Biomedical subjects

L L Walker

Publications and source records attributed to L L Walker.

At least 19 recordsLinked to original sources

PubMed2000

Dependence of mast cell IgE-mediated cytokine production on nuclear factor-kappaB activity.

BACKGROUND: The transcription factor nuclear factor-kappaB (NF-kappaB) has been implicated in the regulation of a number of inflammatory cytokines and has been the proposed target for anti-inflammatory therapeutics. OBJECTIVE: Our purpose was to explore the role of NF-kappaB in the regulation of allergic inflammation. METHODS: To determine whether NF-kappaB is activated during IgE-mediated reactions and what types of mediators it regulates, a mutant form of IkappaB was used to block the ability of NF-kappaB to translocate to the nucleus and promote the transcription of selected genes. RESULTS: Mouse bone marrow-derived mast cells stimulated by IgE receptor cross-linking exhibited an activation of NF-kappaB as assessed by electrophoretic mobility shift assays. Transfected mast cells expressing the mutant IkappaB showed very little NF-kappaB activation. Both control and transfected cells released beta-hexosaminidase after specific antigen challenge, and this release could be potentiated by exogenous adenosine. Transfected mast cells that failed to develop NF-kappaB activation did not produce IL-6 messenger RNA or protein after IgE-mediated stimulation, but these cells retained the ability to produce transcripts for IL-4 and IL-5 in spite of the suppression of NF-kappaB activity. CONCLUSIONS: It appears that NF-kappaB is activated during IgE-mediated allergic inflammation and that this activity is necessary for the production of IL-6, but not IL-4 or IL-5. When considering the use of agents that target NF-kappaB to reduce inflammatory processes, it is important to know precisely which cytokines are under its control.

Animals↗

PubMed2000

Cognitive deficit induced by acute tryptophan depletion in patients with Alzheimer's disease.

OBJECTIVE: The study assessed the effects on global cognitive function and mood of a reduction of brain serotonin by means of acute tryptophan depletion in 16 patients with dementia of the Alzheimer type and in 16 cognitively intact comparison subjects. METHOD: In a double-blind, crossover design, subjects received a tryptophan-free amino acid drink to induce acute tryptophan depletion and, on a separate occasion, a placebo drink containing a balanced mixture of amino acids. On each occasion, ratings of depressed mood were made at baseline and 4 and 7 hours later, and the Modified Mini-Mental State was administered at baseline and 4 hours later. RESULTS: Patients with dementia of the Alzheimer type had a significantly lower mean score on the Modified Mini-Mental State after acute tryptophan depletion than after receiving placebo. The comparison group showed no difference in mean score on the Modified Mini-Mental State after acute tryptophan depletion and after receiving placebo. No significant changes in mood were found in either group. CONCLUSIONS: Acute tryptophan depletion significantly impaired cognitive function in patients with dementia of the Alzheimer type. Compromised serotonergic function, in combination with cholinergic deficit, may make an important contribution to cognitive decline in dementia of the Alzheimer type.

Aged↗

PubMed1999

The effects of angiotensin II on blood perfusion in the rat renal papilla.

1. Systemic infusion of angiotensin II (AII) increased papillary blood perfusion (PBP) measured by laser-Doppler flowmetry in rats, aged about 5 weeks. 2. The mechanisms involved in this response were determined by infusion of AII in the presence of systemic doses of losartan (a type 1 AII receptor antagonist), HOE-140 (a bradykinin B2 receptor antagonist), and an inhibitor of NO production - Nomega-nitro-L-arginine (NOLA). 3. Mean arterial blood pressure (MAP) and PBP increased in a dose-dependent manner in response to intravenous infusions of AII. Infusion of losartan abolished these responses to AII but HOE-140 was without effect. Infusion of NOLA abolished the increase in PBP but did not affect the pressor response to AII. Systemic infusion of sodium nitroprusside restored the response to AII in experiments with NOLA infusion. 4. The results indicate that the increase in PBP caused by AII is mediated via angiotensin AT1 receptors and does not involve bradykinin B2 receptors. The AII-induced increase in PBP is dependent upon the presence of NO, thus providing a mechanism for maintenance of papillary perfusion in the face of generalized renal vasoconstriction due to AII.

Angiotensin II↗

PubMed1999

Imaging of renal medullary interstitial cells in situ by confocal fluorescence microscopy.

Renal medullary interstitial cells are a prevalent and characteristic feature of the inner medulla of the kidney, but the physiological significance of this is unclear. We have developed a method for imaging renal medullary interstitial cells in situ by loading the cells with fluorescent dyes and monitoring their distribution using confocal microscopy. The pH-sensitive probe 2'7'-bis-(carboxyethyl)-5-(and-6)-carboxyfluorescein acetoxymethyl ester was used as a marker of cytoplasmic volume and therefore of cell morphology. Nile Red was used to demonstrate the presence of renal medullary interstitial cell lipid droplets. Papillae were excised from 100 g Sprague-Dawley rats and loaded with the appropriate dye. The papillae were then examined using a Leica TCS 4D confocal microscope and oil immersion lenses. Fluorescence was excited (488 nm) using an argon laser and emission wavelengths above 515 nm collected using a long pass filter. Images of papillae loaded with 2'7'-bis-(carboxyethyl)-5-(and-6)-carboxyfluorescein acetoxymethyl ester clearly demonstrate a ladder-like arrangement of renal medullary interstitial cells. More detailed examination revealed the presence of cytoplasmic extensions that appear to make close contact with adjacent loops of Henle. Three-dimensional reconstructions of serial sections revealed spiral arrangements in some ladders of renal medullary interstitial cells. Nile Red-labelled lipid droplets of 0.5-1.0 microm diameter were located throughout the cytoplasm of renal medullary interstitial cells and especially within the cytoplasmic extensions. These experiments highlight the ability of confocal microscopy to allow investigation of renal medullary interstitial cells in situ.

Animals↗

PubMed1997

Seroprevalence of antibodies to Sarcocystis neurona in horses residing in Oregon.

OBJECTIVE: To determine seroprevalence of antibodies to Sarcocystis neurona in neurologically normal horses residing in 4 regions of Oregon and to describe the effects of age, gender, breed, and housing on seroprevalence within each region. DESIGN: Prevalence survey. SAMPLE POPULATION: Serum samples from 334 horses systematically selected by practicing veterinarians. PROCEDURE: Antibodies to S neurona were measured in sera, using a western blot. Information including age, gender, breed, housing, geographic location, and duration of residence was obtained for each horse. Data were analyzed, using descriptive statistics. RESULTS: 45% (149/334) of horses evaluated were seropositive for antibodies to S neurona with significant differences in the percentage of seropositive horses from different regions of the state. Seroprevalances of antibodies to S neurona in horses in regions I and II, west of the Cascade Range, were 65 and 60%, respectively; whereas seroprevalances in central and eastern Oregon, regions III and IV, were 43 and 22%, respectively. Seroprevalence consistently increased with age of horse for each region. Gender, breed, and housing were not associated with significant differences in seroprevalence of antibodies to S neurona in the overall sample population, or in comparisons of samples obtained from horses within a particular region, or among samples obtained from horses residing in different regions. CLINICAL IMPLICATIONS: The high seroprevalence of antibodies to S neurona in neurologically normal horses indicates that analysis of serum alone would not be useful for definitive diagnosis of equine protozoal myeloencephalitis in horses in Oregon.

Age Distribution↗

PubMed1997

Mast cell adenosine induced calcium mobilization via Gi3 and Gq proteins.

Adenosine is an important mediator of mast cell secretory responses. Adenosine appears to act through one or more adenosine receptor subtypes to activate several signal transduction pathways; however, the specific mechanisms involved are not clearly defined. We studied the pathways involved in adenosine receptor-mediated calcium fluxes in RBL-2H3 cells, a mucosal mast cell-like line. The role of endogenous heterotrimeric G proteins in adenosine mediated calcium mobilization was investigated by microinjection of inhibitory antibodies that block specific G protein subtype function. The calcium transients associated with adenosine and antigen stimulation were compared in noninjected cells and cells that were microinjected with affinity purified neutralizing antibodies to the alpha subunits of Gi3, Gq, or Gs. The percentage of cells responding to adenosine was decreased in the presence of antibodies to Gi3 and Gq, but not Gs. Pertussis toxin decreased the percentage of cells responding to adenosine, but not antigen. These studies demonstrated a functional requirement for the pertussis toxin sensitive Gi3 protein and the pertussis toxin insensitive Gq protein in adenosine mediated calcium mobilization in mast cells.

Adenosine↗

PubMed1996

The phosphatidylinositol 3-kinase inhibitor wortmannin blocks mast cell exocytosis but not IL-6 production.

Phosphatidylinositol 3-kinase (PI3-kinase) activity has been shown to be important in cellular signaling via receptors associated with tyrosine kinases and receptors coupled to small or heterotrimeric G proteins. The importance of this activity in mast cell degranulation, leukotriene C4 generation, and IL-6 production was examined in mouse bone marrow-derived mast cells stimulated by high affinity IgE receptor cross-linking, direct influx of calcium, and/or adenosine receptor agonist exposure. Wortmannin, a fungal metabolite that at nanomolar concentrations inhibits PI3-kinase relatively specifically, blocked the release of granule-associated mast cell mediators independent of the secretory stimulus used. This inhibition was most prominent after a 2- to 5-min preincubation with wortmannin and was equally effective in cells additionally treated with exogenous N-ethylcarboxamidoadenosine to potentiate preformed mediator release. Mast cell production of leukotriene C4 20 min after activation or IL-6 16 h after activation was unaffected by up to 100 nM of wortmannin exposure. Mast cells preincubated with wortmannin failed to develop the classic electronmicroscopic evidence of granule swelling and fusion, increased membrane ruffling, or exocytosis upon Ag challenge. Activation of PI3-kinase appears to be critical for mast cell degranulation but is not required for arachidonic acid metabolism or cytokine production to occur. Furthermore, the inhibition of mast cell secretion by wortmannin is not stimulus specific but is evident for both IgE receptor cross-linking and direct calcium influx.

Androstadienes↗

PubMed1994

Cloning of two adenosine receptor subtypes from mouse bone marrow-derived mast cells.

Adenosine potentiates the stimulated release of mast cell mediators. Pharmacologic studies suggest the presence of two adenosine receptors, one positively coupled to adenylate cyclase and the other coupled to phospholipase C activation. To identify mast cell adenosine receptor subtypes, cDNAs for the A1 and A2a adenosine receptors were obtained by screening a mouse brain cDNA library with the use of PCR-derived probes. Mouse bone marrow-derived mast cell cDNA libraries were constructed and screened with the use of A1 and A2a cDNA probes, which revealed the presence of A2a, but not A1, receptor clones. A putative A2b receptor was identified by using low stringency mast cell library screening. Northern blotting of mast cell poly(A)+ RNA with the use of receptor subtype probes labeled single mRNA bands of 2.4 kb and 1.8 kb for the A2a and A2b receptors, respectively. In situ cells. An A2a receptor-specific agonist failed to enhance mast cell mediator release, which suggests that the secretory process is modulated through the A2b and/or another receptor subtype. By using RNase protection assays, we found that mast cells that had been cultured in the presence of N-ethylcarboxamidoadenosine for 24 h exhibited a decrease in both A2a and A2b receptor RNA levels. Cells that had been cultured for 1 to 2 days in the presence of dexamethasone demonstrated increased amounts of A2a receptor mRNA, but no identifiable change in A2b receptor mRNA. Mast cells possess at least two adenosine receptor subtypes that may be differentially regulated.

Adenosine↗

PubMed1994

Inhibition of protein kinase A fails to alter mast cell adenosine responsiveness.

Adenosine activates adenylate cyclase and phospholipase C in mast cells and potentiates stimulated mediator release. To determine whether activation of adenylate cyclase is necessary for the effects of adenosine on the mast cell secretory process, a specific inhibitor of cAMP-dependent protein kinase, KT5720, was used. Antigen and adenosine each induced a rapid increase in mast cell cAMP-dependent protein kinase activity within 30 s. Preincubation with KT5720 (100 nM-10 microM) suppressed cAMP-dependent protein kinase activity and inhibited antigen-stimulated beta-hexosaminidase and leukotriene C4 releases. Adenosine retained its ability to potentiate beta-hexosaminidase release in antigen- and A23187-stimulated cells even in the presence of complete cAMP-dependent protein kinase inhibition. Mast cells rendered unresponsive to adenosine-related signals by preincubation with adenosine analogs maintained this hyporesponsiveness after incubation with KT5720. It appears that the abilities of adenosine to augment mast cell degranulation and induce receptor hyporesponsiveness are independent of changes in cAMP.

Adenosine↗

PubMed1993

Mast cell desensitization to IgE fails to induce a parallel adenosine receptor desensitization.

Desensitization induced by challenge of mast cells with antigen is specific for IgE-dependent signals. During the secretory process mast cells release adenosine, which can induce a desensitization of adenosine receptors. To determine whether adenosine receptors may be desensitized from a previous antigen challenge, mast cells were sensitized with anti-DNP IgE antibody, challenged with DNP-BSA antigen, returned to culture overnight, resensitized, and rechallenged. Previously challenged cells exhibited increased spontaneous beta-hexosaminidase release, but adenosine retained its ability to augment beta-hexosaminidase release. Adenosine enhanced A23187-stimulated release of beta-hexosaminidase in control and previously challenged cells. Leukotriene C4 generation followed a similar pattern. Mastoparan, a direct G protein activator and mast cells secretagogue, produced a doubling of beta-hexosaminidase release in previously challenged cells. Results using other G protein activators were equivocal. Degranulation alone is insufficient to induce adenosine receptor hyposensitization. Whether the hyperresponsiveness to mastoparan is a consequence of uncoupling of IgE receptors from G proteins remains uncertain.

Adenosine↗

PubMed1993

Altered blood pressure, renal tubular function and levels of circulating inhibitors of Na-K-ATPase in mature SHR exposed to low salt diet in the perinatal period.

1. It has been shown previously that low salt diet, confined to the perinatal period only, reduces blood pressure (BP) and sodium retention in spontaneously hypertensive rat (SHR) offspring at maturity when compared with those given high salt diet. 2. In this study it is shown that such high salt diets are associated with an increased level of circulating Na-K-ATPase inhibitor (CINK) activity. 3. Animals given perinatal high salt diet have a significantly greater tubular reabsorptive capacity when compared with those given low salt diet. 4. The finding of a high level of circulating Na-K-ATPase inhibitory material in the face of increased renal tubular capacity and blood pressure suggests that while this inhibitory material may play a role in the elevated blood pressure of animals given high salt diet, it cannot cause the elevated rate of fluid reabsorption.

Animals↗

PubMed1993

Activity of phenazine analogs against Mycobacterium leprae infections in mice.

Twenty-five compounds structurally related to clofazimine were tested for their ability to inhibit the growth of Mycobacterium leprae using the kinetic method of drug evaluation in the mouse foot pad model of leprosy. Seven of the phenazine derivatives displayed anti-M. leprae activity comparable to that of clofazimine when administered at a concentration of 0.01% (w/w) in the diet. Three of the compounds, B746, B4087, and B4101, were active when administered at 0.001% in the diet. At a dietary concentration of 0.0001%, B4087 and B4101 were slightly more active than clofazimine, while B746 was less active. In the kinetic method of drug evaluation, greater anti-M. leprae activity of phenazine derivatives was generally associated with greater pigmentation of abdominal fat. Of the compounds which did not cause pigmentation when fed at a concentration of 0.01% in the diet B4090 was the most active. This compound also inhibits the growth of a clofazimine-resistant M. smegmatis strain.

Abdomen↗

PubMed1993

Clarithromycin is bactericidal against strains of Mycobacterium leprae resistant and susceptible to dapsone and rifampin.

The anti-Mycobacterium leprae activity of clarithromycin when administered alone and in combination with rifampin and dapsone in the diet was determined using the kinetic method of drug evaluation in mice. Clarithromycin when administered at a concentration of 0.1% (w/w) in the diet completely prevented growth of 2 pan-susceptible, 3 dapsone-resistant, 2 rifampin-resistant, and 2 rifampin and dapsone double resistant strains of M. leprae. A 0.03% (w/w) concentration also completely prevented growth of M. leprae in all mice infected with 2 of 7 strains tested, but in only some of the mice infected with the remaining 5 strains. No antagonistic drug interactions were observed between clarithromycin and dapsone or rifampin. The addition of clarithromycin to the currently recommended multidrug regimen should improve the rate of killing of M. leprae and help to prevent the growth of dapsone-resistant and rifampin-resistant strains.

Administration, Oral↗

PubMed1991

Lysophosphatidylcholine induces mast cell secretion and protein kinase C activation.

Lysophosphatidylcholine (lyso-PC), a natural product of phospholipase A2 activity, induced the secretion of both granule-associated beta-hexosaminidase and newly generated leukotriene C4 from mouse bone marrow-derived mast cells. Micromolar concentrations of lyso-PC potentiated the release of beta-hexosaminidase induced by specific antigen but not the calcium ionophore, A23187. Exogenous adenosine was relatively ineffective in enhancing beta-hexosaminidase release from cells challenged with lyso PC. Lyso-PC caused a marked increase in intracellular free-calcium levels and induced the activation of protein kinase C (PKC). These effects could not be abrogated by a prolonged preincubation with pertussis toxin. Staurosporine, an inhibitor of PKC, partially inhibited the abilities of antigen and A23187 to induce beta-hexosaminidase release but was ineffective when lyso-PC was the secretagogue. Lyso-PC appears to activate mast cell PKC, but its ability to stimulate mast cell mediator release appears to be related to its ability to elevate intracellular free calcium concentrations.

Adenosine↗

PubMed1991

Evaluation of the oral vitamin E absorption test in horses.

An oral vitamin E absorption test used in human beings was modified for use in horses. The most appropriate techniques with which to measure gastrointestinal tract absorption of vitamin E (alpha-tocopherol) in horses were developed. Vitamin E was administered orally, and serum values of alpha-tocopherol were measured by use of high-performance liquid chromatography at 0, 3, 6, 9, 12, and 24 hours after vitamin E administration. Variables included comparison of 2 dosages (45 and 90 IU/kg of body weight), routes of administration, and absorption dynamics of 3 preparations of dl-alpha-tocopherol. Absorption of the 2 doses of dl-alpha-tocopherol acetate indicated a dose response; the area under the curve at 24 hours (AUC24) was 4.3 micrograms.h/ml for the 45-IU/kg dose and 32.2 micrograms.h/ml (P less than 0.01) for the 90-IU/kg dose. Maximal absorption was apparent when vitamin E was naturally consumed in grain, compared with administration of identical preparations by stomach tube or paste. In the same horses, dl-alpha-tocopherol and dl-alpha-tocopherol acetate plus polyethylene glycol had statistically similar absorption curves and both had significantly greater AUC24, compared with dl-alpha-tocopherol acetate; values for the 3 compounds were 23.6, 25.8, and 12.6 micrograms.h/ml, respectively. The AUC24 varied between individual horses, but time of peak value was consistently observed between 6 and 9 hours.(ABSTRACT TRUNCATED AT 250 WORDS)

Administration, Oral↗

PubMed1990

Modulation of mast cell responses to adenosine by agents that alter protein kinase C activity.

The acute incubation of mouse bone marrow-derived mast cells with low concentrations of agents known to activate protein kinase C [phorbol myristate acetate (PMA), 1,2-dioctanoyl-sn-glycerol (diC8), and 1-oleoyl-2-acetyl-glycerol (OAG)] caused an enhancement of beta-hexosaminidase release stimulated by the calcium ionophore A23187. Higher concentrations of protein kinase C activators tended to inhibit A23187- or antigen-induced preformed mediator release. All concentrations studied induced a striking mast cell hyporesponsiveness to the mediator release augmenting effect of adenosine. Agents that have been reported to block protein kinase C activity [1-(5-isoquinolinesulfonyl)-2-methyl-piperazine dihydrochloride (H-7) and sphingosine] demonstrated diverse responses in this system. Up to 100 microM H-7 failed to affect mast cell beta-hexosaminidase release in the presence or absence of PMA and secretagogue. Sphingosine (10 microM) was a potent inhibitor of antigen- or A23187-induced mediator release as well as adenosine responsiveness. Sphingosine also blocked the effects of PMA noted above in a dose-dependent fashion. The generation of leukotriene C4 (LTC4) by stimulated mast cells surprisingly was not affected by concentrations of diC8 that significantly inhibited granule-associated mediator release. Translocation of protein kinase C activity from the cytosol to the mast cell membrane was evident in cells briefly pretreated with A23187, adenosine alone, and diC8 in the presence of Tyrode's buffer, A23187, or adenosine. These findings lend further support to the contention that signal transduction from mast cell adenosine receptors to processes that regulate degranulation may involve protein kinase C.

Adenosine↗

PubMed1990

Myelitis in a cat infected with Toxoplasma gondii and feline immunodeficiency virus.

Severe necrotizing myelitis secondary to localization and reactivation of Toxoplasma gondii within the spinal cord of a domestic shorthair cat was diagnosed by use of light and electron microscopy and immunohistochemistry. The cat also was infected with feline immunodeficiency virus. This case may have useful comparative features to T gondii infections in human patients with acquired immunodeficiency syndrome.

Animals↗

PubMed1990

Filtration failure induced by p-aminophenol in rats is due to raised intratubular pressure and not changes in glomerular function.

1. The p-aminophenol (pAP) model of tubular necrosis displays elevated tubular pressures equivalent to 'stop-flow', with low glomerular filtration rate (GFR) but maintained blood flow and urine output. Renal function, micropuncture, and morphological studies were performed in anaesthetized rats to examine the causes of filtration failure. 2. At the height of pAP-induced renal failure proximal tubular fluid reabsorption (Jv(a] was markedly reduced while proximal and distal free-flow rates measured by tubular fluid collections during venting of the nephron were not significantly different from saline-injected controls. Renal blood flow was maintained over the 4 h observation period despite extensive and selective proximal tubular necrosis. There was no temporal relationship between increased tubular pressure and cast formation. 3. Maintained blood and tubular fluid flow rates indicate that activation of tubuloglomerular feedback plays little or no part in pAP-induced renal failure, which is apparently due to high fluid flow resistance in the region of the connecting tubule, late distal convolution or collecting ducts. Morphological appearances were consistent with compression of these segments.

Absorption↗