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Biomedical subjects

W Gibson

Publications and source records attributed to W Gibson.

At least 127 records · Page 7Linked to original sources
PubMed1985

Isolation of human cytomegalovirus intranuclear capsids, characterization of their protein constituents, and demonstration that the B-capsid assembly protein is also abundant in noninfectious enveloped particles.

Two types of intranuclear capsids have been recovered from human cytomegalovirus (HCMV, strain AD169)-infected cells. By analogy with strain Colburn (simian CMV) particles, these have been designated as A- and B-capsids. Both types of capsids are composed of proteins with molecular weights of 153,000 (major capsid protein), 34,000 (minor capsid protein), 28,000, and 11,000 (smallest capsid protein). In addition to these species, B-capsids contain a 36,000-molecular-weight (36K) protein which has been designated as the HCMV "assembly protein," based on its similarities to counterparts in strain Colburn CMV (i.e., 37K protein) and herpes simplex virus (i.e., VP22a/p40/NC-3/ICP35e). Peptide comparisons established that the assembly protein of HCMV B-capsids and the 36K protein that distinguishes HCMV noninfectious enveloped particles from virions are the same, providing direct evidence that noninfectious enveloped particles are enveloped B-capsids.

Capsid↗
PubMed1984

D,L-alpha-difluoromethylornithine inhibits human cytomegalovirus replication.

D,L-alpha-Difluoromethylornithine (DFMO) is an inhibitor of ornithine decarboxylase, the first enzyme in the polyamine biosynthetic pathway. Exposure of human foreskin fibroblast cells to DFMO before their infection with human strains of cytomegalovirus (CMV) resulted in a reduction in the amount of infectious virus produced. A 3-day exposure to the drug was required to elicit maximal antiviral effect. Cells exposed to DFMO at the time of infection produced normal amounts of infectious virus. Preexposure to the drug for 1, 2, or 3 days before infection resulted in at least 10-, 100-, or 1,000-fold decreases, respectively, in the amount of infectious virus produced. This decrease paralleled the loss of intracellular spermidine and was partially spared by the addition of exogenous putrescine, spermidine, or spermine (10 microM). When added 3 days before infection, DFMO depressed production of herpes simplex virus and simian CMV, as well as wild-type and laboratory prototype strains of human CMV. Although some antiviral effect was observed at a drug concentration of 1 mM, 10 mM gave a stronger effect and was the amount routinely used. At 30 mM DFMO, growth of noninfected cells was slowed but not arrested. Studies to investigate the level at which DFMO interferes with CMV replication showed that DFMO-treated, infected cells (i) exhibit a typical CMV-specific cytopathic effect, (ii) synthesize both viral proteins and viral DNA, (iii) contain at least some capsid forms, and (iv) shed greatly reduced amounts of virus particles into the growth medium. Since CMV virions, like those of herpes simplex virus, contain the polyamines spermidine and spermine, and since DFMO essentially eliminates the pool of intracellular spermidine, the possibility is suggested that this drug may exert its antiviral effect by interfering with virus assembly, perhaps at the level of DNA packaging or capsid envelopment or both.

Cells, Cultured↗
PubMed1984

Sphenoid sinus revisited.

J.H. Maxwell, M.D., in 1959 presented a paper on, "The Diagnosis of Chronic Inflammatory Lesions of the Sphenoid Sinus." In that paper he reviewed chronic inflammatory lesions of the sphenoid sinus and demonstrated through means of case presentations various techniques required for proper diagnosis. During the past five years at the Geisinger Medical Center some interesting noninflammatory lesions of the sphenoid sinuses have been observed and will be reviewed. The symptoms of these sphenoid sinus lesions are usually related to abnormalities of the orbit and/or nasal passages. Cerebrospinal fluid rhinorrhea and meningitis may be present. Pain, when present, is generally not as severe as that described in inflammatory lesions, but is still localized in a similar manner. The diagnosis of sphenoid sinus disease may be suspected on the history and x-rays of the paranasal sinuses. Poytomography and computerized tomography scanning have aided markedly in the identification of the sphenoid lesions. Surgery in these cases included either a transnasal or sublabial transseptal approach to the sphenoid sinus and is recommended as a method to gain access to this rather obscure area.

Adult↗
PubMed1984

Selection of particles and proteins for use as human cytomegalovirus subunit vaccines.

Uncertainties about the ultimate biologic consequences of using live virus vaccines to confer immunologic protection against CMV have focused attention on the use of noninfectious subunit vaccines. At least two classes of such preparations have been demonstrated to be effective in other systems. The first is virus particles bearing the relevant antigens but lacking nucleic acid (eg, hepatitis vaccine [31]). And the second class is biologically or chemically synthesized proteins or peptides with appropriate immunogenicity (eg, foot and mouth disease virus vaccine [32]). In this paper, two noninfectious CMV particles and several viral proteins have been discussed in view of their potential for use as such a vaccine. The two noninfectious virus particles discussed are referred to as dense bodies and NIEPs. The use of dense bodies for vaccine purposes has been suggested by others [14], but the simplicity of their composition has only recently been established [17]. Two characteristics of these particles make them attractive prospects for vaccine purposes. First, neither contains more than trace amounts of DNA or infectivity (ie, less than or equal to 0.1% that of virions). Thus, the concerns about possible adverse consequences of introducing DNA with the vaccine are greatly reduced. Second, both NIEPs and dense bodies contain all of the glycoprotein species present in virions and in approximately the same relative amounts. If, as anticipated, these proteins are important in eliciting the immune response to CMV, then NIEPs and dense bodies may be as effective as virions in that capacity. The fact that NIEPs contain the full complement of virion proteins, and in approximately the same relative amounts, suggests that they may produce a more complete immunologic response than dense bodies, which lack all of the capsid and most of the tegument proteins of the virion. Although NIEPs normally represent only a small percentage of the extracellular particles (eg, less than 1%), we have found that strain AD169 produces them in amounts nearly equivalent to virions. More importantly, we have shown here that NIEP production is essentially unaffected following treatment of infected cells with a concentration of hydroxyurea that reduced virion and dense body production by more than 90% (Fig. 6). Thus, by using strain AD169 to infect cells and hydroxyurea treatment for selective enrichment, it is possible to produce NIEPs in relatively large amounts and with theoretically very low levels of contaminating virions (ie, less than or equal to 0.01% infectivity of equivalent amount of virions).

Cytomegalovirus↗
PubMed1983

Isolation and characterization of a noninfectious virion-like particle released from cells infected with human strains of cytomegalovirus.

Three types of virus particles have been recovered from the culture medium of human foreskin fibroblasts infected with human strains of cytomegalovirus (HCMV). Two of these, virions and dense bodies, are routinely observed and have been described by others. The third, produced in lesser amounts, has not been previously characterized. This particle, separable from virions by rate-velocity sedimentation, is morphologically distinguished from them only by core structure. Radiolabeling and biological assays have established that these particles, like dense bodies, lack DNA and are not infectious. Based on these properties, we have designated this virion-like structure as a noninfectious enveloped particle (NIEP). Comparisons of the protein constituents of these three particles has shown that dense bodies have the simplest composition. Approximately 95% of their protein mass is represented by a 69,000 Da (69K) matrix-like protein. While dense bodies appear to have a normal complement of virion glycoproteins, they completely lack other predominant virion species. The protein compositions of virions and NIEPs are more complex than that of dense bodies, and are distinguished from one another by the presence in NIEPs of a 35,000 Da (35K) protein absent from the two other particles. Biosynthetic radiolabeling and cell fractionation experiments have demonstrated that this 35K protein is produced only in infected cells, is phosphorylated and partitions with the nuclear fraction. These and other results suggest that this protein is the HCMV counterpart of the previously described B-capsid proteins VP22a of herpes simplex and 37K of CMV (strain Colburn). NIEPs are produced by all HCMV strains examined and have not been observed in preparations of herpes simplex virus- or Old World monkey CMV-infected cells. Although this particle is generally present in much lower amounts than virions, strain AD169 overproduces NIEPs by approximately 10-fold. We have also found that the additional NIEP protein of AD169 has an apparently larger size (i.e., 36K) than the corresponding protein of other strains. The correlation between AD169 NIEP overproduction and its altered protein suggests that the two may be causally related.

Centrifugation, Density Gradient↗
PubMed1983

Protein counterparts of human and simian cytomegaloviruses.

Cytomegalovirus (CMV) proteins from isolates of both human (HCMV) and simian (SCMV) origin have been compared. Three classes were analyzed: the immediate-early (IE) proteins, other infected-cell-specific proteins not present in virus particles, and the proteins that constitute the mature extracellular virion. Comparisons were based on one- and two-dimensional (charge-size) separations in denaturing polyacrylamide gels, and on the selectivity of biosynthetic radiolabeling with [32P]orthophosphate and [3H]glucosamine. Results indicate that most, if not all, of the HCMV and SCMV proteins recognized, have counterparts in strain Colburn. As a group, the simian strains exhibit protein similarities that distinguish them from the human strains. Among the most diagnostic of these are the 205K and 145K virion proteins, each of which is about 7K smaller than its HCMV counterpart, and the predominant IE proteins, which are 10K to 20K (depending upon the strain) larger than their HCMV counterparts. The proteins of strain Colburn are shown to be more like those of the simian isolates than the human, and more like those of a vervet strain than rhesus. Leads provided by experiments using strain Colburn have aided in the identification of a previously unrecognized, abundant virion protein that is a principal phosphate acceptor, both in vivo and in vitro. Three additional phosphorylated proteins are identified in HCMV virions, as well as three glycoproteins. Only two HCMV strain-specific protein differences were detected by comparisons based on separation in SDS-containing polyacrylamide gels--one in the IE protein of strains Towne and Davis; the other in a virus capsid protein of strain AD169.

Animals↗
PubMed1983

Replica images of silver-stained gels using direct duplicating film.

A procedure for preparing duplicate images of polyacrylamide gels containing silver-stained proteins using X-ray direct duplicating film is described. Resulting images are shown to be qualitatively and quantitatively faithful representations of the original gel.

Electrophoresis, Polyacrylamide Gel↗
PubMed1983

Induction by morpholine of lysosomal alpha-mannosidase and acid phosphatase in rabbit alveolar macrophages in vivo and in vitro.

alpha-Mannosidase and acid phosphatase were induced in alveolar macrophages from the lungs of rabbits following exposure to morpholine. The induction was observed after inhalation of morpholine vapor or when macrophages were cultured in the presence of morpholine. Lysosomal hydrolases were not induced uniformly. In vivo, maximum induction of alpha-mannosidase and acid phosphatase in female rabbits was 1.7-fold and 2-fold, respectively, and in male rabbits, 3-fold and unchanged, respectively. In vitro, maximum hydrolase induction of alpha-mannosidase and acid phosphatase in macrophages from male rabbits was 1.4-fold and 1.3-fold, respectively, and from female rabbits, 1.3-fold and 1.5-fold, respectively. The induction of acid hydrolases in vitro was rapid, reaching a maximum within 4 hr in alveolar macrophages from male rabbits and within 8 hr in those from females. The induction was dose dependent. This study indicates that the lysosomal enzyme system of the alveolar macrophage may be stimulated by a low molecular weight xenobiotic.

Acid Phosphatase↗
PubMed1983

Phosphorylation, maturational processing, and relatedness of strain Colburn matrix proteins.

The intracellular 66,000 D (66K) and 69,000 D (69K) matrix-like phosphoproteins of strain Colburn cytomegalovirus (CMV) have been compared with each other and with their electrophoretic counterparts in the virion. Three lines of evidence indicate that the 66K and 69K proteins are products of separate genes, and that the intracellular and virion species are closely related. First, "pulse-chase" radiolabeling experiments showed that these proteins have separate precursors; that modification of each to the mature form correlated with phosphorylation; and that phosphorylation of the 69K precursor occurred more slowly than that of the 66K precursor, and resulted in a more dramatic slowing of its electrophoretic mobility. Second, comparisons of the 66K and 69K proteins based on partial proteolysis of [35S]methionine-labeled proteins, using V8 protease, and complete proteolysis of [32P]orthophosphate-labeled proteins, using trypsin or Pronase, provided no evidence of sequence relatedness. These analyses also suggested that the distribution of phosphorylated residues differs in the two proteins--clustered for the 69K and more disperse for the 66K. Phosphoamino acid analyses showed only phosphoserine in the 66K protein. The 69K protein contained, in addition to phosphoserine, an electrophoretically faster moving, unidentified spot. And third, immunological comparisons showed these proteins to exhibit little or no antigenic cross-reactivity. They did, however, demonstrate that the nuclear proteins are immunologically cross-reactive with their respective virion counterparts. Additional comparisons of these nuclear and virion proteins established that the virion 69K protein (v69) differs in electrophoretic mobility and net charge from the nuclear 69K protein but that it, as well as the virion 66K protein, has a two-dimensional phosphopeptide pattern similar to its nuclear counterpart.

Amino Acid Sequence↗
PubMed1981

Laryngeal abductor paralysis in multiple system atrophy. A report on three necropsied cases, with observations on the laryngeal muscles and the nuclei ambigui.

Clinical and pathological findings are reported in three cases of Shy-Drager syndrome with laryngeal stridor severe enough to require tracheostomy. Indirect laryngoscopy in all three cases revealed a failure to abduct the vocal cords properly with a resulting narrowing of the glottic airway. Histological studies showed a marked atrophy of the posterior crico-arytenoid muscles and little atrophy of the other laryngeal muscles. The posterior crico-arytenoid muscles have the sole responsibility of abducting the vocal cords, so it is concluded that the patient's stridor was caused by the selective paralysis of these muscles. Histological studies of the posterior crico-arytenoid muscles were suggestive of denervation but despite this finding, no clear evidence of any motor cell loss in the nuclei ambigui was obtained. Other possible causes of the selective posterior crico-arytenoid atrophy are discussed.

Autonomic Nervous System Diseases↗