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Biomedical subjects

D Deykin

Publications and source records attributed to D Deykin.

At least 91 records · Page 5Linked to original sources
PubMed1980

Platelet arachidonic acid metabolism and platelet function in ten patients with chronic myelogenous leukemia.

We have compared the pathways of arachidonic acid (C 20:4) metabolism in platelets from ten patients with Philadelphia chromosome-positive CML with those of seven normal subjects. Platelets were incubated with 3H-arachidonic acid, gel-filtered, and treated with thrombin (5 U/ml). The cyclooxygenase and lipoxygenase-derived products and free arachidonic acid released from the platelets were separated by high pressure liquid chromatography and their radioactivity determined. The total uptake of 3H-C 20:4 by platelets from CML patients did not differ from controls, but the release of radioactivity in response to thrombin was significantly lower (p < 0.01) in CML patients (32.3% +/- 4.9% of total radioactivity was released from control platelets; 19.0% +/- 7.4% from CML platelets). Both cyclooxygenase and lipoxygenase-derived products were reduced, but there was no specific pattern of abnormality. Although there was no direct correlation between either the WBC or platelet count and impairment of platelet C 20:4 metabolism, the platelets from three patients with accelerated disease released the lowest total amount of 3H-C 20:4 metabolites. In a single patient, studied before and after successful chemotherapy (hydroxyurea), severe abnormalities in platelet arachidonic acid metabolism returned to normal after treatment.

Adult↗
PubMed1979

Specificity of phospholipases in methylcholanthrene-transformed mouse fibroblasts activated by bradykinin, thrombin, serum, and ionophore A23187.

Methylcholanthrene-transformed mouse fibroblasts synthesize prostaglandins in response to bradykinin, thrombin, serum, and the ionophore A23187. These agents activate phospholipases, thereby releasing fatty acids from phospholipids. To examine the phospholipid specificity of the phospholipases activated by bradykinin, thrombin, serum, and A23187, cells were labeled with [14C]arachidonic acid and stimulated with these agents in the presence of delipidated bovine serum albumin. Phospholipid classes were resolved by two-dimensional chromatography on silica gel-coated paper. Only phosphatidylinositol and phosphatidylcholine lost radioactivity upon stimulation. To characterize the fatty acid specificity of the phospholipases, cells were incubated with 14C-labeled stearic, oleic, linoleic, eicosatrienoic, or arachidonic acid and then exposed to the stimuli. Bradykinin, thrombin, and serum caused specific release of radioactivity into the medium only from cells labeled with arachidonic acid or eicosatrienoic acid, whereas A23187 caused release from cells labeled with any one of the five fatty acids. We conclude that bradykinin, thrombin, and serum activate phospholipases that specifically hydrolyze arachidonyl and eicosatrienoyl phosphatidylinositol and phosphatidylcholine, whereas A23187 is less specific activator of phospholipases.

Animals↗
PubMed1979

Arachidonic acid level in cellular lipids determines the amount of prostaglandins synthesized during cell growth in tissue culture.

Methylcholanthrene transformed mouse fibroblast cells can be induced to synthesize prostaglandins by a short term incubation with various vasoactive agents including serum, bradykinin and thrombin or in response to mechanical detachment from the culture dish. The ability of the cells to synthesize prostaglandins upon stimulation changes during growth of the culture on the dish; the response is maximal on the first day after inoculation and decreased sharply thereafter. Feeding of the cells with fresh growth medium enhances prostaglandin production induced by all stimuli. The difference in the cell response during growth is probably not due to change of prostaglandin synthetase activity since the specific enzyme activities assayed with microsomal preparations of cells harvested from the first and third day culture are similar. However, analysis of the cellular content of arachidonic acid after saponification of the total lipid extract of cells harvested at different days of growth reveals that the level of arachidonic acid per cell during growth is parallel to the response to stimuli. It is maximal on the first day and decreases sharply on the second day and stays low on the third day. Our study suggests that the level of arachidonic acid in the cell governs the extent of prostaglandin synthesis upon stimulation.

Animals↗
PubMed1979

The use of high pressure liquid chromatography (HPLC) for the separation of radiolabeled arachidonic acid and its metabolities produced by thrombin-treated human platelets. I. The validation of the technique.

We have developed a technique for the rapid separation and quantitative collection of thromboxane B2 (TXB2), PGE2, PGD2, PGF2 alpha, 12-hydroxy-5,8,10 heptadecatrienoic acid (HHT), 12-L-hydroxy-5,8,10,14 eicosatetraenoic acid (HETE), and arachidonic acid released from thrombin treated human platelets. Platelets were pre-labeled with 3H-arachidonic acid and then isolated by gel filtration. They were then exposed to thrombin for various intervals and separated by centrifugation. Aliquots of the cell-free medium were applied directly to a high pressure liquid chromatograph containing a fatty acid column as the stationary phase. A quarternary solvent system containing tetrahydrofuran (THF), acetonitrile (CH3CN), water and acetic acid (HOAC) resolved and eluted the arachidonic acid metabolites within 30 minutes. Since no sample preparation is required and since the solvent system does not quench the counting efficiency of a standard liquid scintillation fluor the technique permits rapid separation and quantitation of radiolabeled arachidonic acid and its metabolites.

Arachidonic Acids↗
PubMed1979

The use of high pressure liquid chromatography (HPLC) for the separation of radiolabeled arachidonic acid and its metabolites produced by thrombin-treated human platelets. II. Establishment of optimal assay conditions.

We have utilized HPLC to develop optimal conditions for assaying the transformation of arachidonic acid in thrombin-treated human platelets. In the presence of increasing amounts of albumin, the total amount of radioactivity released from thrombin-treated platelets pre-labeled with 3H-arachidonic acid is first enhanced and then inhibited. Maximal release, reflecting primarily enhanced amounts of free labeled arachidonic acid, occurs at a final albumin concentration of 0.5 mg/ml. Calcium promoted the release of all radiolabeled metabolites, but it specifically enhanced HETE formation and release. Magnesium was without effect. Cyclo-oxygenase derived products constituted the bulk of released label at short time intervals, but after ten minutes exposure to thrombin in the presence of albumin (0.5 mg/ml) and 3 mM calcium, radioactivity in the released products was equally distributed among cyclo-oxygenase derived products (TXB2 + PGD2 + HHT), HETE and free arachidonic acid.

Albumins↗
PubMed1979

Antithrombotic therapy. Rationale and application.

Arterial thrombosis and venous thrombosis differ in pathogenesis, morphology, and response to antithrombotic therapy. Antiplatelet therapy usually is aimed at arterial thrombi, in whose formation platelet-mediated reactions predominate, while anticoagulant therapy is effective against venous thrombi, in whose formation coagulation predominates.

Anticoagulants↗
PubMed1979

The hemostatic defect of chronic liver disease. Kinetic studies using 75Se-selenomethionine.

With the use of cohort labeling with 75Se-selenomethionine, simultaneous platelet, fibrinogen, and plasminogen survival studies were carried out in 8 patients with chronic alcoholic liver disease and in 5 normal subjects. Clinical features, liver function tests, coagulation and fibrinolytic system activities, and platelet function were also assessed. On the basis of platelet survival, the patients could be divided into two groups. Three patients had shortened platelet survival; they were all thrombocytopenic and had greater prolongation of the prothrombin time (PT) and activated partial thromboplastin time (PTT) than the other 5 patients. However, platelet turnover was decreased in all the patients, and there was no difference between the two groups with regard to fibrinogen or plasminogen survival nor in the in vitro evidence of disseminated intravascular coagulation (DIC). Fibrinogen survival was increased in 5 of the 8 patients. Plasminogen survival was normal in 6 patients and prolonged in 2 patients with very low plasminogen levels. The absence of increased fibrinogen turnover in the patients studied indicates that the abnormalities in coagulation tests were not due to consumption coagulopathy. The authors' studies suggest that, at least for patients with chronic stable alcoholic liver disease, the concept that the coagulopathy of liver disease is due to increased utilization of clotting factors should be revised with caution.

Blood Coagulation Disorders↗
PubMed1978

The activation by Ca2+ of platelet phospholipase A2. Effects of dibutyryl cyclic adenosine monophosphate and 8-(N,N-diethylamino)-octyl-3,4,5-trimethoxybenzoate.

Thrombin-induced release of arachidonic acid from human platelet phosphatidylcholine is found to be more than 90% impaired by incubation of platelets with 1 mM dibutyryl cyclic adenosine monophosphate (Bt2 cyclic AMP) or with 0.6 mM 8-(N,N-diethylamino)-octyl-3,4,5-trimethoxybenzoate (TMB-8), an intracellular calcium antagonist. Incorporation of arachidonic acid into platelet phospholipids is not enhanced by Bt2 cyclic AMP. The addition of external Ca2+ to thrombin-treated platelets incubated with Bt2 cyclic AMP or TMB-8 does not counteract the observed inhibition. However, when divalent cation ionophore A23187 is employed as an activating agent, much less inhibition is produced by Bt2 cyclic AMP or TMB-8. The inhibition which does result can be overcome by added Ca2+. Inhibition of arachidonic acid liberation by Bt2 cyclic AMP, but not by TMB-8, can be overcome by high concentrations of A23187. When Mg2+ is substituted for Ca2+, ionophore-induced release of arachidonic acid from phosphatidylcholine of inhibitor-free controls is depressed and inhibition by Bt2 cyclic AMP is slightly enhanced. The phospholipase A2 activity of platelet lysates is increased by the presence of added Ca2+, however, the addition of either A23187 or Bt2 cyclic AMP is without effect on this activity. We suggest that Bt2 cyclic AMP may promote a compartmentalization of Ca2+, thereby inhibiting phospholipase A activity. The compartmentalization may be overcome by ionophore. By contrast, TMB-8 may immobilize platelet Ca2+ stores in situ or restrict access of Ca2+ to phospholipase A in a manner not susceptible to reversal by high concentrations of ionophore.

Arachidonic Acids↗
PubMed1978

Clinical trial of a new bleeding-time device.

The authors report a clinical trial comparing a new bleeding time device (Simplate II) with the Mielke Template. Bleeding times were determined at the same time on the same arm using the two devices. Subjects of the study were 24 healthy volunteers, before and two hours after ingestion of 975 mg aspirin, and 28 patients. For the normal subjects the mean pre-aspirin bleeding times were 4.75 +/- 1.42 minutes (1 SD) with the Simplate II and 3.65 +/- 1.22 minutes with the Mielke Template. The mean bleeding times two hours after ingestion of aspirin were 7.86 +/- 2.76 minutes with the Simplate II and 7.84 +/- 2.94 minutes with the Template. The pre- and the post-aspirin values with the two devices were not significantly different from each other, nor were the bleeding times obtained in the patients with the two devices. The extents of scarring were similar with the two devices. The results were highly reproducible by both methods. The new device was simpler and more rapid to use.

Aspirin↗
PubMed1978

Quantitative abnormality of an Aalpha chain molecular weight form in the fibrinogen of cirrhotic patients.

The molecular weight heterogeneity of fibrinogen from the whole plasma of 12 normal and seven cirrhotic individuals was examined by means of a novel two-dimensional sodium dodecyl suphate (SDS) gel electrophoretic technique. Fibrinogen was first separated from other plasma proteins on a large pore gel, cut out of the gel, reduced, and separated into its component Aalpha, Bbeta and gamma chains on a second gel. Fibrinogen was resolved into two major bands, I and II, on the first gel. The ratio of fibrinogen II to fibrinogens I plus II was approximately 0.3 (range 0.2-0.35) for both normals and cirrhotic patients. Two major molecular weight (mol wt) forms of Aalpha chain were present in normal fibrinogen I: Aalpha/I and/or Aalpha/2, mol wt 7 X 10(4) and 6.7 X 10(4) respectively. Normal fibrinogen II contained either one of these Aalpha chains plus one of the smaller Aalpha chains, Aalpha/6--10, accounting for the 3--4 X 10(4) mol wt difference between bands I and II. Aalpha/2 comprised 33% of the total Aalpha chains (range 27--41%) in normal fibrinogen I and approximately 25% of the Aalpha chains in normal fibrinogen II. In contrast, fibrinogen I from six out of the seven patients contained a lower percentage of Aalpha/2 (range 10--25%). Similarly fibrinogen II from these patients was deficient in Aalpha/2, although the protein contained normal levels of lower mol wt Aalpha derivatives. No correlation was found between per cent fibrinogen II and per cent Aalpha/2 in either normal or cirrhotic subjects. These results suggest that at least two independent processes are responsible for the observed levels of Aalpha heterogeneity in normals and cirrhotics and that the process controlling Aalpha/2 production is a abnormal in cirrhotic individuals. This decrease in Aalpha/2 does not affect the coagulability of fibrinogen. Fibrin monomer aggregation studies indicate that a serum component is, in part, responsible for the abnormally transparent clot formed from the plasma of cirrhotics.

Blood Coagulation Tests↗
PubMed1978

Intermediate syndrome of platelet dysfunction.

We studied eight patients with intermittent bleeding episodes usually following trauma and associated with the ingestion of medicine known to interfere with platelet function. All patients had a normal or minimally prolonged baseline bleeding time. All had a normal platelet count, glass bead retention test, and platelet serotonin content and a variable pattern of abnormalities in prothrombin consumption and platelet factor 3 availability. However, all showed abnormal platelet aggregation reactions using epinephrine, adenosine diphosphate, and collagen. Following the administration of 975 mg aspirin, our patients' bleeding times became prolonged to a greater extent than the bleeding times of normal controls (range 13 to greater than 20 min). Review of the literature showed approximately 5% of "normal" controls had findings similar to those we report. We believe we are describing a group of individuals with an intermediate form of platelet dysfunction. Although their bleeding diathesis is not as severe as that of patients with platelet dysfunction syndromes previoulsy described, they do bleed significantly when subjected to trauma following the ingestion of drugs such as aspirin. We propose that this defect is common and should be screened for. The aspirin tolerance test is a simple test for detecting these patients.

Adolescent↗