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J B Smith

Publications and source records attributed to J B Smith.

At least 163 records · Page 9Linked to original sources
PubMed1992

Predictors for live birth after unexplained spontaneous abortions: correlations between immunologic test results, obstetric histories, and outcome of next pregnancy without treatment.

Our objective was to determine whether results from tests for maternal serum antinuclear antibodies, cytotoxic antibodies to paternal lymphocytes, parental histocompatibility types, and blocking factors for maternal-paternal mixed lymphocyte reactions were predictive of pregnancy outcome without immunologic treatment. Pregnancy outcome data from 95 women with a history of unexplained recurrent spontaneous abortions who underwent immunologic tests at Jefferson Medical College were evaluated with multiple logistic regression analyses. The number of prior spontaneous abortions, history of another relevant diagnosis, parental sharing of one histocompatibility antigen, and maternal age were related to the outcome of the next pregnancy in women given no immunologic treatment (p = 0.05). No significant correlation was found between results from the immunologic tests or other history characteristics evaluated and outcome of the next pregnancy. The immunologic tests evaluated were not clinically useful predictors of pregnancy outcome.

Abortion, Habitual↗
PubMed1992

Deriving a 67-nucleotide trans-cleaving ribozyme from the hepatitis delta virus antigenomic RNA.

RNAs derived from the genomic and antigenomic hepatitis delta virus are capable of self-cleavage, and thus have the potential for serving as ribozymes in a trans-cleaving reaction. Because the catalytic core of such an enzymatic RNA was not evident from phylogenetic data, we took a step-wise approach to identifying the core, reducing the RNA in size, and characterizing various properties for each size class. Thus, a 186-nucleotide antigenomic RNA (termed Ag180) was found to be capable of cleaving well in 20 M formamide (Smith and Dinter-Gottlieb, 1991), and this unusual stability in formamide was lost by reducing the 3' end of the molecule, leaving a 140-nucleotide RNA (Ag 140). Both RNAs showed only intramolecular cleavage at a wide range of concentrations, and a number of conformers could be seen in the Ag140 RNA, some of which were resistant to cleavage at 37 degrees C. Since Ag140 could not cleave in 20 M formamide, the 5' and 3' termini of Ag180 were truncated and produced Ag5-84, which cleaved to 100% at 37 degrees C in less than 0.25 min. Internal deletions of the Stem IV region resulted in Ag5-73, still capable of efficient cleavage, although with a lessened stability in formamide. A trans-cleaving enzyme-substrate pair was finally derived from this RNA, and it consisted of a 67-nucleotide enzyme that cleaved a 13-nucleotide RNA substrate.

Base Sequence↗
PubMed1992

Ca2+ influx via Na(+)-Ca2+ exchange in immortalized aortic myocytes. I. Dependence on [Na+]i and inhibition by external Na+.

We have found that immortalized aortic myocytes have high Na(+)-Ca2+ exchange activity that is similar in amount to that of low-passage myocyte cultures. Replacing all external Na+ with K+ slightly increased 45Ca2+ uptake in cells with basal Na+ and greatly increased 45Ca2+ uptake in cells with increased intracellular Na+. External Na+ competitively inhibited 45Ca2+ uptake (inhibition constant approximately 10 mM). The dependence of exchange activity on intracellular Na+ was sigmoidal. The 50% maximum concentration (K0.5) for Na+ was 33 +/- 1 mmol/l cell water space. The Hill coefficient was 2.9 +/- 0.2, which is consistent with a stoichiometry of 3 Na+/1 Ca2+. Cytosolic free Na+ was estimated from the ratio of sodium-binding benzofuran isophthalate (SBFI) fluorescence and an in vivo calibration curve. Sulfinpyrazone, an inhibitor of anion transport, markedly increased the SBFI content of the cells without affecting cell Na+. Cytosolic free Na+ was 9 mM under basal conditions. Because free Na+ did not differ significantly from total Na+, little or no Na+ is bound or compartmentalized in these cells. The complete replacement of extracellular Na+ with K+ fails to evoke appreciable Ca2+ influx, at least in part, because of the low cell Na+ concentration relative to the K0.5 of the exchanger and its sigmoid dependence on Na+.

Animals↗
PubMed1992

Ca2+ influx via Na(+)-Ca2+ exchange in immortalized aortic myocytes. II. Feedback inhibition by [Ca2+]i.

Depolarization with 50 mM K+ evoked a spike in cytosolic free Ca2+ ([Ca2+]i) and increased 45Ca2+ uptake in immortalized aortic myocytes. The following evidence indicates that the electrogenic Na(+)-Ca2+ exchanger caused the Ca2+ influx that was evoked by K+ depolarization. First, K+ depolarization had no effect on [Ca2+]i and 45Ca2+ uptake in cells with basal Na+ but strikingly increased both in Na(+)-loaded cells. Second, the [Ca2+]i increases produced by K+ depolarization depended hyperbolically on external Ca2+ (50% maximum concentration = 1.5 mM). Third, the increases in [Ca2+]i and 45Ca2+ uptake were greater when external Na+ was replaced with K+ rather than with N-methyl-D-glucamine or choline. A series of K+ depolarizations elicited a sequence of [Ca2+]i spikes, provided there was a short incubation at 5 mM K+ between the depolarizations. A prior K+ depolarization almost abolished the 45Ca2+ uptake response to K+ depolarization. The inhibition of exchange activity by a prior K+ depolarization required external Ca2+ and was completely reversible. A prior incubation with angiotensin II, platelet-derived growth factor, or ionomycin also inhibited exchange activity. Moderate [Ca2+]i increases probably feedback inhibit Ca2+ influx via the exchanger by a kinetic mechanism. Inactivation of the exchanger, together with Ca2+ extrusion or sequestration, causes the rapid decrease in [Ca2+]i from the peak evoked by depolarization.

Animals↗
PubMed1992

Expression and regulation of L-selectin on eosinophils from human adults and neonates.

L-Selectin, previously known as LEC.CAM-1, LECAM-1, LAM1, and as the MEL-14, Leu-8, TQ1, and DREG-56 antigens, is a leukocyte membrane protein that participates in adhesion to endothelium. We studied its expression on eosinophils using flow cytometry and the MAb Dreg-56 and Leu-8. Unstimulated peripheral blood eosinophils from healthy adults expressed about one third the level of L-selectin as neutrophils (mean +/- SD specific fluorescence: 20.9 +/- 3.2 versus 54.5 +/- 8.4, p = 0.0001, n = 18). After stimulation with A23187, L-selectin expression on eosinophils was rapidly lost. This was temporally correlated with increased expression of Mac-1 (CD11b/CD18); the kinetics on eosinophils and neutrophils were similar. Eosinophil expression of L-selectin decreased modestly after stimulation with platelet activating factor, but was minimally affected by N-formyl-methionyl-leucyl-phenylalanine, leukotriene B4, or C5a compared with their effects on neutrophils. Eosinophils from cord blood of healthy neonates born at term expressed less L-selectin than adult eosinophils (10.4 +/- 3.8 versus 19.4 +/- 2.7, p = 0.0001, n = 9); the relative reduction was the same as on cord blood neutrophils (36.4 +/- 8.2 versus 55.5 +/- 4.8, p = 0.0001, n = 9). Relative to baseline expression, the responses of neonatal and adult cells to stimulation did not differ. We conclude that neonatal eosinophils have abnormalities in L-selectin expression similar to neonatal neutrophils and suggest that decreased expression of L-selectin and a diminished responsiveness to direct stimulation with chemotactic factors are possible mechanisms that may limit the exudation of eosinophils.

Adult↗
PubMed1992

The incidence and prevalence of dermatitis herpetiformis in Utah.

BACKGROUND AND DESIGN: The incidence and prevalence of dermatitis herpetiformis has never been formally evaluated in any area of the United States. Several northern European studies have shown prevalence rates ranging from 1.2 per 100,000 to 39.2 per 100,000. The present study was performed to evaluate the incidence and prevalence of dermatitis herpetiformis in Utah. Information from 240 patients diagnosed with dermatitis herpetiformis was compiled from hospital records throughout Utah, as well as the sole private dermatopathologist in the state, and from the university referral center of the state. Criteria for inclusion in the study were a clinical diagnosis of dermatitis herpetiformis plus granular deposition of IgA in dermal papillae by direct immunofluorescence of uninvolved skin, or histopathologic findings consistent with the disease. Clinical diagnosis and response to dapsone alone was considered insufficient for inclusion in the study. On the basis of these criteria, as well as exclusion of non-Utah residents, 188 of the original 240 patients qualified for the study. RESULTS: The prevalence of dermatitis herpetiformis in Utah in 1987 was 11.2 per 100,000. The mean incidence for the years 1978 through 1987 was 0.98 per 100,000 per year. The mean age at onset of symptoms for male patients was 40.1 years, and that for female patients was 36.2 years. The male-female ratio was 1.44:1. CONCLUSIONS: This represents the first evaluation of the incidence and prevalence of dermatitis herpetiformis in the United States. These results are similar to those of the previous studies, probably because of Utah's largely northern European ancestry. This population base, plus a much smaller than average black and Oriental population, is likely to have produced a higher incidence and prevalence in Utah than would be seen in other areas of the United States.

Adult↗
PubMed1991

Sodium-calcium exchange in membrane vesicles from aortic myocytes: stimulation by endogenous proteolysis masks inactivation during vesicle preparation.

Plasma membrane vesicles were purified from rat aortic myocytes by centrifugation in a discontinuous sucrose gradient. Vesicles were prepared in the presence or absence of five proteinase inhibitors (aprotinin, benzamidine, leupeptin, pepstatin A and phenylmethylsulfonyl fluoride). The proteinase inhibitors decreased the Vmax by 3.4-fold and had no effect on the Km for Ca2+ of Na+ gradient-dependent 45Ca2+ influx. The proteinase inhibitors had no direct effect on exchange activity, and they had no effect on membrane purity as indicated by 5'-nucleotidase activity. Removing the proteinase inhibitors or adding trypsin or chymotrypsin increased exchange activity approx. 2-fold. The Vmax of exchange activity in intact aortic myocytes is approx. 10-fold higher than the Vmax in plasma membrane vesicles prepared in the presence of proteinase inhibitors. Exchange activity in plasma membrane vesicles is only a sixtieth of the expected value, because the vesicles have approx. 7-fold higher 5'-nucleotidase activity and approx. 6-fold higher specific exchange activity than the crude homogenate. The large loss of exchange activity may be caused by a change in a regulatory domain of the exchanger because endogenous proteolysis restores some of the activity lost during vesicle preparation.

5'-Nucleotidase↗
PubMed1991

Identification of tyrosine 67 in bovine brain myelin basic protein as a specific phosphorylation site for thymus p56lck.

The protein-tyrosine kinase p56lck exhibits a restricted substrate specificity in vitro but can efficiently phosphorylate bovine myelin basic protein (MBP). Results obtained from both peptide mapping and fast atom bombardment mass spectrometry indicate that tyrosine 67 in the sequence -Thr-Thr-His-Tyr67-Gly-Ser-Leu-Pro-Gln-Lys- in bovine MBP is the specific phosphorylation site. p56lck does not phosphorylate the acidic, cytoplasmic domain of erythrocyte band 3. In contrast, p40, another protein-tyrosine kinase purified from bovine thymus that readily phosphorylates band 3, does not phosphorylate MBP. Therefore, MBP and band 3 may prove to be useful substrates for distinguishing between various tyrosine kinases on the basis of substrate specificity. In addition, identification of the recognition sequence in MBP for p56lck may contribute to an understanding of the structural features of physiological substrates for this kinase.

Amino Acid Sequence↗
PubMed1991

Identification of 50 kDa snake venom proteins which specifically inhibit platelet adhesion to collagen.

Of 32 snake venoms tested, the crude venoms of four (B.atrox, B.jararaca, A.halys blomhoffi and C.basiliscus) showed strong inhibitory activity in an assay of platelet adhesion to collagen. Active 50 kDa proteins were purified to homogeneity from each venom and found to be rich in cysteine on amino acid analysis. A monoclonal antibody raised against the purified B. atrox protein crossreacted strongly with the 50 kDa proteins from B.jararaca and A.halys blomhoffi and weakly with the protein from C.basiliscus, indicating that all four proteins possess a similar epitope. The proteins inhibited platelet aggregation induced by collagen but not by other agonists.

Adenosine Diphosphate↗
PubMed1991

Antigenomic Hepatitis delta virus ribozymes self-cleave in 18 M formamide.

The ribozymes derived from Hepatitis delta virus (HDV) RNA appear unique in their sequence requirements for self-cleavage. While truncating the 1679 nucleotide antigenomic HDV RNA, we have characterized the cleavage requirements of a number of ribozymes of intermediate length. Two of these, containing 186 and 106 HDV nucleotides respectively, cleaved to completion in the presence of 18 M formamide. The 186 nucleotide ribozyme also cleaved to completion in 10 M urea. Removal of an additional 10 nts from the 3' terminus of the 106 nt ribozyme resulted in a loss of the ability to cleave in high concentrations of the denaturants. The interaction of nucleotides near the cleavage site with a sequence within this 10 base region may confer unusual stability on these ribozymes.

Base Sequence↗
PubMed1991

Elucidation of the primary structures of proteins by mass spectrometry.

A combination of new mass spectrometric methods that can be used to determine the primary structures of proteins, including post-translational modifications, with unprecedented speed and accuracy is described. Structural characterization of alpha-crystallins from bovine lenses has been used to illustrate the methods. The molecular weights of alpha-crystallins fractionated, but not to homogeneity, by reversed-phase HPLC were determined with an uncertainty of 0.01% which is at least 100 times more accurate than is possible using conventional methods. This information was used to identify the primary gene product as well as its phosphorylated and truncated forms. Molecular weight maps of proteolytic digests of these proteins were determined by directly coupled capillary HPLC fast atom bombardment-mass spectrometry. From these maps, the entire amino acid sequence was confirmed, and the phosphorylated peptide was identified. The MS/MS daughter ion mass spectrum of the phosphorylated peptide provided sufficient information to determine which residue was phosphorylated. Because protein structure, including post-translational modifications, is determined on the basis of molecular weight, this method has broad application and will be useful for a variety of diverse and challenging problems in protein structure elucidation.

Amino Acid Sequence↗
PubMed1991

Inhibition of sodium-calcium and sodium-proton exchangers by amiloride congeners in arterial muscle cells.

The inhibitory potencies of several amiloride congeners towards Na(+)-Ca2+ and Na(+)-H+ exchange were compared in rat aortic myocytes. N-(2,4-Dimethylbenzyl)amiloride (DMB) was 10 times more potent towards Na(+)-Ca2+ than Na(+)-H+ exchange. Amiloride and ethylisopropylamiloride were about 5,000 and 10,000 times more potent toward Na(+)-H+ than Na(+)-Ca2+ exchange respectively. N-(3,4-Dichlorobenzyl)amiloride was almost equipotent towards both exchangers. About 40 nM ethylisopropylamiloride inhibited Na(+)-H+ exchange by 50%. Ethylisopropylamiloride (10 microM) had no effect on basal or angiotensin-evoked 45Ca2+ efflux or net Ca2+ efflux. In contrast to ethylisopropylamiloride, 25-50 microM DMB, which strongly inhibits Na(+)-Ca2+ exchange, markedly decreased both 45Ca2+ efflux and net Ca2+ efflux produced by angiotensin. Replacing extracellular Na+ with N-methyl-D-glucamine inhibited angiotensin-evoked 45Ca2+ efflux similarly to DMB. Neither DMB nor Na+ placement had any effect on basal or angiotensin-evoked production of [3H]inositol phosphates. These findings suggest that Na(+)-H+ exchange has no major influence on short-term Ca2+ regulation and provide evidence that Na(+)-Ca2+ exchange is a major pathway of rapid Ca2+ efflux in stimulated arterial muscle cells.

Amiloride↗
PubMed1991

Facile platelet adhesion to collagen requires metabolic energy and actin polymerization and evokes intracellular free calcium mobilization.

The attachment of platelets to collagen-coated microtiter plates at 20 degrees C was inhibited strongly by depletion of metabolic energy or by addition of cytochalasins and was slightly inhibited by the intracellular Ca2+ chelator BAPTA. In keeping with their respective potencies as inhibitors of actin polymerization, cytochalasins D and H were the most potent inhibitors of adhesion, while cytochalasin B was the least potent. Energy depletion, cytochalasin D or, to a much lesser extent, BAPTA also inhibited platelet adhesion to collagen in a suspension assay system at 37 degrees C. Collagen-induced platelet cytosolic Ca2+ mobilization was inhibited up to 70% by cytochalasin D and abolished by energy depletion or BAPTA. Elevation of intracellular platelet calcium by treatment with ionomycin had little effect on platelet adhesion to collagen. We propose that rapid platelet spreading along collagen fibers is both energy- and actin-dependent and necessary to produce maximal adhesion needed to elicit Ca2+ mobilization required for subsequent responses.

Actins↗
PubMed1991

Sodium-calcium exchange in renal epithelial cells: dependence on cell sodium and competitive inhibition by magnesium.

Kinetic properties of Na(+)-Ca2+ exchange in a renal epithelial cell line (LLC-MK2) were assessed by measuring cytosolic free Ca2+ with fura-2 and 45Ca2+ influx. Replacing external Na+ with K+ produced relatively small increases in free Ca2+ and 45Ca2+ uptake unless the cells were incubated with ouabain. Ouabain markedly increased cell Na+ and strongly potentiated the effect of replacing external Na+ with K+ on free Ca2+ and 45Ca2+ uptake. 45Ca2+ influx in 140 mM K+ or N-methyl-D-glucamine minus influx in 140 mM Na+ was used to quantify Na(+)-Ca2+ exchange activity of Na(+)-loaded cells. The dependence of exchange on cell Na+ was sigmoidal; the K0.5 was 26 +/- 3 mmol/liter cell water space, and the Hill coefficient was 3.1 +/- 0.2. The kinetic features of the dependence of exchange on cell Na+ partly account for the small increase in Ca2+ influx when all external Na+ is replaced by K+. Besides raising cell Na+ ouabain appears to activate the exchanger. Magnesium competitively inhibited exchange activity. The potency of Mg2+ was 8.2-fold lower with potassium instead of N-methyl-D-glucamine or choline as the replacement for external Na+. Potassium also increased the Vmax of exchange by 86% and had no effect on the Km for Ca2+. The exchanger does not cause detectable 22Na(+)-Mg2+ exchange and does not appear to require K+ or transport 86Rb+. Although exchange activity was plentiful in the epithelial cells from monkey kidney, others from amphibian, canine, opossum, and porcine kidney had no detectable exchange activity. All of the measured kinetic properties of Na(+)-Ca2+ exchange in the renal epithelial cells are very similar to those of the exchanger in rat aortic myocytes.

Animals↗
PubMed1991

Situational specificity of tolerance to decreased operant responding by morphine and l-nantradol.

In one experiment, key pressing of rats was maintained under a fixed-ratio schedule of food presentation in a first daily session in one environmental situation, and interruption of a photobeam was maintained under a continuous shock avoidance schedule in a second daily session in another environmental situation. After receiving acute injections of the cannabinoid l-nantradol (0.01-0.3 mg/kg), rats received daily administration of a rate-decreasing dose of the drug after the second session, then before the second session, and then before the first session. Tolerance that developed to decreased avoidance responding in the second daily session did not extend to decreased fixed-ratio responding in the first daily session, but was specific to circumstances coinciding with the pharmacological actions of l-nantradol. In a second experiment lever pressing of squirrel monkeys was maintained under an identical fixed-interval schedule of food delivery in two separate daily sessions in different experimental situations. After receiving once-weekly acute injections of morphine (0.3-3.0 mg/kg), monkeys received daily administration of a rate-decreasing dose of morphine in a counter-balanced order before each session. Just as for experiment 1, tolerance that developed in the environment coinciding with the pharmacological actions of morphine did not immediately generalize to operants in the other environmental situation. Instead, tolerance depended on both pharmacologic action as well as concurrently operating behavioral processes.

Analgesics↗
PubMed1991

Situational specificity of tolerance to effects of phencyclidine on responding of rats under fixed-ratio and spaced-responding schedules.

Responding of rats (n = 5) was maintained under DRL (lever) and Time-Delay (nose-key) schedules of food presentation in different experimental chambers during two separate daily sessions. Tolerance that developed to rate-decreasing effects of phencyclidine for nose-key pressing under the Time-Delay schedule did not extend to effects of phencyclidine on lever pressing under the DRL schedule. In a second experiment, both lever and nose-key pressing of rats were maintained under individual and multiple fixed-ratio schedules. One group of animals (n = 5) experienced both the individual and the multiple schedules in the same experimental chamber and another group (n = 5) experienced the individual and the multiple schedules in different experimental chambers. Tolerance that developed to behavioral effects of phencyclidine during the individual schedule did not extend to responding on even the same manipulandum under the multiple schedule in a different experimental chamber. In contrast, tolerance that developed to behavioral effects of phencyclidine during the individual schedule did extend to responding on even the different manipulandum under the multiple schedule in the same experimental chamber. Thus, tolerance that developed in the environment that was coincident with the pharmacologic actions of phencyclidine did not extend to similar operants in a different environmental condition, but did extend even to a different operant and schedule context in the same environmental condition.

Animals↗