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Biomedical subjects

J M Gerrard

Publications and source records attributed to J M Gerrard.

At least 73 records · Page 4Linked to original sources
PubMed1989

Production of inositol-phospholipid-derived second messengers in a rat natural killer cell line exposed to susceptible tumor targets.

It is currently believed that natural killer (NK) cells kill bound target cells by exocytosis of cytotoxic granules via a calcium-dependent process. After confirming that NK-mediated killing was indeed dependent upon extracellular calcium, we investigated the production of inositol-phospholipid-derived second messengers in a rat NK cell line, RNK, upon exposure to susceptible target cells. These messengers, inositol trisphosphate (IP3) and diacylglycerol (DAG), are associated with calcium-dependent secretory processes in a number of cell types. When RNK cells were exposed to susceptible YAC-1 tumor targets significant amounts of both IP3 and DAG were produced. The levels of the membrane phospholipid parent molecules of these second messengers declined in similarly stimulated RNK cells over a comparable time period. Using three different target cell lines, it was found that the levels of DAG that RNK produced in response to the different targets followed the same rank order as their susceptibility to RNK-mediated lysis. These data suggest that IP3 and DAG are produced in NK cells in response to tumor target cells, and these second messengers may have a functional role in NK-mediated killing.

Animals↗
PubMed1989

A reassessment of the bleeding time: association of age, hematocrit, platelet function, von Willebrand factor, and bleeding time thromboxane B2 with the length of the bleeding time.

In order to provide an overview of the relative contribution of platelet, von Willebrand factor, and other abnormalities to patients with clinical bleeding difficulties, we performed a retrospective survey of coagulation studies on 569 individuals referred to the University of Manitoba coagulation laboratory because they, or a closely related family member, showed clinical evidence of a bleeding disorder. There was a highly significant (p less than 0.001) negative correlation between the bleeding time and each of the following parameters: the platelet count; the hematocrit; the percent aggregation to collagen, epinephrine, ADP, and arachidonic acid; and the logarithm of von Willebrand factor antigen and a measure of its activity (ristocetin cofactor). A significant and independent inverse relationship between the length of the bleeding time and the extent of platelet adhesion to glass beads, patient age, and prothrombin consumption were also observed. Multivariate analysis of the ability of all parameters to predict the bleeding time showed an r2 of only 0.33. Bleeding time thromboxane B2, in a second smaller study of 70 patients, showed a negative correlation with the length of the bleeding time (p = 0.0001), and, when used together with the above parameters, significantly enhanced the ability to predict the length of the bleeding time (r2 = 0.55). Defects in platelet function, as measured in vitro, and significant enough to have an effect on the bleeding time, occurred with greater frequency than defects in von Willebrand factor in the Manitoba patients evaluated.

Adolescent↗
PubMed1988

Growth-promoting activity of desmopressin in murine leukemia cells treated in vitro.

The synthetic vasopressin analogue, desmopressin (dDAVP), has been shown to influence membrane transport of melphalan in murine L5178Y lymphoblasts. Accordingly, the effect of dDAVP on the cytocidal activity of melphalan in L5178Y cells was evaluated. dDAVP did not affect the cytocidal activity of melphalan in these cells, but significantly affected the cloning efficiency of stationary phase or slowly dividing L5178Y cells over a range of concentrations. In particular, stationary phase cells showed an increase in cloning efficiency from 4.3 +/- 0.5% in control cells to 7.0 +/- 0.3% in cells treated with 25 nM dDAVP (P less than 0.001), whereas cells doubling every 26 h showed an increase from 10.8 +/- 1.2% in control cells to 21.0 +/- 2.0% in cells treated with 150 nM dDAVP (P less than 0.001). This phenomenon was associated with significant elevations of 1,2[3H] diacylglycerol after incubation with dDAVP for 9 min (P less than 0.01) and of total [3H]diacylglycerols after incubation for both 3 min (P less than 0.05) and 9 min (P less than 0.02). Within 10 s of treatment with 100 nM dDAVP, there was a marked decrease in the levels of inositol 1,4,5-trisphosphate and inositol 1-phosphate, but subsequently no change was observed for up to 9 min after treatment. We postulate that the increase of diacylglycerol content produced by dDAVP might be primarily from a phosphatidylcholine source and that the growth-promoting activity of desmopressin may be a consequence of activation of protein kinase C.

Animals↗
PubMed1988

Evaluation of the careers of graduates of the University of Manitoba's BSc (Medicine) program.

The careers of graduates who had taken the BSc (Medicine) (BScMed) program at the University of Manitoba, Winnipeg, between 1950 and 1975 were compared with those of matched classmate controls to determine whether the program had any influence on the research careers of the graduates. More BScMed graduates than control subjects chose an academic career (49% v. 21%), achieved specialty certification (83% v. 65%), and obtained grants (51% v. 18%) and personal awards (37% v. 18%). The BScMed graduates also had significantly more publications than the control subjects. Although part of the difference between the two groups may be explained by the tendency of students who were more inclined toward an academic career to enter the BScMed program, it was evident that the program has a substantial effect on promoting the development of clinical investigators.

Adolescent↗
PubMed1988

Correlation of the antiproliferative action of diphenylmethane-derivative antiestrogen binding site ligands with antagonism of histamine binding but not of protein kinase C-mediated phosphorylation.

The nonestrogen receptor-mediated antiproliferative action of antiestrogen binding site (AEBS) ligands, including triphenylethylene antiestrogens and phenothiazines, has been linked to their ability to inhibit protein kinase C (PKC). Recent studies indicate that some diphenylmethane derivatives inhibit growth, are potent AEBS ligands, and antagonize histamine binding at an AEBS-related histamine site different from H1 and H2. Three novel diphenylmethane derivatives, N,N-diethyl-2-[4-(phenylmethyl)phenoxy]ethanamine.HCI (DPPE), 4-decanoyl-DPPE (dec-DPPE), and 4-benzylphenyl decanoate (BPD) were studied in an attempt to determine whether PKC or histamine interactions best correlate with their antiproliferative effects. Platelet aggregation and the phosphorylation of a platelet Mr 47,000 protein (p47) induced by phorbol-12-myristate-13-acetate (PMA) represent two processes mediated by PKC. DPPE inhibits PMA-induced aggregation [50% inhibitory concentration (IC50) = 31.2 +/- 2.4 (SEM) x 10(-6) M] but does not significantly inhibit either PMA-induced phosphorylation of Mr 47,000 protein (IC50 greater than 500 x 10(-6) M), or binding of [3H]phorbol dibutyrate to platelets. dec-DPPE is a more potent inhibitor of PMA-induced platelet aggregation (IC50 = 18.8 +/- 0.7 x 10(-6) M), a weak inhibitor of Mr 47,000 phosphorylation (IC50 = 80-200 x 10(-6) M), but is without effect on [3H]phorbol dibutyrate binding. BPD, which lacks the alkylaminoethoxy side chain necessary for binding to the AEBS/DPPE site, is devoid of anti-PMA effects. These results are compared to the inhibition of [3H]histamine binding in rat cortex membranes (Ki value for DPPE = 0.83 +/- 0.62 x 10(-6) M; Ki value for dec-DPPE = 6.6 +/- 3.5 x 10(-6) M; BPD is inactive) and growth inhibition of MCF-7 cells (IC50 value for DPPE = 4.5 x 10(-6) M; IC50 value for dec-DPPE = 1.5 x 10(-5) M; BPD is ineffective at all concentrations tested). Thus, while dec-DPPE is a more potent inhibitor of PKC-mediated phosphorylation, DPPE is a more potent inhibitor of histamine binding and is correspondingly more antiproliferative than dec-DPPE. The results support a relationship between antagonism of histamine binding and growth inhibition but argue against an association between the antiproliferative effects of DPPE and dec-DPPE and inhibition of PKC. The findings for DPPE suggest that platelet response to PMA, antagonized by diphenylmethane-type AEBS-ligands, may be mediated, at least in part, by mechanisms other than activation of protein kinase C-dependent phosphorylation.

Animals↗
PubMed1988

Prostaglandins as reducing agents: a model of adenylate cyclase activation?

It has been suggested that adenylate cyclase activation involves reduction of a disulfide linkage. Prostaglandin E1 (PGE1), prostaglandin E2 (PGE2), prostaglandin I2 (PGI2) and prostaglandin F2 alpha (PGF2 alpha) were tested for their ability to act as reducing agents with either cytochrome c, or the disulfide 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB), the latter with a catalytic amount of ferric chloride. PGE1, PGE2, and PGI2 significantly reduced cytochrome c while PGF2 alpha did not. PGE1, PGE2 and PGI2 reduced DTNB while PGF2 alpha did not. The results are consistent with the postulate that prostaglandins which are effective in activating adenylate cyclase can act as reducing agents and might be involved in reductive activation of adenylate cyclase.

Adenylyl Cyclases↗
PubMed1988

Phosphorylation of tyrosine enhances its electron transfer capability: a model of redox modulation as oncogene expression?

The influence of tyrosine and o-phosphotyrosine on the transfer of electrons to nitrobluetetrazolium (NBT) was studied. Tyrosine phosphate was found to strongly promote the transfer of electrons from ferrous iron to NBT, while tyrosine was inhibitory. The enhancement of NBT reduction by tyrosine phosphate was blocked by superoxide dismutase (SOD). The results suggest a role for phosphorylated tyrosine residues to promote intracellular redox reactions. We suggest that the role of tyrosine protein kinases in cell proliferation and transformation may be to regulate electron transport in as yet undefined cellular systems. Consistent with a unique role for phosphotyrosine, the other commonly occurring phosphoamino acids, o-phosphoserine and o-phosphothreonine were not effective electron transfer agents.

Electron Transport↗
PubMed1988

Response of human bone marrow progenitor cells to X-rays in vitro.

In 25 out of 33 cases the survival response of myeloid progenitor cells from fresh human bone marrows, X-irradiated in vitro in phosphate-buffered saline at 23 degrees C, was characterized by a two-term exponential relationship, with D0 values of 0.42 +/- 0.19 Gy and 1.38 +/- 0.37 Gy, respectively. In the remaining eight cases for which the colony-forming efficiency was 10 times lower, survival followed a single exponential function with D0 = 1.18 +/- 0.15 Gy. The biphasic response at 23 degrees C became a single exponential response when the temperature at irradiation was 30 or 37 degrees C.

Bone Marrow↗
PubMed1987

Reduction of a metal or disulfide bond associated with the receptor: a general hypothesis for the mechanism of action of adrenergic agents.

Epinephrine and norepinephrine have been found to reduce Fe3+-heme to Fe2+-heme. Evaluation of the influence of these and structurally similar compounds on heme reduction and platelet aggregation led to the concept that epinephrine may exert its effects on platelets by first binding to a receptor and then reducing a nearby membrane heme group to transmit its agonist stimulus. Similar to alpha adrenergic agonists, we now report that beta agonists can reduce heme. Multiplying 1/kd for receptor binding for each agonist times the reduction of heme at 1.5 X 10(-4)M agonist gives a figure for each drug which closely parallels the potency of these drugs on beta 1 agonists (r = 0.994) and on beta 2 agonists (r = 0.853). Epinephrine and isoproterenol were also found to reduce heme in cytochrome c providing evidence that this mechanism could work well in an intact protein. The results are supportive of the hypothesis that all adrenergic agonists first bind to a membrane receptor and then reduce a heme or similar metal group to transmit the activating signal.

Adrenergic alpha-Agonists↗
PubMed1987

Reduction of a disulfide bond by beta-adrenergic agonists: evidence in support of a general "reductive activation" hypothesis for the mechanism of action of adrenergic agents.

Beta-adrenergic agonists, but not antagonists, were found to reduce the disulfide bond of 5,5'-dithiobis-2-nitrobenzoic acid (DTNB). The extent of DTNB reduction was proportional to the intrinsic activity for these agonists. The results suggest a novel mechanism for transmission of the signal when a beta-adrenergic agonist occupies its receptor. We proposed that beta-adrenergic agonists exert their effects to activate the adenylate cyclase by reducing a disulfide bond in the receptor (R) or guanyl nucleotide binding protein (G) component of the adenylate cyclase complex leading to tight binding of GTP to G and activation of G.

Adrenergic beta-Agonists↗
PubMed1987

Glycation of platelet protein in diabetes mellitus: lack of correlation with platelet function.

The relationship of nonenzymatic glycation of platelet proteins to altered platelet function was studied in 33 diabetic patients. Platelets isolated from diabetic patients were glycated to a greater extent than those isolated from nondiabetic controls. No relationship was found between the level of glycation of platelets in diabetics to parameters commonly used to monitor glycemic control (glycated hemoglobin, glycated albumin, fasting blood glucose). Platelets isolated from diabetics did not show an increased level of aggregation and Thromboxane B2 production as compared to nondiabetic controls. No significant relationship was found between the level of glycation and percent aggregation of platelets. The lack of a relationship between glycation and aggregation suggests that the former may not be responsible for the functional changes in platelets seen in diabetics.

Blood Glucose↗
PubMed1987

The influence of lysophosphatidic acid on platelet protein phosphorylation.

Lysophosphatidic acid (LPA) is a lysophospholipid that is produced during thrombin stimulation of platelets, which can promote platelet aggregation. The mechanism of the effect of LPA was explored in normal platelets and in platelets from a patient with a storage pool deficiency (SPD). A comparison with other lysophospholipids showed that only LPA exerted significant effects to cause or potentiate platelet aggregation. Aspirin, an inhibitor of prostaglandin endoperoxide synthetase, had little effect on LPA-induced aggregation, but completely blocked LPA-induced serotonin secretion. LPA also promoted phosphorylation of myosin light chain (MLC), a 47 kilodalton (kDa) protein, and actin-binding protein. Aspirin significantly inhibited the phosphorylation of the 47-kDa and actin-binding proteins at 3-8 min after the addition of LPA, but had no effect on protein phosphorylation within the 1st min and had no significant effect on MLC phosphorylation. In SPD platelets, aspirin partially inhibited both aggregation and phosphorylation of the 47-kDa protein (less than 30% inhibition) and MLC (less than 40% inhibition) at time points of 1 min or less. The addition of ADP to SPD platelets enhanced the LPA response in platelets either pretreated or not pretreated with aspirin. Studies with SPD platelets indicate that thromboxane and secreted ADP contribute to, but are not necessary for, LPA-induced aggregation and phosphorylation. A23187 (a calcium ionophore) and LPA showed some selectivity to promote MLC as opposed to the 47-kDa protein phosphorylation, particularly at low concentrations of agonists and at earlier time points. The protein phosphorylation changes seen are consistent with a role for MLC phosphorylation in the granule centralization promoted with LPA.

Adult↗
PubMed1987

Comparison of arachidonic acid metabolism in nasal polyps and eosinophils.

Nasal polyp tissue is characterized by its frequent infiltration by large numbers of eosinophils. We have studied the metabolism of 14C-labeled arachidonic acid by both nasal polyp tissue and by eosinophils. The major metabolite produced by both is 15-hydroxyeicosatetraenoic acid (HETE) with lesser amounts of 12-HETE. No cyclooxygenase pathway products were found. The 15-lipoxygenase activity of nasal polyps averages 30 times that of normal nasal mucosa or chronically inflamed sinus mucosa. Nasal polyps which contained significant numbers of eosinophils averaged 7 times the 15-lipoxygenase activity of nasal polyps without eosinophilia. Thus, eosinophils appear to be a major source of 15-lipoxygenase activity in nasal polyps.

Arachidonic Acid↗
PubMed1987

Contractile activity of neonatal platelets.

Platelet contractile activity was evaluated by observation of tension development during isometric contraction of platelet-fibrin clots. Cylindrical clots were made with platelet-rich plasma obtained from cord blood or from adult controls. These clots were allowed to contract isometrically at 37 degrees C while attached to a transducer to record tension development. The rate of tension development was dependent on platelet concentration but was equivalent for neonatal and adult platelet clots. Although abnormalities in neonatal platelet aggregation and secretion have been well documented the platelet functions required for clot contraction such as fibrin binding and actin-myosin interaction appear to be intact in neonatal platelets.

Adult↗
PubMed1986

Differential effects of spermine on aggregation, inositol phosphate formation and protein phosphorylation in human platelets in response to thrombin, arachidonic acid and lysophosphatidic acid.

Platelet aggregation stimulated by thrombin, arachidonic acid or lysophosphatidic acid is associated with rapid phosphorylation of two platelet proteins, myosin light chain and a 47 kDa protein. The polyamine, spermine, inhibited platelet aggregation stimulated by all three agents. Spermine inhibited thrombin-stimulated phosphorylation of myosin light chain and the 47 kDa proteins as well as thrombin-induced production of the inositol phosphates and phosphatidic acid. In contrast, spermine did not inhibit phosphorylation of either protein or the formation of inositol phosphates and phosphatidic acid in response to arachidonic acid or lysophosphatidic acid. Although spermine has been demonstrated to inhibit both phosphatidylinositol-specific phospholipase C and calcium-dependent protein kinases in cell free systems, these results suggest that, in the intact platelet, spermine does not directly inhibit these enzymes. Inhibition of aggregation stimulated by arachidonic acid and lysophosphatidic acid is secondary to interference with platelet-platelet interaction but not with platelet activation. In contrast, spermine inhibits thrombin-induced platelet activation. This thrombin-specific inhibition may be related to interference with the binding of thrombin to its receptor or to its catalytic substrate on the cell surface.

Arachidonic Acid↗