Use of a 3D camera for automated patient positioning for chest-abdomen-pelvis CT scans: Effect on positioning accuracy and patient dose.

INTRODUCTION: 3D positioning cameras that automate the positioning of patients with respect to the CT isocentre have been developed and are in common use in CT departments. This study aimed to compare the performance of radiographers and a 3D camera system with respect to positioning accuracy and the effect on patient radiation dose for chest-abdomen-pelvis scans. METHODS: Patient positioning and dose data obtained from a dose management system was evaluated over a two-month period for patients positioned with (CAMon) and without (CAMoff) the positioning camera. Median vertical and lateral offset values were compared between the groups whilst doses were evaluated as a function of patient water equivalent diameter (WED) for the thorax and abdomen-pelvis acquisitions for both cohorts. RESULTS: Radiographers demonstrated high levels of positioning accuracy, however significant improvements in median vertical offset were identified for the CAMon cohort for both thorax (8 mm vs. 17 mm (p = 0.001)) and abdomen-pelvis (7 mm vs. 16 mm (p = 0.003)) scans. The percentage of patients positioned within 5 mm of the isocentre was 39.0% and 16.1% for the CAMon and CAMoff cohorts. For CAMoff scans, 77.4% of patients were positioned below the isocentre, but this was reduced to 45.8% for CAMon scans. No significant changes in dose as a function of WED were identified related to the camera use (thorax: p = 0.569, abdomen-pelvis: p = 0.760). CONCLUSION: Use of a 3D camera delivered significant improvements in the accuracy and reproducibility of patient positioning when compared with radiographers. IMPLICATIONS FOR PRACTICE: Improvements in positioning accuracy were observed at the research site and hence positioning camera use has the potential to become standard practice in CT to help ensure appropriate doses are delivered to patients according to their size.

Genomic cloning and characterization of the human eukaryotic initiation factor-2beta promoter.

The translation initiation factor eIF2 consists of three subunits that are present in equal molar amounts. The genomic DNA containing the gene for eIF2beta and its promoter were cloned and sequenced to characterize further the mechanism of their regulated synthesis. Whereas Southern blot analysis indicated that a number of copies of the gene may exist, only one full-length intron-containing copy was identified. Similar to the eIF2alpha promoter, the eIF2beta promoter is TATA-less, CAAT-less, and GC-rich and contains an alpha-Pal binding motif. Mutation of the alpha-Pal binding sequence resulted in an 8-fold decrease in activity when assayed by the luciferase reporter gene constructs. The data suggest a common mechanism of transcriptional control for the two cloned subunits of eIF2.

Formation of alpha-Pal/Max heterodimers synergistically activates the eIF2-alpha promoter.

The transcription factor alpha-Pal recognizes two tandem palindromic repeats within the promoter of eukaryotic translation initiation factor 2-alpha (eIF2-alpha). Whereas both binding sites have the same "core domain" sequence (CGCATGCG), they differ with respect to their flanking sequences. Of the two sites, the 5'-cap proximal site has a higher binding affinity for alpha-Pal than does the 5'-cap distal site (Jacob, W. F., Silverman, T. A., Cohen, R. B., and Safer, B. (1989) J. Biol. Chem. 264, 20372-20384). The well characterized transcription factor Max binds to sequences that are remarkably similar to the core domain that alpha-Pal recognizes. To date, all of the Max heterodimer partners lack DNA binding domains and are thus dependent on Max interacting with DNA. Here we report that the two alpha-Pal sites have very different binding activities with respect to the E-box-binding protein Max. The 5'-cap distal or low alpha-Pal affinity site binds both alpha-Pal and Max. Furthermore, both heterodimers and homodimers of each of these proteins bind to this site. In contrast to the low affinity site, the high affinity site does not bind Max as a homodimer. This is the first documented case where Max heterodimerizes with a transcription factor that has affinity for DNA independent of Max.

Augmentation of 1-beta-D-arabinofuranosylcytosine (Ara-C) cytotoxicity in leukaemia cells by co-administration with antisignalling drugs.

The ribonucleotide reductase inhibitors hydroxyurea (HU), arabinosyl-2-fluoroadenine (F-Ara-A) and 2-chlorodeoxyadenosine (2-CdA) and the antisignalling drugs all-trans retinoic acid (ATRA), staurosporine and quercetin have been reported to enhance the cytotoxicity of 1-beta-D-arabinofuranosylcytosine (ara-C). We tested the hypothesis that the ara-C-sensitising potency of the antisignalling agents is equipotent with that of the ribonucleotide inhibitors. The cytotoxicity, determined by the 3-(4,5 dimethylthiazol-2-yl-)5 diphenyltetrazolium bromide (MTT) assay, of combinations of ara-C with the agents named above was compared in the leukaemia cell lines HL-60, ara-C-resistant HL-60 (HL-60/ara-C) and U937. Furthermore, a range of protein tyrosine kinase inhibitors, genistein, CGP 52411, tyrphostin A48 and nordihydroguaiaretic acid (NDGA), for which ara-C-sensitisation has hitherto not been described, were included in the study. All three cell types acquired increased sensitivity to ara-C when co-incubated with HU or ATRA, but their ara-C sensitivity was not affected by quercetin or genistein. 2-CdA, CGP 52411, tyrphostin A48, staurosporine and NDGA were active as sensitisers against ara-C in HL-60 cells, CGP 52411 and tyrphostin A48 also in HL-60/ara-C cells, and 2-CdA, staurosporine and NDGA also in U937 cells. F-Ara-A increased ara-C toxicity in HL-60/ara-C and U937 cells. To address the mechanism of the observed sensitisation, the influence of agents with ara-C-sensitising properties on ara-C-induced apoptosis was investigated in HL-60 cells as measured by cell shrinkage, DNA loss and DNA fragmentation. HU, ATRA, tyrphostin A48 and NDGA augmented apoptosis induced by ara-C as assessed by all three indicators. CGP 52411 decreased the effect of ara-C on apoptotic indicators after incubation for 4 h, but not after 12 h. The results suggest that ATRA, CGP 52411, tyrphostin A48, staurosporine and NDGA may be suitable alternatives to the clinically applied ribonucleotide reductase inhibitors as modifiers of ara-C cytotoxicity in the treatment of acute myeloid leukaemia.

Modulation of apoptosis in rat thymocytes by analogs of staurosporine: lack of direct association with inhibition of protein kinase C.

Protein kinase C (PKC) is an important constituent of the signaling pathways involved in apoptosis. The PKC inhibitor staurosporine induces apoptosis in many cell types. We characterized the role of PKC in the induction of apoptosis in immature rat thymocytes by investigating the effects of staurosporine with those of five analogs. Four of them, the indolocarbazoles CGP 41251 and UCN-01 and the bisindolylmaleimides RO 31-8220 and GF 109203X, possess high PKC-inhibitory specificity and potency, whereas one, the UCN-01 stereoisomer UCN-02, is a weak PKC inhibitor. Apoptosis was examined by flow cytometry, internucleosomal DNA cleavage, and formation of large DNA fragments. Staurosporine, UCN-01, and UCN-02 induced a concentration- and time-dependent increase in apoptosis, whereas neither CGP 41251, RO 31-8220, nor GF 109203X induced apoptosis. The mechanism of induction of apoptosis by staurosporine, UCN-01, and UCN-02 was clearly different from the mechanism that mediates induction of apoptosis by etoposide and dexamethasone, as judged by differential effects of modulators of apoptosis. Staurosporine, UCN-01, and UCN-02 at concentrations of a hundredth to a thousandth of those at which they induced apoptosis, and RO 31-8220 inhibited apoptosis elicited by thapsigargin but not apoptosis caused by dexamethasone or etoposide. The results suggest that (i) UCN-01 and UCN-02 mimic staurosporine as inducers of thymocyte apoptosis; (ii) staurosporine, UCN-01 and UCN-02 share a biphasic effect on apoptosis in rat thymocytes, being inhibitory at low concentrations and stimulatory at high concentrations; and (iii) inhibition of PKC alone is insufficient for induction of apoptosis in thymocytes.

The effects of immunoglobulin isotype and antibody affinity on complement-mediated inhibition of immune precipitation and solubilization.

We have studied complement-mediated prevention of precipitation (PIP) and solubilization of immune complexes (IC) formed with DNP19-BSA and murine monoclonal antibodies (MCAs) of different isotypes and affinities. PIP was effective for IC formed with IgG1 and IgM antibodies, but not for IgA MCA. For IgG1 MCAs, affinity appeared to exert a minor effect on PIP, and IC formed in antibody excess or at equivalence were retained in solution more readily than those formed in antigen excess. For IgM MCAs affinity and antigen-antibody ratio did not affect PIP. As PIP did not occur in Mg-EGTA, it was concluded that PIP was entirely classical pathway dependent. Solubilization of IC containing IgG1 MCAs occurred more rapidly and to a greater extent with low affinity antibodies and an inverse relationship between affinity and the extent of solubilization was observed. Complexes formed with IgG1 MCAs were solubilized relatively poorly when formed in antigen excess. In contrast, affinity and antigen-antibody ratio did not influence the rate and extent of solubilization of IC containing IgM MCAs. IC formed with IgG2b were solubilized rapidly whereas those formed with IgG2a or IgA were solubilized poorly. The relative contributions of the classical and the alternative pathways to solubilization varied with each antibody and the effect of antigen-antibody ratio on these relative contributions was inconsistent.