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Biomedical subjects

E A Hall

Publications and source records attributed to E A Hall.

At least 19 recordsLinked to original sources
PubMed2005

Detection of the steroidogenic acute regulatory protein, StAR, in human liver cells.

Overexpressing StAR (a mitochondrial cholesterol transporter) increases (>5-fold) the rate of 27-hydroxylation of cholesterol and the rates of bile acid synthesis in primary rat hepatocytes; suggesting that the transport of cholesterol into mitochondria is rate-limiting for bile acid biosynthesis via the CYP27A1 initiated 'acidic' pathway. Our objective was to determine the level of StAR expression in human liver and whether changes in StAR would correlate with changes in CYP27A1 activity/bile acid synthesis rates in human liver tissues. StAR mRNA and protein were detected in primary human hepatocytes and HepG2 cells by RT-PCR/Northern analysis and by Western analysis, respectively. In immunocompetition assays, liver StAR was competed away with the addition of purified human adrenal StAR. Overexpressing CYP27A1 in both cell types led to >2-fold increases in liver StAR concentration. StAR protein levels also increased approximately 2-fold with the addition of 27-hydroxycholesterol to HepG2 cell culture medium. Overexpressing StAR increased the rates of 27-hydroxylation of cholesterol/bile acid synthesis in both cell lines and increased intracellular levels of 27-hydroxycholesterol. In conclusion, human liver cells contain regulable StAR protein whose level of expression appears capable of regulating cellular cholesterol homeostasis, representing a potential therapeutic target in the management of hyperlipidemia.

Bile Acids and Salts↗
PubMed2005

Mitochondrial cholesterol transport: a possible target in the management of hyperlipidemia.

Sterol 27-hydroxylase (CYP27A1) may defend cells against accumulation of excess cholesterol, making this enzyme a possible target in the management of hyperlipidemia. The study objective was to analyze cholesterol homeostatic responses to increases in CYP27A1 activity in HepG2 cells and primary human hepatocytes. Increasing CYP27A1 activity by increasing enzyme expression led to significant increases in bile acid synthesis with compensatory increases in HMG-CoA reductase (HMGR) activity/protein, LDL receptor (LDLR) mRNA, and LDLR-mediated cholesterol uptake. Under these conditions, only a small increase in cellular 27-hydroxycholesterol (27OH-Chol) concentration was observed. No changes were detected in mature sterol regulatory element-binding proteins (SREBP) 1 or 2. Increasing CYP27A1 activity by increasing mitochondrial cholesterol transport (i.e., substrate availability) led to greater increases in bile acid synthesis with significant increases in cellular 27OH-Chol concentration. Mature SREBP 2 protein decreased significantly with compensatory decreases in HMGR protein. No change was detected in mature SREBP 1 protein. Despite increasing 27OH-Chol and lowering SREBP 2 protein concentrations, LDLR mRNA increased significantly, suggesting alternative mechanisms of LDLR transcriptional regulation. These findings suggest that regulation of liver mitochondrial cholesterol transport represents a potential therapeutic strategy in the treatment of hyperlipidemia and atherosclerosis.

Adenoviridae↗
PubMed2001

Assessment of the fifth ligand-binding repeat (LR5) of the LDL receptor as an analytical reagent for LDL binding.

The fifth ligand binding repeat (LR5) of the low density lipoprotein (LDL) receptor was assessed ex vivo as an 'analytical reagent' to distinguish LDL state, in atherosclerosis risk monitoring. LR5 was immobilized to mercaptoundecanoic acid modified gold surfaces via a glycine linker. Surface plasmon resonance (SPR) was used to monitor LDL binding. Unfolded LR5 was ineffectual as an affinity ligand for LDL but refolded LR5 showed a high affinity for native LDL but little affinity for oxidized LDL. LR5 refolded in the presence of calcium or EDTA gave the equivalent LDL binding capacity. However, EDTA-LR5 was less stable than Ca-LR5 at pH 5 and, from tryptophan fluorescence evidence, they appeared to involve different regions of LR5 and/or LDL in the binding. Involvement of amino acid residues of the calcium cage of LR5 was tested in LDL binding by monitoring calcium ion release with a calcium ionophore. The results were consistent with approximately 7-8 LR5 binding per LDL, of which only some induce calcium release (a maximum of approximately 25 mol% calcium, based on LR5, was released during LDL binding). For LDL binding to the LDL receptor in vivo more than one ligand-binding repeat is needed and this may be consistent with LR5 acting here also at binding sites which other LRs normally occupy in the LDL-LDL receptor complex. This initial study is encouraging for the use of a minimum peptide repeat array based on the conserved region of the LRs as an affinity surface for atherosclerosis risk monitoring.

Amino Acid Sequence↗
PubMed2000

Producing "self-plasticizing" ion-selective membranes.

Polymer membranes have been explored for the analysis of ions that do not require plasticizers and with photocurable properties. This work was focused on investigating the viability of the methacrylic-acrylic copolymers as new self-plasticizing membrane matrixes for ion-selective electrodes or other ion-sensor applications. Copolymers with glass transition temperatures ranging from -20 to -44 degrees C could be prepared without added plasticizer and were found to be functional as ion-selective membranes. Both free-radical solution polymerization and photopolymerization could be used, and "self-plasticizing" behavior of copolymers was observed with a high alkylacrylate (R = C4) content. This was found to be compatible with most commercially available ionophores, and sensors for potassium, sodium, calcium and pH were fabricated entirely by photocure procedures; single-step procedures for the immobilization of benzo-15-crown-5 ionophore on these self-plasticizing copolymer matrixes were also developed. Even though the ionophore was immobilized, potentiometric studies revealed that the ionophore remained functional, and thus, these copolymers have the advantage of suffering neither leaching of ionophore nor plasticizers. All these sensors exhibited a Nernstian or near Nernstian response with selectivity comparable to plasticized PVC membranes or other plasticized and photocurable polymer membranes. The long-term response of the potassium sensor with immobilized ionophore and the sodium sensor showed little deterioration for as long as one month and three months, respectively, under continuous use.

Acrylates↗
PubMed2000

Detection of PCR products using PNA strand invasion.

The unique ability of homopyrimidine peptide nucleic acid (PNA) to strand invade homopurine sites of duplex DNA offers a potential alternative to existing techniques for rapid detection of PCR products. From gel shift studies, PNA was found to specifically strand invade homopurine sites that had been incorporated into an amplicon during the PCR cycle. This was achieved by adding a homopyrimidine sequence to the 5'-terminus of a PCR primer. The position of the strand invasion sites at the termini of the DNA duplex offers kinetic advantages for PNA strand invasion, since the termini of DNA duplexes are known to be unwound. This unwound state was demonstrated using a novel assay that determined single-stranded regions within the amplicon. The presence of the PNA moiety in strand invasion complexes was confirmed by a novel electroblot, an Enzyme Linked Nucleic Acid assay and by an increase in stability as demonstrated by T(m)studies with the Idaho RapidCycler. Since the strand invasion sites can be controlled through selection of the homopurine sequence there is great flexibility for designing strand invasion motifs unique to a particular PCR amplicon, thus providing a huge potential for differentiating and detecting multiplex PCR products.

Base Sequence↗
PubMed2000

Phasor transform to extract glucose and ascorbic acid data in an amperometric sensor.

A method for separating the signals from glucose and ascorbic acid on a single recognition surface using an ac immittance technique is presented. It is proposed that each oxidation process can be represented by a unique vector based on psi and YO, and that the concentration of each analyte can be determined by monitoring the change in the admittance magnitude in the direction of the characteristic angle for that particular species. The total Faradaic admittance (YF,total) for all electroactive species present is given by a linear combination of the independent vectors from the different species. In the system tested, the analytes are glucose and ascorbic acid, the former being estimated via the measurand, hydrogen peroxide. Thus, one of the electroactive species (hydrogen peroxide) is not a bulk solution species, but is 'generated' in the enzyme matrix. The admittance measurements from ascorbic acid and the enzyme-generated hydrogen peroxide showed the characteristic phase angles of each oxidation signal, allowing for good spatial resolution. The behaviour of each of these analytes is presented and calibration curves tested. Based on the calibration curves and the basis vectors, samples containing both glucose and ascorbic acid were measured by transforming the measured total admittance from the complex Cartesian space into 'analyte space', where the X-Y axes are given by the basis vectors ŷEGHP,GOD and ŷAA,GOD, respectively.

Ascorbic Acid↗
PubMed1999

Dipicolinic acid (DPA) assay revisited and appraised for spore detection.

Delayed gate fluorescence detection of dipicolinic acid (DPA), a universal and specific component of bacterial spores, has been appraised for use in a rapid analytical method for the detection of low concentrations of bacterial spores. DPA was assayed by fluorimetric detection of its chelates with lanthanide metals. The influence of the choice and concentration of lanthanide and buffer ions on the fluorescence assay was studied as well as the effects of pH and temperature. The optimal system quantified the fluorescence of terbium monodipicolinate in a solution of 10 microM terbium chloride buffered with 1 M sodium acetate, pH 5.6 and had a detection limit of 2 nM DPA. This assay allowed the first real-time monitoring of the germination of bacterial spores by continuously quantifying exuded DPA. A detection limit of 10(4) Bacillus subtilis spores ml-1 was reached, representing a substantial improvement over previous rapid tests.

Bacillus subtilis↗
PubMed1998

Surface plasmon resonance sensor for heparin measurements in blood plasma.

Surface plasmon resonance (SPR) was used as an affinity biosensor to determine absolute heparin concentrations in human blood plasma samples. Protamine and polyethylene imine (PEI) were evaluated as heparin affinity surfaces. Heparin adsorption onto protamine in blood plasma was specific with a lowest detection limit of 0.2 U/ml and a linear window of 0.2-2 U/ml. Although heparin adsorption onto PEI in buffer solution had indicated superior sensitivity to that on protamine, in blood plasma it was not specific for heparin and adsorbed plasma species to a steady-state equilibrium. By reducing the incubation time and diluting the plasma samples with buffer to 50%, the non-specific adsorption of plasma could be controlled and a PEI pre-treated with blood plasma could be used successfully for heparin determination. Heparin adsorption in 50% plasma was linear between 0.05 and 1 U/ml so that heparin plasma levels of 0.1-2 U/ml could be determined within a relative error of 11% and an accuracy of 0.05 U/ml.

Biosensing Techniques↗
PubMed1998

A lactulose sensor based on coupled enzyme reactions with a ring electrode fabricated from tetrathiafulvalen-tetracyanoquinodimetane.

A dual enzyme electrode is explored for measuring lactulose in milk. A ring electrode (diameter = 3 mm; ring width = 10-20 microns) is proposed onto which tetrathiafulvalen-tetracyanoquinodimetane (TTF-TCNQ) salt was physically packed. The electrode is a band electrode with dimensions approaching those for micro electrodes, so that the improved faradaic current/charging current ratio lead to improved detection limits. Fructose dehydrogenase (FDH) and beta-galactosidase (beta-gal) were immobilized by covering the electrode surface with a dialysis membrane. Lactulose was hydrolyzed to D-fructose and D-galactose by beta-gal. The hydrolyzed D-fructose was oxidized by FDH which was simultaneously reduced to the reduced form (FDH-PQQH2). The FDH-PQQH2 was directly reoxidized by TTF-TCNQ on the ring electrode, whose current was monitored at 200 mV vs Ag/AgCl. The detection limit of the lactulose sensor was 1.0 microM and the selectivity for lactulose was at least 1000 times higher than that for lactose. Pasteurized, UHT and sterilized milks were applied to the lactulose sensor, showing good accuracy and precision and, furthermore, good correlation to a reference photometric method, even though no rigorous procedure for the electrode fabrication has presently been addressed.

Animals↗
PubMed1998

Platinum-catalyzed enzyme electrodes immobilized on gold using self-assembled layers.

The bonding of enzymes to self-assembled monolayers (SAMs) of alkanethiols onto gold electrode surfaces is exploited to produce an enzyme biosensor. The attachment of glucose oxidase to a SAM of 3-mercaptopropionic acid was achieved using carbodiimide coupling. The resultant biosensor showed good sensitivity to glucose and a large dynamic range when measured amperometrically via the p-benzoquinone mediator. On the other hand, subsequent platinization of the enzyme-SAM electrode allowed hydrogen peroxide produced in the enzyme reaction to be detected directly, thus obviating the need for an artificial redox mediator. The performance of such sensors constructed on bulk gold electrodes was evaluated and finally compared to that of some preliminary thin-film gold electrodes. Biosensors constructed using the two alternative electrode surfaces have quite different sensitivities, thus reflecting the influence of the anchoring surface on the performance of the biosensor.

3-Mercaptopropionic Acid↗
PubMed1997

Clinical characteristics and management of acute stroke in patients with atrial fibrillation admitted to US university hospitals.

UNLABELLED: The optimal evaluation and management of patients with atrial fibrillation who suffer an acute ischemic stroke remains controversial. METHODS: Medical records of 171 consecutive patients with atrial fibrillation and acute stroke at six U.S. university hospitals were reviewed. Data collected included the use of antithrombotic therapy, brain and cardiac imaging, bleeding complications, stroke risk factors, and contraindications to anticoagulation. RESULTS: Mean age was 75.4 years. Cardiovascular risk factors associated with increased stroke risk were present in 87%; 35% had at least one contraindication to anticoagulation. Half of the patients with stroke risk factors and no contraindications to anticoagulation were not receiving any antithrombotic therapy at the time of admission. Of the 22 patients who were treated with warfarin, and had INR values on admission, 16 had levels of < 2.0; only six had INR values between 2.0 and 3.0. Transthoracic echocardiography was performed in 107 patients (63%); intracardiac thrombi were visualized in only 5%. Initial brain imaging revealed hemorrhagic transformation in nine. Heparin was used in 93 patients (54%), usually within 48 hours of stroke onset. Patients who received delayed heparin typically did not have repeat brain imaging prior to starting heparin. One patient had a delayed symptomatic cerebral hemorrhage. Of the survivors, 47% were discharged and treated with warfarin (or warfarin plus aspirin), 28% with ASA, 7% with other antithrombotic therapies, and 18% with no antithrombotic therapy. CONCLUSION: Antithrombotic therapy was underutilized and inadequately monitored in atrial fibrillation patients prior to stroke onset. After hospital admission, a wide range of diagnostic and management strategies, which often did not follow current recommendations, were employed.

Acute Disease↗
PubMed1996

Status of antithrombotic therapy for patients with atrial fibrillation in university hospitals.

BACKGROUND: The risk of stroke in patients with atrial fibrillation can be significantly reduced with antithrombotic therapy. Despite this, many physicians remain hesitant to prescribe warfarin sodium or aspirin therapy for patients with atrial fibrillation. OBJECTIVE: To assess the use of antithrombotic therapy in patients with atrial fibrillation at 6 academic hospitals in the United States. METHODS: Records were reviewed from consecutive hospital admissions of 309 patients with atrial fibrillation at 6 members of the University Health System Consortium, Oak Brook, III, which is a member driven alliance of 70 academic health centers in the United States. Risk factors for stroke, contraindications to anticoagulant therapy, and use of antithrombotic therapy at admission and discharge were recorded. RESULTS: The mean age of patients was 71.6 years, 54% had chronic, 22% paroxysmal, and 24% new-onset atrial fibrillation. Eighty-two percent of the patients had cardiovascular risk factors that have been associated with increased risk of stroke. At least 1 relative contraindication to anticoagulant therapy was present in 44%. At the time of admission. 32% of the patients with previously diagnosed atrial fibrillation (n = 235) were receiving warfarin (or warfarin plus aspirin), 31% were receiving aspirin alone, and 36% were receiving no antithrombotic therapy. At discharge (n = 230), 41% of these patients were taking warfarin (or warfarin plus aspirin) and 36% were taking aspirin. Forty-four percent of the patients with risk factors for stroke and no contraindications to anticoagulation (n = 134) were discharged on a regimen of warfarin (or warfarin plus aspirin), 34% were discharged on a regimen of aspirin, and 22% received no antithrombotic therapy. CONCLUSIONS: About half of the patients with atrial fibrillation admitted to these academic hospitals had clinical risk factors that are associated with increased risk of stroke and no contraindications to anticoagulation. Antithrombotic therapy was underused in these patients.

Academic Medical Centers↗
PubMed1996

Postoperative pulmonary changes after laparoscopic cholecystectomy.

Laparoscopic cholecystectomy (LC) has been widely used in recent years because of short postoperative hospital stays and low morbidity. In this study, 24 patients were prospectively evaluated with preoperative and postoperative spirometry, arterial blood gas determinations, and chest radiographs to quantitate the magnitude of postoperative pulmonary changes after LC. Statistically significant reductions were noted in forced vital capacity (FVC) (mean decrease, 810 mL) and forced expired volume in 1 second (FEV1) (mean decrease, 420 mL). Clinically important changes in arterial blood gas values did not occur. Of 20 postoperative chest films, 7 showed the development of atelectasis or effusion and 9 showed persistence of subdiaphragmatic free air 24 hours after LC. In summary, LC caused mean decreases of 23% in FVC and 16% in FEV1 24 hours after surgery. The physiologic derangements that follow LC are sufficiently small that all but the most severely impaired patients with pulmonary disease should be able to tolerate this operation.

Blood Gas Analysis↗
PubMed1991

Flow injection analysis with immobilized reagents.

Immobilized reagent phase flow injection analysis can be configured as discrete reagent cells upstream of the sensor element or as an integral reagent/transduction system (flow injection analysis-biosensor). The former approach has attracted greater attention because several assays can be assembled with greater versatility in reagent column units employing a single sensor, than can be co-immobilized on the surface of a transducer.

Animals↗
PubMed1988

The nature of biosensor technology.

The biosensor exploits the unique specificity of biological recognition events by coupling an enzyme, antibody or other biorecognition species to a transducing device. Interaction of the biocomponent with substrate or antigen is thus converted into a suitable quantitative output. The development of these biosensors is a multidisciplinary effort, exploiting many of the emerging semiconductor and optics technologies and integrating many traditional assay techniques. The impact of these biosensors is likely to be wide-ranging, with applications in medical and industrial environments as well as a variety of other fields.

Biological Assay↗