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Biomedical subjects

M S Read

Publications and source records attributed to M S Read.

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

Gene therapy for hemophilia B: host immunosuppression prolongs the therapeutic effect of adenovirus-mediated factor IX expression.

Hemophilia B is caused by a deficiency of blood clotting factor IX (FIX). Previous studies have shown that the delivery of a recombinant adenoviral vector expressing canine FIX (cFIX) resulted in a complete correction of hemophilia B in FIX-deficient dogs, but that cFIX expression decreased to only about 1-2% of normal levels 3 weeks after treatment. In the present study, therapeutic levels of cFIX expression capable of producing a partial correction of hemophilia B were maintained for at least 6 months after the coadministration of the cFIX-expressing adenovirus and the immunosuppressive agent cyclosporin A (CsA). These findings support a recent report (Yang et al., 1994) that host T-cell-mediated immunity against virally transduced cells is a major contributing factor to the transient nature of adenovirus-mediated gene expression in immunocompetent animals. Although a second administration of the cFIX-expressing adenovirus 6 months after the first infusion had only a minimal effect on plasma FIX levels in a dog that had been continuously treated with CsA, the prolonged expression of the transgene indicates that immunosuppression may be applicable in attaining long-term treatment of clinically relevant disorders.

Adenoviridae↗
PubMed1995

Thrombotic thrombocytopenia induced in dogs and pigs. The role of plasma and platelet vWF in animal models of thrombotic thrombocytopenic purpura.

Thrombotic thrombocytopenia with severe depletion of plasma von Willebrand factor (vWF) was induced in normal large animals (5 dogs and 2 pigs) by botrocetin, a Bothrops factor requiring vWF for platelet agglutination. Botrocetin (90 to 100 U/kg, 2.14 to 2.38 mg/kg, in a single i.v. injection) reduced plasma vWF activity to < 0.1 U/mL for 24 hours. During this period, multimeric analysis of plasma vWF antigen (Ag) revealed the loss of intermediate- and high-molecular-weight forms with a concomitant increase in lower molecular weight forms. A moderate reduction in factor VIII (FVIII) activity was observed. The vWF reduction was accompanied by transient thrombocytopenia and prolonged bleeding times during the deficiency state. Occlusive platelet thrombi were detected by transmission electron microscopy in the microcirculation of lung and spleen but not kidney or brain 30 minutes after the botrocetin injection. Recovery of plasma vWF and platelet count occurred within 48 hours and was associated with the appearance in the plasma of unusually large forms of vWF:Ag multimers. The vWF:Ag multimer distribution was normal at 72 hours. The ultrastructural distribution of vWF in unstimulated normal porcine and canine platelets was examined by using immunogold staining. VWF was detected in the alpha-granules of normal pig platelets but was not observed in platelets from normal dogs. However, both animals developed thrombotic thrombocytopenia when injected with botrocetin. A second group of animals (2 dogs and 3 pigs) with von Willebrand disease (vWD) was given a single botrocetin injection (90 to 100 U/kg). No thrombocytopenia occurred.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Extradural sufentanil by patient-controlled analgesia or nurse-administered compared with optimal morphine in a high dependency unit: effects on oxygenation and pain relief after abdominal surgery.

The incidence of apnoeic episodes (> 12 s) was measured in 30 surgical patients allocated randomly to one of three analgesic regimens and all nursed in a high dependency unit. Ten patients received i.m. morphine (mean 52 (range 30-80) mg), administered on request. The remaining 20 patients received extradural sufentanil as an initial bolus dose of 50 micrograms followed by a bolus of 10 micrograms, either self-administered using PCA (10 patients: mean 275 (range 130-450) micrograms) or administered by a nurse on request (10 patients: 144 (70-200) micrograms). With i.m. morphine apnoeic episodes were maximal 2-3 h after administration while after extradural sufentanil, apnoeas were maximal within a few minutes.

Adolescent↗
PubMed1993

In vivo gene therapy of hemophilia B: sustained partial correction in factor IX-deficient dogs.

The liver represents a model organ for gene therapy. A method has been developed for hepatic gene transfer in vivo by the direct infusion of recombinant retroviral vectors into the portal vasculature, which results in the persistent expression of exogenous genes. To determine if these technologies are applicable for the treatment of hemophilia B patients, preclinical efficacy studies were done in a hemophilia B dog model. When the canine factor IX complementary DNA was transduced directly into the hepatocytes of affected dogs in vivo, the animals constitutively expressed low levels of canine factor IX for more than 5 months. Persistent expression of the clotting factor resulted in reductions of whole blood clotting and partial thromboplastin times of the treated animals. Thus, long-term treatment of hemophilia B patients may be feasible by direct hepatic gene therapy in vivo.

Animals↗
PubMed1993

The roles of von Willebrand factor and factor VIII in arterial thrombosis: studies in canine von Willebrand disease and hemophilia A.

We have studied the roles of von Willebrand factor (vWF) and factor VIII in arterial thrombosis in four canine phenotypes: normal (n = 6), hemophilia A (n = 11), von Willebrand disease (vWD) (n = 9), and hemophilia A/vWD (n = 1). vWF activity was determined by botrocetin-induced agglutination of fixed human platelets and vWF antigen (vWF:Ag) by Laurell electroimmunoassay and crossed immunoelectrophoresis. Plasma from normal dogs and those with hemophilia A had vWF activity, vWF:Ag, and a full range of vWF:Ag multimers on gel electrophoresis equivalent to normal canine plasma pool. Platelet cytosol contents were isolated by freezing and thawing, triton X-100 solubilization, or sonication of washed platelets with and without protease inhibitors and inhibitors of platelet activation. Washed platelets were also stimulated with calcium ionophore and MgCl2. There was no measurable vWF activity or vWF:Ag in platelet lysates or releasates in any dog regardless of phenotype. All dogs were studied using a standard arterial stenosis and injury procedure to induce arterial thrombosis. Thromboses were detected by cyclic reductions in Doppler blood flow velocity. Vessels were examined by light and scanning electron microscopy. Thrombosis developed in the arteries of normal (9 of 10) and hemophilia A dogs (16 of 16) but in none of the vWD dogs (0 of 10). Infusion of canine vWF cryoprecipitate into vWD dogs markedly shortened bleeding time but did not support thrombosis as seen in dogs with vWF in the plasma and subendothelium. Thrombosis, then, fails to occur when vWF is absent from the plasma and subendothelial compartments or present only in the plasma compartment. These data are consistent with the hypothesis that vWF in the plasma and subendothelium supports thrombosis. Neither plasma FVIII nor platelet vWF is essential for thrombosis in this model.

Animals↗
PubMed1992

Porcine von Willebrand disease and atherosclerosis. Influence of polymorphism in apolipoprotein B100 genotype.

The relationship of apolipoprotein-B genotype (Lpb) to diet-induced hypercholesterolemia and atherosclerosis was studied in von Willebrand disease (vWD) and normal pigs. Von Willebrand and normal pigs developed comparable levels of hypercholesterolemia (respectively, 757.9 +/- 49.4 versus 772.8 +/- 47.9 mg/dl, P = 0.95). Pigs with Lpb1/5 and Lpb5/8 genotypes, however, developed significantly higher serum cholesterol levels than those with other Lpb genotypes (866.1 +/- 64.0 mg/dl, P = 0.0343). Coronary and aortic atherosclerosis, measured by computer-assisted automated image analyzer, were not significantly different between vWD and normal pigs. Pigs with an Lpb5 allele developed significantly more atherosclerosis than those with the Lpb3/8 or Lpb8/8 genotypes or the rare Lpb1 allele (r greater than or equal to 0.434, P less than or equal to 0.05). Polymorphism in apolipoprotein B100 genotype, then, significantly influenced the severity of diet-induced hypercholesterolemia and atherosclerotic plaque formation in vWD and normal swine without regard to the vWD genotype.

Animals↗
PubMed1991

von Willebrand factor and animal models: contributions to gene therapy, thrombotic thrombocytopenic purpura, and coronary artery thrombosis.

Use of animal models of von Willebrand factor (vWF) deficiency, both inherited and induced, continues to advance the knowledge of vWF-related diseases. Three examples are reviewed in this article--von Willebrand's disease (vWD), thrombotic thrombocytopenic purpura, and coronary artery thrombosis. The success of gene transfer by liver and bone marrow transplantation in porcine vWD and canine hemophilia A, with a change in phenotype that establishes improved hemostasis, portends imminent testing of gene therapy in these models. With use of recombinant technology, the phenotype of hemophilia B fibroblasts has been transformed to normal, as evidenced by secretion of the normal hemostatically active protein. This result is a prelude to implantation in hemophilic animals. Thrombotic thrombocytopenic purpura is characterized by qualitative and quantitative alterations in vWF. A new animal model induced by the venom factor botrocetin, a cofactor of vWF, closely mimics the human syndrome. A proposed pathophysiologic mechanism for thrombotic thrombocytopenic purpura is outlined. The third contribution is recognition that occlusive coronary thrombosis is a vWF-dependent condition. Without vWF, as in porcine vWD or normal pigs treated with a monoclonal anti-vWF antibody, occlusive thrombi do not develop, even with luminal stenosis. The thrombogenicity of coronary atheromas, including those with fissures of the fibrous cap, is also vWF-dependent.

Animals↗
PubMed1991

Glycoprotein Ib bioassays. Activity levels in Bernard-Soulier syndrome and in stored blood bank platelets.

Glycoprotein Ib (GP-Ib) is a major platelet receptor protein concerned with von Willebrand-factor binding, platelet agglutination, and platelet adhesion, and it is required for normal hemostasis. By the use of botrocetin (venom coagglutinin), both quantitative and semiquantitative assays for GP-Ib activity were developed. The latter assay uses limiting dilutions of botrocetin as a measure of GP-Ib activity. Platelets, stored up to 23 days under blood bank conditions, were assayed by the limiting dilution test. Values of GP-Ib were progressively diminished after 9 to 10 days of storage, reaching levels of less than 10% at 23 days. Platelets from a subject with Bernard-Soulier syndrome showed less than 10% GP-Ib activity. These assays appear to be a specific measure of functional GP-Ib activity, and, when combined with GP-Ib antigen measurement by other methods, they provide a means for further characterizing GP-Ib abnormalities.

Bernard-Soulier Syndrome↗
PubMed1991

Role of von Willebrand factor in arterial thrombosis. Studies in normal and von Willebrand disease pigs.

With normal and von Willebrand disease (vWD) pigs, we studied the role of von Willebrand factor (vWF) in platelet-vessel wall interactions and occlusive arterial thrombosis. Two methods of arterial injury have been used to determine the thrombotic response of flowing blood in vivo. The first involves balloon catheter injury. After superficial denudation of endothelium from coronary intima, platelets adhere to the subendothelium in a monolayer. Similar numbers of adherent platelets are found in both phenotypes, but platelets in vWD pigs have impaired pseudopod formation and are less well spread morphological indexes of limited platelet activation. Deeper injury, which involves the media, produces nonocclusive platelet-fibrin microthrombi. The second injury method involves pinching the artery at a site of superimposed stenosis, a procedure that almost always exposes media. This procedure induces platelet-fibrin microthrombi in normal and vWD pigs, but only normal pigs develop occlusive thrombosis. Both methods of arterial injury have also been performed in normal and vWD pigs with diet-induced hypercholesterolemia and atherosclerosis. Atherosclerosis promotes platelet spread in vWD pigs but does not abolish the protection from stenosis and injury-induced occlusive thrombosis. In addition, neutralization of vWF activity in normal pigs by a monoclonal antibody prevents the induction of occlusive thrombosis by the stenosis and pinch-injury procedure. This monoclonal antibody also causes performed platelet aggregates to break up. These experimental models of inducing arterial thrombosis have been used in normal and vWD pigs to demonstrate interactions between normal and atherosclerotic vessel wall constituents, circulating platelets and vWF that are fundamental in the process of arterial thrombosis.

Animals↗
PubMed1990

Phenotypic correction of factor IX deficiency in skin fibroblasts of hemophilic dogs.

Primary skin fibroblasts from hemophilic dogs were transduced by recombinant retrovirus (LNCdF9L) containing a canine factor IX cDNA. High levels of biologically active canine factor IX (1.0 micrograms per 10(6) cells per 24 hr) were secreted in the medium. The level of factor IX produced increased substantially if the cells were stimulated by basic fibroblast growth factor during infection. Additionally, we also report that endothelial cells transduced by this virus can produce high levels of biologically active factor IX. We propose that skin fibroblasts and endothelial cells from hemophilia B dogs may serve as potential venues for the development and testing of models for treatment of hemophilia B by retrovirally mediated gene replacement therapy.

Amino Acid Sequence↗
PubMed1990

Painful onset of intrathecal blockade.

A case is described in which a woman with well controlled epilepsy and minor abnormalities of peripheral nerve function in the lower limbs experienced intense pain during the onset of a spinal anaesthetic for operation on burns on her thighs. Possible mechanisms and treatments are discussed.

Anesthesia, Spinal↗