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Biomedical subjects

F M Booyse

Publications and source records attributed to F M Booyse.

At least 55 records · Page 3Linked to original sources

Effects of chronic oral consumption of nicotine on the rabbit aortic endothelium.

New Zealand white rabbits (10) were administered daily doses of nicotine (2.4 mg/kg/day) in their drinking water for 25 weeks. Nicotine-treated rabbits were compared with control rabbits (10) in terms of blood serum biochemistry and lipid profiles, blood cells counts, changes in aortic endothelial cell morphologic characteristic and distribution, and vessel wall permeability (Evans blue dye uptake). Fasting serum levels of glucose, triglycerides, total cholesterol, and LDL-cholesterol were elevated in nicotine-treated rabbits. No significant differences (nicotine vs control) were seen in leukocyte, erythrocyte and platelet counts, or hematocrit and hemoglobin. Control and nicotine-treated rabbit aortas showed similar focal areas of increased Evans blue dye uptake; staining was localized primarily to aortic arch areas. Endothelial cells (luminal surface) from non-Evans blue and Evans blue arch areas were examined by a combination of Häutchen preparation (silver-stained vessels) and scanning and transmission electron microscopy. Endothelial cells from nicotine-treated arch areas (Evans-blue-stained) showed extensive changes such as: increased cytoplasmic silver deposition, increased formation of microvilli, and numerous focal areas of "ruffled" endothelium (projections on cell surfaces). These data indicate that nicotine, administered orally to rabbits, has a demonstrable in vivo morphologic effect on endothelial cells in the aortic arch.

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Cultured aortic endothelial cells from pigs with von Willebrand disease: in vitro model for studying the molecular defect(s) of the disease.

Aortic endothelial cells from normal pigs and pigs with von Willebrand disease have been established in long-term cultures. Both cultures appeared similar in terms of general growth characteristics, morphologic features and ultrastructure. Immunofluorescent staining of these cultures with chicken (or rabbit) antiporcine ristocetin-Willebrand factor sera (or IgG) resulted in extensive perinuclear staining of the cells in both cultures. Additionally, staining of semiconfluent cultures of normal cells for ristocetin-Willebrand factor revealed an extensive meshwork of distinct, immunologically identifiable ristocetin-Willebrand factor-containing filaments between cells. Immunoreactive material was considerably decreased and more diffuse between cells in semiconfluent cultures from affected pigs. Through immunocytochemical staining with peroxidase-coupled antiserum, the filaments (of indeterminate length) were found to have a diameter of approximately 300 A. Finally, washed porcine platelets interacted extensively with scrape-damaged cultures of affected endothelial cells. This interaction of platelets with damaged normal cultures was abolished by pretreatment of the cultures with rabbit antiporcine ristocetin-Willebrand factor IgG.

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Comparison of two types of endothelial cells in long term culture.

Bovine aortic endothelial cells have been serially subcultured and maintained for ten months (24 passages) without any apparent loss of endothelial-like properties. There appear to be two endothelial cell types; one type is mononucleated, 40-60 mu in diameter, and the other (giant) cell type, multinucleate and 300-600 mu in diameter. Both cell types were identified as endothelial cells by established criteria; 1. ultrastructurally, by the presence of Weibel-Palade bodies, microfilaments and pinocytotic vesicles and 2. immunologically, by the presence of factor VIII antigen and thrombosthenin.

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Adenosine cyclic 3',5'-monophosphate-dependent protein kinase from human platelets.

A single cyclic AMP-dependent protein kinase (EC 2.7.1.37) has been isolated from human platelets by using DEAE-cellulose ion-exchange chromatography and Sephadex G-150 gel filtration. The molecular weight of the protein kinase was estimated to be 86 490. In the presence of cyclic AMP, the protein kinase could be dissociated into a catalytic subunit of molecular weight 50 000, and either one regulatory subunit of molecular weight 110 000 or two regulatory subunits of molecular weights 110 000 and 38 100, depending on the pH used. Recombination of either of the regulatory subunits with the catalytic subunit restored cyclic AMP-dependency in the catalytic subunit. The apparent Km for ATP in the presence of 10 muM Mg2+ was 4 muM (plus cyclic AMP) and 4.3 muM (minus cyclic AMP). The concentration of cyclic AMP needed for half-maximal stimulation of the protein kinase was 0.172 muM and apparent dissociation constants of 3.7 nM (absence of MgATP) and 0.18 muM (presence of MgATP) were exhibited by the "protein kinase-cyclic AMP complex". The enzyme required Mg2+ for maximum activity and showed a pH optimum of 6.2 with histone as substrate. In addition to four major endogenous platelet protein acceptors of apparent molecular weights 45 000, 28000, 18 500, and 11 100, the platelet protein kinase also phosphorylated the exogenous acceptor proteins thrombin, collagen and histone, all capable of inducing platelet aggregation. Prothrombin, a nonaggregating agent, was not phosphorylated.

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Culture of arterial endothelial cells: characterization and growth of bovine aortic cells.

Arterial endothelial cells were obtained from bovine aortae by mild treatment with collagenase and medium perfusion. These cells were cultured in RPMI-1640 medium containing 15mM Hepes buffer and 35% fetal calf serum at pH 7.35. Essentially all (90-95%) the effluent cells were viable and 80% of these cells attached to the substratum within 1 hour. Small patches of attached cells coalesced to form confluent monolayers in 3-5 days. Confluent monolayers of endothelial cells consisted of a homogeneous population of tightly packed, polygonal cells. Selected cultures were serially subcultured (trypsin-EDTA) for 12-14 months (30-35 passages) without any apparent change in morphology or loss of growth characteristics. Primary and three-month old (15 passages) cultures had population doubling times of 32-34 hours and 29-31 hours, respectively. These cells (primary and subcultures) did not require a minimum cell number to become established in culture. Bovine endothelial cells (primary, first, fifth and thirteenth passages) were characterized ultrastructurally by the presence of Weibel-Palade bodies, pinocytotic vesicles and microfilaments and immunologically by the presence of thrombosthenin-like contractile proteins and Factor VIII antigen. The intercellular junctions of post-confluenct cultures stained specifically with silver nitrate. From these data, we concluded that identifiable endothelial cells could be obtained from bovine aortae and cultured and maintained for prolonged periods of time.

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