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Biomedical subjects

J B Smith

Publications and source records attributed to J B Smith.

At least 235 records · Page 13Linked to original sources
PubMed1987

Evidence for the release of arachidonic acid through the selective action of phospholipase A2 in thrombin-stimulated human platelets.

The release of arachidonic acid from thrombin-stimulated platelets can be attributed to the action of phospholipase A2 on membrane phospholipid. Previously, analysis of individual subclasses of phospholipid demonstrated that 1-acyl-2-[3H]arachidonoyl-sn-glycerophosphocholine and to a lesser degree 1-acyl-2-[3H]arachidonoyl-sn-glycerophosphoethanolamine were the main source of [3H]arachidonic acid in thrombin-stimulated cells. In the present work, 1,2-diacyl phospholipid subclasses were analyzed as 1,2-diacylglycerobenzoates by high-pressure liquid chromatography in order to analyze arachidonate release as mass changes in individual molecular species of phospholipid. Following thrombin stimulation (5 U/ml, 5 min, 37 degrees C) all arachidonoyl-containing molecular species of 1,2-diacyl-sn-glycerophosphocholine decreased in mass and [3H]arachidonate content by almost 50%, while those of 1,2-diacyl-sn-glycerophosphoethanolamine decreased by 20%. The mass change was substantial and indicated that these phospholipids are a major source of arachidonate in stimulated cells. No variation was seen in the other non-arachidonate-containing molecular species of either subclass. Thus, deacylation of membrane 1,2-diacylglycerophosphocholine and 1,2-diacylglycerophosphoethanolamine by phospholipase A2 is selective for those molecular species of phospholipid containing arachidonic acid, suggesting that a certain proportion of arachidonoyl-containing molecular species of phospholipid are compartmentalized with the platelet membrane proximal to the site of action of this enzyme. These studies demonstrate that the human platelet is a cell poised and specialized to release rapidly substantial amounts of arachidonic acid upon stimulation.

Arachidonic Acid↗
PubMed1987

Formation and metabolism of inositol 1,4,5-trisphosphate in human platelets.

1. myo-[3H]Inositol 1,4,5-trisphosphate [Ins(1,4,5)P3], when added to lysed platelets, was rapidly converted into [3H]inositol 1,3,4,5-tetrakisphosphate [Ins(1,3,4,5)P4], which was in turn converted into [3H]inositol 1,3,4-trisphosphate [Ins(1,3,4)P3]. This result demonstrates that platelets have the same metabolic pathways for interconversion of inositol polyphosphates that are found in other cells. 2. Labelling of platelets with [32P]Pi, followed by h.p.l.c., was used to measure thrombin-induced changes in the three inositol polyphosphates. Interfering compounds were removed by a combination of enzymic and non-enzymic techniques. 3. Ins(1,4,5)P3 was formed rapidly, and reached a maximum at about 4 s. It was also rapidly degraded, and was no longer detectable after 30-60 s. 4. Formation of Ins(1,3,4,5)P4 was almost as rapid as that of Ins(1,4,5)P3, and it remained detectable for a longer time. 5. Ins(1,3,4)P3 was formed after an initial lag, and this isomer reached its maximum, which was 10-fold higher than that of Ins(1,4,5)P3, at 30 s. 6. Comparison of the intracellular Ca2+ concentration as measured with fura-2 indicates that agents other than Ins(1,4,5)P3 are responsible for the sustained maintenance of a high concentration of intracellular Ca2+. It is proposed that either Ins(1,3,4)P3 or Ins(1,3,4,5)P4 may also be Ca2+-mobilizing agents.

Blood Platelets↗
PubMed1987

Bryostatins mimic the effects of phorbol esters in intact human platelets.

Bryostatin-7 induces aggregation of human platelets and the phosphorylation of specific platelet proteins. Both the rate and extent of aggregation are similar to that induced by the tumor promoter phorbol ester 12-myristate 13-acetate (PMA); however, the rate of response is markedly reduced compared to that induced by thrombin. The addition of bryostatin-7 to 32P-labeled platelets results in a time-dependent incorporation of 32P into proteins of 20, 47 and 250 kDa; proteins of similar molecular mass are phosphorylated in response to the addition of thrombin or PMA. The time courses and dose responses of the phosphorylations induced by bryostatin-7 are similar to those found with PMA. In addition, bryostatin-7 increases the level of 32P incorporation into platelet polyphosphoinositides, which also occurs in response to PMA. These results suggest that, in intact human platelets, bryostatin-7 mimics the phorbol ester tumor promoter by directly activating protein kinase C.

Blood Platelets↗
PubMed1987

Preliminary evidence for genetic determination of intraerythrocytic sodium concentration in Utah pedigrees.

Intraerythrocytic sodium concentration has been reported to be increased in hypertensive individuals and some of their normotensive relatives. We investigated the "familiality" of this trait in 572 nonhypertensive subjects from 34 Utah kindreds. Most of these kindreds were selected because of a high incidence of hypertension, heart attack, or stroke. Intraerythrocytic sodium concentration was correlated with the sex of the subject and suprailiac skinfold thickness. Intraerythrocytic sodium concentration was adjusted by linear regression for these 2 factors. The residual values were positively correlated in parent-offspring pairs and among sibs, both those presumed to be living together and those presumed to be living apart. The spouse-spouse correlation was not significant. Pedigree analysis suggested a mixed major-gene/polygenic model of inheritance, but these data did not allow us to distinguish between dominant and recessive inheritance for the major gene component. Total heritability due to both major-gene and polygenic components was estimated to be 90-95%; the proportion of the total variance due to polygenes was estimated to be 60-75%. These results suggest that further study of the relationship between the inheritance of intraerythrocytic sodium and the pathophysiology of hypertension is warranted.

Epidemiologic Methods↗
PubMed1987

Familial correlations from genes and shared environment for urine, plasma, and intraerythrocytic sodium.

Spouse-spouse, sib-sib, and parent-offspring correlations were calculated for urinary, plasma, and intracellular sodium levels on over 1,900 persons aged 3-86 years in 98 Utah kindreds. For 36 hours prior to their clinic visit, 31% of the sample was salt-loaded with salt tablets, while the rest followed their normal diet. For those on their normal diet, urine creatine-, age-, and sex-adjusted urinary sodium excretion from a timed 12-hour overnight sample showed similar and significant correlations between spouses (r = .29), sibs less than 20 years old (r = .38), and parent-offspring pairs for offspring less than 20 years old (r = .29). This contrasted with the lower correlations between sibs 20 years of age and older (r = .10) and parent-offspring pairs for offspring 20 years of age and older (r = .13), presumed to live in different households. Adult plasma sodium sib-sib (r = .13) and parent-offspring (r = .15) correlations were similar to the urinary sodium correlations, while the spouse-spouse (r = .48), the sib-sib (r = .64), and the parent-offspring (r = .63) correlations for those presumed to live in the same household nearly doubled. Intracellular sodium correlations for the adult sibs (r = .32) and offspring (r = .36) were over twice as large as for urinary or plasma sodium, although the spouse-spouse correlation (r = .37) remained large also.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1987

Anticardiolipin antibodies in patients with rheumatic diseases.

Recent attention has focused on the presence of anticardiolipin (ACL) antibodies and their possible role in recurrent thrombosis and abortions in patients with systemic lupus erythematosus. We analyzed ACL antibodies in 243 consecutive patients to determine their frequency in patients with different rheumatic disorders. A significantly elevated frequency was found in patients with systemic lupus erythematosus (38%), rheumatoid arthritis (33%), and psoriatic arthritis (28%). No correlation could be found between ACL antibody levels and recurrent thrombosis. In patients with rheumatoid arthritis there was a significant correlation between ACL antibodies and a history of repeated abortions. No significant association was found between ACL antibodies and other autoantibodies except in patients with rheumatoid arthritis; ACL antibody-positive rheumatoid arthritis patients were much more likely to be antinuclear antibody-positive (P less than 0.0002).

Abortion, Habitual↗
PubMed1987

Effects of fixed-interval duration on the development of tolerance to decreased responding by l-nantradol.

Key pecking of pigeons was maintained under a chained schedule in which the first response after 2 h (fixed interval) was followed by the opportunity to obtain food after sequences of 30 responses (fixed ratio), and animals received acute injections of l-nantradol (0.001-0.1 mg/kg). Smallest doses were ineffective; largest doses decreased all responding; and intermediate doses decreased fixed-interval, but not fixed-ratio, responding. When animals subsequently received daily administration of 0.1 mg/kg l-nantradol prior to experimental sessions, initially decreased fixed-interval responding did not recover during 21 sessions of l-nantradol administration, whereas fixed-ratio responding was demonstrated on several occasions by automatic advancement to the fixed-ratio component. When the schedule was changed so that the same total amount of food was available after ten shorter fixed intervals, fixed-interval responding resumed within a single session, and when the schedule was changed back so that all food was available only after 2 h had elapsed, fixed-interval responding diminished within a few sessions and virtually no responding occurred for 21 additional sessions. The behavioral effects of chronic l-nantradol depended on both the schedule and the parameter of reinforcement.

Animals↗
PubMed1987

Na+/K+/Cl- cotransport in cultured vascular smooth muscle cells: stimulation by angiotensin II and calcium ionophores, inhibition by cyclic AMP and calmodulin antagonists.

The specific activity of the Na+/K+/Cl cotransporter was assayed by measuring the initial rates of furosemide-inhibitable 86Rb+ influx and efflux. The presence of all three ions in the external medium was essential for cotransport activity. In cultured smooth muscle cells furosemide and bumetanide inhibited influx by 50% at 5 and 0.2 microM, respectively. The dependence of furosemide-inhibitable 86Rb+ influx on external Na+ and K+ was hyperbolic with apparent Km values of 46 and 4 mM, respectively. The dependence on Cl was sigmoidal. Assuming a stoichiometry of 1:1:2 for Na+/K+/Cl-, a Km of 78 mM was obtained for Cl. In quiescent smooth muscle cells cotransport activity was approximately equal to Na+ pump activity with each pathway accounting for 30% of total 86Rb+ influx. Growing muscle cells had approximately 3 times higher cotransport activity than quiescent ones. Na+ pump activity was not significantly different in the growing and quiescent cultures. Angiotensin II (ANG) stimulated cotransport activity as did two calcium-transporting ionophores. A23187 and ionomycin. The removal of external Ca2+ prevented A23187, but not ANG, from stimulating the cotransporter. Calmodulin antagonists selectively inhibited 86Rb+ influx via the cotransporter. Beta-adrenoreceptor stimulation with isoproterenol, like other treatments which increase cAMP, inhibited cotransport activity. Cultured porcine endothelial cells had 3 times higher cotransport activity than growing muscle cells. Calmodulin antagonists inhibited cotransport activity, but agents which increase cAMP or calcium had no effect on cotransport activity in the endothelial cells.

Angiotensin II↗
PubMed1987

Resolution of radiolabeled molecular species of phospholipid in human platelets: effect of thrombin.

Resolution of individual molecular species of human platelet 1,2-diradyl-sn-glycero-3-phosphocholines and 1,2-diradyl-sn-glycero-3-phosphoethanolamines by reverse phase high pressure liquid chromatography (HPLC) allowed a thorough analysis of those phospholipids labeled with [3H]arachidonic acid. Approximately 54% and 16% of the total incorporated radiolabel was found in choline glycerophospholipids and ethanolamine glycerophospholipids, respectively, with ca. 90% of this being found in the 1,2-diacyl molecular species. Eighty percent of [3H]-arachidonic acid incorporated into 1-acyl-2-arachidonoyl-sn-glycero-3-phosphocholine in resting platelets was equally distributed between 1-palmitoyl-2-arachidonoyl and 1-stearoyl-2-arachidonoyl-sn-glycero-3-phosphocholine, while 70% of the radiolabel in 1-acyl-2-arachidonoyl-sn-glycero-3-phosphoethanolamine was found in 1-stearoyl-2-arachidonoyl-sn-glycero-3-phosphoethanolamine. Thrombin stimulation (5 U/ml for 5 min) resulted in deacylation of all 1-acyl-2-[3H]arachidonoyl molecular species of 1-acyl-2-arachidonoyl-sn-glycero-3-phosphocholine and 1-acyl-2-arachidonoyl-sn-glycero-3-ethanolamine. There was also a slight increase in 1-O-alkyl-2-[3H]arachidonoyl-sn-glycero-3-phosphocholine and a significant increase in 1-O-alk-1'-enyl-2-[3H]arachidonoyl-sn-glycero-3-phosphoethanolamine molecular species of over 300%. Thus, HPLC methodology indicates that arachidonoyl-containing molecular species of phosphatidylcholine and phosphatidylethanolamine are the major source of arachidonic acid in thrombin-stimulated human platelets, while certain ether phospholipid molecular species become enriched in arachidonate.

Arachidonic Acid↗
PubMed1987

Effects of cyclophosphamide alone and scheduled methotrexate-5-fluorouracil combination chemotherapy on transplantable R-3327 prostatic adenocarcinoma in F1 hybrid male rat.

Male F1 hybrid rats bearing the R-3327 transplantable adenocarcinoma demonstrating similar growth patterns within the original sample of animals were carefully separated into control and treatment groups for each growth pattern. This assured treatment of tumors with similar cell kinetics within each group. In one group, cyclophosphamide (100 mg/kg) was injected intraperitoneally once every four weeks for eight weeks (total of two doses). In the second group, methotrexate (7.5 mg/kg) followed in ninety minutes by 5-fluorouracil (50 mg/kg) were injected intraperitoneally once each week for eight weeks (total of eight doses). These drug dosages did not depress the white cell count of the animals, and they tolerated the medicines well. Excellent suppression of tumor growth was obtained in each group with the cyclophosphamide treatment group being significant at the 0.01 level and the methotrexate-5-fluorouracil treatment group being significant at the 0.05 level.

Adenocarcinoma↗
PubMed1987

Constriction of cat coronary arteries by synthetic thromboxane A2 and its antagonism.

Synthetic thromboxane A2 (TxA2S) induced rapid, concentration-dependent constriction of isolated perfused cat coronary arteries. Its potency was approximately 30 times that of the prostaglandin endoperoxide, PGH2. Pinane thromboxane A2 (PTA2), BM-13.505 and SQ-29,548, compounds previously shown to antagonize the effects of stable prostaglandin endoperoxide analogs, inhibited the constriction induced by TxA2S in a concentration-dependent fashion. These experiments provide further evidence that the oxetane structure proposed for TxA2 is correct and show that compounds that inhibit the effects of prostaglandin endoperoxides also antagonize the effects of TxA2.

Animals↗
PubMed1987

Angiotensin decreases cyclic GMP accumulation produced by atrial natriuretic factor.

Atrial natriuretic factor (ANF) produced rapid increases in cyclic GMP (cGMP) in cultured aortic smooth muscle cells. Angiotensin II (ANG II) markedly decreased the accumulation of cGMP that was evoked by ANF. Arginine vasopressin and ATP, which evoke transient increases in free Ca2+ similarly to ANG II, also inhibited cGMP accumulation. The effect of the calcium mobilizing neurohormones was mimicked by the divalent cation ionophore, A23187. The cyclic nucleotide phosphodiesterase inhibitor, 3-isobutyl-1-methylxanthine, prevented ANG II from inhibiting ANF-evoked cGMP accumulation. ANG II also inhibited cGMP accumulation induced by nitroprusside, a compound that activates cytosolic guanylate cyclase. These findings support the hypothesis that ANG II decreases cGMP accumulation by stimulating cGMP hydrolysis, apparently via a Ca2+-activated cGMP phosphodiesterase.

1-Methyl-3-isobutylxanthine↗
PubMed1987

Receptor-mediated effects of a PGH2 analogue (U 46619) on human platelets.

The specific effects of U 46619 (9,11-dideoxy,9 alpha-11 alpha-methanoepoxyprostaglandin F2 alpha), thromboxane A2-prostaglandin H2 (TxA2/PGH2) analogue, on human platelet shape change, myosin light-chain phosphorylation, serotonin release, fibrinogen receptor exposure, and platelet aggregation were measured and compared with binding of [3H]U 46619 to platelets. Shape change and myosin light-chain phosphorylation were found to be saturable and dose dependent, having effective concentration producing 50% of the maximum response (EC50) values of 0.035 +/- 0.005 and 0.057 +/- 0.021 microM, respectively (mean +/- SE). These two effects were competitively inhibited by specific antagonists of TxA2/PGH2 receptors (BM 13177, PTA-OH, and 1.PTA-OH) indicating that they are receptor mediated. Binding of [3H]U 46619 showed two components. Occupancy of high-affinity binding sites [dissociation constant (Kd) = 0.041 +/- 0.009 microM, maximum binding site (Bmax) = 19.4 +/- 5.3 fmol/10(7) platelets, with 1,166 +/- 310 sites/platelet; n = 12] correlated with platelet shape change and myosin light-chain phosphorylation. We propose that a second component with an apparent Kd of 1.46 +/- 0.47 microM (n = 12) represents a second, low-affinity site. Mean EC50 values for U 46619-induced serotonin release, platelet aggregation, and fibrinogen receptor exposure were 0.54 +/- 0.13. 1.31 +/- 0.34 and 0.53 +/- 0.21 microM, respectively. Therefore, the platelet release reaction was not directly correlated with occupancy of high-affinity receptors but could be related to the second binding component of U 46619. Fibrinogen receptor exposure and platelet aggregation caused by U 46619 appeared to be events mediated by the release of adenosine diphosphate from platelet-dense granules.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗
PubMed1987

Ionophore-stimulated rat basophilic leukemia cells produce PAF-acether.

Ionophore (A23187) stimulated rat basophilic leukemia (RBL-1) cells produce a lipid mediator which caused rabbit platelets to aggregate and which by using platelet-activating factor (PAF)-acether antagonists and high-pressure liquid chromotography was shown to be PAF-acether. Thus RBL-1 cells represent a cell type suitable for studying the coordinated release of three mediators of anaphylaxis: histamine, leukotrienes C4/D4 (slow-reacting substances) and PAF-acether.

Animals↗
PubMed1987

Pharmacology of thromboxane synthetase inhibitors.

Several selective and potent thromboxane synthetase inhibitors have now been described. In this paper I shall summarize their effects on platelet aggregation, endotoxin- and thrombin-induced pulmonary hypertension, the Forssman reaction-induced thrombocytopenia, and the rat tail vein bleeding time. Although any clinical utility of thromboxane synthetase inhibition remains to be demonstrated, it may be useful in conditions where vasoconstriction or vasospasm are involved, particularly where platelet activation is a contributing factor.

Animals↗
PubMed1986

ADP stimulates IP3 formation in human platelets.

Aspirinated human platelets labeled with 32PO4 showed a 1.7-fold increase in [32P]IP3 when stimulated with ADP. ADP-stimulated mobilization of internal Ca2+ and phosphorylation of myosin were enhanced in the presence of extracellular Ca2+ but the increase in IP3 was not significantly affected by external Ca2+. The Ca2+ ionophore, ionomycin, elevated internal Ca2+ and induced myosin phosphorylation without a detectable change in IP3. These results indicate that the mechanism of ADP stimulation of human platelets is similar to that of other platelet agonists and supports the theory that IP3 functions to liberate internal Ca2+.

Adenosine Diphosphate↗