Characteristics identified by deans as essential to success.
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Achievement↗
Biomedical subjects
W J O'Brien
Publications and source records attributed to W J O'Brien.
Maintenance of replicative intermediates in ganciclovir-treated human cytomegalovirus-infected retinal glia.
OBJECTIVES: To characterize the molecular structure of the human cytomegalovirus (HCMV) DNA maintained in cultures of human retinal glia following ganciclovir treatment and to determine the biological activity of the DNA. METHODS: Cultures of human retinal glia were established, infected with HCMV, treated with ganciclovir, and embedded in agarose, and the viral DNA was analyzed by field inversion gel electrophoresis. RESULTS: The HCMV DNA was found to persist in cultures of infected, ganciclovir-treated retinal glial cells in the form of replicative intermediates. After removal of ganciclovir, processed forms of DNA in the 500-to 1000-kilobase range were found as well as 230-kb unit length genome. Infectious virus was recovered after termination of ganciclovir treatment. CONCLUSION: The data are consistent with the concept that ganciclovir's virostatic nature permits maintenance of HCMV DNA in retinal glia in a biologically active form that is capable of replication after removal of the drug.
Antiviral Agents↗
Production of ISG-15, an interferon-inducible protein, in human corneal cells.
ISG-15, a 15-kDa protein encoded by an interferon (IFN)-stimulated gene (ISG), was produced in human corneal cell cultures prepared from donor corneas in response to each of three major types of IFN, IFN-alpha, IFN-beta, and IFN-gamma. IFN-alpha and IFN-beta induced more ISG-15 in the first 24 h of treatment than did IFN-gamma. ISG-15 was detectable within the first 3 h of treatment with either type I IFN, and production peaked at 24 h, whereas IFN-gamma did not induce detectable ISG-15 until 16 h and did not induce peak production until 48 h. Conjugates of ISG-15 to cellular proteins were detectable by Western blot beginning at 9 h after IFN-alpha or IFN-beta treatment. ISG-15 persisted in IFN-treated cells for as long as 96 h. Free, unconjugated, ISG-15 was secreted by keratocytes into cell culture medium. The extent and kinetics of production, the conjugation of ISG-15 to cell proteins, and the secretion of ISG-15 in human fibroblast derived from corneas were IFN type dependent and dose dependent.
Cells, Cultured↗
Herpetic stromal disease: response to acyclovir/steroid therapy.
The efficacy of combined acyclovir and steroid therapy in the treatment of herpetic stromal disease was evaluated by clinical evaluation of disease, the rebound of disease following termination of therapy, and the recovery of virus and viral DNA from corneas in a rabbit model. Therapy with acyclovir alone produced a significant reduction in corneal thickness in 10% of eyes. Addition of steroid to acyclovir therapy decreased the severity of stromal disease as measured by corneal thickness and increased the frequency of response to treatment to 63% of eyes. All eyes receiving acyclovir alone experienced rebound of disease following termination of therapy. Combined therapy increased the severity of rebound of corneal disease. Virus was recovered from cell cultures established after recovery from rebound in 50% of untreated and treated eyes. Viral DNA was detected by PCR in five of the nine corneal cultures which did not produce infectious virus.
Acyclovir↗
Assessment of antiviral activity, efficacy, and toxicity of prostaglandin A2 in a rabbit model of herpetic keratitis.
Prostaglandin A2 (PGA2) inhibited the replication of herpes simplex virus type 1 in rabbit and human cornea stromal cells at concentrations of 1 to 5 microM while causing significant toxicity at 55 to 150 microM. Despite favorable therapeutic indices in cultured cells, PGA2 was not effective as a therapeutic agent in the treatment of herpetic keratitis in a rabbit model. The sequelae of disease appeared more severe in animals receiving PGA2 than in untreated or placebo-treated controls. The recovery of virus from tissues of latently infected rabbits was not affected by therapy. PGA2 therapy alone induced breakdown of the blood-aqueous barrier, indicating that pharmacologically active concentrations of drug were achieved in the eye. Thus, PGA2 had antiviral activity, but its proinflammatory effects appeared to be more detrimental than beneficial in the treatment of herpetic keratitis.
Animals↗
Endothelial barrier function and Na+/K(+)-ATPase pump density in herpetic stromal disease.
PURPOSE: Corneal edema is a significant component of the various forms of herpes simplex virus type 1 (HSV-1)-induced stromal disease. Maintenance of corneal thickness, a reflection of corneal hydration, depends on a physical barrier formed by endothelial cell-cell junctions and by the activity of Na+/K(+)-ATPase pumps that regulate ion flux and thus influence water movement through this cell layer. These functions were measured in corneas with increased corneal thickness caused by HSV-1-induced stromal disease to determine their contribution to the pathogenesis of the edema. METHODS: Stromal disease with corneal edema was induced in rabbits by intrastromal injection of the RE strain of HSV-1. At various times after infection, during the development of and recovery from stromal disease, endothelial barrier function and Na+/K(+)-ATPase pump sites were measured in excised rabbit corneas. RESULTS: The endothelial permeability coefficient, Ktrans, for 14C-dextran, 3H-inulin, and 14C-mannitol, were not altered significantly during periods of maximal corneal edema and stromal disease. Endothelial Na+/K(+)-ATPase pump density, as measured by ouabain binding, showed a statistically significant (P < 0.05) decrease in HSV-1-infected corneas during peak edema compared to mock antigen-injected or uninjected control corneas. Pump density returned to baseline values by 24 days after infection, concurrent with the resolution of corneal edema. CONCLUSIONS: These results indicate that corneal endothelial barrier function was not altered in this form of HSV-1-induced stromal edema; however, pump density was reduced significantly.
Animals↗
Replication of human cytomegalovirus in human retinal glial cells.
PURPOSE: The purpose of these studies was to characterize the replication cycle of human cytomegalovirus (HCMV) in human retinal glial cells in vitro. METHODS: Cultured human retinal glial cells were exposed to HCMV strain AD169 or low-passage clinical isolates for a 2-hour adsorption period and then incubated in the appropriate growth medium at 37 degrees C. Cultures were examined by microscopy for cytopathic effect and by immunofluorescence staining using monoclonal antibodies directed against immediate-early, early, and late HCMV proteins. Viral DNA was analyzed by field inversion gel electrophoresis and detected using Southern blot analysis or the polymerase chain reaction. RESULTS: Immunocytochemical staining revealed that the glial cells expressed all three classes of HCMV proteins and that infectious virus could be transferred from the medium of the infected cultures to susceptible MRC-5 cell monolayers. Less than 1% of the glial cells expressed the S-phase enzyme, thymidine kinase, at the time of infection compared to MRC-5 fibroblasts, of which 81% expressed it. Progeny virus was found to be highly cell associated in glial cells (80%) at peak virus titer compared to MRC-5 cells (39% cell associated at peak titer). Four low-passage clinical isolates of HCMV from patients with acquired immune deficiency virus also productively infected cultures of human retinal glial cells. Field inversion gel electrophoresis of HCMV-infected glial cell lysates was performed to identify the replicative forms of DNA. Southern blots probed with HCMV-specific probes showed that HCMV DNA replication proceeds through high molecular weight intermediates before forming the 230-kb unit length genome. CONCLUSIONS: The full permissive replication of HCMV in human retinal glial cells indicates that glial cells are a likely site of HCMV replication in the retina and thus may play an important role in the pathogenesis of HCMV retinitis.
Adolescent↗
Sepsis increases skeletal muscle sodium, potassium-adenosinetriphosphatase activity without affecting messenger RNA or protein levels.
BACKGROUND: Previous studies suggest that sodium, potassium-adenosinetriphosphatase (Na+, K(+)-ATPase) activity in skeletal muscle is increased during sepsis, but the molecular mechanisms are not well understood. We tested the hypothesis that increased muscle Na+,K(+)-ATPase activity during sepsis is associated with increased expression of messenger RNA for the Na+,K(+)-ATPase alpha-1, alpha-2, and beta-1 subunits and increased amounts of the enzyme protein. STUDY DESIGN: Extensor digitorum longus muscles were harvested from rats 16 hours after induction of sepsis by cecal ligation and puncture or sham operation. The Na+,K(+)-ATPase activity was determined spectrophotometrically. Messenger RNA levels for the alpha-1, alpha-2, and beta-1 subunits of Na+,K(+)-ATPase were determined by Northern blot analysis. Enzyme protein levels were measured by Western blot analysis and tritium-ouabain binding assay. RESULTS: Muscle Na+,K(+)-ATPase activity was 46 percent higher in rats that had sepsis than in rats that underwent sham operation (p < .05). No significant differences between septic and control groups were noted in messenger RNA levels for the Na+,K(+)-ATPase subunits. Western blot analysis and tritium-ouabain binding revealed no difference in enzyme protein expression between septic and control animals. CONCLUSIONS: Results suggest that sepsis increases skeletal muscle Na+,K(+)-ATPase activity without affecting enzyme messenger RNA or protein levels. The results are consistent with an increased catalytic constant for individual Na+,K(+)-ATPase protein units, an atypical mechanism for this enzyme.
Animals↗
Comparison of scleral tensile strength after transscleral retinal cryopexy, diathermy, and diode laser photocoagulation.
OBJECTIVES: To determine the effect of transscleral diode photocoagulation on the tensile strength of sclera in an experimental rabbit model and to compare it with that of transscleral cryotherapy and diathermy. METHODS: Twenty-four Dutch-belted rabbits received one of the following three treatment modalities in one eye: (1) transscleral cryotherapy, (2) transscleral diathermy, or (3) transscleral diode laser photocoagulation. The opposite eye served as a paired control. Tensile strength measurements of scleral strips excised from areas of treatment were obtained 6 weeks later. RESULTS: No statistically significant difference in mean tensile strength was observed between eyes receiving transscleral cryopexy (n = 7) or transscleral diode photocoagulation (n = 8) and their corresponding, paired, control eyes. In contrast, transscleral diathermy reduced mean scleral tensile strength to 26% of that of the paired control eyes (n = 8, P = .0001). CONCLUSIONS: In this rabbit model, scleral weakening is significant following transscleral diathermy while transscleral cryopexy or transscleral diode photocoagulation produces no significant weakening relative to paired, untreated controls.
Animals↗
2',5'-oligoadenylate synthetase in interferon-alpha- and acyclovir-treated herpes simplex virus-infected cells.
The 2',5'-oligoadenylate (2-5A) synthetase pathway, induced by interferon-alpha (IFN-alpha), has been shown to be responsible for the antiviral action of IFN-alpha against some viruses. Studies were done to determine the role of this pathway in the anti-herpes simplex virus (HSV) action of IFN-alpha alone or in combination with acyclovir (ACV), a combination that leads to synergistic anti-HSV activity. Treatment of human corneal cells or Vero cells with 100 IU/ml of IFN-alpha induced expression of 2-5A synthetase mRNA and a 10-fold increase in 2-5A synthetase production compared with untreated cells. HSV infection alone did not induce 2-5A synthetase production, but when IFN-alpha-treated cells were infected with HSV, enzyme level was significantly increased (p < 0.05) compared with that in IFN-alpha-treated, uninfected cells. HSV infection actually decreased the level of 2-5A synthetase mRNA in IFN-alpha-treated cells. Although IFN-alpha treatment induced high levels of 2-5A synthetase with or without HSV infection, no activation of the latent endonuclease was detected by specific cleavage of ribosomal RNA. Treatment of infected cells with 5 microM ACV alone or combined with IFN-alpha did not increase 2-5A synthetase or endonuclease activities above those detected in cells not treated with ACV. The data indicate that the 2-5A synthetase pathway was inducible in corneal cells and Vero cells but did not appear to contribute to the anti-HSV activity of IFN-alpha alone or the synergistic activity of IFN-alpha combined with ACV.
2',5'-Oligoadenylate Synthetase↗
Effect of interferon-alpha on the steady-state levels of thymine, thymidine, guanine, and guanosine metabolites in herpes simplex virus-type 1-infected cells.
Interferon-alpha (IFN-alpha) combined with acyclic guanine analogs synergistically inhibits replication of herpes simplex virus type 1. IFN-alpha treatment influenced the metabolism of exogenously supplied nucleobases and nucleosides in a manner expected to contribute to synergistic activity. IFN-alpha treatment of infected human cornea stromal cells or Vero cells significantly reduced steady-state levels of acid soluble metabolites of thymine, as well as thymidine, that accumulate early during virus replication but did not affect metabolism of thymine and thymidine in uninfected cells. IFN-alpha treatment significantly reduced the ability of uninfected cells to accumulate acid-soluble metabolites from guanine, but not guanosine. The effects of IFN-alpha on nucleobase/nucleoside metabolism could contribute to synergistic antiviral activity by reducing the accumulation of thymidine/thymine metabolites and decreasing the guanine taken into cells.
Acyclovir↗
Dialogue between faith and science: the role of the hospital chaplain.
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Aged↗
Thermal shock resistance of core reinforced all-ceramic crown systems.
The coefficient of thermal expansion of core and veneer porcelains for all ceramic crowns must be compatible to prevent formation of stresses during the firing cycles. The purpose of this study was to use a thermal shock test to evaluate the crazing resistance of six all-ceramic crown systems. The systems tested were: Vita In-Ceram, Vita Hi-Ceram, Dicor veneered with Vitadur N, magnesia core veneered with Ceramco II, magnesia core veneered with Silhouette, and magnesia core veneered with Vita VMK 68. A maxillary central incisor was prepared with 1.0 mm axial and 1.5 mm incisal reduction. Refractory dies were prepared and crowns were fabricated using the manufacturers' recommendations. Six specimens of each system were placed inside a beaker in a furnace at 90 degrees C. After 15 min heat soak, ice water was poured into the beaker through a funnel. The samples were dried in air, reheated, and inspected for crazing using light microscopy. If crazing was observed, this would constitute a failure at a temperature difference (delta T) of 90 degrees C. If no failure occurred, the test was repeated at increasing temperature increments of 10 degrees C until failure. The crazing resistance of each system, expressed as the mean delta T, was: > 200 (Hi-Ceram), 163 +/- 14 (In-Ceram), 152 +/- 19 (Dicor/Vitadur N), 143 +/- 5 (magnesia core/Vita VMK 68), 122 +/- 21 (magnesia core/Silhouette), and 118 +/- 10 (magnesia core/Ceramco II) degrees C. All systems tested resisted crazing due to quenching in ice cold water from 100 degrees C. Systems with lower coefficients of expansion showed significantly greater resistance to thermal shock using ANOVA.
Ceramics↗
Leucite content of selected dental porcelains.
Leucite is a major crystalline component of dental porcelains. The presence of tetragonal leucite in dental porcelains increases their coefficients of thermal expansion due to its high coefficient of thermal expansion (20-25 x 10(-6)/degrees C). This is particularly useful for those porcelains designed for bonding to precious metals and nickel alloys. The purpose of this study was to determine the leucite content of selected commercial dental porcelains in relation to their coefficient of thermal expansion values. The weight fraction of leucite was determined with quantitative x-ray diffraction using copper as an internal standard. Coefficient of thermal expansion values were determined using a thermal dilatometric analyzer. Five commercial body porcelains were studied. Leucite was not detected in samples of Vitadur N and Duceram LFC. An ANOVA showed that there was a significant difference in the weight fraction of leucite for Silhouette, Ceramco II, and Optec HSP porcelains. Linear regression revealed a correlation (R = 0.91) between weight fraction of leucite and the coefficient of thermal expansion for those samples containing leucite. Duceram LFC, which is recommended by the manufacturer for use with metals and leucite-containing porcelains, had no detectable leucite although the coefficient of thermal expansion was found to be 13.2 +/- 0.4 x 10(-6)/degrees C at 25-472 degrees C. A low glass transition temperature contributed to the high average coefficient of thermal expansion value.
Aluminum Silicates↗
Ouabain binding kinetics of the rat alpha two and alpha three isoforms of the sodium-potassium adenosine triphosphate.
The Na,K-ATPase has three alpha isoforms which differ in cardiac glycoside sensitivity and tissue distribution. The rodent alpha 1 isoform is relatively resistant to cardiac glycosides, while the alpha 2 and alpha 3 isoforms are quite sensitive. Because both the alpha 2 and alpha 3 isoforms are generally expressed in the same tissue, it has been difficult to differentiate and accurately determine the kinetics of ouabain binding to these isoforms. To more fully understand the interactions of the alpha 2 and alpha 3 isoforms with cardiac glycosides, the association and dissociation rates of ouabain binding were measured in transfected cell lines. cDNA's coding for the rat alpha 2 and alpha 3 isoforms were transfected into NIH 3T3 cells and characterized by Na,K-ATPase activity and [3H]ouabain binding. By individually expressing the alpha 2 and alpha 3 isoforms in ouabain-insensitive NIH 3T3 cells, the ouabain-binding characteristics of each isoform could be accurately determined. The association rate constants of the alpha 2 and alpha 3 isoforms were similar while the dissociation rate constant was 33 times slower for the alpha 3 isoform than the alpha 2 isoform. Calculation of the dissociation constant (Kd) from these rate constants yielded values of 115 and 1.6 nM for rat alpha 2 and alpha 3 isoforms, respectively. Scatchard analysis of the rat alpha 2 isoform produced a similar value for Kd of 37 +/- 9 nM. Inhibition of Na,K-ATPase activity indicates the rodent alpha 1 isoform has an IC50 1000-fold higher than the alpha 2 or alpha 3 isoform at 4.8 x 10(-5) M. The results are consistent with the hypothesis that the order of ouabain affinity between the rat alpha isoforms of the Na,K-ATPase is alpha 3 > alpha 2 >> alpha 1.
3T3 Cells↗
Traumatic delivery: a risk factor for bleeding during ECMO?
Traumatic delivery is associated with orthopedic, nervous, and soft tissue injuries. Anticoagulation for extracorporeal membrane oxygenation (ECMO) support may predispose to hemorrhage from these birth injuries. A case of delayed hemoperitoneum secondary to hepatic laceration after delivery and ECMO, not previously reported, is detailed herein.
Birth Injuries↗
Catheter-induced urinary bladder rupture presenting with pneumoperitoneum.
Neonatal bladder injury is rare and usually associated with umbilical artery catherization. Patients may present with apparent renal failure, abdominal distension, and respiratory distress. Treatment involves operative closure of the perforation and bladder drainage. A case of Foley catheter-induced bladder rupture in a premature infant, not previously reported in the literature, is detailed.
Female↗
Evaluation of some properties of an opaque porcelain fired simultaneously with the body porcelain.
Recently, a porcelain-fused-to-metal opaque porcelain was introduced that does not require a separate firing before application of the body porcelain. The objective of this study was to determine the properties of this new opaque porcelain and its ability to bond to metal. The properties studied included flexural strength, linear firing shrinkage, coefficient of thermal expansion, powder particle size, and ability to bond to body porcelain and dental alloys. Sintering of this opaque porcelain was complete when fired at 1760 degrees F (960 degrees C) with a linear firing shrinkage of 13.1% +/- 0.2%. No boundary between the opaque and body porcelains could be found with a scanning electron microscope after firing at 1760 degrees F (960 degrees C). The mean flexural strengths were 99 +/- 7 and 101 +/- 8 MPa respectively, for this opaque porcelain and a conventional opaque porcelain, and were not significantly different as assessed with Student's t-test (p = 0.548). The coefficients of thermal expansion for this opaque porcelain was 13.3 +/- 0.2 x 10(-6)/degrees C. Particle size analysis showed a 63% increase in the particles below 5 microns for this opaque porcelain and bonding to two alloys was adequate as indicated by its cohesive failure. Simultaneous firing of this special opaque porcelain and body porcelain produced satisfactory sintering, strength, and bonding to metal.
Chemical Phenomena↗