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Biomedical subjects

W J O'Brien

Publications and source records attributed to W J O'Brien.

At least 55 records · Page 3Linked to original sources
PubMed1994

Regulation of herpes simplex virus thymidine kinase in cells treated with a synergistic antiviral combination of alpha interferon and acyclovir.

Alpha interferon (IFN-alpha) and acyclovir (ACV) are synergistic in their anti-herpes simplex virus activities. IFN-alpha treatment reduced the herpes simplex virus thymidine kinase (TK) activity present in cells 6 h postinfection, while steady-state levels of TK mRNA remained at or above the amount in infected, untreated cells. The inhibition of TK production by IFN-alpha treatment appeared to be transient and translational, not transcriptional.

Acyclovir↗
PubMed1994

Novel, ultrasensitive, Q-beta replicase-amplified hybridization assay for detection of Chlamydia trachomatis.

A sensitive, nonisotopic hybridization assay termed "dual capture" is described. The assay rapidly and specifically detects very low levels of target nucleic acids and organisms. The assay is based on the principles of sandwich hybridization, reversible target capture, and Q-Beta replicase amplification. The assay can be completed in less than 4 h, and in the described model format, it detects Chlamydia trachomatis rRNA or rDNA. Up to 96 samples can be analyzed simultaneously. The assay employs two types of probes: a test-specific capture probe, which mediates the cycling of the target probe complex on and off derivatized magnetic beads, and a replicatable RNA detector molecule containing a sequence complementary to and adjacent to the capture probe site on the target. Following reversible target capture, detection of the signal is accomplished by replication of the detector molecule by Q-Beta replicase in the presence of propidium iodide. A specific assay signal can be detected from as few as 1,000 molecules above the background. In a limited study of 94 urogenital samples the assay detected five of the six culture-positive samples and did not detect the C. trachomatis target in 85 of the 88 culture-negative samples.

Base Sequence↗
PubMed1994

Corneal ablations produced by the neodymium doped yttrium-lithium-fluoride picosecond laser.

This study examines corneal ablations produced by the neodymium doped yttrium-lithium-fluoride (Nd:YLF) picosecond laser. The laser delivers a 1-KHz, 40-ps pulsed, 1,053-nm wavelength beam (with energy measured in microjoules) to a 15-microns diameter spot size. The ablation mechanism is by plasma formation, which generates acoustic shock waves. Using enucleated rabbit (n = 25) and human donor eyes (n = 29), corneas were examined after tissue ablation at energies ranging from 40 to 300 microJ per pulse with various programmed ablation depths and patterns. The histologic data were collected using light microscopy and transmission electron microscopy. The tissue effects and Nd:YLF laser functions studied were ablation thresholds, cutting ability, programmed ablation depth accuracy, and acute endothelial effects. Our study showed histologic ablation thresholds for the following human corneal layers: epithelium = 7.15 +/- 0.05 x 10(11) W/cm2 (34.1 +/- 8.1 J/cm2 per pulse, 50 microJ per pulse); Bowman's layer = 1.33 +/- 0.29 x 10(12) W/cm2 (58.5 +/- 3.3 J/cm2 per pulse, 100-110 microJ per pulse); stroma and endothelium = 7.10 x 10(11) W/cm2 (28.4 J/cm2 per pulse, 50 microJ per pulse). Depth of corneal ablation was found to be directly related to energy and independent of programmed ablation depth. This study shows the endothelial loss in rabbit corneas by energy beams (50 microJ per pulse) focused 100 microns from this layer.

Animals↗
PubMed1994

The mechanism of ablation of corneal tissue by the neodymium doped yttrium-lithium-fluoride picosecond laser.

This study examines the structural changes in cornea resulting from plasma formation and propagated acoustic shock waves produced by the neodymium doped yttrium-lithium-fluoride (Nd:YLF) picosecond laser. Human donor eyes and enucleated rabbit eyes were subjected to various ablation patterns at energies ranging from 40 to 300 microJ per pulse. Two distinctly different patterns were produced depending on the location of initial plasma formation. Plasmas initiated at the corneal surface produced smooth, straight-edged ablations of corneal tissue that consisted of collagen fibril fragmentation, fibril organizational disruption, and possible thermal effect observed along the lateral borders and wound apex. The extent of lateral damage was directly related to the energy applied. The range of acute collagen disorganization observed at the ablation edge in rabbit corneas at various pulsed energies was as follows: 50 microJ = 1.0-12 microns, 150 microJ = 3.8-12.5 microns, 250 microJ = 6.2-23.7 microns, and 300 microJ = 7.5-45.0 microns. Plasma formation initiated within the stroma at or above threshold energies (50-150 microJ per pulse) produced an inter- or intralamellar separation effect with little evidence of ablation or collagen fibril fragmentation. Intrastromal plasmas generated from higher energies (200-300 microJ per pulse) produced tissue ablation, along with ablation or disruption of tissue anterior to the intrastromal target area.

Animals↗
PubMed1993

Amino acid residues of the Na,K-ATPase involved in ouabain sensitivity do not bind the sugar moiety of cardiac glycosides.

The identification of amino acid substitutions in the alpha subunit of the Na,K-ATPase that alter cardiac glycoside sensitivity provides a unique opportunity to examine the role these residues play in binding to the structural domains of these drugs. Substitution of a residue(s) involved in binding to the sugar moiety of cardiac glycosides would be expected to yield similar affinities for ouabain and its aglycone, ouabagenin. Sheep Na,K-ATPase alpha 1 subunit amino acid substitutions previously shown to influence ouabain sensitivity were tested for activity in the presence of ouabain and ouabagenin. These substitutions included both transmembrane and extracellular regions: C104F, D121E, N122D, Q111K, N122K, and Q111R, A112S. Na,K-ATPase activity versus drug concentration curves yielded I50 values over a 1000-fold range for wild type HeLa and HeLa transfectants. Interestingly, the I50 ratio for ouabagenin to ouabain in all mutants tested indicated a 19-24-fold lower affinity of the Na,K-ATPase for ouabagenin. This constant ratio among all mutations tested implies that the first transmembrane region and the first extracellular loop (H1-H2) do not participate in the binding of the sugar moiety of cardiac glycosides.

Amino Acid Sequence↗
PubMed1993

The strengthening mechanism of a magnesia core ceramic.

A high-expansion core material containing magnesia and forsterite may be used to make all-ceramic dental crowns with porcelain-fused-to-metal body porcelains. The purpose of this study was to investigate the strengthening mechanism for the magnesia core material. Six batches of the magnesia core material were made by reacting magnesia with a silica glass with holding times ranging from 17 to 120 min. The flexural strength was measured using three-point loading according to the ISO specification for dental ceramics. The forsterite content was measured using quantitative x-ray diffraction. A statistically significant correlation was found between the forsterite content and flexural strength. The proposed mechanism for strengthening is the precipitation of fine forsterite crystals in the glass matrix surrounding unreacted magnesia. Longer reaction times produced more dissolution of magnesia and subsequent precipitation of forsterite. This method results in a new strengthening mechanism for dental ceramics which have previously relied on the incorporation of alumina, leucite or ceramic whiskers.

Analysis of Variance↗
PubMed1992

Infection with a plasmid-free variant Chlamydia related to Chlamydia trachomatis identified by using multiple assays for nucleic acid detection.

Clinical samples in transport media from 40 patients exhibiting pathologies potentially caused by Chlamydia trachomatis infection were analyzed for chlamydial nucleic acid, and the results were compared with those of culture. Chlamydial culture was performed by a shell vial centrifugation method with HeLa 229 host cells. Polymerase chain reaction (PCR) assays were used to detect either regions on a 7.5-kb plasmid characteristic of C. trachomatis (plasmid-PCR) or a segment of the 16S rRNA genes (rRNA-PCR). All PCR results were confirmed by hybridization with probes for the specific amplified products in either a Southern or a dot blot format. An RNase protection (RNP) assay was used to detect genus-specific chlamydial 16S rRNA directly from the clinical samples. The PCR assays detected C. trachomatis but not other bacteria, including Chlamydia spp. C. trachomatis was isolated from six samples which were positive by the rDNA-PCR and plasmid-PCR assays. Five of the culture-positive specimens were positive by the RNP assay. Twenty-two samples were negative by all criteria. Surprisingly, nine samples were positive by rRNA-PCR and RNP assays only. Nucleic acid sequencing of the rRNA-PCR-amplified products indicated a close relationship between the variants and C. trachomatis. The data may indicate an unrecognized process in C. trachomatis infection or that these patients were infected by a variant strain of C. trachomatis which lacks the C. trachomatis-specific plasmid.

Animals↗
PubMed1992

Persistence of herpes simplex virus DNA in rabbit corneal cells.

Corneal cell cultures were established from the corneas of rabbits killed during a period of latency 118 d after ocular infection with the RE strain of herpes simplex virus (HSV). DNA was isolated from frozen cell pellets of 42 cell cultures that did not develop viral cytopathic effects during 44 d in culture. Using the polymerase chain reaction (PCR) to amplify HSV thymidine kinase (TK) gene sequences, HSV-specific DNA was detected in 15 of 42 culture-negative cell cultures. Subsequent reamplification, using nested primers that were complementary to HSV TK sequences internal to the orginal primers, resulted in eight additional culture-negative samples showing positive hybridization for HSV TK DNA. Twenty three of the 42 virus culture-negative corneal cell cultures tested by PCR were found to contain HSV genetic material. Detailed examination of the clinical histories of the eyes from which the corneal cultures were obtained showed no correlation between increased frequency or severity of epithelial disease, stromal disease, or virus shedding and more frequent isolation of virus or detection of HSV-specific DNA. These studies document that HSV DNA residues in the corneas of HSV-infected rabbits up to 118 d post-infection. About 10% of the eyes contained virus that could be reactivated in culture, whereas an additional 55% of the eyes contained DNA sequences homologous to a portion of the HSV TK gene.

Animals↗
PubMed1991

Inhibition of ribonucleotide reductase by gallium in murine leukemic L1210 cells.

Our previous studies of the mechanism of cell growth inhibition by gallium have suggested that the block in cellular iron uptake induced by transferrin-gallium results in an inhibition of the iron-dependent M2 subunit of ribonucleotide reductase. However, it is not known whether the inhibitory effect of gallium on ribonucleotide reductase is solely the result of limiting iron availability for enzyme activity or whether a direct effect of intracellular gallium on the enzyme is also involved. In the present study, utilizing a cell-free assay, we show that gallium nitrate directly inhibits CDP and ADP reductase activity. Inhibition of DNA synthesis by gallium nitrate thus appears to be due to a combination of a block in iron availability to ribonucleotide reductase and a direct inhibition of the enzyme by gallium.

Animals↗
PubMed1991

Sources of color variation on firing porcelain.

The final color matching of porcelain crowns depends upon the accuracy of the original shade matching by the dentist and variables introduced during processing. Possible sources of processing variables include thickness and color of the opaque, thickness, color, and translucency of the body and enamel layers, firing temperature, and number of firings (Miller, 1987). These processing variables can lead to an error in shade match. The purpose of this study was to quantify, in CIE delta E units: (1) the shade variations when the same batches are fired, (2) the shade variations between different batches, and (3) the differences in color produced by the multiple firing. Three lots of six shades of four commercial brands were included in this study. The color variation of the opaque samples (mean delta E was 0.46) was generally lower than that of the body/opaque samples (mean delta E was 0.86). The average color variation for three different batches of the body/opaque samples was 1.44. The average color difference produced as a result of multiple firings was 1.00 after six firings, compared with the color after three firings.

Analysis of Variance↗
PubMed1991

The development of corneal edema in herpes simplex virus type 1-infected rabbits following termination of therapy for corneal stromal disease.

One complication of combined antiviral/corticosteroid therapy for herpetic stromal disease in patients is rebound of disease upon termination of therapy. To develop a model of steroid rebound, rabbits were injected intrastromally with 10(3) pfu of HSV-1 (RE strain). Therapy with 1% trifluorothymidine (F3TdR) alone or in combination with immunosuppressive agents was initiated 7 days post-infection, at a time when epithelial disease had reached its peak and corneal thickness had begun to increase. Therapy was continued 5 times daily through day 18 post-infection. Following cessation of therapy 13 of 16 eyes receiving both 1% F3TdR and 0.125% prednisolone acetate experienced rebound of disease characterized by an increase in corneal thickness from 514 +/- 106 microns to 743 +/- 189 microns, reaching a maximum at 27 +/- 3 days post-infection. Rabbits receiving therapy with either phosphate buffered saline or F3TdR alone displayed rebound in 2 and 3 of 12 eyes, respectively. Rabbits receiving F3TdR combined with either cyclosporine or deoxycoformycin experienced rebound of disease in 9 of 20 and 6 of 16 eyes, respectively. Cultures of eye washings taken from eyes at the time of rebound were negative in all cases. The data indicate that only steroids significantly increased the proportion of eyes with rebounding stromal disease and corneal edema. These studies document steroid rebound of stromal disease in an animal model.

Animals↗
PubMed1991

Combined anti-herpes virus activity of nucleoside analogs and interferon.

Addition of interferon (IFN) to nucleoside analog therapy for herpetic keratitis has been shown to significantly increase the efficacy of therapy compared to nucleoside alone. We have analysed several nucleoside analogs and recombinant IFN-alpha 2 to determine which combinations have increased anti-herpes simplex virus type 1 (HSV) activity. Synergistic anti-HSV activity between IFN-alpha 2 and the acyclic guanosine analogs, acyclovir (ACV) and ganciclovir (DHPG), was demonstrated in cytopathic effect reduction assay in human corneal cell cultures as well as in Vero cells. In this assay system IFN-alpha 2 alone had little detectable antiviral activity at titers of greater than or equal to 2,000 IU/ml, however, treatment of cells with about 100 IU/ml of IFN-alpha 2 for 24 hrs prior to infection decreased the ED50 of ACV approximately 2- to 3-fold and of DHPG approximately 5- to 6-fold in Vero cells. Combinations of IFN-alpha 2 with bromovinyldeoxyuridine (BVdU) in Vero cells or human corneal stromal cells did not increase the antiviral activity of BVdU. Combinations of IFN-alpha 2 with trifluorothymidine (TFT) also did not increase the effective antiviral activity of this nucleoside and resulted in decreased uptake of TFT from the medium. These studies document that combinations of acyclic nucleoside analogs, ACV and DHPG, with IFN-alpha 2 resulted in synergistic anti-HSV activities in both Vero and human corneal stromal cells, while the pyrimidine analogs, TFT and BVdU, were not synergistic with IFN-alpha 2. IFN-alpha 2 treatment of cells induced modifications of nucleoside (e.g., thymidine and TFT), but not nucleobase (e.g., ACV) uptake. These studies suggest that selective inhibition of nucleoside versus nucleobase uptake may contribute to the mechanism of IFN/nucleobase synergy in the inhibition of HSV replication.

Animals↗
PubMed1991

Therapeutic response of herpes simplex virus-induced corneal edema to trifluridine in combination with immunosuppressive agents.

Herpetic stromal disease often is treated with combinations of antiviral agents and corticosteroids. The addition of steroids to the antiviral treatment regimen frequently increases the efficacy of therapy in patients; however, many complications may arise as a result of corticosteroid therapy. Using a rabbit model, the effects of trifluridine (F3TdR) on corneal edema and stromal disease were examined when combined with each of three immunosuppressive agents. The therapeutic response was evaluated by classifying eyes as either responsive or unresponsive based on the maximum corneal thickness attained during therapy. The data indicate that about 56% of the eyes responded to therapy with 1% F3TdR alone even when therapy was initiated after signs of stromal inflammation had begun to appear and epithelial disease was resolving. Combination of F3TdR with 0.125% prednisolone acetate significantly increased the proportion of responsive eyes to about 78%. Therapy with F3TdR combined with topical 5% cyclosporine A was no better than F3TdR alone, and combination with 0.2% deoxycoformycin and 0.4% 2'-deoxyadenosine significantly decreased the proportion of responsive eyes. These data further document that the responses of stromal disease to therapy must be evaluated on an eye-by-eye basis because the distribution of the data may not be Gaussian in nature. Eyes with corneal edema and stromal disease induced by herpes simplex viral (HSV) infection may respond to therapy with antiviral agents alone, but others require steroid. Still others do not respond to combined therapy. Combining the responses of all eyes in a given treatment group to obtain a "population mean" may be misleading.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1990

Nucleoside metabolism in herpes simplex virus-infected cells following treatment with interferon and acyclovir, a possible mechanism of synergistic antiviral activity.

Alpha interferon (IFN-alpha) and nucleoside analogs have been shown to have synergistic antiherpesvirus activity in cultured cells. The mechanisms responsible for this synergistic activity are not known, but we hypothesize that IFN-alpha-induced alterations of nucleoside metabolism in virus-infected cells may play an important role. Infection of cells with herpes simplex virus type 1 (HSV-1) led to an increase in uptake of thymidine into cells. Treatment of infected cells with recombinant IFN-alpha for 24 h prior to infection resulted in a significant reduction in the uptake of exogenous thymidine but did not reduce the apparent incorporation of exogenous thymidine into DNA. The amount of exogenous thymidine phosphorylated relative to the amount taken up was the same in IFN-alpha-treated and control cultures. IFN-alpha treatment of HSV-1-infected cells also resulted in a reduction in the pool sizes of endogenous deoxyribonucleoside-5'-triphosphates relative to those of untreated HSV-1-infected cultures. Although IFN-alpha affected the metabolism of natural nucleosides in HSV-1-infected cells, it did not significantly reduce the uptake of the antiviral guanosine analog acyclovir into HSV-1-infected cells or the amount of acyclovir-5'-triphosphate accumulated. Therefore, in IFN-alpha-treated cells the concentration of a natural nucleoside, thymidine, was reduced, as were the pools of all deoxyribonucleoside-5'-triphosphates. No decrease in acyclovir or acyclovir-5'-triphosphate concentration was observed, however, when acyclovir-treated cells were exposed to IFN-alpha. These data suggest that IFN-alpha-induced alterations in nucleoside metabolism may be one mechanism whereby IFN-alpha and acyclovir express synergistic antiherpes-virus activity.

Acyclovir↗
PubMed1990

Pathogenesis of corneal oedema associated with herpetic eye disease.

Corneal oedema and stromal disease, induced in rabbits by intrastromal injection of herpes simplex virus, type 1, strain RE (HSV-1, RE), reached a peak of 12-15 days after infection. Corneal oedema as measured by ultrasonic pachymetry, and stromal disease as measured by a subjective scoring system, were closely related for 30 days after infection. Morphometric analysis of wide field specular micrographs showed that no immediate endothelial cell damage occurred in either control or HSV-1 infected corneas. Alizarin red S staining of corneas taken during the period of most severe oedema indicated no significant endothelial cell loss; however, visual inspection indicated numerous staining abnormalities. Scanning and transmission electron microscopy provided evidence of an intact endothelial layer possessing integrated infiltrating cells. Virus antigen could not be detected on endothelial cells by immunoperoxidase staining at any time during development of corneal oedema. The results indicate that corneal oedema associated with HSV-1 induced disease can occur in the absence of detectable virus replication and cytolysis of corneal endothelial cells.

Animals↗
PubMed1990

Correlation between histological and behavioral measures of visual acuity in a zooplanktivorous fish, the white crappie (Pomoxis annularis).

Estimates of visual acuity in a pelagic freshwater zooplanktivorous fish, the white crappie (Pomoxis annularis, Centrarchidae), were made using a behavioral measure, the maximum observed prey pursuit distance (MxPD), and a histological measure, the density of cone cells in the retina. The greatest number of pursuits occurs in the 0-30 degrees wedge of the visual field; 87% of all pursuits occur in the first 40 degrees. The longest pursuits (200 mm) also occur in this area and generally get shorter from 0 to 180 degrees (from forward-directed) in the visual field. Consistent with the behavioral results, the largest number of cone photoreceptors (13,000/mm2) is found in the far temporal retina along the eye's horizontal meridian. Cone cell densities in the corresponding region of the nasal retina are approximately half this value. The number of cones decreases dorsally and ventrally from the horizontal meridian. Although the absolute values of visual acuity calculated from cone cell topography (i.e. MxPDs of 500 mm) are 2-3 times greater than those observed behaviorally (i.e. MxPDs of 200 mm), the trends in visual acuity across the visual field obtained from both measures are consistent. We suggest that overestimates of visual acuity obtained from cone cell counts alone result from this measure's not accounting for, among other properties of the nervous system, cone cell convergence onto ganglion cells and higher brain centers. Behavioral measures of visual acuity are, therefore, likely to yield a more accurate estimate of an animal's visual abilities.

Animals↗
PubMed1990

A new, small-color-difference equation for dental shades.

Traditionally, dental-shade-guide standards are designated in terms of Munsell hue (H), value (V), and chroma (C). However, delta E color differences proposed as ADA tolerances for shade guides are in the CIE L*a*b* system. The purpose of this study was to evaluate a new color-difference equation, delta EM = C delta H/5 + 7 delta V + 4 delta C for estimation of small color differences by Munsell parameters. The published values of the Bioform shade-guide tooth colors determined with a Beckman spectrophotometer were used. Color differences among 276 combinations of the 24 Bioform shade-guide colors were calculated with Eq. 1, with use of the Munsell notation, and also with the CIE L*a*b* equation for delta E. An estimate of the accuracy of Eq. 1 was 0.41 delta E units when delta E CIE was below 4.0. The Vita shade-guide colors were determined with a Beckman spectrophotometer. This data set contained 16 samples, and 120 combinations were used for calculation of color difference. An estimate of the accuracy for this set of data was 0.35 delta E units when delta E CIE was less than 4.0. The new color-difference equation provides a means for estimation of delta E CIE L*a*b* color difference between dental shades with Munsell notation. This equation will be useful for estimation of small delta E CIE L*a*b* values for shade-guide teeth that are designated in terms of Munsell notation.

Color↗