Biomedical subjects

D Keller

Publications and source records attributed to D Keller.

At least 19 recordsLinked to original sources

PubMed2002

World-Wide Web-based graphical user interfaces for laboratory data.

OBJECTIVES: Electronic medical record systems permit collection of large amounts of medical information. Usually, information is presented in a fixed format, either as text or tables. Health care providers have to navigate this fixed format in order to find information useful for a specific patient-provider interaction. The main objective of this work was to allow the provider immediate access to specific laboratory information through the development of a highly customizable, graphical user interface to the Mayo Clinic laboratory information system. METHODS RESULTS: Here we describe this platform-independent, World-Wide-Web-based graphical user interface that allows the provider to see all or a predetermined panel of essential laboratory data in graphical format. Advantages include availability at internet-based workstations, immediate recognition of trends over time, ability to zoom in and out of specific periods of time, and detailed analysis of patient values in relationship to normal values. CONCLUSIONS: Web browser-based user interface allowing graphical display of laboratory data using Java technology was described. The connection to the Mayo Clinic laboratory information system combines cross-platform support for use on virtually any networked machine, interaction through a Web browser for ease of use, and a combination of the Perl and Java languages for powerful data processing and interactivity.

Clinical Laboratory Information Systems↗

PubMed2001

Flexibility and bioactivity of insulin: an NMR investigation of the solution structure and folding of an unusually flexible human insulin mutant with increased biological activity.

The structure and folding of a novel human insulin mutant, [Thr(B27) --> Pro, Pro(B28) --> Thr]insulin (PT insulin), in aqueous solution and in mixtures of water and 2,2,2-trifluoroethanol (TFE) have been studied by NMR spectroscopy. It was found that PT insulin has a highly flexible structure in pure water and is present in at least two different conformations, although with an overall tertiary structure similar to that of native insulin. Furthermore, the native helical structures are poorly defined. Surprisingly, the mutant has a biological activity about 50% higher than native insulin. In contrast, in TFE/water solution the mutant reveals a propensity of forming a well-defined structure at the secondary structure level, similar to monomeric native insulin. Thus, as shown by a detailed determination of the structure from 208 distance restraints and 52 torsion angle restraints by distance geometry, simulated annealing, and restrained energy minimization, the native insulin helices (A2-A7, A13-A19, and B10-B19) as well as the beta-turn (B20-B23) are formed in 35% TFE. However, the amount of tertiary structure is decreased significantly in TFE/water solution. The obtained results suggest that only an overall tertiary fold, as observed for PT insulin in pure water, is necessary for expressing the biological activity of insulin, as long as the molecule is flexible and retains the propensity to form the secondary structure required for its receptor binding. In contrast, a compact secondary structure, as found for native insulin in solution, is unnecessary for the biological activity. A model for the receptor binding of insulin is suggested that relates the increased bioactivity to the enhanced flexibility of the mutant.

Amino Acid Sequence↗

PubMed2001

Is lithium response related to G(s)alpha levels in transformed lymphoblasts from subjects with bipolar disorder?

BACKGROUND: A series of studies from independent laboratories have found increased levels of G(s)alpha in bipolar disorder in postmortem brain and peripheral blood cells. Long-term lithium administration blunts G-protein coupled cAMP signaling and may regulate G(s)alpha levels. METHODS: We measured G(s)alpha in transformed lymphoblasts obtained from subjects with bipolar disorder and compared the findings with 23 age- and sex-matched controls. To reduce patient heterogeneity, we included only patients with an excellent response to lithium prophylaxis. RESULTS: We found no differences in G(s)alpha protein levels measured with immunoblotting. G(s)alpha levels did not correlate with age, age of onset or duration of lithium therapy. LIMITATIONS: This study made use of transformed lymphoblasts, which may not fully represent changes that occur in regionalized brain tissue. Furthermore, the transformed lymphoblasts used in this study were acquired from a select group of bipolar disorder subjects that responded to lithium treatment. Lastly, consideration has to be given to the small sample size of the study. CONCLUSIONS: These results are consistent with recent observations suggesting that mood state and treatment effects may account at least in part for increased G(s)alpha levels in bipolar disorder. CLINICAL RELEVANCE: This study suggests a need to further characterize biological phenotypes in subjects with mood disorders to enhance genetic studies.

Adult↗

PubMed2001

The physics of molecular motors.

Molecular motors convert chemical energy into mechanical force and movement. Operating at energies just above those of the thermal bath, these motors experience large fluctuations, and their physical description must be necessarily stochastic. Here, motor operation is described as a biased diffusion on a potential energy surface defined by the interactions of the motor with its track and its fuel. These ideas are illustrated with a model of the rotary movement of the F(o) motor.

Animals↗

PubMed2001

Ethanol potentiates the function of the human dopamine transporter expressed in Xenopus oocytes.

Ethanol alters a variety of properties of brain dopaminergic neurons including firing rate, synthesis, release, and metabolism. Recent studies suggest that ethanol's action on central dopamine systems may also involve modulation of dopamine transporter (DAT) activity. The human DAT was expressed in Xenopus oocytes to examine directly the effects of ethanol on transporter function. [3H]Dopamine (100 nM) accumulation into DAT-expressing oocytes increased significantly in response to ethanol (10 min; 10-100 mM). In two-electrode voltage-clamp experiments, DAT-mediated currents were also enhanced significantly by ethanol (10-100 mM). The magnitude of the ethanol-induced potentiation of DAT function depended on ethanol exposure time and substrate concentration. Cell surface DAT binding ([3H]WIN 35,428; 4 nM) also increased as a function of ethanol exposure time. Thus, the increase in dopamine uptake was associated with a parallel increase in the number of DAT molecules expressed at the cell surface. These experiments demonstrate that DAT-mediated substrate translocation and substrate-associated ionic conductances are sensitive to intoxicating concentrations of ethanol and suggest that DAT may represent an important site of action for ethanol's effects on central dopaminergic transmission. A potential mechanism by which ethanol acts to enhance DAT function may involve regulation of DAT expression on the cell surface.

Animals↗

PubMed2001

Neonatal mouse model: review of methods and results.

The neonatal mouse model, in various forms, has been used experimentally since 1959 and a large number of chemicals have been tested. The neonatal model is known to be very sensitive for the detection of carcinogens that operate via a genotoxic mode of action. In contrast, it is known not to respond to chemicals that act via epigenetic mechanisms, commonly observed in the two-year carcinogenicity studies. As such, the model has a high sensitivity and specificity in its response. Dose selection for the neonatal model is based on the maximum tolerated or feasible dose. Traditionally, compounds have been tested via the IP route of administration in this model. In some cases, this has limited the amount of material that can be administered because of the low dosing volumes (10 to 20 microL) that can be administered IP. For the ILSI project, the neonatal model was adapted for oral administration, which has the advantages of being the same route for which most pharmaceuticals are administered. In addition, a 10-fold increase in the volume of administration (100 to 200 microL) and the ability to dose drugs in suspension, permits much higher doses to be used as compared to the IP route of administration. The spontaneous tumors in the neonatal model occurred mainly in the liver of male mice and lung of male and female mice with a few tumors observed in the Harderian gland. The positive control, DEN produced a robust, uniform, and reproducible tumor response with the target organs essentially limited to liver and lung. A total of 13 compounds out of the 21 ILSI ACT compounds were evaluated in the neonatal model involving 18 studies with duplicate studies for some compounds. The genotoxic carcinogens including those used as positive controls were clearly positive (cyclophosphamide, diethylnitrosamine, 6-nitrochrysene). The non-genotoxic rodent carcinogens were clearly negative (chlorpromazine, sulfisoxazole, sulfamethoxazole, clofibrate, DEHP, haloperidol, metaproteranol, and phenobarbital). The non-genotoxic human carcinogen (cyclosporin) was clearly negative. The two other human carcinogens phenacetin and DES were negative and interestingly estradiol was negative in one of the two oral studies, but was clearly positive in the other. Considering the mode of action for three of the human carcinogens (DES, cyclosporin and phenacetin), which were negative in this model, the mode of action in humans is likely to be epigenetic. Overall, for the 3 clearly genotoxic chemicals, all were positive. For the 9 clearly non-genotoxic chemicals, all 9 were negative. The two human carcinogens for which genotoxicity may or may not play a role (DES and phenacetin) were negative and estradiol was positive in I of the two oral studies. Overall, the extensive database for compounds tested in the neonatal mouse model would support its use as an alternative model for the assessment of the carcinogenic potential of a chemical. The model responds to chemicals that act via a genotoxic mode of action that represent a greater concern for human cancer risk.

Administration, Oral↗

PubMed2001

Physician Order Entry impact on drug turn-around times.

This paper describes a study of the impact of Physician Order Entry (POE) on pharmacy order turn-around times. The study looked at two surgical services, Neurosurgery and Transplant, of a large Midwestern academic medical center. Pharmacy orders were followed in these units from the time a physician wrote an order to the time the patient received the medication. The first part of the study tracked pharmacy orders for a two-month period before the implementation of POE and the second part of the study tracked pharmacy orders for a two-month period after POE had been implemented. The pre- and post-POE pharmacy turn-around times were compared. It was expected that the data would show a substantial decrease in pharmacy order turn-around times. Our study did, in fact, show a significant reduction in this turn-around-time.

Clinical Pharmacy Information Systems↗

PubMed2000

MDM2 inhibits p300-mediated p53 acetylation and activation by forming a ternary complex with the two proteins.

p300 acetylates and activates the tumor suppressor p53 after DNA damage. Here, we show that MDM2, a negative-feedback regulator of p53, inhibited p300-mediated p53 acetylation by complexing with these two proteins. First, we purified a p300-MDM2-p53 protein complex from HeLa nuclear extracts, which was inactive in p53 acetylation, but active in histone acetylation. Also, wild-type, but not N-terminally deleted, MDM2 inhibited p53 acetylation by p300 in vitro and in vivo. This inhibition was specific for p53, because MDM2 did not affect acetylation of histones or the C terminus of p73 by p300. Consequently, wild-type, but not the mutant, MDM2 repressed the p300-stimulated sequence-specific DNA-binding and transcriptional activities of p53. These results demonstrate that an additional mechanism of p53 inactivation by MDM2 is to inhibit p53 acetylation by p300.

Acetylation↗

PubMed2000

UV but not gamma irradiation accelerates p53-induced apoptosis of teratocarcinoma cells by repressing MDM2 transcription.

Induction of p53 by DNA damage results in apoptosis of teratocarcinoma cells, whereas MDM2, encoded by a p53-responsive gene, can reverse this phenotype by inhibiting p53 function. Here we report that UV (10 or 20 J/m2), but not gamma irradiation (7 or 10 Gy), caused a massive apoptosis of human teratoma Tera-2 or murine testicular carcinoma F9 cells, both of which contain wild-type p53, but not murine p53 null testicular carcinoma EB-16 cells. Most Tera-2 or F9 cells died overnight after UV but not gamma irradiation. Correlated with this phenotype was a dramatic and continuing accumulation of p53 proteins after UV but not gamma irradiation. This was attributable to UV-responsive repression of MDM2 expression, because both its protein and RNA were not detectable after UV irradiation. This UV-induced repression appeared to be specific to MDM2, because expression of other genes, such as p21, p53, or glyceraldehyde-3-phosphate dehydrogenase, was not reduced. Also, RNase protection analysis showed that a DNA region, excluding the p53 binding site, in the MDM2 promoter mediated transcriptional repression in response to UV. Thus, these results suggest that UV but not gamma irradiation can induce p53 by suppressing MDM2 expression in a p53-independent fashion and subsequently, massive cell death.

Animals↗

PubMed2000

Single-molecule studies of the effect of template tension on T7 DNA polymerase activity.

T7 DNA polymerase catalyses DNA replication in vitro at rates of more than 100 bases per second and has a 3'-->5' exonuclease (nucleotide removing) activity at a separate active site. This enzyme possesses a 'right hand' shape which is common to most polymerases with fingers, palm and thumb domains. The rate-limiting step for replication is thought to involve a conformational change between an 'open fingers' state in which the active site samples nucleotides, and a 'closed' state in which nucleotide incorporation occurs. DNA polymerase must function as a molecular motor converting chemical energy into mechanical force as it moves over the template. Here we show, using a single-molecule assay based on the differential elasticity of single-stranded and double-stranded DNA, that mechanical force is generated during the rate-limiting step and that the motor can work against a maximum template tension of approximately 34 pN. Estimates of the mechanical and entropic work done by the enzyme show that T7 DNA polymerase organizes two template bases in the polymerization site during each catalytic cycle. We also find a force-induced 100-fold increase in exonucleolysis above 40 pN.

Bacteriophage T7↗

PubMed2000

Nerve growth factor treatment alters Ca2+ pump levels in PC12 cells.

Nerve growth factor (NGF) treatment converts rapidly dividing PC12 cells into a neuronal phenotype. To understand the Ca2+ sequestration mechanisms accompanying this differentiation, we examined the endoplasmic reticulum Ca2+ (SERCA) pump levels using two different assays: ATP-dependent azide insensitive oxalate stimulated 45Ca2+ uptake by PC12 cells permeabilized with saponin, and Western blots using a monoclonal antibody which reacts with all the SERCA isoforms. We also examined the reaction to an antibody against the plasma membrane Ca2+ (PMCA) pump. NGF treatment decreased the SERCA pump expression but it increased the PMCA pump level. These results are consistent with a greater role of PMCA pumps in neuronal cells than in most other cells and with an increased role of SERCA pumps during cell proliferation.

Animals↗

PubMed2000

The mechanochemistry of molecular motors.

A theory of molecular motors is presented that explains how the energy released in single chemical reactions can generate mechanical motion and force. In the simplest case the fluctuating movements of a motor enzyme are well described by a diffusion process on a two-dimensional potential energy surface, where one dimension is a chemical reaction coordinate and the other is the spatial displacement of the motor. The coupling between chemistry and motion results from the shape of the surface, and motor velocities and forces result from diffusion currents on this surface. This microscopic description is shown to possess an equivalent kinetic mechanism in which the rate constants depend on externally applied forces. By using this equivalence we explore the characteristic properties of several broad classes of motor mechanisms and give general expressions for motor velocity versus load force for any member of each class. We show that in some cases simple plots of 1/velocity vs. 1/concentration can distinguish between classes of motor mechanisms and may be used to determine the step at which movement occurs.

Biomechanical Phenomena↗

PubMed2000

Teaching prevention in pediatrics.

Pediatrics has attempted to inculcate the "culture of prevention" into practice, both through anticipatory guidance in well-child care and through behavioral interventions in sick care. The effectivenesses of many components of well-child care have not been conclusively demonstrated, particularly in health education, counseling, and anticipatory guidance, nor has teaching prevention in pediatrics been thoroughly evaluated. This article reviews methods of teaching prevention in pediatrics and highlights innovative programs. Teaching programs use the wide range of approaches now common in medical education, in a variety of inpatient and outpatient sites. Programs across the country are trying new approaches to teaching traditional topics or are introducing new topics into their curricula. Examples of specific programs are given, organized by the themes of the programs. The field needs to develop in three major directions. First, there is a need to develop competencies and curricula in prevention issues of contemporary importance, including the new morbidities, cross-cultural issues, cost-effectiveness, quality of care, and practice in managed care and other community settings. Second, further work is needed to evaluate programs and measure educational outcomes. This feedback must in turn be used to redefine competencies, curricula, and programs, Third, there needs to be an accessible clearinghouse, and educational tools need to be disseminated. To be effective, a curriculum for prevention in pediatrics cannot stand alone, but must be part of a vertically and horizontally integrated curriculum. Further, creating horizontally and vertically integrated curricula in prevention teaching across disciplines should be the standard.

Child↗

PubMed1999

The p38MAPK inhibitor SB203580 alleviates ultraviolet-induced phosphorylation at serine 389 but not serine 15 and activation of p53.

Phosphorylation of p53 at serine 389 has been shown to be responsive uniquely to UV but not gamma irradiation. This report describes identification of the UV-responsive p38MAPK protein as a serine 389 kinase. The immunoprecipitated p38MAPK from UV-irradiated murine embryonic testicular carcinoma F9 cells phosphorylated the serine 392 residue but not serine 15 of the human p53 protein in vitro and this phosphorylation was inhibited by a p38MAPK-specific chemical inhibitor SB203580. The inhibitor also remarkably alleviated the UV-caused induction and serine 389 but not serine 15 phosphorylation of the murine p53 protein in vivo. Subsequently, this compound suppressed transcriptional activity of p53 and partially retarded UV-induced apoptosis. Moreover, p53 bound to p38 as revealed by immunoprecipitation with anti-p53 antibodies from UV-treated F9 cells. Thus, these results suggest that UV-stimulated p53 phosphorylation at serine 389 is mediated by the stress-responsive p38MAPK.

Animals↗

PubMed1999

Psychometric assessment of patients with chronic upper extremity pain attributed to workplace exposure.

Upper extremity pain attributed to workplace exposure is a growing concern in medicine today. This study was undertaken to investigate the psychological findings in patients with chronic upper extremity pain attributed to workplace exposure. Sixty-three consecutive patients were evaluated. All patients underwent a history and physical examination by a hand surgeon and a psychological evaluation by a licensed psychologist, including a Minnesota Multiphasic Personality Inventory-2 (MMPI-2). Physical and psychological findings were then compared. Forty-one patients (65%) showed abnormalities on MMPI-2 examination (t score > 70 = 97% confidence). This result is far above what generally would be expected in medical patients. When physical findings were grouped with psychological findings, 5 distinct groups were identified: 1 patient had normal physical and psychological results, 6 patients had normal physical findings but abnormal psychological assessments, 21 patients had abnormal physical findings but normal psychological assessments, 28 patients had both abnormal physical and psychological findings, and 7 patients had abnormal physical results but invalid MMPI-2 evaluations due to purposeful attempts to deny psychological symptoms. Identification of these groups may have diagnostic, prognostic, and treatment value. In addition, identification of these groups may help clarify several aspects of the debate over the role of psychosocial factors in the development and maintenance of chronic upper extremity pain that patients attribute to workplace exposure.

Arm↗

PubMed1999

MDM2 suppresses p73 function without promoting p73 degradation.

The newly identified p53 homolog p73 can mimic the transcriptional activation function of p53. We investigated whether p73, like p53, participates in an autoregulatory feedback loop with MDM2. p73 bound to MDM2 both in vivo and in vitro. Wild-type but not mutant MDM2, expressed in human p53 null osteosarcoma Saos-2 cells, inhibited p73- and p53-dependent transcription driven by the MDM2 promoter-derived p53RE motif as measured in transient-transfection and chloramphenicol acetyltransferase assays and also inhibited p73-induced apoptosis in p53-null human lung adenocarcinoma H1299 cells. MDM2 did not promote the degradation of p73 but instead disrupted the interaction of p73, but not of p53, with p300/CBP by competing with p73 for binding to the p300/CBP N terminus. Both p73alpha and p73beta stimulated the expression of the endogenous MDM2 protein. Hence, MDM2 is transcriptionally activated by p73 and, in turn, negatively regulates the function of this activator through a mechanism distinct from that used for p53 inactivation.

Apoptosis↗

PubMed1999

Where to treat? Is the habitual bite the best functional bite relationship?

One important consideration for any dental treatment involves evaluating the relationship of the mandible to the maxilla. Is the habitual bite that the patient possesses the best functional bite? There are a multiplicity of factors that must be considered in this evaluation before the doctor determines whether to treat to the patient's habitual bite position, or should a better bite position be determined.

Biomechanical Phenomena↗

PubMed1998

Characterization of a novel structural member, LukE-LukD, of the bi-component staphylococcal leucotoxins family.

A new member of the staphylococcal bi-component leucotoxins family, LukE (32 kDa) and LukD (34.3 kDa) has been characterized from Staphylococcus aureus strain Newman. LukE was 58-68% identical with the class S proteins, whereas LukD was 71-77% identical with the class F proteins of the family. A partial immunoreactivity with the various affinity-purified antibodies specific for the other proteins was observed. Immunoprecipitation assay and gene probing confirmed a 30% frequency among human clinical isolates, differing from the distribution of the other known leucotoxins (P<0.005). LukE+LukD was as effective as the Panton-Valentine leucocidin for inducing dermonecrosis when injected in the rabbit skin, but not hemolytic and poorly leucotoxic compared to other leucotoxins expressed by Staphylococcus aureus.

Amino Acid Sequence↗