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Biomedical subjects

D P O'Brien

Publications and source records attributed to D P O'Brien.

At least 109 records · Page 6Linked to original sources
PubMed1987

Cloning and expression of human tissue factor cDNA.

Tissue factor is a membrane protein that plays an essential role in the initiation of blood coagulation. When exposed to the circulation, tissue factor interacts with the serine protease factor VII, and the complex triggers fibrin clot formation by activating both factors IX and X of the coagulation cascade. This report describes the cloning and expression of the complementary DNA (cDNA) for human tissue factor. The cDNA encodes a protein of 263 amino acids preceded by a 32 amino acid signal peptide. The predicted protein sequence contains a potential hydrophobic membrane anchoring domain at its carboxy terminus, and bears no significant homology to any other known protein. Tissue factor mRNA of 2400 nucleotides was detected in adipose, adrenal, small intestine and a number of other tissues by Northern blot hybridization analysis. In order to confirm the identity of the cDNA, an expression vector containing the cloned cDNA was used to transfect cultured mammalian cells. These cells produced active tissue factor which was assayed using purified factors VII and X.

Amino Acid Sequence↗
PubMed1987

Pharmacological subtraction of the sensory controls over grasping in rats.

Catecholamine-depletion-induced catalepsy isolates and leaves intact an aggregate of allied reflexes (e.g., righting, standing still, bracing, and clinging) which involve all the body and limb segments in defending stable static equilibrium. Because other movement subsystems (locomotion, orienting, scanning, directed use of mouth or forepaws) are depressed, such animals cling in a vertical position for an abnormally long period of time. As a consequence, grasping reflexes may be studied independently of other responses. Haloperidol, a dopamine antagonist, abolishes visually elicited reaching and grasping, but leaves intact tactile and proprioceptive control of grasping. The grasping of haloperidol-treated rats can be further simplified by the pharmacological removal of the remaining sensory controls. The addition of morphine to haloperidol abolishes tactile grasping, while the addition of diazepam to haloperidol abolishes both tactile and proprioceptive (traction-elicited) grasping. Although visual, tactile, and proprioceptive grasping are abolished by haloperidol-plus-diazepam, some vestibular input to clinging remains: such rats, in response to being held vertically upright in the air, flex their digits with sufficient strength to allow them to cling vertically. The strength of forepaw digit flexion is severely diminished by labyrinthectomy, but the digits of the hindpaws appear to be unaffected. This residual non-labyrinthine digit gripping appears to be induced by proprioceptive inputs from the head, neck and torso in response to the vertical body position. Wrapping an elastic bandage snugly around the head and neck of a labyrinthectomized rat given haloperidol-plus-diazepam further diminishes the strength of forepaw digit flexion, and to a lesser degree hindpaw digit flexion.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1987

Cystitis cystica: an electron and immunofluorescence microscopic study.

Cystitis cystica was studied with the aid of electron and immunofluorescence microscopy. By electron microscopy, the epithelium demonstrated morphologic features suggestive of an active metabolism. Secretory-type granules were seen in the cytoplasm just beneath the luminal membrane of surface columnar cells. Microvilli of the plasma membrane also were seen at the luminal surface. Rough endoplasmic reticulum and Golgi apparatuses were present. The cells were rich in mitochondria. By immunofluorescence microscopy, IgA, secretory piece and IgM were localized in the epithelial cells, especially at the luminal surfaces. IgG was occasionally found. These findings contrast markedly with the transitional cells and their relatively scanty content of secretory-type organelles. In addition, they may explain the large amounts of IgA in the urine of patients with cystitis cystica.

Adult↗
PubMed1986

Multicentric nerve sheath fibrosarcomas of multiple cranial nerve roots in two dogs.

Nerve sheath fibrosarcomas of the left 5th through 8th cranial nerve roots were diagnosed in 1 dog and of the left 4th through 8th cranial nerve roots in another dog. Clinical signs in both dogs included head tilt to the left, circling to the left, left hemiparesis and proprioception deficits, rotary nystagmus, left-sided atrophy of masticatory muscles, and cutaneous hypalgesia of the left side of the face. Cranial nerve roots from both dogs were incorporated in discrete, firm, encapsulated, lobulated, tan masses of various sizes that compressed adjacent brain stem and cranial nerves. There were no regional or distant metastases; however, there was enlargement of nerve roots adjacent to the masses. The masses were composed of bundles and sheets of anaplastic, polymorphic to spindle-shaped cells that infiltrated cranial nerves and ganglia and extended into the brain along nerve roots. Masses contained various amounts of collagen and reticulin fibers, but no mucopolysaccharide ground substance. There was no myelin or S-100 protein associated with neoplastic cells. The tumors appeared to have a multicentric origin from cranial nerve sheaths.

Animals↗
PubMed1986

Nonmetastatic bladder cancer associated with hypercalcemia, thrombocytosis and leukemoid reaction.

We reviewed 4 patients with urothelial bladder cancer and hypercalcemia but without evidence of bony metastasis. Of the patients 2 presented with a leukemoid reaction (1 also had thrombocytosis). None of the patients had evidence of bone metastases or other causes of hypercalcemia, such as hyperparathyroidism, sarcoidosis or vitamin D intoxication. All 4 patients received aggressive therapy for the tumors. In each instance the serum calcium returned to normal following radiation therapy or tumor removal, lending support to the theory of humoral hypercalcemia of cancer. A diagnostic and therapeutic approach to such patients is outlined.

Aged↗
PubMed1986

Condylomata acuminata and verrucous carcinoma of the bladder: case report and literature review.

Condylomata acuminata and verrucous carcinoma are 2 lesions rarely seen in the bladder. Condylomata of the bladder can be treated successfully by local therapy but underlying carcinoma must be considered if the lesion is extensive or progressive in nature. Vigorous biopsies of the base of the lesions are necessary to eliminate the possibility of malignancy. If verrucous carcinoma is found the patient should be treated surgically. A case of condylomata acuminata of the bladder undergoing malignant change to verrucous carcinoma is reported.

Adult↗
PubMed1986

Studies on immunological assay of vitamin-K dependent factors. III. A double monoclonal immunoradiometric assay for factor IX antigen.

Two-site immunoradiometric assays (IRMAs) for factor IX antigen (IX:Ag) were developed using a monoclonal antibody (RFF-IX/1) on the solid-phase and either another monoclonal antibody (RFF-IX/4) or a human polyclonal inhibitor antiserum as tracer (M-M and M-I IRMA respectively). The lower sensitivity limits of these two assays for IX:Ag in normal reference plasma were 4 X 10(-4) (M-M IRMA) and 2 X 10(-4) (M-I IRMA) units/ml. In 20 samples of normal plasma, levels of factor IX coagulation activity (IX:C) and of factor IX antigen measured by both IRMAs were highly correlated. Mean values of approximately 1.0 units/ml were obtained in all three assays. In normal serum, IX:Ag levels were lower with means of 0.84 (M-M IRMA) and 0.83 (M-I IRMA) units/ml. 4/25 patients with haemophilia B were CRM neg., two were CRM + and the remaining 19 patients were CRMr variants. In two of these, IX:Ag was detectable by M-I IRMA whilst IX:C and IX:Ag measured by M-M IRMA were undetectable. In plasma from a fetus subsequently terminated on eugenic grounds, IX:C and IX:Ag by both M-M and M-I IRMA were undetectable. In warfarin-treated plasma (n = 12), the level of IX:C was low (mean 0.39 units/ml). The levels of IX:Ag measured by M-M IRMA (mean of 0.80 units/ml) and by M-I IRMA (0.70 units/ml) showed a discrepancy. M-M IRMA reflects the real amount of IX:Ag in warfarinized plasma because both monoclonal antibodies bind to epitopes distant from the light chain carboxylated region. Western blotting of denatured factor IX demonstrated that RFF-IX/1 binds an epitope that is lost after XIa activation. RFF-IX/4 binds the heavy chain. Antigen measured after activation but without denaturing showed loss of 60% reactivity after XIa activation but no change after RVV activation. These data indicate a binding site for RFF-IX/1 within the activation peptide (residues 146-180).

Antibodies, Monoclonal↗
PubMed1985

Purification of human factor VIII:C and its characterization by Western blotting using monoclonal antibodies.

Human factor VIII:C has been purified over 300 000-fold from cryoprecipitate by polyelectrolyte purification followed by affinity chromatography on Sepharose linked to antibody to factor VIIIR:Ag (monoclonal or polyclonal) and Sepharose linked to monoclonal antibody to factor VIII:C. The purified material has been analyzed by polyacrylamide gel electrophoresis (PAGE) and Western blotting using monoclonal antibodies. PAGE shows predominant bands at 360K (unreduced), 210K, and 90K and an 80K/79K doublet; Western blotting showed all the monoclonal antibodies used bound the 360K form. In a small-scale purification, plasma from blood taken directly into thrombin inhibitor Kabi S-2581 was applied directly to the monoclonal anti-factor VIII:C column. Western blot analysis of this material showed the 360K band on reduction. The purified factor VIII:C could be activated 13-fold by human thrombin. Gel analysis of the activated material showed intensification followed by fading of the band at 90K and generation of bands at 70K/69K, 55K, and 40K. Western blotting shows that the 70K/69K doublet derives from the 80K/79K moiety and the 40K peptide derives from the 90K and is presumed to contain the active site. From these studies an epitope map of the factor VIII:C molecule has been constructed.

Antibodies, Monoclonal↗
PubMed1985

Clinical features of spongy degeneration of the central nervous system in two Labrador retriever littermates.

Spongy degeneration of the CNS was diagnosed in 2 female Labrador Retriever littermates. The dogs had histories of progressive cerebellar ataxia and episodes of extreme extensor rigidity beginning at 4 and 6 months of age. One had a history of hearing difficulties. When examined at 9 months of age, both dogs had moderate ataxia and dysmetria of the head, trunk, and limbs, hyperreflexia with clonus, and extensor rigidity with episodes of exaggerated rigidity and opisthotonos. One dog was euthanatized and the other was monitored for 2 months, during which time her signs progressed to severe dysmetria, weakness, muscle atrophy, and rigidity with marked hyperreflexia. Necropsy of both dogs revealed astrocytic abnormalities and spongy degeneration of the white matter of the CNS. The clinical signs and postmortem findings were similar to the juvenile form of spongy degeneration in man.

Animals↗
PubMed1985

Vestibular versus tail-pinch activation in cats with lateral hypothalamic lesions.

Mild tail-pinch induces eating in animals which are aphagic following lateral hypothalamic lesions. This study compares the specificity of the behavior produced by tail-pinch to that produced by vestibular stimulation in cats which are akinetic and aphagic following lateral hypothalamic lesions. In these cats, tail-pinch elicited licking and biting at food while vestibular stimulation preferentially elicited forward locomotion. These results suggest that such stimuli activate specific movement subsystems rather than producing general, nonspecific arousal.

Animals↗
PubMed1985

Bilateral clear cell papillary cystadenoma of the epididymides presenting as infertility: an early manifestation of von Hippel-Lindau's syndrome.

Papillary cystadenoma of the epididymis is a rare benign tumor, accounting for only 4 per cent of all epididymal tumors. Histologically, it can be confused with metastatic renal cell carcinoma. We report a case of bilateral involvement of the epididymides and infertility. Its diagnostic significance is its association with the von Hippel-Lindau syndrome, necessitating close surveillance.

Adult↗
PubMed1985

An immunoradiometric assay for human factor VIII/von Willebrand factor (VIII:vWF) using a monoclonal antibody that defines a functional epitope.

A murine monoclonal antibody has been produced (RFF-VIII:R/2) that binds specifically to human factor VIII-related antigen (VIII:RAg) in plasma and in vascular endothelial cells but has no reactivity with factor VIII procoagulant antigen (VIII:cAg). This antibody is a potent inhibitor of von Willebrand factor activity (VIII:vWF) in that it can totally neutralize ristocetin-induced aggregation of platelet rich plasma and inhibit platelet adhesion at high flow rates. RFF-VIII:R/2 can be used in a one-stage, fluid phase immunoradiometric assay that can detect VIII:RAg at concentrations of 0.001 u/ml. This method has been used to analyse plasma from patients with von Willebrand's disease (vWD). Results obtained in these patients showed a high degree of correlation between the monoclonally-defined epitope and VIII:vWF levels measured by ristocetin-induced aggregation of washed platelets. This correlation was maintained in those patients with the 'variant' types of vWD who exhibit highly disparate VIII:vWF and VIII:RAg levels when the latter is determined using polyclonal antisera. It appears that this monoclonal antibody recognizes a site on the VIII:RAg molecule which is associated with its interaction with the platelet membrane. Immunoradiometric assays using RFF-VIII:R/2 offer a simplified, reproducible means of detecting functionally-active VIII:RAg as an alternative or supplement to techniques involving platelet interactions.

Animals↗
PubMed1985

Spongy degeneration of the central nervous system in two canine littermates.

Progressive dysmetria was observed at four months and six months of age in two female Labrador retriever littermates. Neurological examinations indicated a cerebellar disorder. Laboratory analyses were normal. The dogs were studied clinically until they were killed for necropsy at nine months and eleven months of age. Both dogs had normal brain size, shape, and calvarial symmetry; the older dog had body weight loss, reduced brain weight, and muscle atrophy. Microscopically, there were vacuoles, hypertrophied fibrous astrocytes, myelin loss, and prominent blood vessels, distributed symmetrically in the subcortical and deep white matter of all lobes of the cerebrum; in the folial and deep white matter of the cerebellum; in the tracts of some cranial nerves; in the thalamic area, midbrain and brainstem; and in the white matter of the spinal cord. There was no significant myelinolysis, inflammation, or axonal degeneration. Ultrastructurally, there were intramyelinic vacuoles with separation of lamellae at intraperiod lines and larger spaces formed by coalescence of ruptured vacuoles. Hypertrophied fibrous astrocytes had abundant glial filaments, edematous cytosol, membrane-bound crystalline inclusions, dilated cytocavitary systems, and abnormal mitochondria. The clinical, histological, and ultrastructural findings resembled those reported for the juvenile form of Canavan's disease (van Bogaert and Bertrand type) in children.

Animals↗
PubMed1983

Monoclonal antibodies to human procoagulant factor VIII.

VIII:C was purified from intermediate-purity concentrate by adsorption on polyelectrolyte E5 and affinity chromatography on Sepharose/anti-VIIIR:Ag. The highly purified VIII:C preparation (sp. act. 1598 U/mg) was used to immunize Balb-C mice. Spleen cells from a mouse with a serum antibody titer of 963 U/ml were fused with P3 NSI mouse myeloma cells. Hybrid clones were screened by a coagulation inhibition assay and by a four-layer antibody adsorption procedure. Nine monoclonal antibodies specific to VIII:C were produced. Five of these antibodies have been cloned and grown in mouse ascitic fluid. Antibody titers from ascitic fluid ranged from 35 to 82,000 BU/ml. The antibodies, when radiolabeled, form a high-molecular-weight complex with antigens present in normal plasma and factor VIII concentrate, but not when incubated with CRM-negative hemophilic plasma. A two-site assay using a combination of monoclonal antibodies is able to detect VIII:CAg in normal plasma and in factor VIII concentrate. Sensitive two-site immunoradiometric assays using monoclonal antibodies as the solid phase have been set up.

Animals↗