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Biomedical subjects

A J Marcus

Publications and source records attributed to A J Marcus.

At least 37 records · Page 2Linked to original sources
PubMed1995

Calreticulin, an antithrombotic agent which binds to vitamin K-dependent coagulation factors, stimulates endothelial nitric oxide production, and limits thrombosis in canine coronary arteries.

Coagulation Factor IX/IXa has been shown to bind to cellular surfaces, and Factor IXa expresses its procoagulant activity by assembling into the intrinsic Factor X activating complex (Factors IXa/VIIIa/X), which also forms on membrane surfaces. This led us to identify cellular proteins which bind Factor IX/IXa; an approximately 55-kDa polypeptide was purified to homogeneity from bovine lung extracts based on its capacity to bind 125I-Factor IX in a dose-dependent and saturable manner. From protein sequence data of the amino terminus and internal peptides, the approximately 55-kDa polypeptide was identified as calreticulin, a previously identified intracellular calcium-binding protein. Recombinant calreticulin bound vitamin K-dependent coagulation factors, 125I-Factor IX, 125I-Factor X, and 125I-prothrombin (Kd values of approximately 2.7, 3.2, and 8.3 nM, respectively), via interaction with its C-domain, although it did not affect the coagulant properties of these proteins. 125I-Calreticulin also bound to endothelial cells in vitro (Kd approximately 7.4 nM), and mouse infusion studies showed an initial rapid phase of clearance in which calreticulin could be localized on the vascular endothelium. Exposure of endothelial cells to calreticulin led to dose-dependent, immediate, and sustained increase in the production of nitric oxide, as measured using a porphyrinic microsensor. In a canine electrically induced thrombosis model, intracoronary infusion of calreticulin (n = 7) prevented occlusion of the left circumflex coronary artery in a dose-dependent manner compared with vehicle-treated controls (n = 5). These results indicate that calreticulin interacts with the endothelium to stimulate release of nitric oxide and inhibit clot formation.

Animals↗
PubMed1995

The value of antegrade stenting for lower ureteric obstruction.

Eleven kidneys in 10 patients (mean age 80 years) presenting over a period of 1 year with obstruction of the lower ureter secondary to malignant obstruction (8 patients), stone disease (1 patient), and bladder wall hypertrophy (1 patient) underwent nephrostomy with subsequent immediate or delayed antegrade stenting. An antegrade placement of the stent was achieved in 10 of the 11 kidneys (90% success). In three cases a combined procedure was performed with cystoscopic assistance. In most cases there was rapid improvement of renal function following stenting. Insertion of ureteric stents in patients admitted to hospital with malignant ureteric obstruction allowed treatment of their renal failure and subsequent discharge without use of permanent external urinary diversion appliances. No significant complications of the procedure were encountered. Medium term follow-up of patients stented for malignant ureteric obstruction suggests that the stents can be left in place without replacement for up to 20 months.

Aged↗
PubMed1995

Thrombosis and inflammation as multicellular processes: significance of cell-cell interactions.

Platelet activation as a result of vascular injury provokes endothelial cells to respond in a manner which limits or reverses the occlusive consequences of platelet accumulation. If the agonistic forces are strong, platelet accumulation is irreversible. In vitro data from our laboratory have repeatedly demonstrated that platelets become unresponsive to all agonists when in proximity to endothelial cells. This unresponsiveness is due to at least three separate endothelial "thromboregulatory" systems: eicosanoids, endothelium-derived relaxing factor (EDRF/NO), and most importantly an endothelial cell ecto-nucleotidase which metabolizes released platelet adenosine diphosphate (ADP) with consequent restoration of platelets to the resting state. This nucleotidase is operative in the complete absence of EDRF/NO and eicosanoids, indicating that the latter two are dispensable thromboregulators. We have solubilized the human endothelial cell ectoADPase, as well as that from placental tissue. Candidate proteins from a purified ADPase fraction are now being studied in further detail. An understanding of the molecular biology of the ADPase gene may lead to development of therapeutic agents such as soluble forms of the enzyme as well as approaches toward up-regulation of ectoADPase activity. This could result in "early thromboregulation", i.e. prevention and/or reversal of platelet accumulation at sites of vascular damage via immediate metabolic removal of the prime platelet agonist-ADP.

Adenosine Diphosphate↗
PubMed1994

Cardiac preservation is enhanced in a heterotopic rat transplant model by supplementing the nitric oxide pathway.

Nitric oxide (NO) is a novel biologic messenger with diverse effects but its role in organ transplantation remains poorly understood. Using a porphyrinic microsensor, the first direct measurements of coronary vascular and endocardial NO production were made. NO was measured directly in the effluent of preserved, heterotopically transplanted rat hearts stimulated with L-arginine and bradykinin; NO concentrations fell from 2.1 +/- 0.4 microM for freshly explanted hearts to 0.7 +/- 0.2 and 0.2 +/- 0.08 microM for hearts preserved for 19 and 38 h, respectively. NO levels were increased by SOD, suggesting a role for superoxide-mediated destruction of NO. Consistent with these data, addition of the NO donor nitroglycerin (NTG) to a balanced salt preservation solution enhanced graft survival in a time- and dose-dependent manner, with 92% of hearts supplemented with NTG surviving 12 h of preservation versus only 17% in its absence. NTG similarly enhanced preservation of hearts stored in University of Wisconsin solution, the clinical standard for preservation. Other stimulators of the NO pathway, including nitroprusside, L-arginine, or 8-bromoguanosine 3',5' monophosphate, also enhanced graft survival, whereas the competitive NO synthase antagonist NG-monomethyl-L-arginine was associated with poor preservation. Likely mechanisms whereby supplementation of the NO pathway enhanced preservation included increased blood flow to the reperfused graft and decreased graft leukostasis. NO was also measured in endothelial cells subjected to hypoxia/reoxygenation and detected based on its ability to inhibit thrombin-mediated platelet aggregation and serotonin release. NO became undetectable in endothelial cells exposed to hypoxia followed by reoxygenation and was restored to normoxic levels on addition of SOD. These studies suggest that the NO pathway fails during preservation/transplantation because of formation of oxygen free radicals during reperfusion, which quench available NO. Augmentation of NO/cGMP-dependent mechanisms enhances vascular function after ischemia and reperfusion and provides a new strategy for transplantation of vascular organs.

Amino Acid Oxidoreductases↗
PubMed1993

Thromboregulation: multicellular modulation of platelet reactivity in hemostasis and thrombosis.

Blood platelets represent the first line of host defense when normal vessels are injured. Platelet adhesion to subendothelium, aggregation, and further platelet recruitment culminate in hemostatic plug formation, which is accompanied by the consolidating effect of fibrin deposition on and between platelets. The process is multicellular in that erythrocytes promote and neutrophils inhibit platelet plug formation. Endothelial cells in proximity possess three protective mechanisms (thrombo-regulators) for limiting the size of the hemostatic plug-ADPase, eicosanoids, endothelium-dependent relaxing factor/NO. We propose that in advanced atherosclerotic blood vessels such as coronary arteries, an ulcer or fissure in the fibrous cap of the atheroma serves as an agonist that transforms the platelet into a major prothrombotic offender. Induction of excessive platelet activation overcomes the normal thromboregulatory mechanisms. Erythrocytes further activate platelets, even in the presence of aspirin, and neutrophil blockage of platelet reactivity is insufficient to prevent impending vascular occlusion. Appreciating that multiple cell types and metabolic pathways are involved in modulation of platelet reactivity in vascular occlusion is a relatively recent concept. Strategies designed to restore processes such as thromboregulation may serve to improve therapeusis in thrombosis, which at present is far from optimal.

Animals↗
PubMed1993

Dietary n-3 fatty acids accelerate catabolism of leukotriene B4 in human granulocytes.

Addition of n-3 fatty acids to a human diet for more than 3 weeks lowers levels of the powerful proinflammatory compound, leukotriene (LT) B4. This can be shown ex vivo after stimulation of human granulocytes with ionophore A23187. In a controlled, randomized, observer-blind study in 14 human volunteers, we investigated the effect of adding 7 g/day of a 85% n-3 fatty acid concentrate to the diet of 7 volunteers (7 served as controls). Levels of LTB4, 20-OH-LTB4, 20-COOH-LTB4 as well as LTB5, 20-OH-LTB5 and 20-COOH-LTB5 were measured by high-pressure liquid chromatography (HPLC) after stimulation and extraction of a platelet-free granulocyte preparation (92% neutrophils). LTB5 and 20-COOH-LTB5 were only detected during n-3 fatty acids, when 20-OH-LTB5 increased from trace amounts to substantial quantities. Importantly, levels of catabolites of LTB4, i.e., 20-OH-LTB4 and 20-COOH-LTB4 were not significantly altered throughout the study. However, the level of LTB4 itself was reduced dramatically after 6 weeks (less so after 1 week) of dietary n-3 fatty acid administration. These data demonstrate that during dietary n-3 fatty acids levels of LTB4 are lowered by a combination of accelerated catabolism and diminished LTB4 generation. This newly observed mechanism of increased LT catabolism may be mediated via induction of peroxisomal enzymes catabolizing leukotrienes B.

Adult↗
PubMed1993

The radiological investigation of occult ectopic ACTH-dependent Cushing's syndrome.

The radiological features of 14 patients with Cushing's syndrome due to 'occult' ectopic ACTH syndrome have been reviewed. In 11 of the 14 patients (79%), the ACTH-producing tumour was located in the thorax. Bronchial carcinoid tumours (eight patients) were the single most common source of 'occult' ectopic ACTH production with thymic carcinoid tumours (two patients) and mediastinal metastases from a medullary carcinoma of the thyroid gland (one patient) accounting for the other intrathoracic tumours. The tumours were usually small with five of the eight bronchial carcinoid tumours measuring between 4 and 10 mm in diameter. Since it remains difficult to distinguish between pituitary-dependent Cushing's disease and 'occult' ectopic ACTH-dependent Cushing's syndrome both clinically and biochemically, the role of radiology remains vital.

ACTH Syndrome, Ectopic↗
PubMed1993

Downregulation of human platelet reactivity by neutrophils. Participation of lipoxygenase derivatives and adhesive proteins.

Unstimulated neutrophils inhibited activation and recruitment of thrombin- or collagen-stimulated platelets in an agonist-specific manner. This occurred under conditions of close physical cell-cell contact, although biochemical adhesion between the cells as mediated by P-selectin was not required. Moreover, in the presence of monoclonal P-selectin antibodies that blocked biochemical platelet-neutrophil adhesion, thrombin-stimulated platelets were more efficiently downregulated by neutrophils. This suggested a prothrombotic role for P-selectin under these circumstances. The neutrophil downregulatory effect on thrombin-stimulated platelets was amplified by lipoxygenase inhibition with 5,8,11,14-eicosatetraynoic acid. In contrast, the neutrophil inhibitory effect on platelets was markedly reduced by platelet-derived 12S-hydroxy-5,8-cis, 10-trans, 14-cis-eicosatetraenoic acid (12S-HETE), as well as by the platelet-neutrophil transcellular product, 12S,20-dihydroxy-5,8,10,14-eicosatetraenoic acid (12S,20-DiHETE), but not by another comparable metabolite, 5S,12S-dihydroxy-6-trans, 8-cis, 10-trans, 14-cis-eicosatetraenoic acid (5S,12S-DiHETE), or the neutrophil-derived hydroxy acid leukotriene B4. The neutrophil downregulatory effect on thrombin-induced platelet reactivity was enhanced by aspirin treatment. This may represent a novel action of aspirin as an inhibitor of platelet function. These results provide in vitro biochemical and functional evidence for the thromboregulatory role of neutrophils and emphasize the multicellular aspect of hemostasis and thrombosis.

Adenosine Diphosphate↗
PubMed1993

Vascular transcellular signaling.

Thus, it is apparent that humoral factors released during inflammation can affect cholesterol metabolism in arterial cells during atherogenesis. These humoral factors released from the macrophage, endothelium, or smooth muscle can modify the cytokine/growth factor/eicosanoid network in the vessel wall in either a paracrine or autocrine manner (6, 40). We also postulate that this could result in alterations in native LDL induced by endothelium (6, 40). Therefore, regulation of the cytokine/growth factor network by eicosanoids may represent an important aspect of arterial responsiveness to injury, as well as progression of intimal hyperplasia and CE deposition in a setting of inflammatory cell activation. Recent understanding of the biochemistry of eicosanoids and the metabolic consequences of these biological response modifiers has helped us to further develop this concept as it relates to mechanisms involving cholesterol delivery and trafficking within the vessel wall during thrombo-atherosclerosis. In this review, we have attempted to highlight recent data which support our classification system for cell-cell interactions, and document that eicosanoids and cytokines released from one cell can activate corresponding receptors on neighboring cells. They can interact with each other in this "cross talk" phenomenon during transmembrane signaling. Recent evidence demonstrating that phosphorylation reactions involving protein kinases A and C and tyrosine protein kinase, coupled with the highly regulated eicosanoid pathway and the DAG-phosphatidylinositol system, appears to have a major impact in our understanding of at least three processes related to atherogenesis: 1) cholesterol delivery, 2) intracellular cholesterol processing, and 3) cholesterol efflux. Identification of these diverse pathways associated with transmembrane signaling have helped us to define processes related to thrombosis since they share common pathways in a complex arteriopathy during atherogenesis.

Arteriosclerosis↗
PubMed1992

The pre-ulcerative phase of carrageenan-induced colonic ulceration in the guinea-pig.

The pre-ulcerative phase of carrageenan-induced colonic ulceration was investigated in guinea-pigs supplied 3% degraded carrageenan as an aqueous solution as drinking fluid for 2 or 3 days during which no ulceration of the bowel was observed with the naked eye or dissecting microscope. Mucosal microscopic changes, from caecum to rectum, were multifocal and included cellular infiltrates, dilatation of glands, crypt abscesses, micro-ulcers and sulphated polysaccharide in the lamina propria. Sulphated polysaccharide was also demonstrated histologically for the first time within the surface epithelium and showed ultrastructural features similar to carrageenan. The results indicate that colonic epithelium in the guinea-pig is capable of macromolecular absorption. Carrageenan, a highly active polyanionic electrolyte, within the surface epithelial cells is most likely a primary factor in the breakdown of mucosal integrity. Macromolecular absorption causing enteropathy of the large bowel is a new pathophysiological concept which may have implications in man, particularly in the pathology of large bowel disease.

Animals↗
PubMed1991

Inhibition of human platelet reactivity by endothelium-derived relaxing factor from human umbilical vein endothelial cells in suspension: blockade of aggregation and secretion by an aspirin-insensitive mechanism.

To determine a role for endothelium-derived relaxing factor/nitric oxide (EDRF/NO) in regulation of human platelet reactivity by human endothelial cells (EC), we studied combined suspensions of human umbilical vein endothelial cells (HU-VEC, passage 2 through 3) and washed human platelets. Confluent HUVEC monolayers were treated with aspirin (1 mmol/L) to prevent prostacyclin (PGI2) formation, washed, and harvested. Aspirin-treated platelets alone (58 x 10(6)) were fully aggregated by thrombin at 0.05 U/mL or more. In the presence of 10(6) HUVEC, however, platelet serotonin release and aggregation in response to thrombin at doses as high as 0.5 U/mL were blocked. We demonstrated for the first time that inhibition of aggregation and serotonin release, due to EDRF/NO, occurred in parallel. HUVEC-dependent inhibition of platelet responsiveness was enhanced by superoxide dismutase (SOD) and reversed by hemoglobin. The inhibitory effect was also reversed by preincubation of HUVEC with NG-monomethyl-L-arginine (NMA) or NG-nitro-L-arginine (NNA) through competitive blockade of arginine metabolism. Pretreatment of platelets with methylene blue indicated that EC-dependent inhibition of platelet reactivity occurred through activation of platelet soluble guanylate cyclase. When platelets and HUVEC were separated by a permeable membrane and both cells were stimulated by thrombin, platelets remained unresponsive. This indicated that inhibition was induced by a fluid-phase mediator, independent of direct cell-cell contact. These data demonstrate that EDRF/NO formation from L-arginine by human EC plays an important role as an aspirin-insensitive fluid-phase inhibitor of human platelet reactivity.

Arginine↗
PubMed1991

Erythrocytes metabolically enhance collagen-induced platelet responsiveness via increased thromboxane production, adenosine diphosphate release, and recruitment.

Erythrocytes promoted platelet reactivity in a plasma medium, as demonstrated in an in vitro system that independently evaluated the biochemistry of platelet activation and recruitment. The prothrombotic erythrocyte effects were metabolically regulated, as evidenced by lack of activity of ATP-depleted or glutaraldehyde-fixed erythrocytes. They occurred in the absence of cell lysis as verified by lactate dehydrogenase assays, and had an absolute requirement for platelet activation. The presence of erythrocytes induced a twofold increase in platelet thromboxane B2 (TXB2) synthesis upon collagen stimulation, indicating that erythrocytes modulated platelet eicosanoid formation. Cell-free releasates from stimulated platelet-erythrocyte suspensions, which exhibited increased recruiting capacity, contained 6.9-fold more ADP and 4.9-fold more ATP than releasates from stimulated platelets alone. Following aspirin ingestion, TXB2 formation was blocked, but erythrocyte promotion of platelet reactivity persisted at those doses of collagen that reinduced platelet activation. Moreover, when platelet mixtures consisted of as little as 10% obtained before aspirin plus 90% obtained post-aspirin ingestion, significant erythrocyte enhancement of platelet reactivity occurred, even at low agonist concentrations. These erythrocyte effects would decrease the therapeutic potential of inhibition of platelet cyclooxygenase by aspirin. The erythrocyte-induced modulation of platelet biochemistry and function emphasizes the importance of cell-cell interactions in stimulus-response coupling.

Adenosine Diphosphate↗
PubMed1991

Enhancement of platelet reactivity and modulation of eicosanoid production by intact erythrocytes. A new approach to platelet activation and recruitment.

Erythrocytes are known to influence hemostasis. Bleeding times are prolonged in anemia and corrected by normalizing the hematocrit. We now demonstrate that intact erythrocytes modulate biochemical and functional responsiveness of activated platelets. A two-stage procedure, permitting studies of cell-cell interactions and independently evaluating platelet activation and recruitment within 1 min of stimulation, was developed. Erythrocytes increased platelet serotonin release despite aspirin treatment, enzymatic adenosine diphosphate removal, protease inhibition, or combinations thereof. The data suggested that erythrocyte enhancement of platelet reactivity can reduce the therapeutic effectiveness of aspirin. Erythrocytes metabolically modified platelet arachidonate or eicosapentaenoate release and eicosanoid formation. They promoted significant increases in cyclooxygenase and lipoxygenase metabolites upon platelet stimulation with collagen or thrombin. However, with ionophore, erythrocytes strongly reduced platelet lipoxygenation. These erythrocyte modulatory effects were stimulus-specific. Activated platelet-erythrocyte mixtures, with or without aspirin, promoted 3-10-fold increases in extracellular free fatty acid, which would be available for transcellular metabolism. Erythrocyte-induced increases in free eicosapentaenoate may contribute to antithrombotic and anti-inflammatory effects of this fish oil derivative. These results provide biochemical insight into erythrocyte contributions to thrombosis and hemostasis, and support the concept of thrombus formation as a multicellular event.

Adenosine Diphosphate↗
PubMed1991

Inhibition of platelet function by an aspirin-insensitive endothelial cell ADPase. Thromboregulation by endothelial cells.

We previously reported that platelets become unresponsive to agonists when stimulated in combined suspension with aspirin-treated human umbilical vein endothelial cells. Inhibition occurred concomitant with metabolism of platelet-derived endoperoxides to prostacyclin by endothelial cells. We now demonstrate that if aspirin-treated platelets which fully respond to appropriate doses of agonists are exposed to aspirin-treated endothelial cells, they remain unresponsive despite absence of prostacyclin. Platelet inhibition is due in large part to ecto-ADPase activity on the endothelial cells. This was established by incubating aspirin-treated endothelial cells with 14C-ADP. Radio-thin layer chromatography and aggregometry demonstrated that 14C-ADP and induction of platelet activation decreased rapidly and concurrently. AMP accumulated transiently, was further metabolized to adenosine, and deaminated to inosine. The apparent Km of the endothelial cell ADPase was 33-42 microM and the Vmax 17-43 nmol/min per 10(6) cells, values in the range of antithrombotic potential. Thus, at least three complementary systems in human endothelial cells control platelet responsiveness: a cell-associated, aspirin-insensitive ADPase which functions in parallel with fluid phase autacoids such as the aspirin-inhibitable eicosanoids, and the aspirin-insensitive endothelium-derived relaxing factor.

Adenosine Diphosphate↗