Search PubMed⌕ Search

Biomedical subjects

M K Taylor

Publications and source records attributed to M K Taylor.

At least 19 recordsLinked to original sources
PubMed2001

A cGMP-dependent protein kinase is implicated in wild-type motility in C. elegans.

In mammals, cyclic GMP and cGMP-dependent protein kinases (cGKs) have been implicated in the regulation of many neuronal functions including long-term potentiation and long-term depression of synaptic efficacy. To develop Caenorhabditis elegans as a model system for studying the neuronal function of the cGKs, we cloned and characterized the cgk-1 gene. A combination of approaches showed that cgk-1 produces three transcripts, which differ in their first exon but are similar in length. Northern analysis of C. elegans RNA, performed with a probe designed to hybridize to all three transcripts, confirmed that a major 3.0 kb cgk-1 transcript is present at all stages of development. To determine if the CGK-1C protein was a cGMP-dependent protein kinase, CGK-1C was expressed in SF:9 cells and purified. CGK-1C shows a K(a) of 190 +/- 14 nM for cGMP and 18.4 +/- 2 microM for cAMP. Furthermore, CGK-1C undergoes autophosphorylation in a cGMP-dependent manner and is inhibited by the commonly used cGK inhibitor, KT5823. To determine which cells expressed CGK-1C, a 2.4-kb DNA fragment from the promoter of CGK-1C was used to drive GFP expression. The CGK-1C reporter construct is strongly expressed in the ventral nerve cord and in several other neurons as well as the marginal cells of the pharynx and intestine. Finally, RNA-mediated interference of CGK-1 resulted in movement defects in nematode larvae. These results provide the first demonstration that cGMP-dependent protein kinase is present in neurons of C. elegans and show that this kinase is required for normal motility.

Animals↗
PubMed2001

Patients' rights on the World Wide Web.

Managed care reform, commonly referred to as "patients' rights" legislation, has become a hot topic. Many groups, including consumers, health care professionals, employers, managed care organizations, political parties, and government agencies, have strong opinions about measures that should be taken and what the outcomes of these measures might be. Those investigating this multidisciplinary topic will want to examine health care administration, ethics, health services research, and political science sources. Web resources covered in this article include: clearinghouses; government agencies; federal legislative and legal sites; and home pages of professional and trade associations, policy research institutes, and consumer advocacy organizations.

Health Care Sector↗
PubMed2000

The amino-terminal cyclic nucleotide binding site of the type II cGMP-dependent protein kinase is essential for full cyclic nucleotide-dependent activation.

For the type I cGMP-dependent protein kinases (cGKIalpha and cGKIbeta), a high affinity interaction exists between the C2 amino group of cGMP and the hydroxyl side chain of a threonine conserved in most cGMP binding sites. To examine the effect of this interaction on ligand binding and kinase activation in the type II isozyme of cGMP-dependent protein kinase (cGKII), alanine was substituted for the conserved threonine or serine. cGKII was found to require the C2 amino group of cGMP and its cognate serine or threonine hydroxyl for efficient cGMP activation. Of the two binding sites, disruption of cGMP-specific binding in the NH(2)-terminal binding site had the greatest effect on cGMP-dependent kinase activation, like cGKI. However, ligand dissociation studies showed that the location of the rapid and slow dissociation sites of cGKII was reversed relative to cGKI. Another set of mutations that prevented cyclic nucleotide binding demonstrated the necessity of the NH(2)-terminal, rapid dissociation binding site for cyclic nucleotide-dependent activation of cGKII. These findings suggest distinct mechanisms of activation for cGKII and cGKI isoforms. Because cGKII mediates the effects of heat-stable enterotoxins via the cystic fibrosis transmembrane regulator Cl(-) channel, these findings define a structural target for drug design.

Amino Acid Sequence↗
PubMed2000

Composite method to quantify powder flow as a screening method in early tablet or capsule formulation development.

The flow properties of typical tablet and capsule formulation excipients, active compounds, and representative formulation blends were tested with current and novel flow measurement techniques to identify a reliable bench test to quantify powder flow as a screening method in early tablet and capsule formulation development. Test methods employed were vibrating spatula, critical orifice, angle of repose, compressibility index, and avalanching analysis. Powder flow results from each method were compiled in a database, sorted, and compared. An empirical composite index was established and powder flow was ranked in accordance with formulator experience. Principal components analyses of the angle of repose, percent compressibility, and critical orifice of the powder materials were also performed. The first principal component accounted for 72.8% of data variability; scores associated with this principal component score can serve as an index of flowability. Data generated from vibrating spatula and avalanching methods were not reproducible and were inconsistent with formulator experience and cited vendor references for flow. Improvements of test instruments and further studies are necessary for better assessment of these approaches.

Capsules↗
PubMed2000

Supercritical fluid extraction of 2,4,6-trichloroanisole from cork stoppers.

2,4,6-Trichloroanisole (TCA) is the compound most often associated with cork taint in wines and has been shown to have a very low sensory threshold ( approximately 5 ng/L in wine). A supercritical fluid extraction (SFE) method for TCA in bark cork stoppers was developed with quantification via gas chromatography-mass spectrometry with selected ion monitoring. Supercritical carbon dioxide functioned as the extracting solvent, and temperature and pressure were optimized for the extraction. The method was validated using the stable isotope (2)H(5)-TCA as the internal standard. Recovery of TCA from spiked corks was found to be within 1-4% of the theoretical concentration with a coefficient of variation ranging from 2.6 to 9.7%. TCA levels in corks pulled from wines described as tainted by experienced judges ranged from 0.13 to 2.11 microg/g of cork. The SFE procedure offers a rapid, quantitative, nearly solvent-free, and automated method for the extraction of TCA from complex solid matrices such as cork.

Anisoles↗
PubMed1999

Genetic structure of the world's polar bear populations.

We studied genetic structure in polar bear (Ursus maritimus) populations by typing a sample of 473 individuals spanning the species distribution at 16 highly variable microsatellite loci. No genetic discontinuities were found that would be consistent with evolutionarily significant periods of isolation between groups. Direct comparison of movement data and genetic data from the Canadian Arctic revealed a highly significant correlation. Genetic data generally supported existing population (management unit) designations, although there were two cases where genetic data failed to differentiate between pairs of populations previously resolved by movement data. A sharp contrast was found between the minimal genetic structure observed among populations surrounding the polar basin and the presence of several marked genetic discontinuities in the Canadian Arctic. The discontinuities in the Canadian Arctic caused the appearance of four genetic clusters of polar bear populations. These clusters vary in total estimated population size from 100 to over 10 000, and the smallest may merit a relatively conservative management strategy in consideration of its apparent isolation. We suggest that the observed pattern of genetic discontinuities has developed in response to differences in the seasonal distribution and pattern of sea ice habitat and the effects of these differences on the distribution and abundance of seals.

Animals↗
PubMed1998

Chlorinated hydrocarbon contaminants in polar bears from eastern Russia, North America, Greenland, and Svalbard: biomonitoring of Arctic pollution.

Adipose tissue samples from polar bears (Ursus maritimus) were obtained by necropsy or biopsy between the spring of 1989 to the spring of 1993 from Wrangel Island in Russia, most of the range of the bear in North America, eastern Greenland, and Svalbard. Samples were divided into 16 regions corresponding as much as possible to known stocks or management zones. Concentrations of dieldrin (DIEL), 4,4'-DDE (DDE), sum of 16 polychlorinated biphenyl congeners (sigma PCB), and sum of 11 chlordane-related compounds and metabolites (sigma CHL) were determined. In order to minimize the effect of age, only data for adults (320 bears age 5 years and older) was used to compare concentrations among regions. Concentrations of sigma PCB were 46% higher in adult males than females, and there was no significant trend with age. Concentrations of sigma CHL were 30% lower in adult males than females. Concentrations of sigma PCB, sigma CHL, and DDE in individual adult female bears were standardized to adult males using factors derived from the least-square means of each sex category, and geometric means of the standardized concentrations on a lipid weight basis were compared among regions. Median geometric mean standardized concentrations (lipid weight basis) and ranges among regions were as follows: sigma PCB, 5,942 (2,763-24,316) micrograms/kg; sigma CHL, 1,952 (727-4,632) micrograms/kg; DDE, 219 (52-560) micrograms/kg; DIEL, 157 (31-335) micrograms/kg. Geometric mean sigma PCB concentrations in bears from Svalbard, East Greenland, and the Arctic Ocean near Prince Patrick Island in Canada were similar (20,256-24,316 micrograms/kg) and significantly higher than most other areas. Atmospheric, oceanic, and ice transport, as well as ecological factors may contribute to these high concentrations of sigma PCB. sigma CHL was more uniformly distributed among regions than the other CHCs. Highest sigma CHL concentrations were found in southeastern Hudson Bay, which also had the highest DDE and DIEL concentrations. In general, concentrations of sigma CHL, DDE, and DIEL were higher in eastern than western regions, suggesting an influence of North American sources. Average sigma PCB concentrations in bears from the Canadian Arctic were similar to those in 1982-84, while average sigma CHL and DDE concentrations were 35-44% lower and DIEL was 90% lower. However, the significance of these temporal trends during the 1980s is not conclusive because of the problems of comparability of data.

Adipose Tissue↗
PubMed1998

Interferon-gamma-secreting T-cell populations in rejecting murine cardiac allografts: assessment by flow cytometry.

Interplay between T-helper-1 (Th1) and T-helper-2 (Th2) cells is considered important in the development of acute allograft rejection and many other immune-mediated disease processes. Existing methods for evaluating expression of Th1 and Th2 cytokines, including reverse transcriptase polymerase chain reaction (RT-PCR), RNase protection assay (RPA), immunohistochemistry, and enzyme-linked immunosorbent assay (ELISA) all have limitations; alternate techniques to quantify cell populations expressing specific cytokine proteins, generate statistically analyzable data, and allow simultaneous identification of cytokine-secreting cell type are needed. To this end, we adapted a flow cytometric technique for intracellular cytokine immunofluorescence staining for use with cells isolated from solid tissue. To demonstrate the utility of the method, we determined the number of CD4+ and CD8+ cells secreting the prototypical Th1 and Th2 cytokines, interferon (IFN)-gamma, and interleukin (IL)-4 in acutely rejecting murine cardiac allografts. We also measured the cytokine production via ELISA, RPA, and semiquantitative competitive RT-PCR. The number of CD4+ cells producing IFN-gamma increased as rejection proceeded, in agreement with previous data; we detected no IL-4 production at any time, although relatively low numbers of IL-10-producing cells were identified. In addition, a high percentage of CD8+ cells, which outnumber CD4+ cells at day 6 after transplant, also produce IFN-gamma, suggesting that cytotoxic lymphocytes contribute significantly to the local cytokine milieu. This new application of intracellular cytokine staining provides a powerful methodology for studying transplantation immunology. The method may also be easily adapted to the study of other immune-mediated processes.

Animals↗
PubMed1998

Comparison of the performance of two sample thieves for the determination of the content uniformity of a powder blend.

The objective of this study was to compare the performance of two sample thieves (plug and grain) to determine the content uniformity of a powder blend. The powder blend was prepared by mixing 2% drug substance with the remaining excipients in a tumble blender for 30 min. Samples were taken at 10 locations in the blender using both thieves. The performance of each sample thief was assessed based on the respective content uniformity values and relative standard deviations obtained for each device, as well as the content uniformity values reported following analysis of the resulting compressed tablets. The relative standard deviation values for blend samples taken with the plug thief were approximately half of those obtained using the grain thief. The superior performance of the plug thief in this study is attributed to the static charge acquired by the microcrystalline cellulose, which leads to poor flow characteristics. This impeded the flow of the blend into the sample chamber of the grain thief resulting in segregation and variable content uniformity results. The plug thief, which does not require powder flow to obtain a sample, performs better for this formulation. The selection of a sampling thief should be assessed on a case-by-case basis. Superior performance is expected for the plug thief when poor flowing, compressible blends are sampled.

Pharmaceutical Preparations↗
PubMed1998

Coronary arteriosclerosis after T-cell-mediated injury in transplanted mouse hearts: role of interferon-gamma.

This study evaluated the contribution of acute parenchymal rejection and interferon (IFN)-gamma to the development of graft arterial disease (GAD) in totally allogeneic murine cardiac transplants. BALB/c (H-2d) hearts were transplanted into wild-type C57BL/6 (B6, H-2b) or B6 IFN-gamma-deficient (GKO) recipient mice. Assessing the role of acute parenchymal rejection in the GAD process involved two different immunosuppression protocols using anti-CD4 and -CD8 monoclonal antibodies (MAbs): virtually complete long-term immunosuppression (denoted as complete immunosuppression) was achieved by administering both MAbs 6, 3, and 1 day before transplantation and weekly thereafter; in contradistinction, a single, early, transient episode of rejection (transient rejection) was attained by administering MAbs beginning 4 days after transplant and then at weekly intervals. The extent and duration of T cell depletion under these two regimens were evaluated using flow cytometric analysis of peripheral blood lymphocytes. After a single injection of MAbs, peripheral blood CD4+ and CD8+ T cell depletion was approximately 98% at 1 week and approximately 88% at 2 weeks. After three injections (analogous to days 6, 3, and 1 before transplant), peripheral blood CD4+ and CD8+ T cell depletion was >98% at 2 weeks and approximately 87% at 4 weeks. Functioning cardiac allografts were removed at 8 and 12 weeks after transplant and analyzed by hematoxylin and eosin, elastic tissue, and immunohistochemical stains, and the severity of parenchymal rejection versus GAD was scored. With complete immunosuppression (antibody before and after transplant), BALB/c allografts showed little parenchymal rejection or GAD, suggesting that persistent depletion of T cells blocked subsequent development of GAD. However, even a single transient acute rejection episode allowed the subsequent development of GAD accompanied by augmented major histocompatibility complex (MHC) class II, VCAM-1, and ICAM-1 expression at 12 weeks; these allografts showed no residual CD4+ or CD8+ T cells. In comparison, allografts undergoing transient rejection in GKO recipients did not develop GAD, despite persistent macrophage and natural killer cell (NK) infiltrates comparable to those seen in wild-type recipients. Moreover, the arterioles of hearts transplanted into GKO recipients showed no or minimal increases in MHC class II, ICAM-1, and VCAM-1 relative to baseline expression. In conclusion, a single episode of allogeneic injury mediated by T cells suffices to evoke subsequent graft arteriosclerosis, even in the absence of additional T-cell-mediated injury, and the process appears to depend on IFN-gamma.

Animals↗
PubMed1997

Interferon-gamma deficiency prevents coronary arteriosclerosis but not myocardial rejection in transplanted mouse hearts.

We have hypothesized that T cell cytokines participate in the pathogenesis of graft arterial disease (GAD). This study tested the consequences of IFN-gamma deficiency on arterial and parenchymal pathology in murine cardiac allografts. Hearts from C-H-2(bm12)KhEg (bm12, H-2(bm12)) were transplanted into C57/B6 (B6, H-2(b)), wild-type, or B6 IFN-gamma-deficient (GKO) recipients after immunosuppression by treatment with anti-CD4 and anti-CD8 mAbs. In wild-type recipients, myocardial rejection peaked at 4 wk, (grade 2. 1+/-0.3 out of 4, mean+/-SEM, n = 9), and by 8-12 wk evolved coronary arteriopathy. At 12 wk, the GAD score was 1.4+/-0.3, and the parenchymal rejection grade was 1.2+/-0.3 (n = 8). In GKO recipients of bm12 allografts, myocardial rejection persisted at 12 wk (grade 2.5+/-0.3, n = 6), but no GAD developed (score: 0.0+/-0.0, n = 6, P < 0.01 vs. wild-type). Mice treated with anti-IFN-gamma mAbs showed similar results. Isografts generally showed no arterial changes. In wild-type recipients, arterial and parenchymal cells showed increased MHC class II molecules, intercellular adhesion molecule-1, and vascular cell adhesion molecule-1 compared to normal or isografted hearts. The allografts in GKO recipients showed attenuated expression of these molecules (n = 6). Thus, development of GAD, but not parenchymal rejection, requires IFN-gamma. Reduced expression of MHC antigens and leukocyte adhesion molecules may contribute to the lack of coronary arteriopathy in hearts allografted into GKO mice.

Animals↗
PubMed1997

Facilitation and inhibition of the acoustic startle reflex in the rat after a momentary increase in background noise level.

Small increments in background noise were shown to increase the amplitude of a subsequently elicited acoustic startle reflex (ASR) in rats by as much as 100% under optimal conditions. Increment lead time (5-160 ms) and level (1.5-15 dB), initial noise level (30-70 dB), startle level (95-125 dB), number of test days (1-5), and drug condition (diazepam or saline ip) were varied in 6 experiments. Prepulse facilitation (PPF), measured by difference scores, was greatest for intermediate increments (3 dB) and lead times (20-40 ms) and was replaced by prepulse inhibition (PPI) for higher values, especially in the later test days. Diazepam reduced baseline ASR and diminished PPI, but it did not affect PPF. These data argue against hypotheses that attribute PPF of this sort to either temporal integration within the ASR pathways or to the elicitation of a nonspecific arousal reaction by the prepulse.

Acoustic Stimulation↗
PubMed1995

Effects of single and repeated exposure to apomorphine on the acoustic startle reflex and its inhibition by a visual prepulse.

The acoustic startle reflex (ASR) is inhibited by startle-irrelevant stimuli that briefly precede reflex elicitation. This effect, prepulse inhibition (PPI), is reduced in strength for animals that have received dopamine agonists, such as apomorphine (APO). Reduction in PPI is most evident for weak masked noise prepulses, thus suggesting that APO disrupts the reception of stimuli to the extent that they present a low signal-to-noise ratio. Here we examine the effect of APO on PPI produced by non-masked visual prepulses. Light flashes were given at two intensities, 40, 70, 110, or 220 ms before ASR elicitation. In phase 1 (5 weeks in duration) half of the animals received one weekly injection of APO (0.5 mg/kg, IP) and one of vehicle (VEH), while the other half received two injections of VEH. Within these groups, half were tested 30 min after the injections, the other half kept test naive (four groups total). In phase 2, following a 4-week rest, all groups were tested after a low dose of APO (0.1 mg/kg) and VEH, 1 week apart. APO eliminated PPI for a dim flash and reduced PPI for a brighter flash to a level normally obtained with the dim flash, while increasing both ASR control values and activity. The bright light was maximally effective at a lead time of 70 ms and APO did not alter this value. Because in general the time of maximal inhibition varies with prepulse intensity for visual stimuli, the finding that the time of the peak remained constant reveals that APO has its effect on inhibition rather than on effective stimulus intensity. In phase 2, APO reduced PPI with no sign of sensitization from past drug exposure. However, APO increased the ASR only in groups previously exposed to APO, indicating behavioral sensitization. The differential effects of repeated exposure on these response measures suggest that neural substrates for the several behavioral effects of APO function at least in part independently.

Animals↗
PubMed1995

Interaction of p72syk with the gamma and beta subunits of the high-affinity receptor for immunoglobulin E, Fc epsilon RI.

Activation of protein tyrosine kinases is one of the initial events following aggregation of the high-affinity receptor for immunoglobulin E (Fc epsilon RI) on RBL-2H3 cells, a model mast cell line. The protein tyrosine kinase p72syk (Syk), which contains two Src homology 2 (SH2) domains, is activated and associates with phosphorylated Fc epsilon RI subunits after receptor aggregation. In this report, we used Syk SH2 domains, expressed in tandem or individually, as fusion proteins to identify Syk-binding proteins in RBL-2H3 lysates. We show that the tandem Syk SH2 domains selectively associate with tyrosine-phosphorylated forms of the gamma and beta subunits of Fc epsilon RI. The isolated carboxy-proximal SH2 domain exhibited a significantly higher affinity for the Fc epsilon RI subunits than did the amino-proximal domain. When in tandem, the Syk SH2 domains showed enhanced binding to phosphorylated gamma and beta subunits. The conserved tyrosine-based activation motifs contained in the cytoplasmic domains of the gamma and beta subunits, characterized by two YXXL/I sequences in tandem, represent potential high-affinity binding sites for the dual SH2 domains of Syk. Peptide competition studies indicated that Syk exhibits a higher affinity for the phosphorylated tyrosine activation motif of the gamma subunit than for that of the beta subunit. In addition, we show that Syk is the major protein in RBL-2H3 cells that is affinity isolated with phosphorylated peptides corresponding to the phosphorylated gamma subunit motif. These data suggest that Syk associates with the gamma subunit of the high-affinity receptor for immunoglobulin E through an interaction between the tandem SH2 domains of SH2 domains of Syk and the phosphorylated tyrosine activation motif of the gamma subunit and that Syk may be the major signaling protein that binds to Fc epsilon RI tyrosine activation motif of the gamma subunit and that Syk may be the major signaling protein that binds to Dc epsilon tyrosine activation motifs in RBL-2H3 cells.

Amino Acid Sequence↗
PubMed1993

Importance of hypothalamic function to stressor-induced responsiveness of the GABAA receptor in the cerebral cortex: a non-corticosterone influence.

Catecholamine terminals in the paraventricular nucleus (PVN) of the hypothalamus of 60-day-old rats were destroyed by the stereotaxic injection of 6-hydroxydopamine into the PVN (6-OHDA; 9 micrograms/1.5 microliters bilaterally), and the rats were tested 2 weeks later. Lesions led to a 70% reduction of norepinephrine in the hypothalamus and a loss of dopamine-beta-hydroxylase immunoreactivity in the PVN. Furthermore, 6-OHDA lesions in the hypothalamus disrupted stressor-induced (15 min of restraint) changes in GABAA receptor function in the cerebral cortex (assessed by measuring chloride-facilitated benzodiazepine binding) but did not alter stressor-induced increases in plasma corticosterone levels. Additionally, the lesion did not change the responsiveness of the GABAA receptor to the corticosterone metabolite, allotetrahydrodeoxycorticosterone. These results indicate that stressor-induced changes in cortical GABAA receptor function are not driven by the stressor-induced release of corticosterone. A separate group of animals were tested for behavioral responses to challenge, and while 6-OHDA-induced lesions did not alter total scores in the test of environment-specific social interaction, the lesions did induce a change in composition of the behavior. Lesioned animals demonstrated increased physical (vigorous contact) interactions, similar to behavior previously observed in younger rats. The results of the behavioral study support a role for the GABAA receptor in the cerebral cortex in mediating appropriate behavioral responses to challenge in the adult rat. Thus, a hypothalamic lesion that prevented challenge-induced changes in GABAA receptor function in the cortex (with no change in the corticosterone response to the stressor) also led to altered behavioral responses to challenge.

Anesthetics↗
PubMed1993

Altered stressor-induced changes in GABAA receptor function in the cerebral cortex of adult rats exposed in utero to diazepam.

Prenatal administration of the anxiolytic drug diazepam (DZP), 2.5 mg/kg) to the pregnant rat over gestational days 14-20 altered function and stressor-induced responsiveness of the GABAA receptor in the cerebral cortex of exposed animals as adults. In Experiment 1, the impact of 15 min of restraint on chloride-facilitated benzodiazepine binding was evaluated in male and female rats at 70-90 days of age. Early exposure to DZP led to an enhanced potency of chloride on binding in both males and females. In Experiment 2, GABA stimulation of 36chloride uptake was measured in male rats at 35 or 70 days of age following 10 min of forced swimming at ambient temperature. In control animals, stressor-induced changes in receptor function were not evident until 70 days, and in DZP-exposed rats the stressor had no effect on receptor function at either age. These changes in GABAA receptor responsiveness induced by early exposure to DZP may underlie the disrupted behavioral responses to environmental challenge that have been previously reported.

Aging↗
PubMed1992

Cell-mediated cytotoxicity.

Progress is being made in determining how cytotoxic cells are activated, the way the lethal hit is delivered and the subsequent events in the target cell. Several factors cloud the issue, including the heterogeneity of cytotoxic cells, differences between fresh cells and cell lines, and the possibility of single cells using multiple cytotoxic mechanisms. The most difficult task will be to define which cytotoxic mechanisms are significant in vivo.

Animals↗