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R M Albrecht

Publications and source records attributed to R M Albrecht.

At least 37 records · Page 2Linked to original sources
PubMed1996

Self-association of bound fibrinogen on platelet surfaces.

Binding of fibrinogen to receptors on the surfaces of activated platelets triggers movement of the ligand-bound receptors. In this study this process was followed by using native and colloidal gold-labeled fibrinogen. Both labeled and unlabeled proteins on platelet surfaces could be visualized by low-voltage, high-resolution scanning electron microscopy. Fibrinogen and gold-conjugated fibrinogen were observed to bind to platelet surfaces and to trigger identical patterns of receptor-ligand complex redistribution. In addition to previously described long- and short-range translocation patterns, fibrinogen, either unlabeled or conjugated to gold particles, formed small, specific, nonfibrillar aggregates after binding to platelet surface receptors. Similar triggering and movement resulted from binding of gold-conjugated antibody to the fibrinogen receptor, but no subsequent self-association of the antibody-gold was observed.

Blood Platelets↗
PubMed1996

Failure of antiseptic bonding to prevent central venous catheter-related infection and sepsis.

Infection associated with the use of triple lumen catheters in hospitals is a frequent and serious complication. The prevailing hypothesis for the origin of catheter-related infection (CRI) is bacterial colonization and subsequent infection of the skin insertion site and catheter interface. The recently released ARROWgard catheter contains a bonded synergistic combination of silver sulfadiazine and chlorhexidine, which is thought to render the catheter resistant to bacterial colonization and subsequent sepsis. The purpose of this study is to compare the incidence of CRI and catheter-related sepsis (CRS) between a standard triple lumen catheter and ARROWgard antiseptic coated catheter in patients receiving total parenteral nutrition (TPN). A randomized, prospective clinical trial was conducted at a community referral center from January 1993 through April 1994. One-hundred-ninety-one patients with need for TPN were randomized to receive either the ARROWgard or a standard triple lumen catheter placed under a strict sterile protocol. CRI was defined as >/= 15 colony forming units by semiquantitative culture technique of the catheter tip or intracutaneous segment. CRS was defined as growth of the same organism on the catheter and at least one peripheral blood culture. All catheters were cultured. Ninety-two patients received the ARROWgard catheter, and 99 patients received the standard catheter. There were no differences between the average age, sex, length of hospital stay, days on TPN, number of catheters/patient, indications for TPN, primary diagnoses, or duration of the central line between the two groups. The overall rate of CRI was 11.5 per cent, and CRS was 8.4 per cent in this study. The rate of CRI for the ARROWgard was 10.9 per cent, compared with 12.9 per cent for the standard catheter (P = NS). The rate of CRS for the ARROWgard was 8.7 per cent, compared with 8.1 per cent for the standard catheter (P = NS). The coating of central venous catheters with silver sulfadiazine and chlorhexidine does not reduce the rate CRI or CRS when compared with standard central venous catheters in patients receiving TPN.

Anti-Infective Agents, Local↗
PubMed1995

Cell-surface receptors and proteins on platelet membranes imaged by scanning force microscopy using immunogold contrast enhancement.

High resolution scanning force microscope (SFM) images of fibrinogen-exposed platelet membranes are presented. Using ultrasharp carbon tips, we are able to obtain submolecular scale resolution of membrane surface features. Corroboration of SFM results is achieved using low voltage, high resolution scanning electron microscopy (LVHRSEM) to image the same protein molecule that is seen in the SFM. We obtain accurate height dimensions by SFM complemented by accurate lateral dimensions obtained by LVHRSEM. The use of 14- and 5-nm gold labels to identify specific membrane-bound biomolecules and to provide contrast enhancement with the SFM is explored as a useful adjunct to observation of unlabeled material. It is shown that the labels are useful for locating specific protein molecules on platelet membrane surfaces and for assessing the distribution of these molecules using the SFM. Fourteen nm labels are shown to be visible over the membrane corrugation, whereas 5-nm labels appear difficult to resolve using the present SFM instrumental configuration. When using the 5-nm labels, collateral use of LVHRSEM allows one to examine SFM images at submolecular resolution and associate function with the structures imaged after the SFM experiment is completed.

Blood Platelets↗
PubMed1995

Use of adhesion-defective mutants of Staphylococcus aureus to define the role of specific plasma proteins in promoting bacterial adhesion to canine arteriovenous shunts.

We used an ex vivo canine arteriovenous shunt model, previously developed to study plasma protein adsorption and thrombogenesis on polymeric biomaterials, to define the role of host proteins in promoting adhesion of Staphylococcus aureus. Either polyethylene or polyvinyl chloride tubings were exposed to canine blood for 5, 15, or 60 min at a flow rate of 300 ml/min and then were flushed in phosphate-buffered saline (PBS), cut into 1.5-cm segments, and stored at -70 degrees C. After thawing, each segment was preincubated in 0.5% albumin in PBS to prevent nonspecific staphylococcal attachment to surfaces that were not exposed to blood. Each segment was then incubated with 4 x 10(6) CFU of [3H]thymidine-labelled S. aureus per ml for 60 min at 37 degrees C in an in vitro adhesion assay. Two site-specific mutants of S. aureus were tested: one specifically defective in adhesion to surface-bound fibronectin (FnAd-def) and the other defective in adhesion to fibrinogen (FgAD-def) [corrected]. Compared with their respective parental strains, the FgAd-def, but not the FnAd-def, mutant of S. aureus showed a strong (> 80%) decrease in attachment to ex vivo tubings. The adhesion of each strain of S. aureus onto polyethylene was consistently more than twofold higher than the adhesion onto polyvinyl chloride segments exposed to flowing blood for 5 or 15 min, but adhesion became similar to that on polyvinyl chloride after 60 min of exposure. In conclusion, the specific adhesion-defective mutants of S. aureus suggested that fibrinogen was the most active adhesion-promoting protein in a short-term blood-material interaction. The experimental approach described in this study should prove useful for screening materials thought to be resistant to protein-mediated staphylococcal adhesion and colonization.

Animals↗
PubMed1995

Failure of methylprednisolone to improve the outcome of spinal cord injuries.

The infusion of methylprednisolone (MP) within 8 hours of injury for spinal cord injuries (SCI) has been advocated to improve the motor function of patients after this catastrophic injury. However, clinical improvement in the outcome of SCI has not been consistently identified, despite the use of MP. We reviewed the outcome of SCI patients with MP to those without MP (No-MP) at two Level I Trauma Centers from 1989-1992. Acute SCI patients were identified from the trauma registries with trauma demographics and hospital data obtained from registry and medical records. Rehabilitation data for Functional Independence Measure (FIM) was obtained from the rehabilitation institute database. Primary outcome parameters were mortality, and for survivors, patient mobility (6 point scale) and FIM scores. There were 145 acute SCI patients: 80 treated with MP and 65 with No-MP. FIM data was available on 45 MP and 25 No-MP patients. There was no difference in the admission trauma score, ICU length of stay (LOS), or hospital LOS between the two groups. The MP patients were significantly younger (30 years vs 38 years, P = < 0.05) and had lower ISS scores (24 vs 31, P = < 0.05). There was no statistically significant difference in mortality (MP, 3.8% vs No-MP, 10.7%) between the two groups. Although admission mobility was not statistically different (MP, 5.99 vs No-MP, 5.90), there was a significantly poorer discharge mobility in the MP group when compared to the No-MP group (MP, 5.16 vs No-MP, 4.67, P = < 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Activities of Daily Living↗
PubMed1994

Modulation of cell surface fibronectin assembly sites by lysophosphatidic acid.

Lysophosphatidic acid is a product of activated platelets and has diverse actions on cells. We have characterized the effect of lysophosphatidic acid on cell-mediated binding and assembly of fibronectin, an extracellular matrix protein. Serum made from whole blood, but neither platelet-poor plasma nor serum made from platelet-poor plasma, caused enhanced binding of fibronectin to cultured fibroblastic cells. The ability of whole blood serum to enhance binding of fibronectin was abolished by phospholipase B. These results indicate that lysophosphatidic acid derived from platelets is the principal component in whole blood serum that is active in the fibronectin binding assay. 1-oleoyl lysophosphatidic acid, 20-200 nM, was as active as 0.1-0.2% whole blood serum. The stimulatory effect of lysophosphatidic acid on the binding of fibronectin or the amino-terminal 70-kD fragment of fibronectin was rapid, sustained, and lost upon removal of lysophosphatidic acid. The stimulatory effect on binding could not be duplicated by bradykinin, platelet-activating factor, bombesin, or a peptide agonist of the thrombin receptor. Enhanced binding of the 70-kD fragment was due to increases in both the number and affinity of binding sites. Enhanced binding and assembly of fibronectin correlated with changes in cell shape and actin-containing cytoskeleton. The binding sites for fibronectin on lysophosphatidic acid-stimulated cells, as assessed by fluorescence, video, and scanning electron microscopy, were on areas of cell membrane containing numerous filopodia that extended between cells or between cells and substratum. These observations suggest that lysophosphatidic acid functions as a powerful and specific modulator of cell shape and early matrix assembly during wound healing.

Actin Cytoskeleton↗
PubMed1994

Biomaterials with permanent hydrophilic surfaces and low protein adsorption properties.

Low protein adsorbing polymer films have been prepared with which to fabricate intravenous containers, designed for compatibility with low concentrations of protein drugs. The material is economically manufactured utilizing physical melt blending of water-soluble surface-modifying polymers (PEO, PEOX, PVA, and PNVP) with a base polymer (EVA, PP, PETG, PMMA, SB, and nylon). Permanency of the hydrophilic surfaces so generated was confirmed by surface contact angle experiments and total organic carbon leachables analysis of the aqueous contacting solutions. Binding of IgG, albumin and insulin was studied. A sixfold reduction of protein adsorption was obtained by adding 5% PVA13K to EVA, for IgG at a bulk concentration of 2.5 ppm. Surface bound protein measured by micro-BCA colorimetry, agreed with the solution protein lost, as determined by the Fluoraldehyde procedure. Imaging of the protein exposed plastic surfaces by silver enhanced protein conjugated gold staining agreed with the quantitative assay determinations.

Adsorption↗
PubMed1993

Integrin receptors and platelet adhesion to synthetic surfaces.

The activation-independent and -dependent integrin receptors--glycoproteins GPIc-IIa (alpha 5-beta 1) and GPIIb-IIIa (alpha IIb-beta 3)--are involved in platelet adhesion and thrombus growth on damaged subendothelium through interactions with fibrinogen, fibronectin, von Willebrand factor, and other adhesive proteins. Because these receptors are used in normal in vivo hemostatic adhesion, they may also have a role for adhesion onto synthetic surfaces in the vasculature. Platelet adhesion in vitro was examined onto Formvar, glass, and four polyurethaneureas with various soft segment chemistries and surface properties. Platelets were pretreated with RGD peptides before and after adhesion. RGD peptide pretreatment inhibited spreading and close contact formation compared to treatment with saline or control RGE peptides, with no observable effect on the number of adherent platelets per area. High-voltage electron microscopy showed abnormally sparse and short microfilament structures with RGD peptide treatment, suggesting an indirect inhibition of actin filament formation. Video-enhanced light microscopy showed a cessation of spreading and a partial reversal of close contacts following RGD peptide application to adherent platelets. Because minimal amounts of plasma proteins are present in column-washed platelet suspensions, and as platelet secretion appeared to be minimal in these experiments, these observations suggest that RGD binding integrin receptors may function in platelet spreading even in the absence of exogenous ligand. As RGD peptides did not affect the numbers of adherent platelets, while producing substantial decreases in the extent of spreading, we suggest that platelet integrins, possibly GPIIb-IIIa, are involved in spreading on synthetic surfaces but not for initial adhesion.

Blood Platelets↗
PubMed1993

Adhesion of Staphylococcus aureus to surface-bound platelets: role of fibrinogen/fibrin and platelet integrins.

Platelets adhering to artificial or biologic surfaces have been implicated in the pathogenesis of catheter infections or endocarditis; however, the ligands involved in Staphylococcus aureus interaction with adherent platelets remain incompletely understood. Radiolabeled S. aureus Cowan I were incubated with purified platelets adherent to polymethylmethacrylate (PMMA) coverslips and washed, and adhesion was determined. Platelets promoted adhesion of S. aureus approximately 30-fold compared with adhesion to albumin-PMMA. In the presence of both plasma (1% vol/vol) and platelets, adhesion was extensively promoted, with 30% (of inoculated) S. aureus adherent (150-fold increase). Platelet pretreatment with anti-GPIIb/IIIa monoclonal antibodies or inhibitors of platelet activation decreased plasma-enhanced adhesion, suggesting a role of platelet activation in S. aureus adhesion. Plasma-enhanced adhesion was sensitive to thrombin antagonists, proteinase inhibitors, heparin, or antifibrinogen antibodies, indicating that fibrinogen/fibrin is necessary for bridging between adherent platelets and S. aureus. In conclusion, S. aureus adhesion to immobilized platelets may play a role in the pathogenesis of invasive bloodstream infections or endocarditis.

Bacterial Adhesion↗
PubMed1993

Ischemia/reperfusion injury of the ascending colon in ponies: a correlative study utilizing microvascular histopathology and corrosion casting.

Volvulus of the ascending colon (ACV) in the horse results in microvascular injury and necrosis of the intestinal mucosa. This study investigated the site and type of microvascular injury which occurs within the mucosa and submucosa following ACV. Histopathology of volvulus treated ponies demonstrated mucosal necrosis with microvascular hemorrhage and thrombosis. Thrombi occurred within the subepithelial capillaries and edema and hemorrhage developed throughout the mucosa and submucosa. Vascular casts allowed 3-D viewing of samples obtained from the entire pelvic flexure and demonstrated two distinct microvascular changes: 1) disruption of the colonic glandular capillary network occurred concomitantly with the mucosal injury, and 2) extensive endothelial leakage from the submucosal microvasculature contributes to edema formation. Thus, microcorrosion casting of the equine pelvic flexure provided an effective means to characterize the location and severity of vascular leakage and visualize the extent and severity of injury to the capillary network not easily depicted by histopathology. Microvascular casting in conjunction with routine histopathology provided additional information on the pathomorphologic changes in this model of ischemia/reperfusion injury.

Animals↗
PubMed1993

Improved tissue corrosion of vascular casts: a quantitative filtration method used to compare tissue corrosion in various concentrations of sodium and potassium hydroxide.

In this study, we compare weights of filter retained material (11 microns particle retention) after solubilization and filtration of unfixed, un-perfused tissue (fat, liver and trachea) in sodium and potassium hydroxide (1%, 5%, 10% and 20% weight/volume, w/v) at 8, 24 and 48 hour time points at 45 degrees C. Three detergents [1% Triton-X-100 (volume/volume, v/v), 1% 7X (v/v), 1% Terg-A-Zyme (w/v)] used in combination with hydroxide were evaluated for use in solubilizing fat. Additionally, vascular casts from mouse kidneys were corroded to test the practical effectiveness of corroding solutions on resin infused tissue. Five percent KOH for eight hours proved to be the most effective concentration and time required to corrode fatty tissue. Liver tissue was corroded most rapidly in 1% to 5% NaOH or in 1% to 20% KOH. Corrosion of trachea tissue showed that 5, 10 and 20% hydroxide (NaOH or KOH) are equally effective after 8 hours of corrosion. Use of detergents improved solubilization of fat when combined with 2%, 3% or 5% NaOH. However, fatty tissue was solubilized more rapidly without the use of detergents in 1% NaOH. Scanning electron microscopy of vascular casts shows that corrosion in 1% NaOH appears equally as effective as corrosion in 15% KOH.

Adipose Tissue↗
PubMed1993

Evidence for two mechanisms of ligand-receptor movement on surface-activated platelets.

The movement of platelet membrane glycoproteins was studied by correlative light and electron microscopy. Two distinct classes of movement were seen. In the first type, binding of colloidal gold-labeled ligand or antibody to the fibrinogen receptor (GPIIb/IIIa) triggered a long-range centripetal movement of the receptor-ligand complexes, in the plane of the membrane, on the surfaces of fully spread platelets. This movement was dependent on the presence of the intact cytoplasmic actin cytoskeleton, as evidenced by its inhibition by treatment of the platelets with cytochalasins D and E to disrupt actin filaments. A second surface receptor, the platelet von Willebrand's factor receptor (GPIb), did not undergo this long range, actin-dependent movement in response to antibody binding. However, in a second class of movement, both GPIb and GPIIb/IIIa were translocated on the platelet membrane. Both receptors, GPIb and GPIIb/IIIa, in response to binding of gold-labeled antibody or ligand, accumulated in the channels of the open canalicular system when these were present, and between platelets, under the platelet margins at the sites of platelet-platelet contact. This type of movement occurred only over a much shorter distance than the centripetal movement seen with ligand-bound GPIIb/IIIa and was not affected by the disruption of filamentous actin by the cytochalasins. These results suggest that there are two active mechanisms driving ligand-receptor movement on surface-activated platelets, and that one receptor type may be driven by either mechanism. However, each mechanism predominates at a different stage in the sequence of platelet activation, adherence, and spreading, consistent with differences in function related to receptor-ligand complex formation at the different stages.

Actins↗
PubMed1992

Cytochalasin D and E: effects on fibrinogen receptor movement and cytoskeletal reorganization in fully spread, surface-activated platelets: a correlative light and electron microscopic investigation.

This study investigates the involvement of actin microfilaments in fibrinogen receptor redistribution and cytoskeletal reorganization that takes place in fully spread, surface-activated platelets. Colloidal gold-labeled fibrinogen (Fgn-Au label) in conjunction with video-enhanced differential interference contrast light microscopy (VDIC) was used to identify fibrinogen binding sites, glycoprotein IIb/IIIb (GPIIb/IIIa), on fully spread platelets. Platelets were treated with cytochalasins D and E (5 x 10(-5) mol/L to 5 x 10(-8) mol/L) for 10 minutes, before or after incubation with Fgn-Au label. Results observed with VDIC were subsequently confirmed by high-voltage transmission and low voltage-high resolution scanning electron microscopic examination of the specimens. Preincubation of activated platelets with cytochalasin D or E (5 x 10(-5) and 5 x 10(-6) mol/L) inhibited fibrinogen receptor redistribution and abolished cytoskeletal reorganization in fully spread platelets. After surface-activated platelets were incubated with Fgn-Au label, treatment with the above concentrations of cytochalasin D or E disrupted cytoskeletal reorganization and caused random movement of previously redistributed receptor-ligand complexes. Incubation of platelets with cytochalasin E 5 x 10(-6) mol/L prevented platelet activation and spreading. Thus, actin filaments appear necessary for platelet spreading from the discoid to the fully spread stage. The ligand-triggered, cytoskeletally directed movement of fibrinogen receptors in fully spread platelets appears to be dependent on the presence of intact, polymerized actin. This movement is distinct from the cytochalasin-insensitive accumulation of GPIIb/IIIa-ligand in the channels of the open canalicular system.

Actins↗
PubMed1991

The effects of substrate-adsorbed albumin on platelet spreading.

Adsorbed albumin appears to passivate nearly all materials, minimizing platelet adhesion and thrombus formation. Since in vitro platelet spreading can be an indicator of in vivo reactivity leading to thrombosis, and as in vitro platelet adhesion investigations are routinely done in the presence of bovine or human serum albumin (BSA or HSA), we examined the influence of albumin on platelet reactivity to material substrates. Platelet spreading was examined subsequent to adherence onto several related polyurethanes, and to Formvar, in the presence of bulk albumin concentrations sufficient to form an adsorbed monolayer or a multilayer. No other exogenous proteins were present. The spreading behavior of adherent platelets was analyzed using generalized linear interactive modeling (GLIM). The models showed that the polymer type always influenced platelet responses, irrespective of the albumin concentration. In many experiments, platelet behavior could be adequately modeled without including the effects of albumin. Thus, the polymer type appeared to be the primary determinant of platelet shape-change with adsorbed albumin producing a secondary effect. Additionally, somewhat different effects on spreading were observed with HSA and BSA, suggesting qualitatively different interactions between human platelets and HSA, than with BSA, which is commonly used in platelet preparations.

Adsorption↗
PubMed1991

Increased acylcarnitine clearance and excretion in septic rats.

Carnitine has been hypothesized to be a semi-essential nutrient in the nutrition of critically ill patients. The purpose of this study was to evaluate the effect of sepsis upon carnitine metabolism in the rat, using the model of cecal ligation and puncture. Three treatment groups, septic, sham, and non-operative controls, were used. The septic rats had significantly increased (p less than 0.05) excretion of acylcarnitine and over six-fold higher urinary acylcarnitine/free carnitine ratio, relative to the other two groups. The septic rats also had significantly higher liver and plasma free and total carnitine compared to the other two groups. A possible explanation for the increased urinary acylcarnitine excretion is that carnitine may be acting to remove toxic metabolites from the body. The septic model of cecal ligation and puncture was suitable for the study of carnitine metabolism during sepsis in the rat.

Animals↗
PubMed1990

Failure of BCAA supplementation to promote nitrogen retention in injured patients.

The purpose of this study was to determine if supplementing total parenteral nutrition (TPN) with lipids or the branched chain amino acids (BCAA) leucine, isoleucine, and valine influences nitrogen balance in the injured patient. Four TPN study solutions were used, with each patient receiving two of the solutions for 4 days each. Group A received solutions consisting of 19% and 44% BCAA, with nonnitrogen calories supplied by 100% carbohydrate. Group B received a 7:3 carbohydrate-to-lipid ratio of nonnitrogen calories as a fuel source. Neither BCAA supplementation nor varying fuel substrates promoted a difference in nitrogen retention. The added cost of BCAA supplementation, along with the lack of an effect upon nitrogen accretion, indicates that greater benefits must be demonstrated before widespread use of BCAA supplementation can be recommended in the injured patient.

Amino Acids, Branched-Chain↗
PubMed1990

Optimizing parameters for correlative immunogold localization by video-enhanced light microscopy, high-voltage transmission electron microscopy, and field emission scanning electron microscopy.

Correlative video-enhanced light microscopy, high-voltage transmission electron microscopy, and low-voltage high resolution scanning electron microscopy were used to examine the binding of colloidal gold-labeled fibrinogen to platelet surfaces. Optimal conditions for the detection of large (18 nm) and small (3 nm) gold particles are described.

Blood Platelets↗