ABSTRACT
AIM: To introduce and assess effectiveness of a radiographer-led screening programme for the detection of unsuspected pulmonary emboli on routine contrast-enhanced computed tomography (CT), and to evaluate radiographer response to this extended role. MATERIALS AND METHODS: A training programme was devised for all radiographic staff working in CT. The screening service was introduced and monthly quality assurance performed with cumulative analysis of the first 2 years. Clinical effectiveness before and after screening was evaluated by comparing the time interval between the scan and the start of a clinical consultation for anticoagulant prescription. A satisfaction survey was sent to all participating staff. RESULTS: Thirty-two radiographers completed the training. During the training period, the radiographer detection rate of incidental pulmonary emboli was 89%. Main, lobar, segmental, and subsegmental emboli were detected. The overall detection rate after full introduction of the programme was 92% for the first 2 years. The time interval between the scan and clinical consultation for anticoagulant prescription dropped from a mean of 1.5 days to a mean of 26 minutes and ensured that treatment was commenced at the same patient attendance. Eighty-four percent of staff completed the satisfaction survey and all were satisfied with the extended role. CONCLUSION: Radiographer screening for incidental pulmonary emboli was effective and accurate. It resulted in immediate communication with the responsible physician and commencement of anticoagulation therapy at the same hospital attendance, creating a "one-stop" service. Radiographer satisfaction with the extended role was high.
Subject(s)
Contrast Media , Incidental Findings , Pulmonary Embolism/diagnostic imaging , Radiographic Image Enhancement/methods , Tomography, X-Ray Computed/methods , Cancer Care Facilities , Humans , Pulmonary Artery/diagnostic imagingABSTRACT
Telomere repeat sequences cap the ends of eucaryotic chromosomes and help stabilize them. At interstitial sites, however, they may destabilize chromosomes, as suggested by cytogenetic studies in mammalian cells that correlate interstitial telomere sequence with sites of spontaneous and radiation-induced chromosome rearrangements. In no instance is the length, purity, or orientation of the telomere repeats at these potentially destabilizing interstitial sites known. To determine the effects of a defined interstitial telomere sequence on chromosome instability, as well as other aspects of DNA metabolism, we deposited 800 bp of the functional vertebrate telomere repeat, TTAGGG, in two orientations in the second intron of the adenosine phosphoribosyltransferase (APRT) gene in Chinese hamster ovary cells. In one orientation, the deposited telomere sequence did not interfere with expression of the APRT gene, whereas in the other it reduced mRNA levels slightly. The telomere sequence did not induce chromosome truncation and the seeding of a new telomere at a frequency above the limits of detection. Similarly, the telomere sequence did not alter the rate or distribution of homologous recombination events. The interstitial telomere repeat sequence in both orientations, however, dramatically increased gene rearrangements some 30-fold. Analysis of individual rearrangements confirmed the involvement of the telomere sequence. These studies define the telomere repeat sequence as a destabilizing element in the interior of chromosomes in mammalian cells.
Subject(s)
Adenine Phosphoribosyltransferase/genetics , Chromosome Fragility/genetics , Mutagenesis, Insertional/genetics , Repetitive Sequences, Nucleic Acid/genetics , Telomere/genetics , Animals , Base Sequence , Blotting, Southern , CHO Cells , Chromosome Deletion , Cricetinae , Gene Targeting , Introns/genetics , Molecular Sequence Data , Phenotype , RNA, Messenger/biosynthesis , RNA, Messenger/genetics , Recombination, Genetic/genetics , Sequence Analysis, DNA , Sequence Homology, Nucleic Acid , Thymidine Kinase/geneticsABSTRACT
Spontaneous recombination between direct repeats at the adenine phosphoribosyltransferase (APRT) locus in ERCC1-deficient cells generates a high frequency of rearrangements that are dependent on the process of homologous recombination, suggesting that rearrangements are formed by misprocessing of recombination intermediates. Given the specificity of the structure-specific Ercc1/Xpf endonuclease, two potential recombination intermediates are substrates for misprocessing in ERCC1(-) cells: heteroduplex loops and heteroduplex intermediates with non-homologous 3' tails. To investigate the roles of each, we constructed repeats that would yield no heteroduplex loops during spontaneous recombination or that would yield two non-homologous 3' tails after treatment with the rare-cutting endonuclease I-SCE:I. Our results indicate that misprocessing of heteroduplex loops is not the major source of recombination-dependent rearrangements in ERCC1-deficient cells. Our results also suggest that the Ercc1/Xpf endonuclease is required for efficient removal of non-homologous 3' tails, like its Rad1/Rad10 counterpart in yeast. Thus, it is likely that misprocessing of non-homologous 3' tails is the primary source of recombination-dependent rearrangements in mammalian cells. We also find an unexpected effect of ERCC1 deficiency on I-SCE:I-stimulated rearrangements, which are not dependent on homologous recombination, suggesting that the ERCC1 gene product may play a role in generating the rearrangements that arise after I-SCE:I-induced double-strand breaks.
Subject(s)
DNA Repair/genetics , DNA-Binding Proteins , DNA/chemistry , DNA/metabolism , Endonucleases , Proteins/metabolism , Recombination, Genetic/genetics , Adenine Phosphoribosyltransferase/genetics , Animals , Blotting, Southern , Cell Line , Crossing Over, Genetic/genetics , DNA/genetics , DNA Damage/genetics , Gene Deletion , Nucleic Acid Conformation , Nucleic Acid Heteroduplexes/chemistry , Nucleic Acid Heteroduplexes/genetics , Nucleic Acid Heteroduplexes/metabolism , Proteins/genetics , Repetitive Sequences, Nucleic Acid/genetics , Substrate Specificity , TransfectionABSTRACT
Nucleotide excision repair proteins have been implicated in genetic recombination by experiments in Saccharomyces cerevisiae and Drosophila melanogaster, but their role, if any, in mammalian cells is undefined. To investigate the role of the nucleotide excision repair gene ERCC1, the hamster homologue to the S. cerevisiae RADIO gene, we disabled the gene by targeted knockout. Partial tandem duplications of the adenine phosphoribosyltransferase (APRT) gene then were constructed at the endogenous APRT locus in ERCC1- and ERCC1+ cells. To detect the full spectrum of gene-altering events, we used a loss-of-function assay in which the parental APRT+ tandem duplication could give rise to APRT- cells by homologous recombination, gene rearrangement, or point mutation. Measurement of rates and analysis of individual APRT- products indicated that gene rearrangements (principally deletions) were increased at least 50-fold, whereas homologous recombination was affected little. The formation of deletions is not caused by a general effect of the ERCC1 deficiency on gene stability, because ERCC1- cell lines with a single wild-type copy of the APRT gene yielded no increase in deletions. Thus, deletion formation is dependent on the tandem duplication, and presumably the process of homologous recombination. Recombination-dependent deletion formation in ERCC1- cells is supported by a significant decrease in a particular class of crossover products that are thought to arise by repair of a heteroduplex intermediate in recombination. We suggest that the ERCC1 gene product in mammalian cells is involved in the processing of heteroduplex intermediates in recombination and that the misprocessed intermediates in ERCC1- cells are repaired by illegitimate recombination.
Subject(s)
DNA Repair , DNA-Binding Proteins , Endonucleases , Proteins/genetics , Recombination, Genetic , Sequence Deletion , Adenine Phosphoribosyltransferase/genetics , Animals , Cell Line , Cricetinae , Thymidine Kinase/geneticsABSTRACT
The retinoblastoma protein (p110RB) interacts with many cellular proteins in complexes potentially important for its growth-suppressing function. We have developed and used an improved version of the yeast two-hybrid system to isolate human cDNAs encoding proteins able to bind p110RB. One clone encodes a novel type 1 protein phosphatase catalytic subunit (PP-1 alpha 2), which differs from the originally defined PP-1 alpha by an amino-terminal 11-amino-acid insert. In vitro-binding assays demonstrated that PP-1 alpha isoforms preferentially bind the hypophosphorylated form of p110RB. Moreover, similar p110RB sequences are required for binding PP-1 alpha 2 and SV40 large T antigen. Cell cycle synchrony experiments revealed that this association occurs from mitosis to early G1. The implications of these findings on the regulation of both proteins are discussed.
Subject(s)
Cell Division/genetics , Cloning, Molecular/methods , Gene Expression Regulation, Fungal , Phosphoprotein Phosphatases/metabolism , Recombinant Fusion Proteins/metabolism , Retinoblastoma Protein/metabolism , Saccharomyces cerevisiae Proteins , Transcription Factors , Antigens, Polyomavirus Transforming/metabolism , Base Sequence , Cell Division/physiology , DNA, Fungal/analysis , DNA-Binding Proteins , Escherichia coli/genetics , Fungal Proteins/genetics , Fungal Proteins/metabolism , Genomic Library , Humans , Isoenzymes/genetics , Macromolecular Substances , Molecular Sequence Data , Phosphoprotein Phosphatases/genetics , Protein Binding , Recombinant Fusion Proteins/genetics , Saccharomyces cerevisiae/genetics , Substrate SpecificityABSTRACT
An assessment is presented of the prototype of a computerized radiological reporting system (SIREP). The system is operated by a radiologist who signals to a computer by touching a glass surface on to which is projected the image of one of 165 slides, each containing words and phrases relevant to the radiographs to be reported. The report is shown on a television screen and is typed automatically when completed. The system was installed in the X-ray Department serving the Accident and Emergency Service of the Bristol Royal Infirmary after a study had been made of the reporting sequences and times using conventional methods. This showed that the major factor influencing the time taken to report a film was whether it was "normal" or "abnormal". The assessment of system looked at its effect on reporting and on the department; its technical aspects and reliability; and its acceptability to radiologists. It was found that because there is no storage or analysis of reports the system directly affected only the radiologist and the secretary. For an average of 100 reports daily it is calculated that a secretary would be saved 64 minutes of typing daily, and that a radiologist would take 63 minutes longer over reporting. In general the system was mechanically reliable and fast in operation, with a maximum slide transit time of three seconds. All radiologists using the machine found that the reporting terminal was simple to operate, but found that reporting abnormalities caused an increased effort and took a longer time. None of the radiologists would use the machine in preference to traditional methods, and in this country no cost or time benefit could be established.