Localization of the interferon-induced, 2-5A-dependent RNase gene (RNS4) to human chromosome 1q25.
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Chromosome Mapping↗
Biomedical subjects
R H Silverman
Publications and source records attributed to R H Silverman.
Corneal pachymetric topography.
PURPOSE: The authors developed a system for producing topographic pachymetric maps of the corneal epithelium and anterior scar tissue. METHOD: The system uses high-frequency ultrasound scanning enhanced by digital signal processing. Ultrasonic echo data from consecutive parallel B-scans of the cornea spaced at 250-microns intervals are digitized and stored. Using the I-scan (obtained by computing the analytic signal magnitude of the deconvolved ultrasound signal), layer thickness measurements are made with a precision of 2 microns (standard deviation) at 120-microns intervals along each scan plane. The data are stored as an array, z(x,y), mapping thickness, z, onto horizontal and vertical (x,y) spatial coordinates. Pachymetric maps are then constructed by plotting local thickness, represented by a color scale, against measurement point position. RESULTS: Examples of a normal cornea, a contact lens-wearing cornea, Reis-Bückler dystrophy, and postphotorefractive keratectomy are presented. Areas with significant subepithelial scarring and general epithelial thickening in a subject with Reis-Bückler dystrophy are mapped. Unevenness in the epithelial thickness profile of the cornea in a subject after photorefractive keratectomy is shown, relative to the fellow (untreated) cornea. CONCLUSION: This technique provides the corneal surgeon with a new tool for the topographic evaluation of the thickness of anterior corneal layers in normal and pathologic corneas with high precision. In addition, the technique is not limited to optically transparent tissue.
Adult↗
Epithelial and corneal thickness measurements by high-frequency ultrasound digital signal processing.
PURPOSE: The authors determine the mean central corneal and epithelial thickness in a group of normal human subjects using a new high-frequency ultrasound technique, incorporating digital signal processing. METHOD: Both eyes of ten volunteers (age range, 23-44 years) were scanned through a normal saline standoff. Digitized ultrasonic echo data were mathematically transformed to produce a plot, the I-scan, which optimally localizes acoustic interfaces to provide improved measurement precision. System precision was determined by analysis of variance of repeated measures. Central epithelial thickness was obtained by averaging multiple measurements. Central corneal thickness was determined by fitting measurements of apparent corneal thickness in consecutive parallel B-scans to a mathematically modeled cornea. A speed of sound of 1640 m/second was used. RESULTS: Epithelial pachymetric precision using A-scan and I-scan was 4.8 and 2.0 microns (standard deviation), respectively. The mean epithelial thicknesses for the right and left eyes were 50.7 +/- 3.7 microns and 50.3 +/- 3.4 microns, respectively. The mean corneal thicknesses in the right and left eyes were 514.6 +/- 38.4 microns and 516.2 +/- 37.8 microns, respectively. The root mean-square differences in epithelial and corneal thickness between the left and right eyes of each subject were 1.3 and 7.7 microns, respectively (neither was statistically significant). CONCLUSION: This system provides a pachymetric precision superior to current optical and ultrasound methods. Epithelial and corneal pachymetry is obtained noninvasively by a method that is not limited to optically clear media.
Adult↗
Fascination with 2-5A-dependent RNase: a unique enzyme that functions in interferon action.
Interferon (IFN) treatment of cells results in the induction of 2-5A-synthetases, double-stranded RNA-activated enzymes that produce unusual 5'-phosphorylated 2',5'-linked oligoadenylates known as 2-5A. 2.5A activates a unique IFN-induced endoribonuclease, the 2-5A-dependent RNase (RNase L), that is capable of degrading both viral and cellular RNA. The expression cloning of 2-5A-dependent RNase is leading to meaningful analysis of the physiological functions of the 2-5A system. For example, expression in mouse cells of a dominant-negative mutant form of 2-5A-dependent RNase suppressed both the antiencephalomyocarditis virus and anticellular activities of IFN. Future investigations into this intriguing ribonuclease pathway promise to provide an intricate view into a molecular pathway of IFN action.
Chromosome Mapping↗
High-frequency ultrasound corneal pachymetry in the assessment of corneal scars for therapeutic planning.
We used high-frequency ultrasound B-scanning with digital signal processing for pachymetric analysis of corneal scars to help determine the optimal management strategy. Four patients were selected for this report. By an automated motor system, 12 consecutive, parallel ultrasound B-scans, each 2.5 to 3 mm wide at 0.25 mm intervals, were obtained from the central corneal area of three patients with anterior corneal scarring. In a fourth patient with near complete corneal scarring obscuring the view of the anterior chamber, a set of 15 mm wide B-scans was obtained. Digitized ultrasound signals were used to produce high-resolution images and I-scans enabling a pachymetric precision of +/- 2 microns (SD). Epithelial, scar, and corneal thickness measurements were made along each scan to determine the most significant zone of pathology. Pachymetry of the cornea and the individual layers was used to assess the suitability for either photorefractive or penetrating keratectomy. B-scan imaging of the full anterior segment provided useful information for the preoperative planning of anterior segment reconstruction and the prognostic evaluation of penetrating keratoplasty. This method provides a powerful tool for the corneal surgeon in management planning.
Adult↗
Expression cloning of 2-5A-dependent RNAase: a uniquely regulated mediator of interferon action.
2-5A-dependent RNAase, an interferon-induced enzyme that is activated by 5'-phosphorylated, 2',5'-linked oligoadenylates (2-5A), is implicated in both the molecular mechanisms of interferon action and the fundamental control of RNA stability in mammalian cells. Here we report the expression cloning and analysis of murine and human 2-5A-dependent RNAases. The 2-5A binding properties and RNAse activities of recombinant and naturally occurring forms of 2-5A-dependent RNAase were identical. Interferon induction of 2-5A-dependent RNAse expression was demonstrated by measuring the mRNA levels in cells treated with interferon and cycloheximide. Analysis of aligned murine and human 2-5A-dependent RNAse sequences revealed several intriguing features, including similarity to RNAase E, which is implicated in the control of mRNA stability in E. coli. Interestingly, a duplicated phosphate-binding loop motif was determined by deletion analysis and site-directed mutagenesis to function in the binding of 2-5A.
Amino Acid Sequence↗
Targeting RNA for degradation with a (2'-5')oligoadenylate-antisense chimera.
Antisense oligonucleotides hold considerable promise both as research tools for inhibiting gene expression and as agents for the treatment of a myriad of human diseases. However, targeted destruction of RNA has been difficult to achieve in a versatile, efficient, and reliable manner. We have developed an effective strategy for cleaving unique RNA sequences with 2-5A-dependent RNase, an endoribonuclease that mediates inhibitory effects of interferon on virus infection and is activated by 5'-phosphorylated 2'-5'-linked oligoadenylates known as 2-5A [pn5' A2'(p5' A2')mp5'A], resulting in the cleavage of single-stranded RNA predominantly after UpUp and UpAp sequences. To direct 2-5A-dependent RNase to cleave unique RNA sequences, p5' A2' p5' A2'p5'A was covalently linked to an antisense oligonucleotide to yield a chimeric molecule (2-5A:AS). The antisense oligonucleotide component of 2-5A:AS bound a specific RNA sequence while the accompanying 2-5A component activated 2-5A-dependent RNase, thereby causing the cleavage of the RNA in the targeted sequence. This strategy was demonstrated by inducing specific cleavage within a modified human immunodeficiency virus type 1 vif mRNA in a cell-free system from human lymphoblastoid cells. Because 2-5A-dependent RNase is present in most mammalian cells, the control of gene expression based on this technology--including therapies for cancer, viral infections, and certain genetic diseases--can be envisioned.
Adenine Nucleotides↗
High-frequency ultrasound digital signal processing for biometry of the cornea in planning phototherapeutic keratectomy.
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Adult↗
High-frequency ultrasound quantitative analyses of corneal scarring following excimer laser keratectomy.
OBJECTIVE: To report objective grading and analysis of excimer laser-induced scarring using high-frequency ultrasound. METHODS: High-frequency ultrasound (50 MHz) corneal examination was performed on eight New Zealand white rabbits at different time points (10, 17, 24, and 74 days following phototherapeutic keratectomy). We used biometry and signal-processing techniques to determine corneal, epithelial, and scar thicknesses and to quantify the acoustic backscatter. RESULTS: Excimer laser-induced scarring showed an irregularly distributed acoustic hyperreflectivity that decreased through day 74. Corneal thickness remained reduced after ablation (mean, 318 microns compared with 419 microns for controls). Epithelial thickness averaged 62 microns and scar thickness, 87 microns. Scar peak and average anterior stroma acoustic backscatter ratios decreased from day 10 to day 74 (19.65 to 2.76, and 6.42 to 1.32, respectively). Histopathologic study showed increased keratocyte activity at early time points that correlated with acoustic backscatter ratios and imaging pattern. CONCLUSIONS: High-frequency ultrasound signal processing is a noninvasive method that quantitatively grades excimer laser-induced corneal scarring.
Animals↗
A dominant negative mutant of 2-5A-dependent RNase suppresses antiproliferative and antiviral effects of interferon.
2-5A-dependent RNase is the terminal factor in the interferon-regulated 2-5A system thought to function in both the molecular mechanism of interferon action and in the general control of RNA stability. However, direct evidence for specific functions of 2-5A-dependent RNase has been generally lacking. Therefore, we developed a strategy to block the 2-5A system using a truncated form of 2-5A-dependent RNase which retains 2-5A binding activity while lacking RNase activity. When the truncated RNase was stably expressed to high levels in murine cells, it prevented specific rRNA cleavage in response to 2-5A transfection and the cells were unresponsive to the antiviral activity of interferon alpha/beta for encephalomyocarditis virus. Remarkably, cells expressing the truncated RNase were also resistant to the antiproliferative activity of interferon. The truncated RNase is a dominant negative mutant that binds 2-5A and that may interfere with normal protein-protein interactions through nine ankyrin-like repeats.
3T3 Cells↗
The role of fibronectin in tumor implantation at surgical sites.
Fibronectins are a family of glycoproteins with modular functional domains. They mediate cell-cell and cell-matrix interactions which are important in embryogenesis, wound healing, metastasis and other processes. We present data on the influence of fibronectin on wound implantation of a murine mammary carcinoma line, TA3Ha. Fibronectin used in these studies was derived from bovine plasma, human serum, human foreskin fibroblasts, and mouse embryo cultures. TA3Ha cells rarely form tumors in the liver of syngeneic mice when injected intravenously but after hepatic wedge resection, 45% (107/240) of the mice develop tumors in the hepatic wound. Wound implantation is markedly reduced when the cells are pre-exposed to 200 micrograms/ml bovine plasma fibronectin (13%, P = 0.007), human serum fibronectin (0%, P = 0.02), human cellular fibronectin (0%, P = 0.02), or mouse cellular fibronectin (0%, P = 0.04). Lung colonization is also reduced by these fibronectins. These effects are not due to a cytotoxic action of fibronectin, since intraperitoneally injected fibronectin-treated cells form ascites tumor as effectively as do control untreated cells. Local application of a solution containing 0.25 mg/ml mouse cellular fibronectin to the hepatic wound reduces the frequency of tumor implantation from 45% to 5% (1/21, P = 0.001). No tumor implantation inhibition is seen when only suspending medium or albumin in suspending medium is used. The mechanism by which topical application of fibronectin reduces hepatic wound implantation of tumor cells is unclear, but this finding raises an exciting possibility of preventing local recurrence of cancer.
Adenocarcinoma↗
High-frequency ultrasound imaging and spectral analysis in traumatic hyphema.
PURPOSE: High-frequency (50-MHz) ultrasound allows high-resolution imaging of anterior ocular structures. Spectrum analysis of acoustic backscatter is sensitive to the concentration, size, and density of tissue inhomogeneities. The authors sought to determine whether acoustic imaging and spectrum analysis of hyphema would allow them to distinguish organized from fluid hyphema and recent from old hemorrhage in the eye. METHODS: Trauma-induced hyphemas were followed by slit-lamp photography and high-frequency ultrasonography in six New Zealand white rabbits. The blood collections were analyzed using the normalized power spectra of the digitized radio frequency ultrasound data and compared with in vitro references. RESULTS: The 50-MHz acoustic images permitted differentiation between fluid (diffuse) and clotted (organized) blood. Spectrum analysis allowed quantitative characterization of the degree of blood organization. Significant changes were observed in spectral properties during the time course of absorption both for initial and after-rebleeding hemorrhages. The characteristics of a human postsurgical hyphema also were examined and found to be similar to those seen in the experimental model. CONCLUSION: Spectrum analysis of high-frequency ultrasound data was able to distinguish organized from recent hemorrhage, which is clinically helpful for planning hyphema therapy.
Animals↗
Tissue-related and species-specific differences in the 2-5A oligomer size requirement for activation of 2-5A-dependent RNase.
2',5'-Oligoadenylate (2-5A)-dependent RNase (L or F) is the final enzyme in the 2-5A pathway and a key component in the molecular mechanism of interferon (IFN) action. Here we demonstrate differences in the 2-5A oligomer size requirement between rabbit 2-5A-dependent RNase from reticulocytes and from cultured kidney cells. The rabbit reticulocyte enzyme was activated by tetramer 2-5A, whereas the ribonuclease from rabbit kidney cells required only trimer 2-5A. Interestingly, in contrast to the 2-5A-dependent RNase from rabbit reticulocytes, that from murine reticulocytes could be activated by trimer 2-5A. Partial proteolysis of affinity-labeled, 80-kD 2-5A-dependent RNase from rabbit reticulocytes and rabbit kidney cells resulted in the same pattern of labeled peptides. However, the affinity labeling reaction with a 32P-labeled 2-5A analog did produce some different labeled polypeptides in rabbit kidney cell extract and rabbit reticulocyte lysate. These results could indicate specialized functions for the 2-5A system in different organ systems.
Adenine Nucleotides↗
Measurement of ocular tumor volumes from serial, cross-sectional ultrasound scans.
A system has been developed to determine intraocular tumor volume, a characteristic that has been estimated in previous studies from linear dimensions using a variety of models. Volume was determined by tracing tumor boundaries in sequential, parallel ultrasound scans, adding the areas and multiplying by the interslice interval. The in vitro accuracy of the technique was within +/- 2%. The mean difference between volumes determined from replicate in vivo scans of intraocular tumors was 4.3%. Serial scan volumes were, on average, 19% smaller than volumes computed from an ellipsoidal model and 13% smaller than area-rotational volumes. Differences of as much as 50% were observed between serial scan volumes and volumes computed with these models. The results indicate that methods based on either linear measurements or the tumor area in a single cross-section will tend to both systematically overestimate tumor volume and suffer from unpredictable, nonsystematic errors.
Humans↗
Using high-frequency ultrasound to characterize intraocular foreign bodies.
We scanned uniformly-sized particles of brass, aluminum, glass, and plastic at ultrasonic frequencies of 10 and 50 MHz to determine the effects of material type and transducer frequency on the ultrasonographic appearance of these particles. The particles were scanned in vitro against both anechoic and echoic backgrounds and in vivo, implanted in the angle of rabbit eyes. Both the metals and glass produced reverberations against an anechoic background; the plastic particles appeared as discrete objects, with no reverberations. Against an echoic background, the plastic particles created prominent shadows, while the metal ones did not. When placed in the angle of a rabbit eye, with the exception of the plastic at 10 MHz, all the materials could be differentiated from surrounding tissues at both frequencies. However, the appearance of the foreign body and the surrounding tissues, as well as their exact location and size, were most obvious with the 50-MHz system. In general, the particles produced more reverberations at 10 than at 50 MHz, and more prominent shadowing at 50 than at 10 MHz. In summary, high-frequency imaging enabled superior characterization of all the particles.
Animals↗
New perspectives: 3-D volume rendering of ocular tumors.
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Eye↗
Ophthalmic ultrasonography.
Definition and differentiation of ocular structures and abnormalities necessitates the use of high-frequency, high-resolution, and high-definition ultrasound. Frequencies of 10 to 100 MHz are being employed. Color Doppler imaging, tissue characterization, parameter image staining, and three-dimensional volume rendering are new and useful adjuncts to ocular diagnosis.
Eye↗
Ultrasonic tissue characterization of uveal melanoma and prediction of patient survival after enucleation and brachytherapy.
We performed survival studies on 136 patients with uveal malignant melanoma who were examined with ultrasonic tissue characterization before treatment with cobalt-60 brachytherapy (74 patients) or enucleation (62 patients). Mean follow-up time was 58.7 months for the brachytherapy group and 59.0 months for the enucleated group. The maximal follow-up time was nearly ten years. Univariate survival analysis showed that patients with small tumors (less than 250 mm3, 49 patients) had a higher five-year survival when treated with brachytherapy than when treated with enucleation. No patients in this study with tumor volumes greater than 1,500 mm3 (13 patients) were treated with brachytherapy. For tumors of intermediate size (74 patients), survival analysis did not indicate appreciable differences between the treatment options. However, multivariate models including two ultrasonic tissue characterization variables, scatterer size and concentration, showed appreciable intergroup differences in the dependence of survival on these factors. Results suggest that tissue properties detectable with ultrasonic techniques are related to differences in patient survival and may be used for treatment planning for tumors of intermediate size.
Brachytherapy↗