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Biomedical subjects

D S Williams

Publications and source records attributed to D S Williams.

At least 19 recordsLinked to original sources
PubMed1992

31P-NMR spectroscopy of perifused rat hepatocytes immobilized in agarose threads: application to chemical-induced hepatotoxicity.

A system consisting of isolated rat hepatocytes immobilized in agarose threads continuously perifused with oxygenated Krebs-Henseleit (KH) solution has been found to maintain cell viability with excellent metabolic activity for more than 6 h. The hepatocytes were monitored by phosphorus-31 nuclear magnetic resonance (31P-NMR) spectroscopy at 4.7 Tesla, by measurement of oxygen consumption and by the leakage of lactate dehydrogenase (LD) and alanine aminotransferase (ALT). The data obtained were comparable to those found for an isolated perfused whole liver in vitro. The effects of allyl alcohol (AA), ethanol, and 4-acetaminophenol (AP) were examined. A solution of 225 microM AA perifused for 90 min caused the disappearance of the beta-phosphate resonance of adenosine triphosphate (ATP) in the 31P-NMR spectra, a 7-fold increase in LD leakage and a 70% reduction in oxygen consumption. Ethanol (1.0 M) perifused for 90 min reduced the beta-ATP signal intensity ratio by 20%, the phosphomonoester (PME) signal by 50% and inorganic phosphate (Pi) by 33% (P less than 0.05). AP (10 mM) caused only mild liver-cell damage. The results demonstrate that perifused immobilized hepatocytes can be used as a liver model to assess the effects of a wide range of chemicals and other xenobiotics by NMR spectroscopy.

Animals↗
PubMed1992

Infections due to beta-lactamase-producing, high-level gentamicin-resistant Enterococcus faecalis.

OBJECTIVE: To investigate the risk factors, clinical features, molecular epidemiology, and treatment outcomes associated with an outbreak of infections due to beta-lactamase-producing, high-level gentamicin-resistant Enterococcus faecalis. DESIGN: Case-control and molecular genetics study. SETTING: Tertiary care Veterans Affairs hospital. PATIENTS: Sixty-five patients infected or colonized with beta-lactamase-producing high-level gentamicin-resistant E. faecalis (case patients) were matched and compared with 65 randomly selected patients infected or colonized with beta-lactamase-negative, gentamicin-susceptible E. faecalis. MEASUREMENTS AND MAIN RESULTS: During the 20-month study period, 124 of 1506 isolates (8.2%) of E. faecalis from 70 patients were found to produce beta-lactamase. Univariate analysis showed older age, higher APACHE II score, nosocomial acquisition, recent surgical procedure, total parenteral nutrition, and antibiotic treatment to be significantly associated with the acquisition of beta-lactamase-producing, high-level gentamicin-resistant E. faecalis. A multivariate analysis, done using stepwise multiple logistic regressions, showed that only two variables remained significant: an APACHE II score greater than 6 (odds ratio, 9.5; 95% CI, 4.4 to 20.3) and antibiotic treatment (odds ratio, 10.2; CI, 4.5 to 23.2). The mortality rate for case patients was 7.7% (5 of 65 patients; CI, 1.2% to 14.2%); no control patient died. Of 23 infections occurring in case patients, 13 were treated with "inappropriate" antibiotics (regimens that included a beta-lactamase-unstable antibiotic); 2 patients improved and 11 had complete resolution of disease. "Appropriate" treatments (regimens that included a beta-lactamase-stable antibiotic) were used in 10 patients; 5 of 10 infections were fatal. Restriction enzyme digests of total chromosomal DNA showed nearly identical patterns for selected isolates of beta-lactamase-producing, high-level gentamicin-resistant E. faecalis, suggesting dissemination through the hospital of a single strain of E. faecalis. CONCLUSIONS: Fatal infections were observed despite treatment with beta-lactamase-stable antibiotics. The risk for infection or colonization with beta-lactamase-producing, high-level gentamicin-resistant E. faecalis was strongly associated with severe underlying disease (acute physiology and chronic health evaluation [APACHE] II score, greater than 6) and previous antibiotic treatment. These results may be useful in targeting high-risk patients for infection-control interventions.

Case-Control Studies↗
PubMed1992

Magnetic resonance imaging of perfusion using spin inversion of arterial water.

A technique has been developed for proton magnetic resonance imaging (MRI) of perfusion, using water as a freely diffusable tracer, and its application to the measurement of cerebral blood flow (CBF) in the rat is demonstrated. The method involves labeling the inflowing water proton spins in the arterial blood by inverting them continuously at the neck region and observing the effects of inversion on the intensity of brain MRI. Solution to the Bloch equations, modified to include the effects of flow, allows regional perfusion rates to be measured from an image with spin inversion, a control image, and a T1 image. Continuous spin inversion labeling the arterial blood water was accomplished, using principles of adiabatic fast passage by applying continuous-wave radiofrequency power in the presence of a magnetic field gradient in the direction of arterial flow. In the detection slice used to measure perfusion, whole brain CBF averaged 1.39 +/- 0.19 ml.g-1.min-1 (mean +/- SEM, n = 5). The technique's sensitivity to changes in CBF was measured by using graded hypercarbia, a condition that is known to increase brain perfusion. CBF vs. pCO2 data yield a best-fit straight line described by CBF (ml.g-1.min-1) = 0.052pCO2 (mm Hg) - 0.173, in excellent agreement with values in the literature. Finally, perfusion images of a freeze-injured rat brain have been obtained, demonstrating the technique's ability to detect regional abnormalities in perfusion.

Animals↗
PubMed1992

Perfusion imaging.

Measurement of tissue perfusion is important for the functional assessment of organs in vivo. Here we report the use of 1H NMR imaging to generate perfusion maps in the rat brain at 4.7 T. Blood water flowing to the brain is saturated in the neck region with a slice-selective saturation imaging sequence, creating an endogenous tracer in the form of proximally saturated spins. Because proton T1 times are relatively long, particularly at high field strengths, saturated spins exchange with bulk water in the brain and a steady state is created where the regional concentration of saturated spins is determined by the regional blood flow and regional T1. Distal saturation applied equidistantly outside the brain serves as a control for effects of the saturation pulses. Average cerebral blood flow in normocapnic rat brain under halothane anesthesia was determined to be 105 +/- 16 cc.100 g-1.min-1 (mean +/- SEM, n = 3), in good agreement with values reported in the literature, and was sensitive to increases in arterial pCO2. This technique allows regional perfusion maps to be measured noninvasively, with the resolution of 1H MRI, and should be readily applicable to human studies.

Animals↗
PubMed1992

Measurement of brain perfusion by volume-localized NMR spectroscopy using inversion of arterial water spins: accounting for transit time and cross-relaxation.

The theoretical model for perfusion measurement by NMR using arterial labeling of endogenous water is extended to include the effects of transit time and cross-relaxation of tissue water with macromolecules. Water magnetization in rat brain is monitored using the STEAM method to simultaneously determine the transit time, magnetization transfer rate constant, and perfusion. The results show that the transit time in rat brain is quite short, and thus its effect on perfusion measurement is small. It is also demonstrated both theoretically and experimentally that the steady-state effects of cross-relaxation with macromolecules on perfusion measurement are accounted for by a proper control experiment.

Animals↗
PubMed1992

Localization of peripherin/rds in the disk membranes of cone and rod photoreceptors: relationship to disk membrane morphogenesis and retinal degeneration.

The outer segments of vertebrate rod photoreceptor cells consist of an ordered stack of membrane disks, which, except for a few nascent disks at the base of the outer segment, is surrounded by a separate plasma membrane. Previous studies indicate that the protein, peripherin or peripherin/rds, is localized along the rim of mature disks of rod outer segments. A mutation in the gene for this protein has been reported to be responsible for retinal degeneration in the rds mouse. In the present study, we have shown by immunogold labeling of rat and ground squirrel retinas that peripherin/rds is present in the disk rims of cone outer segments as well as rod outer segments. Additionally, in the basal regions of rod and cone outer segments, where disk morphogenesis occurs, we have found that the distribution of peripherin/rds is restricted to a region that is adjacent to the cilium. Extension of its distribution from the cilium coincides with the formation of the disk rim. These results support the model of disk membrane morphogenesis that predicts rim formation to be a second stage of growth, after the first stage in which the ciliary plasma membrane evaginates to form open nascent disks. The results also indicate how the proteins of the outer segment plasma membrane and the disk membranes are sorted into their separate domains: different sets of proteins may be incorporated into membrane outgrowths during different growth stages of disk morphogenesis. Finally, the presence of peripherin/rds protein in both cone and rod outer segment disks, together with the phenotype of the rds mouse, which is characterized by the failure of both rod and cone outer segment formation, suggest that the same rds gene is expressed in both types of photoreceptor cells.

Animals↗
PubMed1992

Differential localizations of and requirements for the two Drosophila ninaC kinase/myosins in photoreceptor cells.

The ninaC gene encodes two retinal specific proteins (p132 and p174) consisting of a protein kinase domain joined to a domain homologous to the head region of the myosin heavy chain. The putative myosin domain of p174 is linked at the COOH-terminus to a tail which has some similarities to myosin-I tails. In the current report, we demonstrate that the ninaC mutation results in light- and age-dependent retinal degeneration. We also show that ninaC flies display an electrophysiological phenotype before any discernible retinal degeneration indicating that the electrophysiological defect is the primary effect of the mutation. This suggests that ninaC has a role in phototransduction and that the retinal degeneration is a secondary effect resulting from the defect in phototransduction. To examine the requirements for the individual ninaC isoforms, mutant alleles were generated which express only p132 or p174. Elimination of p174 resulted in a ninaC phenotype as strong as the null allele; however, elimination of p132 had little if any effect. As a first step in investigating the basis for the difference in requirements for p174 and p132 we performed immuno-localization at the electron microscopic level and found that the two isoforms display different subcellular distributions in the photoreceptor cells. The p132 protein is restricted primarily to the cytoplasm and p174 to the rhabdomeres, the microvillar structure which is the site of action of many of the steps in phototransduction. This suggests that the p174 myosin-I type tail is the domain responsible for association with the rhabdomeres and that the substrate for the p174 putative kinase may be a rhabdomeric protein important in photo-transduction.

Aging↗
PubMed1992

Distribution of the myosin I-like ninaC proteins in the Drosophila retina and ultrastructural analysis of mutant phenotypes.

The Drosophila ninaC gene encodes for two head-specific proteins of 132 kDa and 174 kDa. Their predicted amino acid sequences indicate that they may have myosin I and kinase properties. We have: (1) determined the cellular and subcellular distributions of the ninaC proteins in the Drosophila retina by electron microscopic immunocytochemistry with an antibody specific for epitopes shared by both proteins; (2) characterized the ultrastructure of the mutant phenotype. The proteins were detected only in the photoreceptor cells, but were detected in all classes of the compound eye photoreceptors. Within the photoreceptors, they were found in the rhabdomeral microvilli and the cytoplasm adjacent to the rhabdomeres. This distribution coincides with that shown previously for actin filaments. Immunolabelling of tissue from the ninaC P221 mutant, which lacks the 174 kDa protein, and two mutants whose rhabdomeres degenerate, suggests that the 132 kDa protein is present primarily in the cytoplasm adjacent to the rhabdomeres, and that the 174 kDa protein is concentrated in the rhabdomeres. Our ultrastructural analysis showed that the axial cytoskeleton of the rhabdomeral microvilli (which contains filamentous actin) was absent in both the null and P221 mutants. In the photoreceptor cell cytoplasm, the number of multivesicular bodies in the null mutant, but not the P221 mutant, was 3-fold greater in comparison with wild-type.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Association of myosin with the connecting cilium of rod photoreceptors.

The cilium of a vertebrate photoreceptor cell connects the phototransductive outer segment of the cell to the inner segment. Previous studies have shown that, within the connecting cilium, there is a small cluster of actin filaments, which play a critical role in the formation of new disk membranes. Here, we have detected a polypeptide in rat rod outer segments that is recognized by myosin heavy chain antibodies and was found to possess other characteristics of conventional non-muscle myosin heavy chain: it comigrates in SDS-PAGE with non-muscle myosin heavy chain; it associates with the cytoskeleton of rod outer segments in an ATP-sensitive manner; and it binds to purified actin filaments in the absence of ATP. Myosin ATPase activity was also detected in isolated rod outer segments. Electron immunomicroscopy revealed that myosin is present in the small actin-containing domain within the connecting cilium at the site of disk membrane morphogenesis. These results pose the possibility that an actin-myosin contractile mechanism functions in the formation of new photoreceptor disk membranes.

Actins↗
PubMed1992

Modification of the daily photoreceptor membrane shedding response in vitro by antioxidants.

The shedding of rod outer segment disc membrane was investigated in a commonly used frog eyecup preparation, with respect to length of time in vitro. It was found that the amplitude, but not the timing, of the "dawn" (ie, when the lights are turned on) shedding response was increased threefold if the time in vitro was increased from 1.5-2 hr to 12-13 hr before dawn. After the overnight incubation, morphologic deterioration of the photoreceptors was not apparent by light or electron microscopy. The addition of the antioxidants, vitamin C or alpha-tocopherol (vitamin E), inhibited the increase in shedding, suggesting that it was induced by oxidation. These results help define conditions necessary for the maintenance of normal retinal metabolism in vitro. They also suggest a link between the amplitude of the dawn disc membrane shedding response and oxidation.

Animals↗
PubMed1991

Involvement of protein kinase C in the phosphorylation of rhodopsin.

The possibility that protein kinase C is involved in phototransduction by phosphorylating rhodopsin was explored in situ and in vitro. Pretreatment of intact retinas with phorbol myristate acetate markedly increased the light-dependent phosphorylation of rhodopsin, with the greatest effects observed at lower light levels. Phorbol myristate acetate treatment did not affect rhodopsin phosphorylation in retinas not exposed to light, suggesting that protein kinase C modulates the phosphorylation state of rhodopsin in a light-dependent manner. Limited proteolysis of rhodopsin phosphorylated in situ indicates that protein kinase C modifies rhodopsin on a domain distinct from that recognized by rhodopsin kinase. In vitro, protein kinase C purified from bovine retinas phosphorylated unbleached and bleached rhodopsin. Our results are consistent with protein kinase C phosphorylating unbleached rhodopsin in response to low light, suggesting that protein kinase C plays a role in light adaptation.

Amino Acid Sequence↗
PubMed1991

Are universal precautions effective in reducing the number of occupational exposures among health care workers? A prospective study of physicians on a medical service.

Using a daily questionnaire, we prospectively studied 277 physicians from two hospital medical services for incidents of exposure to blood and body fluids and barrier use before and after the implementation of universal precautions. We found that implementation significantly increased the frequency of barrier use during exposure incidents from 54% before implementation to 73% after implementation of universal precautions. Implementation led to a decrease in the number of exposure incidents that resulted in direct contact with blood and body fluids (actual exposures), from 5.07 to 2.66 exposures per physician per patient care month, and to an increase in averted exposures in which direct contact was prevented by the use of barrier devices, from 3.41 exposures per patient care month before implementation to 5.90 exposures per patient care month after implementation. Implementation affected neither the types of body fluid or procedures involved nor the overall rate of exposure incidents (8.5 per patient care month) but, through an increase in barrier use, it did prevent direct contact with blood and body fluids and thus converted what would have been an actual exposure into an averted one. We conclude that universal precautions were effective in reducing the risk of occupational exposures among physicians on a medical service.

Body Fluids↗
PubMed1991

Actin filaments and photoreceptor membrane turnover.

The shape and turnover of photoreceptor membranes appears to depend on associated actin filaments. In dipterans, the photoreceptor membrane is microvillar. It is turned over by the addition of new membrane at the bases of the microvilli and by subsequent shedding, mostly from the distal ends. Each microvillus contains actin filaments as a component of its cytoskeletal core. Two myosin I-like proteins co-localize with the actin filaments. It is suggested that one of the myosin I-like proteins might be linked to the microvillar membrane. By interacting with the actin filaments, this motor should move the membrane of a microvillus in a distal direction, thus providing a possible mechanism for the turnover of the membrane. A vertebrate photoreceptor cell contains a small cluster of actin filaments in its connecting cilium at the site where new transductive disk membranes are formed. Disruption of the actin filaments perturbs disk morphogenesis. The most likely explanation for this perturbation is that the process of initiating a new disk is inhibited. Conventional myosin (myosin II) is found in the connecting cilium with the same distribution as actin. A simple model is proposed to illustrate how the actin-myosin system of the connecting cilium might function to initiate the morphogenesis of a disk membrane.

Actins↗
PubMed1991

Alpha-actinin and actin in the outer retina: a double immunoelectron microscopic study.

Actin has many diverse functions in the outer retina. To help elucidate its organization in this area, we have investigated the extent of its association with the actin cross-linking protein alpha-actinin. Ultrathin sections of chicken retina were double-immunolabelled with monospecific antibodies against actin and alpha-actinin. The highest relative amount of alpha-actinin to actin label was measured in the adherens junctions between the individual retinal pigmented epithelial (RPE) cells and between the photoreceptor and Mueller cells; in the photoreceptor myoid; and in the RPE basal microvilli. The lowest amount was in the Mueller cell microvilli, the RPE apical processes, and in the photoreceptor ellipsoid. It is likely that the areas containing the highest ratio of alpha-actinin to actin labelling are where the actin filaments are most highly cross-linked into bundles and linked to the plasma membrane by alpha-actinin. Actin filaments terminate in these areas, and, except for the myoid region, they are involved in cell-cell or cell-substrate adherens junctions.

Actinin↗
PubMed1991

An evaluation of the integrity of the blood-testis barrier by magnetic resonance imaging.

This present investigation was initiated to noninvasively evaluate the usefulness of intravenous gadopentetate dimeglumine (Gd-PTDM)-enhanced magnetic resonance imaging (MRI) in assessing the integrity of the blood-testis barrier (BTB). The intensity of different image slices was measured in pre- and post-Gd-PTDM images in rats receiving cytochalasin D or alcohol treatment. It was found that MRI can be used to assess the integrity of the BTB and may be a useful tool in the evaluation of potentially toxic agents that affect the testis, particularly those that affect the BTB.

Animals↗
PubMed1991

Cyclosporine and liver regeneration studied by in vivo 31P nuclear magnetic resonance spectroscopy.

The changes in fructose-1-phosphate (F-1-P), intracellular pH, and ATP content of the liver after a fructose challenge were investigated noninvasively in vivo using phosphorus-31 nuclear magnetic resonance spectroscopy of dog liver four days after a portacaval shunt (PCS) with or without portal venous infusion of cyclosporin (CsA). The F-1-P metabolism was slower in PCS dogs (N = 2) as compared to either the normal (N = 2) or PCS + CsA-treated dogs (N = 3) (P less than 0.05). The intracellular pH temporarily decreased from 7.3 +/- 0.05 to 7.0 +/- 0.05 during the fructose challenge. The regenerative indexes were increased in the PCS + CsA group (P less than 0.01). These data obtained in vivo using 31P-NMR spectroscopy in the liver following a portacaval shunt, suggest that: (1) the energy status of the liver and the metabolic response to fructose are reduced in PCS compared to normal animals and (2) CsA treatment enhances the regenerative response of the liver and prevents the reduction in hepatic function associated with portacaval shunting.

Acid-Base Equilibrium↗
PubMed1991

Epithelial cell-substrate adhesion in the cornea: localization of actin, talin, integrin, and fibronection [corrected].

During corneal wound healing, epithelial cells migrate and spread over a basement membrane to cover the denuded area. We have initiated studies on the proteins involved in this cell-substrate interaction. In the present report, we show the distribution of actin, talin, integrin and fibronectin in the unwounded chicken corneal epithelium. Rhodamine-phalloidin and antibodies against talin, the beta-subunit of integrin, and fibronectin were used to fluorescently label semi-thin cryosections of chicken cornea. Phalloidin labeling indicated the presence of f-actin around the boundaries of all epithelial cells. Antitalin labeled the most basal aspect of the basal cells. Diffuse cytoplasmic labeling of the basal cells was also seen. Integrin was detected by the polyclonal anti-CSAT and monoclonal 30B6 antibodies. With both antibodies, the basal cells were outlined by label. These cells were similarly labeled by antifibronectin. Less distinct labeling of fibronectin was present around the boundaries of the outer epithelial cells. Our results indicate that many of the proteins associated with cell-substrate adherens junctions in other systems are also present in cell-substrate adhesion of the corneal epithelium. Details of the distribution of some of the proteins appear to be somewhat unusual, however.

Actins↗