4 based) medium, Vs averaged -85.8 mV. Elevation of serosal bath [HCO3-] at constant CO2 increased Vs; reduction of the [HCO3-] reduced Vs and increased the fractional resistance of the serosal membrane. When serosal bath HCO3- and CO2 were completely replaced, Vs was -44.8 mV. The serosal membrane was also depolarized on complete replacement of medium HCO3- and CO2 when bath pH was buffered with phosphate. 4,4'-Dinitro-2,2'-stilbene disulfonate (DNDS) and acetazolamide blocked the decline in Vs produced by lowering serosal medium [HCO3-]. Replacement of serosal Na+ reduced Vs 32.7 mV. DNDS blocked this response. It is concluded that a Na+-dependent, stilbene-sensitive HCO3- exit pathway resides in the basolateral membrane of the intestinal cells. Possibly there is also a parallel, conductive pathway for HCO3-.

Acetazolamide↗
PubMed1989

Characteristics of chloride ion influx in Amphiuma small intestine.

Net Cl- absorption by Amphiuma small intestine is electrogenic but associated with the secretion of HCO3-. To define the mechanisms of Cl- entry into the enterocytes the initial rate of uptake of 36Cl into isolated segments of small intestine was measured. Luminal extracellular space was measured using [3H]inulin. Cl- influx was saturable with a Km of 5.3 mM. When the mucosal medium Cl- concentration was 20 mM influx was linear for 5 min. Cl- influx in 5 min (JiCl) was not reduced by 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid added to the serosal medium, although the Cl- current was abolished. Hence the luminal membrane was the barrier to Cl- uptake. Monovalent anions blocked Cl- influx in the order I- = SCN- = NO3- greater than Br- greater than F-. Anoxia and dinitrophenol reduced JiCl 33 and 71%, respectively. Substitution of medium Na+ with choline or N-methyl glucamine reduced JiCl 60-70%. Removal of medium K+ reduced influx 51%. After medium Na+ and K+ were both replaced influx was stimulated upon reexposure to (Na+ + K+); Na+ alone did not stimulate. JiCl was reduced 34% by furosemide. Neither amiloride nor SITS in the mucosal medium altered influx. JiCl was reduced by replacement of the HCO3- -CO2 buffer with either phosphate or N-2-hydroxyethyl-piperazine-N'-2-ethanesulfonic acid and by exposure to acetazolamide. Theophylline reduced influx 60%, whereas the Ca ionophore A23187 reduced net Cl- absorption and lowered JiCl by 17%. Norepinephrine (10(-5) M) in the serosal bathing medium stimulated Cl- influx 51%. These results indicate that Cl- influx into the intestinal mucosa occurs by a Na+- and, possibly, K+-dependent pathway. Cl- entry is under adrenergic influence.

Acetazolamide↗
PubMed1989

A cluster analysis of institutionalized mentally retarded individuals.

The scores of two samples of 200 subjects each on five factors of a measure of adaptive behavior were subjected to cluster analysis using two hierarchical agglomerative clustering methods. Three clusters were found that proved stable across clustering methods, subject samples, and time points 3 years apart. In addition, cluster membership was shown to be meaningfully associated with a number of demographic characteristics and neurological and sensory handicaps. The three-cluster solution thus possesses some desirable features, but its practical utility remains to be demonstrated.

Activities of Daily Living↗
PubMed1989

Prevalence and prediction of psychotropic drug use in California developmental centers.

The extent of psychotropic drug use was evaluated in California's institutionalized developmentally disabled population (N = 6,450). Mean psychotropic drug use was 35.4% for the entire population (range = 13.7% to 63.6% across institutions), with antipsychotic drug use at 26.8% (range = 11.0% to 59.6%). Psychotropic drug use was positively associated with aggression, IQ, depression, and self-injurious behavior, ps less than .0001; it was inversely associated with cerebral palsy and epilepsy. When the influence of demographic and behavioral factors was controlled, significant institutional differences in drug use remained.

Adult↗