Biomedical subjects

D T Brown

Publications and source records attributed to D T Brown.

At least 73 records · Page 4Linked to original sources

PubMed1994

Changes in vertical tooth position and face height related to long term anterior repositioning splint therapy.

This study evaluates whether extended full-time wear of a partial coverage mandibular anterior repositioning splint (MORA) causes intrusion of posterior teeth and determines the effect on jaw position. Sixty-four patients from two private orthodontic practices were studied using cephalometric radiographs to measure vertical change in position of the anterior and posterior teeth and the mandible. The splint wear time ranged from a minimum of one half year to a maximum of 4.8 years, with a mean of 1.33 years. No significant change was recorded in the distance from the mandibular molar to the mandibular plane. On average, the maxillary incisor and maxillary molar extruded about 1 mm, while the mandibular molar was unchanged and the mandibular incisor intruded about 0.6 mm. Posterior face height increased an average of 1.6 mm, and anterior face height increased an average of 2.7 mms. In 20% of the patients, intrusion of the mandibular molars of 1 mm or more occurred. In 41%, extrusion of the maxillary incisors of 1 mm or more was noted. Intrusion of the upper molars or extrusion of the lower incisors occurred in only 5% of the patients. The data indicates that only a very small proportion of patients having long term splint therapy using the MORA have clinically significant molar intrusion. Change in mandibular position was expressed in a vertical increase in posterior and anterior face height. Only very small changes occurred in antero-posterior position.

Adolescent↗

PubMed1994

Outcome measurement for treated and untreated TMD patients using the TMJ scale.

Changes in symptom profiles of two groups of treated and untreated temporomandibular disorders (TMD) patients were measured using the TMJ Scale. Forty-nine consecutive TMD patients were included in the study. Of these, 29 patients were treated using splint therapy, physical therapy, counseling, and non-steroidal anti-inflammatory (NSAID) medications. Patients completed the TMJ Scale questionnaire prior to treatment and at completion of the TMD therapy. Sixty-five patients were not treated in any manner, having completed the questionnaire during an initial diagnostic study and having declined treatment. They were contacted by mail at least four months later and asked to complete the second questionnaire if they had not received any treatment. Twenty untreated patients returned the completed questionnaire. The initial scale scores for the two groups were similar. Comparison of the two tests of both groups revealed the treated group improved significantly, while the untreated group remained unchanged, except for a statistically non-significant increase of scale scores (increased symptom severity) for joint dysfunction and range of motion limitation. Younger patients responded more favorably to therapy than older patients. The findings do not support the conclusions of the few previous studies that found TMD are often self-limiting and rarely progress to chronic intracapsular derangement. The findings strongly support the hypothesis that symptom improvement was related to treatment and not solely to the passage of time.

Adolescent↗

PubMed1994

Formation and rearrangement of disulfide bonds during maturation of the Sindbis virus E1 glycoprotein.

The rigidly ordered icosahedral lattice of the Sindbis virus envelope is composed of a host-derived membrane bilayer in which the viral glycoproteins E1 and E2 reside. E1-E1 interactions stabilized by intramolecular disulfide bridges play a significant role in maintaining the envelope's structural integrity (R. P. Anthony and D. T. Brown, J. Virol. 65:1187-1194, 1991; R. P. Anthony, A. M. Paredes, and D. T. Brown, Virology 190:330-336, 1992). We have examined the acquisition of disulfide bridges within E1 during its maturation. Prior to exit from the endoplasmic reticulum, E1 folds via at least three intermediates, differing in the number and/or arrangement of their disulfides, into a single, compact form. This E1 species remains stable with respect to its disulfides until late in the secretory pathway, when E1 attains a metastable conformation. At this point, when appropriately triggered, intramolecular thiol-disulfide exchange reactions within E1 can occur, resulting in the generation of alternative E1 species. This metastable nature of mature E1 may have important implications for the mechanism of virus disassembly during the initial stages of the infection process (B. Abell and D. T. Brown, J. Virol. 67:5496-5501, 1993).

Animals↗

PubMed1993

Three-dimensional structure of a membrane-containing virus.

The structure of Sindbis virus was determined by electron cryomicroscopy. The virion contains two icosahedral shells of viral-encoded proteins separated by a membrane bilayer of cellular origin. The three-dimensional structure of the ice-embedded intact Sindbis virus, reconstructed from electron images, unambiguously shows that proteins in both shells are arranged with the same icosahedral lattice of triangulation number T = 4. These studies also provide structural evidence of contact between the glycoprotein and the nucleocapsid protein across the membrane bilayer. The structural organization of Sindbis virus has profound implications for the morphogenesis of the alphaviruses. The observed interactions confirm stoichiometric and specific protein associations that may be crucial for virion stability and predict a mechanism for assembly.

Animals↗

PubMed1993

Identification through overexpression and tagging of the variant type of the mouse H1e and H1c genes.

Multiple, nonallelic forms of histone H1 are found in most mammalian tissues. Attempts to determine a functional significance to this diversity is hindered by a paucity of primary amino acid sequence data. Although numerous H1 genes have been cloned, the type of variant they encode cannot be determined by sequence analysis alone. We used transformation and overexpression methods to determine that two cloned mouse H1 genes, MH143 and MH175, encode H1c and H1e, respectively. Since these genes have been completely sequenced, these results establish the amino acid sequence of these variants. Assignment was accomplished by mutagenically "tagging" the genes by incorporation of a codon for methionine, which is not found in the major somatic H1 variants. These genes were placed under the control of the mouse metallothionein I promoter and introduced into 3T3 cells. Products of these mutagenized genes were detected by [35S]methionine labeling and identified by high performance liquid chromatography and sodium dodecyl sulfate-acid polyacrylamide gel electrophoresis. We were also able to induce major alterations in the normal stoichiometry of chromatin-associated H1 variants through overproduction of H1c and H1e. This had little effect on the growth properties of these transformants.

3T3 Cells↗

PubMed1993

Purification and characterization of a Sindbis virus-induced peptide which stimulates its own production and blocks virus RNA synthesis.

The antiviral activity described by Riedel and Brown (J. Virol. 29, 51-60) has been purified to homogeneity. A study of the effects of this pure peptide on mosquito cell growth and virus replication has been undertaken. We have found that treatment of noninfected Aedes albopictus cells with this antiviral peptide results in a temporary arrest of cell division accompanied by acquisition of virus resistance and the constitutive production of the antiviral protein.

Aedes↗

PubMed1993

Phosphorylation and dephosphorylation events play critical roles in Sindbis virus maturation.

We have examined the effects of various inhibitors of protein kinases and phosphatases on Sindbis virus maturation in BHK cells. 2-aminopurine, a nonspecific protein kinase inhibitor, N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride (W-7), a specific inhibitor of calmodulin/Ca(2+)-dependent protein kinase, and okadaic acid (OKA), a protein phosphatase inhibitor, dose-dependently inhibited Sindbis virus maturation. Although virus production was inhibited, the membrane glycoprotein precursors PE2/E1 were exported from the endoplasmic reticulum and PE2 was converted to E2 at normal kinetic rates. The glycoproteins were delivered to the plasma membrane in conformations which rendered them competent for low pH-mediated cell-cell fusion from within. Electron microscopy showed that in the presence of W-7, virus nucleocapsids were free in the cell cytoplasm, while in the presence of OKA, the nucleocapsids were associated with cell membranes. Metabolic labeling of Sindbis virus-infected cells with [32P]orthophosphate in the presence of OKA resulted in the specific labeling of the PE2/E2 glycoprotein. We have previously shown that the carboxyl terminus of the PE2 glycoprotein is initially buried in cell membranes and is then exposed to the cytoplasm at some later stage in virus maturation. The data shown are consistent with the hypothesis that phosphorylation and dephosphorylation play a critical role in a late stage in Sindbis virus maturation, possibly in releasing of the E2 tail from cell membranes.

2-Aminopurine↗

PubMed1993

Recent graduates' and current dental students' evaluation of their prosthodontic curriculum.

This survey of recent graduates and third- and fourth-year dental students at Indiana University School of Dentistry determined whether the prosthodontic curriculum adequately prepared students for the prosthodontic portion of dental practice and whether there was material that should be added to or omitted from the prosthodontic curriculum. The survey evaluated 106 topics in the prosthodontic curriculum and areas related to prosthodontics. On the basis of their importance to the respondents, 59.4% of the topics were considered adequately emphasized, 2.9% of the topics were overemphasized and should be reduced in emphasis or eliminated, and 37.7% of the topics were underemphasized, which indicated the need to increase time devoted to these topics. Most of the underemphasized topics dealt with new materials, alternate techniques, private practice and its management, and the prosthodontic needs of special types of patients.

Curriculum↗

PubMed1993

Transient translocation of the cytoplasmic (endo) domain of a type I membrane glycoprotein into cellular membranes.

The E2 glycoprotein of the alphavirus Sindbis is a typical type I membrane protein with a single membrane spanning domain and a cytoplasmic tail (endo domain) containing 33 amino acids. The carboxyl terminal domain of the tail has been implicated as (a) attachment site for nucleocapsid protein, and (b) signal sequence for integration of the other alpha-virus membrane proteins 6K and E1. These two functions require that the carboxyl terminus be exposed in the cell cytoplasm (a) and exposed in the lumen of the endoplasmic reticulum (b). We have investigated the orientation of this glycoprotein domain with respect to cell membranes by substituting a tyrosine for the normally occurring serine, four amino acids upstream of the carboxyl terminus. Using radioiodination of this tyrosine as an indication of the exposure of the glycoprotein tail, we have provided evidence that this domain is initially translocated into a membrane and is returned to the cytoplasm after export from the ER. This is the first demonstration of such a transient translocation of a single domain of an integral membrane protein and this rearrangement explains some important aspects of alphavirus assembly.

Amino Acid Sequence↗

PubMed1993

The distribution of Sindbis virus proteins in mosquito cells as determined by immunofluorescence and immunoelectron microscopy.

Two Aedes albopictus (mosquito) subclones, C7-10 and C6/36, were examined by immunofluorescence and immunoelectron microscopy for the distribution of Sindbis virus structural and non-structural proteins. Both the viral glycoproteins, E1 and E2, and the non-structural proteins, nsP1 and nsP2, were found within vesicles and electron-dense, amorphous matrices associated with Sindbis virus infection. The labelling patterns indicated that both replication of viral RNA and production of virus particles were localized within the same structures in the infected cell. The data support previous reports that alphavirus infection is contained within specific structures in the cytoplasm and provide additional evidence that the C6/36 and C7-10 subclones may represent different tissue types in the adult insect.

Aedes↗

PubMed1993

Purification and properties of cyanide hydratase from Fusarium lateritium and analysis of the corresponding chy1 gene.

The filamentous fungus Fusarium lateritium is cyanide tolerant, due, at least in part, to the induction by cyanide of the enzyme formamide hydrolyase (EC 4.2.1.66). This enzyme, more commonly known as cyanide hydratase, catalyses the hydration of cyanide to formamide. The enzyme was purified from F. lateritium and showed a subunit molecular mass of 43 kDa (as judged by SDS-PAGE), while the native protein appeared to form aggregates of up to 1217 kDa (as judged by gel-filtration and non-denaturing PAGE). mRNA samples from cultures grown with and without cyanide were in vitro translated and immunoprecipitated. This demonstrated that, in this species, the gene encoding the enzyme designated chy1, is cyanide inducible. Differential screening was used to isolate a cyanide hydratase cDNA clone which was subsequently used to obtain the corresponding genomic clone. A fragment of the cDNA clone encoding all but the first seven amino acids of the protein was expressed in E. coli using the expression vector pGEX-2T. Features of F. lateritium cyanide hydratase together with an analysis of the nucleotide sequence encoding this enzyme are presented.

Amino Acid Sequence↗

PubMed1993

Sindbis virus membrane fusion is mediated by reduction of glycoprotein disulfide bridges at the cell surface.

We have examined the role of thiol-disulfide exchange reactions during the penetration of cells by Sindbis virus. The protein-protein association that form the rigid icosahedral lattice of the Sindbis virus envelope have been shown to be stabilized by disulfide bridges, and reduction of these critical disulfide bridges during cell penetration may be the mechanism by which the rigid protein lattice is disrupted prior to fusion (R. Anthony and D. T. Brown, J. Virol. 65:1187-1194, 1991; R. Anthony, A. Paredes, and D. T. Brown, Virology 190:330-336, 1992). Reduction of disulfide bridges occurs at near neutral pHs via thiol-disulfide exchange reactions, and these reactions can be blocked by covalent modification of the thiol involved. In this study, the effects of the reducing agent 2-mercaptoethanol on Sindbis virus-mediated cell-cell fusion from without and the effects of the membrane-impermeable thiol-alkylating reagent 5,5'-dithiobis(2-nitrobenzoic acid) on Sindbis virus penetration were determined. The presence of exogenous reducing agent was found to induce fusion from without under conditions unfavorable to both typical Sindbis virus-mediated fusion from without and cysteine-mediated thiol-disulfide exchange reactions. In addition, the thiol-alkylating reagent was found to inhibit Sindbis virus entry when present during infection. These results are consistent with a model for Sindbis virus entry in which reduction of critical disulfide bridges at the cell surface disrupts the rigid protein-protein associations of the envelope, allowing membrane fusion and release of the viral genome into the cell.

Animals↗

PubMed1993

Force-moment systems on single maxillary anterior implants: effects of incisal guidance, fixture orientation, and loss of bone support.

An engineering model of maxillary single anterior implants was developed to calculate the force-moment systems resulting from occlusal contacts. Analysis of a clinical example showed that flattening the incisal guidance and/or placing the implant in a more upright position reduced the moment and the labial component of the force but increased the intrusive component. The calculations are applicable to the design of implants and may help in the long-term evaluation of clinical success.

Algorithms↗

PubMed1992

Effects of environmental factors on maxillofacial elastomers: Part II--Report of survey.

Part II of a four-part report presents an international survey that was conducted to determine the most frequently used materials in the fabrication of facial and somatoprostheses. The survey also solicited information about advantages and disadvantages, methods of coloring, and the perceived properties of an "ideal" material. The results of the survey indicated that the majority of prosthodontists and prosthetists are using room temperature-vulcanized silicone products, intrinsically colored with dry earth pigments or artist's pigments. The need for further research in materials for external prostheses was clearly indicated.

Color↗

PubMed1992

The mass of the Sindbis virus nucleocapsid suggests it has T = 4 icosahedral symmetry.

The mass of the nucleocapsid of the alphavirus Sindbis was determined by scanning transmission electron microscopy. The nucleocapsid was found to have an average mass of 10.54 x 10(6) Da with the largest percentage of particles showing a mass of 10.7 to 10.9 x 10(6) Da. These data predict that the nucleocapsid contains 240 copies of protein and is a T = 4 icosahedron.

Capsid↗

PubMed1992

Disulfide bonds are essential for the stability of the Sindbis virus envelope.

Sindbis virus is a membrane-containing virus which has two glycoproteins organized in an icosahedral lattice. Protein-protein associations have been identified which participate in the formation of the icosahedron and these associations are stabilized by intramolecular disulfide bridges (Anthony, R. P., and Brown, D. T., 1990, J. Virol. 65, 1187-1194). The present study further examines the role of disulfides in the structure and function of Sindbis virus by following the effect of dithiothreitol on the protease sensitivity of envelope proteins as well as the electron microscopic appearance and infectivity of Sindbis virus. Treatment of isolated virus with 5 mM dithiothreitol for 6 hr causes a marked increase in trypsin sensitivity of both E1 and E2, profound morphological alterations in the viral envelope, increased susceptibility of the nucleocapsid to RNase, and 95% loss of infectivity. These effects are greatly enhanced and accelerated when treatment with DTT is preceded by a brief exposure of the virus to pH 5.3, suggesting that acid-induced conformational changes render structurally critical disulfides more accessible to reductive cleavage by DTT. When compared to other manipulations known to change the conformation of the viral envelope, such as heating to 51 or 60 degrees or exposure to acid pH, only the exposure to DTT with or without prior acid treatment caused marked structural changes correlated with a loss of infectivity. These data provide electron microscopic and functional evidence that intact disulfide bonds are critical for the stability of the virus envelope and suggest that the cleavage of critical disulfide(s) may play a role in the process of virus infection.

Animals↗

PubMed1992

The effect of monoclonal antibodies to calcitonin gene-related peptide (CGRP) on CGRP-induced vasodilatation in pig coronary artery rings.

1. The modification of the vasodilator effect of calcitonin gene-related peptide (CGRP) by a panel of monoclonal antibodies (MAbs), which map to discrete epitopes on the CGRP molecule, was investigated in pig coronary artery rings (PCA). The preparations were pre-constricted with acetylcholine (3 x 10(-7) M) and concentration-response curves to CGRP (2 x 10(-10)-2.56 x 10(-8) M) were obtained in the presence or absence of each MAb. 2. CGRP caused a concentration-dependent relaxation of PCAs which reached a maximum (98.2 +/- 4.8%, n = 25) at 1.28 x 10(-8) M and gave an EC50 of 3.8 +/- 0.8 x 10(-9) M. 3. Two MAbs which map to the N-terminal, CN1 and CRA3, did not affect the CGRP response whilst a third, CRA5, significantly inhibited its effect. 4. The C-terminal MAb, CRA2, did not modify the CGRP response whilst, in contrast, CB3 (C-terminal) potentiated its effect. A similar augmentation of the CGRP-induced vasodilatation was seen in the presence of the middle-region MAb, CRA8. 5. These results suggest that regional specific MAbs can modify the vasodilator effect of CGRP causing either inhibition (CRA5, N-terminal) or potentiation (CB3, C-terminal; CRA8, middle region).

Analysis of Variance↗

PubMed1992

Morphogenesis of Sindbis virus in three subclones of Aedes albopictus (mosquito) cells.

The morphogenesis of Sindbis virus in three Aedes albopictus subcloned cell lines was examined. Each line was distinguishable with respect to morphology, cytopathic response to infection, and progeny yield. C7-10 cells, which produced the highest titers of virus and exhibited the most severe cytopathic response, were characterized ultrastructurally by the presence of budding particles at the cell surface and at the membranes of internal vesicles. C6/36 cells, which displayed a moderate cytotoxic response, manifested similar features in response to Sindbis virus infection. Both cell types also produced a structure composed of an electron-dense matrix in which nucleocapsids were embedded. Internally matured virions were released by exocytosis from these cells. In addition to a lack of cytopathic effect, u4.4 cells also failed to exhibit obvious morphogenetic changes upon infection. Virus particles were occasionally seen within vesicles, but budding at the cell surface was not detected. The mechanism of release of internally matured virions was not apparent. These studies provide further evidence that these three subcloned mosquito cell lines represent different tissues in the larval or adult insect.

Aedes↗