Establishment and characterization of a megakaryoblast cell line with amplification of MLL

A new cell line with megakaryoblastic features, designated UoC-M1, was established from the malignant cells of a 68-year-old patient with acute myeloid leukemia. The patient's leukemic cells reacted with alpha-naphthyl acetate esterase and acid phosphatase and expressed CD7, CD24, CD34, CD38, CD45, HLA-DR and CD61. Cytogenetic analysis of the patient's malignant cells (and of the UoC-M1 cells) showed a human, male hypodiploid karyotype with many chromosome rearrangements and marker chromosomes. Spectral karyotyping (SKY) analysis complemented the G-banded karyotyping and clarified several chromosomal translocations and identified the marker chromosomes. Fluorescence in situ hybridization (FISH) and SKY analysis demonstrated that one marker chromosome contained three segments of chromosome 9 interspersed with three segments of chromosome 11, as well as a portion of chromosome 19. FISH analysis with a probe for MLL revealed that the UoC-M1 cells contained four copies of the MLL gene. Southern blot analysis determined that the MLL gene had a germline profile while Northern and Western analyses showed that the MLL mRNAs and protein were of the appropriate sizes. This is the first report of amplification of the MLL gene which may be an additional mechanism of leukemogenesis or disease progression.     

Accuracy of technicians versus pharmacists in checking syringes prepared for a dialysis program

The accuracy of pharmacy technicians versus pharmacists in checking drug doses prepared in syringes for a dialysis program was studied. Three pharmacy technicians from the pharmacy of a regional kidney disease program in Minnesota participated in the study after completing a training program and after common preparation errors had been identified by pharmacists. From November 1995 to April 1996, the technicians used labels printed from a database of pharmacist-verified orders to prepare and label i.v. syringes. Four medications were used-epoetin alfa, calcitriol, heparin prepared from beef lung, and heparin prepared from porcine intestinal mucosa. Each syringe was checked by one of nine pharmacists for accuracy of dose and medication, and all errors were recorded. The technicians checked syringes prepared by other technicians and also recorded errors. Accuracy rates (percentages of syringes correctly evaluated) for pharmacists and technicians were compared. A total of 10,608 syringes were checked. Accuracy rates for pharmacists and pharmacy technicians were 99.86% and 99.83%, respectively. Accuracy rates in checking syringes did not differ significantly between pharmacists and technicians in this study setting.     

Regional myocardial motion and thickening assessed at rest by ECG-gated 99mTc-MIBI emission tomography and by magnetic resonance imaging

We have validated ECG-gated emission tomography using technetium-99m methoxyisobutylisonitrile for the assessment of regional ventricular function by comparing it with cine magnetic resonance imaging (MRI). Gated tomography was performed at rest in 24 patients referred for myocardial perfusion imaging [17 males and seven females with a mean age of 58 years, nine of whom had had a previous myocardial infarction (MI)]. Scores were assigned to each of nine myocardial segments for wall motion and for thickening. Cine MRI was analysed in an identical fashion. Four out of 216 (2%) segments were uninterpretable by gated tomography because of inadequate tracer uptake. In eight patients without coronary artery disease (CAD), wall motion and thickening were normal by both methods. Gated tomography showed abnormal wall motion or thickening in all patients with previous MI and in five of seven patients with CAD but no prior MI. Association between wall motion and thickening was good (rs=0. 86). Overall, there was good agreement between gated tomography and MRI for both wall motion (178/212 segments, kappa=0.66) and wall thickening (184/212 segments, kappa=0.69). In segments with severely reduced perfusion, however, there was poorer agreement (kappa=0.31). Interobserver and intraobserver agreement was high (kappa from 0.61 to 0.78). Thus, in patients investigated for CAD, there is good overall agreement between gated tomography and MRI but the agreement is lower in segments with severe perfusion defects.     

Flicker photometric sensitivity and chromatic adaptation in color-deficient and normal observers

Congenital red-green color deficients and observers with normal color vision were asked to make flicker photometric matches on a red background, on a green background, and with no background present. Adaptation had no effect on the matches made by color deficients, but clearly affected the matches of normal observers. There were small differences between the matches of different observers. We attempted to characterize these differences by fitting the R and G curves of Smith and Pokorny to the data. Two parameters, Λ_max and optical density, were varied to obtain the best fit. Extremely good fits were obtained, suggesting that individual differences can be characterized with these two parameters.     

Postnatal development of respiratory function in lambs studied serially between birth and 8 weeks

The molecular structure of the telomeric region at the left arm of the second chromosome of the mosquito Anopheles gambiae has been determined in the transformed strain G418 that contains a pUChsneo transgene attached at the 2L chromosome end, and in the Pink eye laboratory strain (PE). Both strains contain the same complex satellite positioned distal to a unique region. FIGE mapping of the telomeric region of the PE strain revealed distinct DNA fragment lengths that segregated with individual chromosomes. Genomic DNA fragments were cloned from the 2L telomeric region, which accounted for about half of 2L chromosomes in the PE population. In all three cases studied, long fragments of different middle repetitive sequences were found attached to the distal ends of the 2L satellite. We propose that random fragments of DNA may be occasionally added during recombination between complex satellite repeats at the chromosome ends.     

Modulation of antigen processing and presentation by covalently linked complement C3b fragment

Ligands such as complement fragments (C3, C4), IgG or alpha 2-macroglobulin, which bind antigen (Ag) before their uptake by antigen-presenting cells (APC), are likely to modulate the different steps of Ag processing and presentation. These ligands contribute to internalization and endosomal targeting of Ag; they also influence its processing and, consequently, the binding of resulting peptides to major histocompatibility complex (MHC) class II molecules before presentation to T cells. Complement protein C3 contains, like other members of the alpha 2-macroglobulin family, an intrachain thiolester bond. Conformational alteration or limited proteolysis of C3 into C3b leads to breaking of the thiolester with transient capacity of the revealed carbonyl group to esterify hydroxyl groups of Ag. Ester-linked complexes including tetanus toxin (TT) and C3b were prepared to analyse the influence of bound C3b on TT processing and presentation by APC. Covalent binding of C3b to TT resulted in increased and prolonged stimulation of specific T-cell proliferation. This effect was observed with non-specific B cells, as well as with a TT-specific B-cell clone, as APC. On the other hand, SDS-PAGE analysis of proteolysates of TT or C3b-TT, obtained with endosome/lysosome-enriched subcellular fractions prepared from human Epstein-Barr virus (EBV)-transformed B cells, indicated a delay of TT proteolysis when TT was associated to C3b. Treatment of APC with protease inhibitors, before and during exposure of the cells to Ag, resulted in differences in the inhibition of TT and C3b-TT proteolysis. Using purified cathepsins B and D, we demonstrated that covalent binding of C3b to TT totally abolished TT proteolysis by cathepsin D, while proteolysis by cathepsin B was preserved. This finding and the absence of cathepsin B in endosomes may explain a delay in TT processing when it is associated to C3b. Confirming these data, presentation by formaldehyde-fixed cells of C3b-TT proteolysates showed higher stimulation of specific T-cell clones than formaldehyde-fixed TT proteolysates.