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D Doyle

Publications and source records attributed to D Doyle.

At least 91 records · Page 5Linked to original sources
PubMed1988

Identification of a complex of the three forms of the rat liver asialoglycoprotein receptor.

We have generated antibodies against synthetic peptides which represent the carboxyl terminus of either the major, or the two minor, forms of the rat hepatic lectin which recognizes galactose-terminated glycoproteins (asialoglycoproteins). The antibodies were shown to be specific for the form of the lectin containing the immunizing peptide sequence by the following: reaction with purified lectin after sodium dodecyl sulfate-polyacrylamide gel electrophoresis, immunoprecipitation of sodium dodecyl sulfate-denatured lectin, immunoprecipitation of lectin synthesized in vitro. These antibodies, however, precipitated all three rat hepatic lectin forms from nonionic detergent extracts of hepatocytes labeled with 125I via the lactoperoxidase catalyzed technique. A similar result was obtained if antibody was bound to intact cells prior to solubilization with detergent and collection of the immune complexes. We conclude that at least the plasma membrane-associated fraction of the rat hepatic lectin forms exists as a heterotypic complex.

Animals↗
PubMed1988

Differential phosphorylation of murine class I major histocompatibility antigens.

Recent studies have shown that the H-2K and H-2D transplantation antigens are expressed differentially in different tissues of mouse. Our previous investigations also established that in thioglycolate-stimulated peritoneal macrophages the H-2Dk antigen exists in distinct cell surface and intracellular forms. These two forms are glycosylated differently. In this report, we have found that (1) H-2Dk antigen is phosphorylated whereas H-2Kk antigen is not, and (2) only the cell surface form of H-2Dk antigen is phosphorylated in thioglycolate-stimulated macrophages derived from C3H/Heha mice. This differential phosphorylation of H-2 antigens will provide a model system for further studies on the molecular mechanism and function of phosphorylation of H-2 antigens.

Animals↗
PubMed1988

Analysis of renal and hippocampal type I and type II receptors by fast protein liquid chromatography.

Type I and Type II adrenal steroid receptors from rat renal and hippocampal cytosols were studied by the technique of Fast Protein Liquid Chromatography. Type I receptors were labelled with [3H]aldosterone plus excess RU26988, and Type II receptors with [3H]dexamethasone. On a Mono Q anion exchange column the molybdate-stabilized renal and hippocampal Type I receptors both eluted as single symmetrical peaks at 0.27 M NaCl, with a recovery of approximately 90% and 60-fold purification (renal) and 10-15-fold (hippocampal). Molybdate-stabilized Type II binding sites from both hippocampal and renal cytosols co-eluted with the Type I sites. On Superose gel filtration renal Type I receptor-steroid complexes consistently eluted two fractions later than hippocampal Type I complexes, suggesting that the renal complexes are smaller; Type II receptor-steroid complexes from both cytosols co-eluted, consistently one fraction behind hippocampal Type I sites. Sequential gel filtration and anion exchange chromatography achieved a 1000-fold purification of renal Type I binding sites, with an overall recovery of 10%.

Androstanols↗
PubMed1988

Asialoglycoprotein receptor genes are linked on chromosome 11 in the mouse.

The asialoglycoprotein receptor on the hepatocyte plasma membrane recognizes galactose-terminated glycoproteins and internalizes them for subsequent degradation in lysosomes. The rat receptor, also known as rat hepatic lectin (RHL), is comprised of three protein subunits called RHL-1, RHL-2, and RHL-3; two genes code for RHL-1 and RHL-2/3, respectively. We have cloned and sequenced the gene for RHL-2/3, and demonstrated that homologous asialoglycoprotein receptor genes exist in the mouse genome. Biochemical studies have demonstrated that receptor subunits exhibit the same temporal expression during development and function in a coordinate manner. This study asks if mouse receptor genes are linked and thus could possibly respond to shared cis-acting regulatory elements. Using restriction fragment length polymorphisms (RFLPs) and recombinant inbred lines, we mapped two closely linked mouse hepatic lectin (MHL) genes to chromosome 11. Asgr is designated to name asialoglycoprotein gene loci. Coordinate regulation of this linked gene family is discussed.

Animals↗
PubMed1988

Brain damage in fatal non-missile head injury without high intracranial pressure.

As part of a comprehensive study of brain damage in 635 fatal non-missile head injuries, the type and prevalence of brain damage occurring in the absence of high intracranial pressure were analysed. Of 71 such cases, 53 sustained their injury as a result of a road traffic accident; only 25 experienced a lucid interval. Thirty eight had a fractured skull, a mean total contusion index of 12.9 and diffuse axonal injury in 29: severe to moderate ischaemic damage was present in the cerebral cortex in 25, brain swelling in 13, and acute bacterial meningitis in nine. The prevalence and range of brain damage that may occur in the absence of high intracranial pressure are important to forensic pathologists in the medicolegal interpretation of cases of fatal head injury.

Adolescent↗
PubMed1988

Postviral fatigue syndrome: persistence of enterovirus RNA in muscle and elevated creatine kinase.

Enterovirus-specific probes have been prepared by reverse transcription of conserved sequences in purified Coxsackie B2 virus genomic RNA and molecular cloning techniques. These probes were used in quantitative slot blot hybridizations to test for the presence of enterovirus-specific RNA in skeletal muscle biopsy specimens from 96 patients who had suffered from the postviral fatigue syndrome myalgic encephalomyelitis for up to 20 years. Biopsy specimens from 20 patients were positive for the presence of virus-specific RNA with hybridization signals more than three standard deviations greater than the mean of the normal muscle controls. Biopsies from the remaining 76 patients were indistinguishable from the controls. These data show that enterovirus RNA is present in skeletal muscle of some patients with postviral fatigue syndrome up to 20 years after onset of disease and suggest that a persistent virus infection has an aetiological role.

Adolescent↗
PubMed1988

Medtronic-Hall prosthesis: valve related deaths and complications.

The purpose of this retrospective study was to assess the clinical performance of the Medtronic-Hall prosthesis based on a review of all valve related complications over a period of nine years. From January 1978 until December 1986, 502 prostheses were implanted in 450 patients. There were 200 aortic valve replacements, 198 mitral valve replacements and 52 double valve replacements. There were 234 associated procedures performed, the most frequent being coronary grafting in 129 patients and tricuspid annuloplasty in 35 patients. The mean age of the patients (230 men and 220 women) was 54.08 +/- 11.7 years. Preoperatively, 92.2% were in NYHA class III or IV. Early mortality was 7.1% and late mortality 16.7%. The follow-up totalled 1733 patient-years (mean 49.8 +/- 31.7 months). Over a period of nine years, the valve related complication rate was 4.4% per patient-year. Thromboembolic events occurred in 37 patients (2.1% per patient-year), anticoagulant related hemorrhage in 23 patients (1.3% per patient-year), endocarditis in 11 patients (0.6% per patient-year), perivalvular leak in six patients (0.3% per patient-year) and death and reoperation in 28 patients (1.6% per patient-year). At eight and one-half years, the survival rate was 71.28 +/- 2.7% for the whole group. The actuarial rate of patients free from all valve related complications was 75.92 +/- 2.7%, rate of freedom from thromboembolism was 86.01 +/- 2.4%, freedom from anticoagulant related hemorrhage was 92.7 +/- 1.6%, freedom from endocarditis was 97.17 +/- 0.8%, freedom from perivalvular leak was 98.27 +/- 0.7% and freedom from death and reoperation was 90.70 +/- 1.9%.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1987

Isolation and characterization of a Mr = 110,000 glycoprotein localized to the hepatocyte bile canaliculus.

A Mr = 110,000 glycoprotein, GP 110, was partially purified using wheat germ agglutinin-Sepharose affinity chromatography from a bile canalicular-enriched membrane fraction denoted N2u of rat liver. This fraction was subjected to preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis and the Mr = 110,000 polypeptide was excised and used as an immunogen in rabbits. The antisera were found to specifically recognize a Mr = 110,000 polypeptide, named GP 110, in the N2u membrane fraction. In isolated hepatocytes, GP 110 was readily accessible to cell surface iodination catalyzed by lactoperoxidase at 4 degrees C and was judged by immunoprecipitation studies to contain about 2% of total radioactivity incorporated into externally oriented proteins of the cell. Immunoprecipitated GP 110 was shown by two-dimensional polyacrylamide gel electrophoresis to migrate with an approximate pI of 4.9. Indirect immunofluorescence on frozen liver sections demonstrated that GP 110 was primarily localized in the bile canaliculus. In corroborative studies employing subcellular fractionation, it was found that GP 110 was enriched nearly 19-fold in P2, a plasma membrane fraction primarily derived from the sinusoidal domain, and 44-fold in N2u. In contrast, only low levels of GP 110 were present in endoplasmic reticulum, mitochondrial, cytosolic, and nuclear-enriched fractions of liver. The physiological function of GP 110 is as yet unknown; antisera to it did not immunoprecipitate other known bile canalicular proteins of similar molecular weights. GP 110 was found to be extensively glycosylated relative to other known membrane proteins; approximately 33% of the apparent molecular weight appear to be carbohydrate. In agreement, limited removal of N-linked carbohydrate chains indicated that there are approximately eight chains/GP 110 polypeptide. Neuraminidase treatment of GP 110 resulted in a desialylated Mr = 85,000 polypeptide suggesting that the majority of carbohydrate chains on GP 110 are of the complex type.

Animals↗
PubMed1987

Biosynthesis and turnover of a Mr = 110,000 glycoprotein localized to the hepatocyte bile canaliculus.

Antiserum was raised in rabbits against a bile canalicular glycoprotein of Mr = 110,000 purified to homogeneity from of rat liver. The antisera specifically immunoprecipitated a Mr = 110,000 polypeptide from hepatocytes metabolically labeled with [35S]methionine. When hepatocytes in primary culture were incubated with tunicamycin before labeling with [35S]methionine in the presence of tunicamycin, the major polypeptide immunoprecipitated by the specific antiserum from Triton X-100 extracts of cells had a molecular weight of 59,000. Enzymatic removal of N-linked carbohydrates from the Mr = 110,000 glycoprotein by N-glycanase digestion also yielded a polypeptide with minimum Mr = 59,000. In pulse-chase experiments using [35S]methionine, the Mr = 110,000 protein detected by the specific antisera first appears as Mr = 85,000 and 75,000 intermediate species which are endoglycosidase H sensitive. The Mr = 85,000 intermediate form is lost first with time followed by the Mr = 75,000 form giving rise to the Mr = 110,000 form that is endoglycosidase H resistant. Neuraminidase digestion of the Mr = 110,000 form generated an Mr 85,000 form but with a different carbohydrate structure than the intermediate Mr 85,000 form detected in the pulse-chase experiments. The time required to accomplish the processing of the Mr = 85,000 and 75,000 forms is relatively slow. Finally, the terminal sugars are added and the mature Mr = 110,000 glycoprotein is rapidly transported to the cell surface. A minimum time of 90 min is required for the Mr = 110,000 bile canalicular glycoprotein to be synthesized, processed, and reach the cell surface which is long relative to the time required (10 min) for another domain-specific protein, the receptor for asialoglycoproteins, to reach the sinusoidal surface. The Mr = 110,000 bile canalicular glycoprotein turns over in the bile canalicular domain with a half-life of 43 h while the asialoglycoprotein receptor turns over in the sinusoidal domain with a half-life of 23 h.

Animals↗
PubMed1987

Two-dimensional polyacrylamide gel electrophoresis analysis of phosphorylated, membrane-localized ras p21 proteins.

The ras p21 oncogene product migrates as a heterogeneous series of polypeptides as resolved by both one- and two-dimensional polyacrylamide gel electrophoresis (PAGE). We have prepared polyclonal rat serum antibody to ras p21 and used this as well as monoclonal antibodies to immunoprecipitate forms of p21 synthesized in vivo in transformed NIH3T3 cells. Two-dimensional PAGE of p21 resolved two distinct groups of polypeptides, one acidic (pI 4.8-5.3) which we call the "A" forms, and one less acidic or more basic (pI 6.5-7.0) which we call the "B" forms. It is the membrane-localized, B forms of v-ras-Ha p21 that are predominantly phosphorylated in vivo.

Animals↗
PubMed1987

Microcephaly: genetic counselling and antenatal diagnosis after the birth of an affected child.

We describe the clinical and genetic details of a series of microcephalic patients who were referred to the Genetic Counselling Service for the West of Scotland. There were 29 isolated cases of microcephaly and 9 families with recurrent microcephaly. The sib recurrence risk was 19%, which reflects the high incidence of autosomal recessive microcephaly in this series. There was evidence for several varieties of recessive microcephaly. The most frequent, affecting 5 sib pairs, was associated with spastic quadriplegia, seizures, and profound mental handicap. In 15 families with one microcephalic child, prenatal diagnosis by serial ultrasound scans was undertaken in 21 subsequent pregnancies. Four recurrences of microcephaly were detected in the third trimester and one recurrence was missed because no scans were performed after 24 wk gestation when the ultrasound measurements indicated satisfactory head growth. The main reason for late diagnosis of affected fetuses was that head growth did not slow appreciably until the last trimester. The high recurrence risk in this prospective series emphasizes the contribution of autosomal recessive inheritance of microcephaly amongst patients of our Genetic Counselling Service.

Female↗
PubMed1987

Isolation of domains of the plasma membrane of hepatocytes.

Several recent studies have demonstrated the ability of techniques based on immunoadsorption to selectively isolate specialized subregions of membranes, termed domains, which are derived from a larger more complex parent membrane like the plasma membrane. The immunoadsorbent is directed against a specific antigen that resides exclusively or predominantly in the membrane domain to be isolated. Thus, a monospecific antibody to the domain-specific antigen is required. In the present study we developed a method employing a modified immunoblotting strategy which could utilize polyspecific antibodies to isolate membrane vesicles derived from a specific membrane domain of the hepatocyte plasma membrane. We also used specific cell surface labeling of the hepatocyte plasma membrane by lactoperoxidase-catalyzed iodination at 4 degrees C and preparation of different sized vesicles by sonication to facilitate isolation of the specific domain. For this study, polyspecific antisera were raised in goats against a membrane fraction, denoted N2u, which is enriched in bile canalicular proteins. This antiserum recognizes, among other antigens, a 110,000 Mr polypeptide previously shown to be localized in the bile canaliculus (J. Cook et al. (1983) J. Cell. Biol. 97, 1823-1833). A monospecific antiserum was raised in rabbits against the rat hepatocyte asialoglycoprotein receptor, a sinusoidal domain-specific set of glycoproteins whose major form has a Mr of 43,000. These antisera were each coupled indirectly to different pieces of nitrocellulose by the immunoblotting protocol and were used to isolate membrane vesicles from a crude extract of liver plasma membrane prepared by sonication. The ratio of iodinated asialoglycoprotein receptor to the 110,000 Mr polypeptide in vesicles isolated by the affinity nitrocellulose immunoadsorbent method indicate a 10- to 15-fold enrichment of sinusoidal-derived vesicles relative to bile canalicular-derived membrane vesicles. These results show that the affinity nitrocellulose immunoadsorbent method can be used to isolate domain-specific vesicles. Further, the affinity immunoadsorbent method described here for the isolation of domains of the plasma membrane is an integrative one allowing isolation of vesicles present in relatively small concentration in crude cell extracts and it requires minimal ultracentrifugation time.

Affinity Labels↗
PubMed1987

cDNA cloning for a bile canaliculus domain-specific membrane glycoprotein of rat hepatocytes.

Hepatocytes are polarized cells with distinct sinusoidal, bile canalicular, and basolateral plasma membrane domains. Each domain contains proteins that are specific for it. We have isolated three cDNA clones encoding a rat liver bile canaliculus domain-specific glycoprotein with Mr 110,000 (gp110) by immunologically screening a rat kidney lambda gt11 cDNA library with a rabbit polyclonal antiserum directed against purified gp110. The authenticity of these clones was verified as follows. (i) The antiserum recognizes specifically isopropyl beta-D-thiogalactoside-induced fusion proteins on electrophoretic transfer blots of total lysogen lysates containing these cDNA clones. (ii) Antibodies epitope-selected by these clones are able to interact with gp110 on electrophoretic transfer blots. (iii) The amino acid sequencing derived from the DNA sequence was confirmed by amino acid sequencing of a tryptic peptide of gp110. Rescreening of the same library with the cDNA clones identified a full-length cDNA clone for this glycoprotein. Sequence analysis indicates that the N-linked carbohydrate chains are concentrated on the N-terminal part of this highly glycosylated protein.

Amino Acid Sequence↗
PubMed1987

Magnetic resonance imaging in the management of resistant focal epilepsy: pathological case report and experience of 12 cases.

Clinical improvement in epilepsy following temporal lobectomy is more often obtained when an abnormality is found on subsequent histological examination. Pre-operative MRI demonstrated an abnormal signal in the temporal lobe of a patient with pathologically proven mesial temporal sclerosis with microvascular anomaly. MRI may therefore be helpful in the selection of patients for temporal lobectomy. MRI findings of 12 patients with resistant focal epilepsy are reviewed. A wide range of T1 and T2 weighting is suggested to maximise selection of patients.

Adult↗
PubMed1987

Rhino-orbital-cerebral mucormycosis--a clinico-pathological report of two cases.

Rhino-orbital-cerebral mucormycosis is an acute fungal infection of the oropharynx, paranasal sinuses, orbit and intracranial structures. It is rare, occurring mainly in diabetics in ketoacidosis. The clinical presentation is highly suggestive of the diagnosis. Current treatment has resulted in a greatly improved prognosis for survival and clinical awareness of this disease is important for early initiation of treatment with optimal effect. A clinico-pathological report of two cases is presented.

Adult↗