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W L Smith

Publications and source records attributed to W L Smith.

At least 163 records · Page 9Linked to original sources
PubMed1993

PGH synthase isoenzyme selectivity: the potential for safer nonsteroidal antiinflammatory drugs.

With the recent cloning of a second gene coding for the prostaglandin endoperoxide (PGH) synthase (cyclooxygenase), it has become obvious that mammalian cells contain two related, but unique, isozymes of PGH synthase. Both of these isozymes catalyze the conversion of arachidonic acid to PGH2, leading to production of biologically active prostaglandins. Although the first of these isozymes, PGH synthase-1 (PGHS-1), has long been thought to be the primary and sole site of action of nonsteroidal antiinflammatory drugs (NSAIDs), it is now known that the second isozyme, PGH synthase-2 (PGHS-2), is also sensitive to NSAIDs. Cloning of complementary DNAs for murine PGHS-1 and PGHS-2 has permitted individual expression of these two isozymes in the cos-1 cell system and comparison of their relative inhibition by several common NSAIDs in vitro. These studies have demonstrated that the two mouse isozymes, PGHS-1 and PGHS-2, are pharmacologically distinct. PGHS-1 is a constitutively expressed enzyme that early observations indicate is the principal enzyme involved in producing prostaglandins that regulate cellular housekeeping functions, such as gastric cytoprotection, vascular homeostasis, and kidney function. In contrast, PGHS-2 appears only to be expressed in inflamed tissue or following exposure to growth factors, lymphokines, or other mediators of inflammation. Expression of PGHS-2 is inhibited by antiinflammatory glucocorticoids, lending further support to the hypothesis that this enzyme produces prostaglandins involved in inflammation. We have identified NSAIDs that preferentially inhibit murine PGHS-1 or PGHS-2 or inhibit both isozymes equally. The finding that the two isozymes can be differentially inhibited provides a possible mechanism for identifying safer, more effective NSAIDs. Screening for drugs that preferentially inhibit PGHS-2 may allow identification of NSAIDs that reduce inflammation, but spare renal and gastric prostaglandin synthesis, thus reducing the untoward side effects commonly associated with most NSAIDs. Thus far, nabumetone is the only NSAID identified that preferentially inhibits murine PGHS-2.

Animals↗
PubMed1993

Cloning of an adenosine A1 receptor-encoding gene from rabbit.

A partial cDNA encoding the A1 adenosine receptor (A1AR), which lacks nucleotides coding for the first 74 amino acids (aa), was isolated from a rabbit kidney cDNA library. The missing 5' end sequence was obtained from an overlapping rabbit genomic clone which was found to contain the flanking 5' untranslated region (5'UTR), the first exon and part of the first intron. Together, the cDNA and genomic clones provide the entire open reading frame (ORF) encoding rabbit A1AR. The deduced aa sequence is highly homologous to the canine, rat and bovine A1ARs. These data also indicate that the A1AR gene belongs to the family of intron-containing G-protein-linked receptor genes.

Adenosine↗
PubMed1993

The effect of the 5-HT3 receptor antagonist, RS-42358-197, in animal models of anxiety.

The S-isomer of the novel 5-HT3 receptor antagonist RS-42358 ((S)-N-(1-azabicyclo[2.2.2]oct-3-yl)-2,4,5,6-tetrahydro-1-H- benzo[de]isoquinolin-1-one, RS-42358-197) disinhibited behaviour in the mouse suppressed by the aversive situation of the light/dark test box. RS-42358-197 was effective at sub-ng/kg dose levels and the efficacy was maintained over a 100 million-fold dose range. In contrast, the R-isomer was ineffective at all doses studied. The S-isomer also disinhibited a suppressed behaviour in social interaction and elevated X-maze tests in the rat and reduced anxiety-related behaviours in a marmoset human threat test. RS-42358-197 prevented the exacerbation of the suppression of behaviour in the mouse light/dark test following withdrawal from treatment with alcohol, nicotine, cocaine and diazepam. Thus, the S-isomer of RS-42358 has a consistent non-sedating anxiolytic profile in rodent and primate models. It is exceptionally potent and a maintained efficacy at high doses distinguishes its actions from many other 5-HT3 receptor antagonists.

Animals↗
PubMed1993

Differential inhibition of prostaglandin endoperoxide synthase (cyclooxygenase) isozymes by aspirin and other non-steroidal anti-inflammatory drugs.

Murine prostaglandin endoperoxide (PGH) synthase-1 and PGH synthase-2 expressed in cos-1 cells were found to be differentially sensitive to inhibition by common nonsteroidal anti-inflammatory drugs (NSAIDs). Aspirin completely inhibited bis-oxygenation of arachidonate by PGH synthase-1; in contrast, aspirin-treated PGH synthase-2 metabolized arachidonate primarily to 15-hydroxyeicosatetraenoic acid (15-HETE) instead of PGH2. ID50 values were determined for a panel of common NSAIDs by measuring instantaneous inhibition of cyclooxygenase activity using an oxygen electrode. Among common NSAIDs tested, indomethacin, sulindac sulfide, and piroxicam preferentially inhibited PGH synthase-1; ibuprofen, flurbiprofen, and meclofenamate inhibited both enzymes with comparable potencies; and 6-methoxy-2-naphthylacetic acid preferentially inhibited PGH synthase-2. These results demonstrate that the two PGH synthases are pharmacologically distinct and indicate that it may be possible to develop isozyme-specific cyclooxygenase inhibitors useful both for anti-inflammatory therapy and for delineating between the biological roles of the PGH synthase isozymes.

Animals↗
PubMed1993

Subcellular localization of prostaglandin endoperoxide synthase-2 in murine 3T3 cells.

Polyclonal antisera specific for prostaglandin endoperoxide (PGH) synthases-1 and -2 were used to determine the subcellular locations of each PGH synthase isozyme in detergent-permeabilized mouse 3T3 fibroblasts by indirect immunocytofluorescence. Antiserum to PGH synthase-1 demonstrated a mottled pattern of cytoplasmic and perinuclear staining of both serum-starved and serum-stimulated 3T3 cells. This pattern of staining is consistent with the results of earlier studies which demonstrated that PGH synthase-1 is associated with the endoplasmic reticulum and nuclear envelope of these cells. As expected, antibodies directed against a peptide unique to PGH synthase-2 failed to stain serum-starved cells, which lack appreciable levels of this second form of the enzyme. However, serum-stimulated 3T3 cells, which do express PGH synthase-2, showed the same pattern of staining with PGH synthase-2 antibodies as was observed with anti-PGH synthase-1 serum--mottled cytoplasmic staining and perinuclear staining. We conclude that the subcellular location of PGH synthase-2 is the same as PGH synthase-1 in murine 3T3 cells. Thus, the notable differences in the primary amino acid sequence--the signal peptide and the additional 18 amino acid C-terminal segment in PGH synthase-2--do not cause a change in localization. Colocalization of PGH synthases-1 and -2 implies that the source of arachidonate substrate, the site of PGH2 and prostanoid formation, and the mechanism of product transport from the inside to the outside of the cell are the same for these isozymes.

3T3 Cells↗
PubMed1993

Evaluation of resident applicants by letters of recommendation: a comparison of traditional and behavior-based formats.

RATIONALE AND OBJECTIVES: Traditional, narrative letters of recommendation solicited by medical students applying for radiology residency are widely used as a selection tool. Letters of recommendation are considered a source of reliable information about the attitudes and behaviors (non-cognitive variables) of the resident applicant. However, in many instances, this information is not present or is highly encoded and cannot be extracted. This study attempted to document the deficiencies of traditional letters of recommendation and determine the effectiveness of a structured letter of recommendation in obtaining information regarding noncognitive variables. METHODS: One hundred thirteen randomly selected letters of recommendation were analyzed by two radiologists with experience in residency selection. Deficiencies in inclusion of information or the ability to extract information about noncognitive variables were documented. A standard behavioral assessment was sent to the writers of these letters of recommendation and these results tabulated. RESULTS: The traditional letters of recommendation were frequently deficient in data regarding the noncognitive variables. In letters that contained such data, two experienced reviewers could not reliably extract the information. The structured form produced clearly identifiable information about the letter writer's assessment of noncognitive variables. CONCLUSIONS: Traditional letters of information are frequently deficient in data regarding noncognitive variables. A standardized statement is effective in eliciting information on noncognitive variables related to applicant performance.

Internship and Residency↗
PubMed1993

Joseph E. Whitley Award. The effect of preradiology clinical training on the performance of radiology residents.

RATIONALE AND OBJECTIVES: A multi-institutional study was developed to evaluate whether having preradiology clinical training has an effect on the quality of performance of radiology residents assessed both after 1 year of training and at the end of the 4-year residency. MATERIAL AND METHODS: Five faculty members from each of nine collaborating institutions that accept residents both directly from medical school and with postgraduate clinical training completed 1,015 evaluation forms on their 203 residents who had just completed their first year of radiology training or had finished training in the past 2 years. Each resident was evaluated by two senior faculty, two junior faculty, and the residency program director using a cross-institutionally validated, behavior-oriented, standardized instrument that assessed five behavioral characteristics: conscientiousness, curiosity, interpersonal skills, confidence, and recognition of limits. The differences between residents who did and did not have preradiology clinical training were evaluated by analysis of variance and by simple effects testing. RESULTS: At the conclusion of training, no significant differences in resident performance were found between residents who did and did not have postgraduate clinical training before radiology residency. Among residents who had just finished their first year of training, those who had had clinical training scored significantly higher on four of the five behavioral scales. CONCLUSIONS: Clinical training before radiology residency does not correlate with any measurable benefit in performance at the end of radiology residency. Training programs should consider elimination of this requirement.

Adult↗
PubMed1993

Evaluation of milk antibiotic residue screening tests in cattle with naturally occurring clinical mastitis.

Milk from 172 commercial cows with mild to moderate clinical mastitis was tested with five antibiotic residue detection assay systems. One hundred cows were treated with one of two intramammary beta-lactam antibiotics, and the remaining 72 cows were treated with intramuscular oxytocin. Milk samples were collected pretreatment, twice after therapy, and again 21 d following the initiation of treatment. Presumptive false-positive assay results were tabulated from all pretreatment and 21-d milk samples and from samples collected following oxytocin therapy. The percentage of false-positive results was 43.6, 37.7, 81.7, 2.6, and 18.8% for the CITE probe (beta-lactam), Delvotest-P, Charm Farm, LacTek (beta-lactam), and Bacillus stearothermophilus var. calidolactis disk assay, respectively. In four of the assay systems, average SCC were significantly higher in samples yielding false-positive results than in those with negative results. Specificity and sensitivity were estimated for each assay system, and, based on these estimates, positive and negative predictive value curves were graphed as the prevalence of milk samples containing detectable concentrations of exogenous antibiotic residues in the sample population was varied from 0 to 100%.

Animals↗
PubMed1993

Desferrioxamine and Alzheimer's disease: video home behavior assessment of clinical course and measures of brain aluminum.

Drug trials designed to modify the progression of Alzheimer's disease (AD) have required the development of mental state and behavior evaluation instruments that are sensitive to cognitive decline and measure skills useful in everyday living. We describe a videotaped home behavior (VHB) assessment instrument with high construct validity and reliability and a strong relationship to criterion references. The VHB was employed to test the hypothesis that aluminum is an important pathogenic factor in AD. The trivalent chelating agent desferrioxamine (DFO), 125 mg i.m. twice daily five days per week, was used in a randomized single-blind, oral lecithin, placebo-controlled clinical trial in 48 patients with AD. Analysis showed that the treatment and no-treatment groups were closely matched at entry into the trial but that the rate of decline, as measured by the VHB over 2 years of observation, was twice as rapid in the no-treatment group compared with the DFO-treated group. Furthermore, trace-metal analysis of autopsied brain confirmed that extended treatment with DFO lowered neocortical brain aluminum concentrations to near control concentrations. Aluminum ion-specific chelation may be a useful palliative treatment for AD, and further clinical trials are indicated.

Aged↗
PubMed1993

RS 42358-197, a novel and potent 5-HT3 receptor antagonist, in vitro and in vivo.

The pharmacological activity of RS 42358-197, a novel 5-HT3 receptor antagonist has been evaluated in vitro and in vivo. In functional experiments in vitro, RS 42358-197 behaved as a competitive antagonist against 5-HT-induced contractions in the guinea pig ileum (low-potency phase), yielding a pA2 estimate of 8.1. RS 42358-197 was devoid of any agonistic or antagonistic activity at 5-HT1-like receptors (contraction of canine saphenous vein), 5-HT2 receptors (contraction of rabbit aorta) or 5-HT4 receptors (contraction of guinea pig ileum, high-potency phase). RS 42358-197 failed to affect the concentration-effect curve to substance P in guinea pig ileum. In anesthetized rats. RS 42358-197, administered by the intravenous, intraduodenal or transdermal route, dose-dependently inhibited the Bezold-Jarisch reflex induced by 2-methyl 5-HT (ID50:0.05 micrograms/kg; i.v., 5.7 micrograms/kg; i.d., and 11.6 micrograms/chamber, respectively). In this regard, when administered intraduodenally, RS 42358-197 was more potent and exhibited a longer duration of action than either ondansetron or granisetron. In dogs, RS 42358-197, administered either intravenously or orally, dose-dependently inhibited the emesis induced by cisplatin, actinomycin and cyclophosphamide, but not that induced by apomorphine. When tested at maximally effective doses against cisplatin-induced emesis in dogs, RS 42358-197 had a longer duration of antiemetic activity (> 6 h) than ondansetron (2 h). RS 42358-197, administered orally, also afforded protection against cisplatin-induced emesis in ferrets. At doses that showed marked anti-emetic activity in dogs (10-100 micrograms/kg; i.v. and 100-1000 micrograms/kg; i.d.), RS 42358-197 did not produce any hemodynamic changes.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Milk antimicrobial drug residue assay results in cattle with experimental, endotoxin-induced mastitis.

Antimicrobial drug residue testing was performed on milk samples obtained from 8 cows with experimental endotoxin-induced mastitis, using 4 commercially available assay kits. Although none of the cows in the study received antimicrobials, only 1 of the 4 assay procedures, assay C, had consistently negative results (specificity = 1.00). The proportion of positive assay results varied from 0 to 1.00 among combinations of sampling time, sample status (endotoxin-infused quarter vs composite noninfused sample). The proportion of positive results found when assay C was used (0) differed significantly from the proportion found when the 3 other assays were used. The proportion of positive results did not differ significantly between assay A (0.45) and assay B (0.48); however, both assays had a significantly lower proportion of positive assays than did assay D (0.86). Logistic regression models were developed predicting positive milk antimicrobial drug residue assay results as a function of assay kit, sample status, and time interval following experimental challenge exposure. Using assay A as a baseline risk, assay B and assay D were more likely to have positive assay results, and assay C had a decreased risk of positive assay results. Milk samples from endotoxin-infused quarters were at increased risk for positive assay results, compared with noninfused composite samples. Samples collected from endotoxin-infused quarters or control quarters were at increased risk for positive assay results following the intramammary infusion of endotoxin. Our findings suggest that specificity of milk antimicrobial drug residue assays varies greatly among assay kits and that intramammary inflammation may increase the proportion of false-positive assay results.

Animals↗
PubMed1992

Distribution and characterization of cyclooxygenase immunoreactivity in the ovine brain.

Evidence from tissue culture studies suggests that glial cells are the principal source of prostaglandins in the brain. We have used immunohistochemistry, Western blot analysis, and enzyme activity assays to localize cyclooxygenase (COX), the enzyme responsible for the conversion of arachidonic acid to prostaglandins, in situ in the normal ovine brain. We observed very few immunoreactive glial cells. In contrast, an extensive distribution of COX-like immunoreactive (ir) neuronal cell bodies and dendrites and a corresponding pattern of COX enzyme activity were observed. COXir neurons were most abundant in forebrain sites involved in complex, integrative functions and autonomic regulation such as the cerebral cortex, hippocampus, amygdala, bed nucleus of the stria terminalis, substantia innominata, dorsomedial nucleus of the hypothalamus, and tuberomammillary nucleus. Moderate populations were observed in other regions of the central nervous system implicated in sensory afferent processing, including the dorsal column nuclei, spinal trigeminal nucleus, and superior colliculus, and in structures involved in autonomic regulation, such as the nucleus of the solitary tract, parabrachial nucleus, and the periaqueductal gray matter. We did not observe COXir axons or terminal fields, however. Our results suggest that neurons may use prostaglandins as intracellular or perhaps paracrine, but probably not synaptic, mediators in the normal brain.

Acetylcholinesterase↗
PubMed1992

Prostaglandin endoperoxide synthase. The aspirin acetylation region.

Aspirin selectively acetylates Ser-530 of prostaglandin endoperoxide (PGH) synthase-1. This causes inactivation of the cyclooxygenase activity of the enzyme, but does not appreciably affect its peroxidase activity. Although the aspirin-acetylated enzyme is inactive, we found that PGH synthase-1 in which Ser-530 had been replaced with an alanine was catalytically active; accordingly, we proposed that aspirin inhibits cyclooxygenase activity by placing a larger than normal side chain at position 530 thereby interfering with arachidonate binding (DeWitt, D.L., El-Harith, E. A., Kraemer, S. A., Andrews, M. J., Yao, E. F., Armstrong, R. L., and Smith, W. L. (1990) J. Biol. Chem. 265, 5192-5198). As a further test of this hypothesis we have used site-directed mutagenesis and transient expression in cos-1 cells to prepare and characterize five additional substitutions of Ser-530. Consistent with our proposal, the presence of amino acids with bulky side chains at position 530 inhibited cyclooxygenase activity and decreased the apparent affinity of the enzyme for arachidonate. In related work, we characterized a series of mutant PGH synthases-1 having substitutions at residues adjoining Ser-530, including Phe-529, Leu-531, Lys-532, and Gly-533, in order to evaluate the contributions of each residue to cyclooxygenase catalysis. The most significant conclusion of this part of the study is that residues 529-533 all are important for the peroxidase activity as well as the cyclooxygenase activity of PGH synthase-1. Phe-529, in particular, was found to be critical for PGH synthase-1 structure and catalysis; some substitutions at this position led to the production of proteins lacking about 100 amino acids from their COOH termini.

Acetylation↗
PubMed1992

A lipoxygenase pathway of arachidonic acid metabolism mediates FMRFamide activation of a potassium current in an identified neuron of Helisoma.

The neuropeptide FMRFamide causes a presynaptic inhibition of neurotransmitter release from neuron B5 of Helisoma. In this study we demonstrate that one of FMRFamide's actions is to activate an outwardly rectifying potassium current. Arachidonic acid also activates an outward current in B5. The phospholipase A2 inhibitor, 4-bromophenacylbromide (BPB), and nordihydroguaiaretic acid (NDGA), an inhibitor of arachidonic acid metabolism, but not indomethacin, block FMRFamide's activation of the potassium current. Taken together these data demonstrate that one of FMRFamide's presynaptic actions is to activate a potassium current through a lipoxygenase pathway of arachidonic acid metabolism.

Acetophenones↗
PubMed1992

Expression of prostaglandin endoperoxide synthase-1 in a baculovirus system.

A cDNA coding for ovine prostaglandin endoperoxide (PGH) synthase-1 was used to construct a recombinant baculovirus which was expressed in Spodoptera frugiperda (Sf9) insect cells. Two proteins reactive with anti-PGH synthase antibody were produced. A larger protein (Mr = 72,000) coelectrophoresed with native enzyme; a smaller, more abundant protein (Mr = 66,000) was unglycosylated enzyme. About 90% of both the immunoreactivity and the cyclooxygenase activity were present in a low speed (10(5) g x min) pellet; variable but low peroxidase activities were observed in this fraction. The specific cyclooxygenase activity of solubilized PGH synthase-1 from Sf9 cells was 56 units/mg versus 112 units/mg for the same cDNA expressed in cos-1 cells. The baculovirus-insect cell system is not ideal for generating large amounts of active PGH synthase-1 apparently because of inefficient N-glycosylation.

Animals↗
PubMed1992

Contrast enema diagnosis of necrotizing enterocolitis.

Contrast enema was performed in a select group of 126 neonates with ambiguous diagnosis of necrotizing enterocolitis (NEC). Enema findings were compared with that on plain abdominal radiographs (AXR) and the clinical outcome. Thirty-one of the 34 patients with a discharge diagnosis of NEC were interpreted to have a positive contrast enema, and 91 of the 92 patients without a discharge diagnosis of NEC were interpreted to have a negative contrast enema. The AXR was interpreted as positive in 57 patients and as negative or indeterminate in 69. Not a single case of perforation was produced. Judicious use of contrast enema can improve specificity of diagnosis in cases with discordant clinical and plain radiographic findings.

Barium Sulfate↗