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Biomedical subjects

G B Caughman

Publications and source records attributed to G B Caughman.

12 recordsLinked to original sources
PubMed1992

Characterization of the regulatory functions of the equine herpesvirus 1 immediate-early gene product.

Use of the translation-inhibiting drug cycloheximide has indicated that the equine herpesvirus 1 (EHV-1) immediate-early (IE) gene, the sole EHV-1 IE gene, encodes a major viral regulatory protein since IE mRNA translation is a prerequisite for all further viral gene expression (W.L. Gray, R. P. Baumann, A. T. Robertson, G. B. Caughman, D. J. O'Callaghan, and J. Staczek, Virology 158:79-87, 1987). An EHV-1 IE gene expression vector (pSVIE) in combination with chimeric EHV-1 promoter-chloramphenicol acetyltransferase (CAT) reporter constructs was used in transient transfection assays to characterize the regulatory functions of the IE gene product. These experiments demonstrated that (i) the EHV-1 IE gene product is a bifunctional protein capable of both positive and negative modulation of gene expression; (ii) the IE gene product possesses an autoregulatory function which represses the IE promoter; (iii) IE autoregulation is dependent on IE promoter sequences mapping within positions -288 to +73 relative to the transcription initiation site (+1) of the IE gene; (iv) the IE gene product can independently activate the EHV-1 tk promoter (an early promoter) by as much as 60-fold; (v) two EHV-1 beta-gamma (leaky late) promoters, those of IR5 (gene 5 in the inverted repeat) and the glycoprotein D gene, demonstrate a requirement for both the IE gene product as well as a gene product encoded within the EHV-1 XbaI G fragment for significant activation; and (vi) the IE gene product is capable of activating heterologous viral promoters.

Animals↗
PubMed1991

Correlation of cytotoxicity, filler loading and curing time of dental composites.

Previous studies have shown that dental resin composites tested in cell culture produce cytotoxic effects on human gingival tissues. In this study, the cytotoxic potential of resin composites on primary human gingival fibroblast cultures was evaluated, based on inhibition of cellular protein synthesis measured by [35S] methionine incorporation. Both resin content and percentage of monomer conversion were considered as potential causes of cytotoxicity. Three resin composites were selected to provide a range of filler content from 45 to 86 wt%. Duplicate sample discs (1 mm thick x 10 mm diameter) of each composite were polymerized for 15, 30 and 60 s, followed by heat (110 degrees C, 10 min), and the degree of monomer conversion for each sample group was measured using Fourier transform infrared spectrophotometry. Identically fabricated discs were placed into 35 mm culture dishes with gingival fibroblasts and incubated for 24 h at 37 degrees C. The cell monolayers then were labelled at 24 h with [35S] methionine, washed and solubilized; then incorporated radioactivity was quantitated by liquid scintillation spectrometry. For each composite, as the percentage of monomer conversion increased, cellular toxicity decreased. In comparing different composites having similar monomer conversions, it was found that the filler/resin ratio was not the only factor determining the composite's relative toxicity.

Biocompatible Materials↗
PubMed1991

Altered lipid metabolism in parvovirus-infected cells.

A broad spectrum of cell lipid alterations are known to occur as a consequence of various viral infections. These changes include inhibition of lipid synthesis, stimulation of lipid synthesis and changes in the proportions of various lipids. The current study examined the effects of two parvoviruses on lipids of rat kidney (NRK) cells. Cells were infected with H-1 or Kilham rat virus (KRV) and the effects on 14C-acetate incorporation determined. Results showed that H-1 virus rapidly inhibited lipid formation (in 1 h) while KRV produced a similar effect beginning around 8 h. Pretreatment of the cells with cycloheximide did not alter this response. Fatty acid analysis by gas chromatography did not reveal major alterations in this component of total cell lipids although some fatty acids became undetectable by 18 h post-infection. The data suggest that these parvoviruses, especially H-1 virus, are able to rapidly alter lipid formation following infection and that this effect may be mediated by a virion component.

Animals↗
PubMed1990

Glass ionomer and composite resin cements: effects on oral cells.

Because the cement interfaces of restorations can approximate the periodontium, it is critical to determine the biocompatibility of cements. In this study, the cytotoxic potential of resin luting agents on cultures of gingival fibroblasts and oral epithelial cells was evaluated for direct microscopic cytotoxicity, cell morbidity, impaired adherence, and inhibition of macromolecular synthesis. Visible effects ranged from severe toxicity with inadequately polymerized composite resin to no detectable morphologic cell damage by a glass ionomer cement, but inhibition of protein and RNA synthesis varied with the material and cell type. The glass ionomer cement demonstrated no morphologic damage, but exhibited inhibition of macromolecular synthesis in gingival fibroblasts. These results confirmed that in vitro metabolic assays are appropriate for examining the biologic effects of materials.

Cells, Cultured↗
PubMed1990

Temporal regulation of equine herpesvirus type 3 transcription.

The transcription of equine herpesvirus type 3 (EHV-3; equine coital exanthema virus) has been examined and found to be temporally regulated into three classes: immediate early (IE), early (E), and late (L). Hybridization of in vivo 32PO4-labeled transcripts revealed that IE transcript(s) are derived exclusively from the inverted repeat segments (IRs) of the viral genome, while E and L transcripts are not restricted to any specific region of the genome. Northern blot analysis of EHV-3 IE RNA revealed a single transcript of approximately 5.7 kb (3.8 MDa). We have previously shown that transcription of equine herpesvirus type 1 (EHV-1) DNA is temporally regulated and produces a single 6 kb IE RNA which is derived from the IRs segments. In this paper, we show that the EHV-1 and EHV-3 IE RNA species are homologous, reflecting the colinearity of the genomes of these two related viruses. While four IE polypeptides are synthesized in EHV-1 infected cells in the presence of actinomycin D following the removal of a cycloheximide block, only one major IE polypeptide (180 kDa) is detectable in EHV-3 infected cells under these conditions. However, immunoprecipitation of EHV-3 infected cell extracts with polyvalent rabbit antisera to IE1 of EHV-1 revealed at least two other viral specific IE polypeptides.

Animals↗
PubMed1989

Disinfection of visible-light-curing devices.

Assays were developed for evaluating disinfection of visible-light-curing devices which were deliberately contaminated with an indicator organism, Streptococcus mutans, and devices which were contaminated during routine clinical use. Results indicated that wiping the surface with a substituted phenolic agent followed by wrapping in gauze saturated with the same solution was most effective for disinfecting the device handle and tip surfaces. Longer contact with the disinfectant is recommended to ensure virucidal/sporicidal action.

Dental Equipment↗
PubMed1988

Characterization of equine herpesvirus type 1 immediate early proteins.

EHV-1 immediate early (IE) gene expression in lytic infection results in the production of four high mol wt immediate early polypeptides (IEPs), designated IE1, IE2, IE2, and IE4; however, IE transcription is limited to the synthesis of a single 6-kb mRNA. Together, these findings raised questions as to whether the four IEPs were related products of the same gene. In the present study the IEPs were characterized with respect to their structural similarities, antigenic relatedness, and postsynthetic modifications. IE1 was the most abundant IEP, in that it accounted for approximately 80% of the IEP-incorporated radiolabel in infected rabbit kidney cells labeled under IE conditions with [35S]methionine or 14C-labeled amino acid mixtures. IE1 also was the major phosphorylated species. Limited proteolytic digestion of isolated radiolabeled IEP bands with Staph V8 protease yielded virtually identical fragment profiles in SDS-PAGE, as did digestions with chymotrypsin and N-chlorosuccinimide. Monospecific rabbit antisera raised against each of the four isolated IEPs reacted with all the IEP species in immunoblotting assays. Pulse-chase experiments indicated that all the IEPs were detectable immediately after a 15-min pulse and that several alterations in the IEP profile occurred during subsequent chase periods. Thus, the EHV-1 IEPs are closely related structurally and antigenically and appeared to be either produced simultaneously or processed to yield the individual forms immediately.

Antibodies, Viral↗
PubMed1988

Analysis of the in vitro translation products of the equine herpesvirus type 1 immediate early mRNA.

Equine herpesvirus type 1 (EHV-1) gene expression is coordinately regulated in an alpha, beta, gamma fashion. Viral alpha gene products include a 6.0-kb immediate early (IE) mRNA species (W. L. Gray et al., 1987, Virology 158, 79-87) and at least four closely related IE polypeptides (IEPs) (G.B. Caughman et al., 1985, Virology 145, 49-61). In this report, we describe results obtained from a series of in vitro translation experiments which were performed in an effort to characterize the IEPs and identify the mechanism by which individual IE protein species are generated. Our data indicate that a family of IEPs is generated in vitro from the 6.0-kb mRNA size class and that these IEPs correspond in overall size and antigenicity to those synthesized in infected cells. Using time-course/pulse-chase analyses, we show that production of three of the major IEPs [IE1' (193 kDa), IE3' (166 kDa), and IE4' (130 kDA)] occurs concomitantly, that none of these protein species can be chased completely into another, and that at least two additional minor species appear to be processed following synthesis. Finally, we show that the 6.0-kb mRNA species isolated during early or late stages of the infection cycle can be translated to yield all of the major IE proteins, indicating that production of the family of IEPs is not dependent upon accumulation of the IE mRNA which occurs during a cycloheximide blocked infection cycle. The implications of these findings are discussed as they relate to the origin and production of the IEPs both in vivo and in vitro.

Blotting, Northern↗
PubMed1985

Equine herpesvirus type 1 infected cell polypeptides: evidence for immediate early/early/late regulation of viral gene expression.

EHV-1 polypeptide synthesis was examined in productively infected rabbit kidney and hamster embryo cells. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analyses of extracts from [35S]methionine- and 3H-amino acid-labeled-infected and mock-infected cultures revealed the presence of 30 infected cell-specific polypeptides (ICPs) which ranged in apparent molecular weights from 16.5K to 213K. Twenty-two of these ICPs comigrated with virion structural proteins. Four ICPs (203K, 176K, 151K, 129K) were detected in extracts of infected cultures labeled in the presence or absence of actinomycin D (Act D) immediately after release from a 4-hr treatment with cycloheximide (CH). These polypeptides, which were designated as EHV-1 immediate early (alpha) ICPs, were not detected in unblocked (non-CH-treated) infected cells. The most abundant ICP was a 31.5K nonstructural protein which, in addition to a 74K protein, was detected in unblocked infected cells at 2-3 hr postinfection. These proteins appeared to be regulated as early (beta) ICPs, since neither protein was observed in Act D-treated cultures released from CH block. Twelve ICPs were classified as late (gamma) polypeptides on the basis of their reduced synthesis in cultures in which viral DNA replication was inhibited by phosphonoacetic acid. All but one (40K) of these late ICPs corresponded to virion structural proteins.

Animals↗
PubMedDate not supplied

Effects of denture base resins on oral epithelial cells.

The current proliferation of light-polymerized denture base resins and the continual modification of their formulations make standardized biocompatibility testing a necessity. The biocompatibilities of three light-polymerized denture base resins were compared using an in vitro epithelial cell culture system. The effect of varied lengths of polymerization of denture base resin on cell toxicity was examined. This study indicated that light-polymerized denture base resins have an effect on oral epithelial cells that appears to be related to the specific formulation of the material and not to the type of polymerization required. Varying the polymerization time or light-polymerization unit appeared to have little effect.

Acrylic Resins↗