Identical abnormality of the short arm of chromosome 18 in two Philadelphia-positive chronic myelocytic leukemia patients with erythroblastic transformation,resulting in duplication of BCR-ABL1 fusion

Two patients with Ph-positive chronic myelocytic leukemia in erythroblastic transformation and rearrangement of the short arm of chromosome 18 are reported. Fluorescence in situ hybridization studies showed that the 18p rearrangement resulted from translocation of the main part of chromosome 22 long arm to 18p, including BCR-ABL1 fusion. The 18p abnormality resulted, thus, in loss of 18p and duplication of BCR-ABL1 in both patients. The possible relation to the erythroblastic type of blastic phase is briefly discussed. In addition an apparently intact germline ABL1 gene was duplicated and inserted into chromosome 6 at band p21 in one of these patients.     

Differential stability and fusion activity of Lyssavirus glycoprotein trimers

The oligomeric structure and the fusion activity of lyssavirus glycoprotein (G) was studied by comparing G from Mokola virus (GMok) and rabies virus (PV strain) (GPV), which are highly divergent lyssaviruses. G expressed at the surface of BSR cells upon either plasmid transfection or virus infection are shown to be mainly trimeric after cross-linking experiments. However, solubilization by a detergent (CHAPS) and analysis in sucrose sedimentation gradient evidenced that GMok trimer is less stable than GPV trimer. A chimeric glycoprotein (G Mok-PV) associating the N-terminal half of GMok to the C-terminal half part of GPV formed trimers with an intermediate stability, indicating that the G C-terminal domain is essential in trimer stability. A cell to cell fusion assay revealed that GMok (and not G Mok-PV) was able to induce fusion at a higher pH (0.5 pH unit) than GPV. Such differences in the oligomeric structure stability and in the fusion activity of lyssavirus glycoproteins may partly account for the previously reported differences of their immunogenic and pathogenic properties.     

Critical stages of tumor growth regulation in transgenic mice harboring a hepatocellular carcinoma revealed by distinct patterns of tumor necrosis factor-alpha and transforming growth factor-beta mRNA production

There is now good evidence that cytokines contribute to the regulation of tumor growth. The cytokine-driven modulation of tumor growth was investigated during the progression of a hepatocellular carcinoma (HCC) in SV40 large T tumor antigen transgenic mice. In vivo, an increased rate of liver growth correlated with increased transforming growth factor (TGF)-beta 1 mRNA expression, while the greatest amounts of tumor necrosis factor (TNF)-alpha mRNA were detected earlier during tumor development. Conversely, no particular alteration of IL-1 alpha, IL-1 beta, IL-6, IL-2, IL-4 and IFN-gamma mRNA production could be reported. In vitro, hepatocyte-like tumor cell lines established at two stages, either before or after HCC differentiation, were characterized. The early-stage-derived cell line produced TNF-alpha mRNA, but had barely detectable expression of TGF-beta 1 mRNA, while later-stage- derived cell lines showed the reciprocal pattern. All cell lines displayed a lack of sensitivity to TNF-alpha, although some degree of sensitivity to TNF-alpha could be observed in the presence of actinomycin-D or after treatment with IFN-gamma. The early-stage- derived cell line was sensitive to the growth inhibitory effects of TGF- beta 1, but late-stage-derived tumor cell lines displayed a loss of sensitivity to TGF-beta 1 which correlated with the increased expression of TGF-beta 1 mRNA. Altogether, this suggests that tumor cells contribute to the discrete TNF-alpha and TGF-beta 1 expression patterns during HCC progression. This model of HCC could be of valuable interest to assess the impact of various immunotherapeutic strategies on modulation of tumor growth.      

Arrhythmogenic right ventricular cardiomyopathy versus fatty replacement of the right ventricle. An autopsy case report

We report an autopsy case of a cardiomyopathy characterized by fatty replacement of the right ventricular myocardium and compare its clinical and histologic characteristics with those of the arrhythmogenic right ventricular cardiomyopathy. A 39-year old male died suddenly in a hospital room. He had an alcoholic cirrhosis with ascitis, but the clinical examination and the biology showed no abnormalities explaining the death. Histologically, in the right ventricle, large areas of cardiomyocytes were replaced by fat, but there was no fibrosis. In contrast, fibrosis is present in association with fat in arrhythmogenic right ventricular cardiomyopathy. Fatty replacement of the right ventricle is likely to be a distinct entity. Right ventricular failure has been shown to be a possible complication. Sudden death is probably rare and is likely to occur when other arrhythmogenic factors are associated.     

Axonal and glial currents activated during the post-tetanic hyperpolarization in non-myelinated nerve

 Changes in membrane potential and potassium concentration in the extracellular space ([K⁺]_e) of rabbit vagus nerve were measured simultaneously during electrical activity and during the period of recovery using a modified sucrose-gap method and potassium-sensitive microelectrodes. After stimulation for 15 s at 15 Hz the main activity-induced increase in [K⁺]_e reached 16.9 mM. This increase in [K⁺]_e was paralleled by a depolarization of the preparation. The period of activity was followed by a post-tetanic hyperpolarization (PTH) lasting tens of seconds, generated by the axonal electrogenic Na⁺-K⁺ pump and to a lesser extent by the pump of the surrounding Schwann cells. The amplitude of the PTH dramatically increased in experiments in which inward currents were blocked by removal of Cl^– or after application of Cs⁺ or Ba²⁺, indicating that under normal conditions the current generated by the Na⁺-K⁺ pump is strongly short-circuited. A pharmacological and kinetic study showed that these currents are: (1) the hyperpolarization-activated current I _h, and (2) the inwardly rectifying I _KIR current. The results show that the latter originates from Schwann cells. Our data indicate that in non-myelinated nerves there is a subtle association of inward ionic channels which (1) helps the cell to maintain an optimal membrane potential after a period of activity, and (2) contributes to the removal of excess K⁺ from the extracellular space. Received: 7 August 1997 / Received after revision 6 April 1998 / Accepted: 15 April 1998     

Fully automatic quantification of microarray image data

DNA microarrays are now widely used to measure expression levels and DNA copy number in biological samples. Ratios of relative abundance of nucleic acids are derived from images of regular arrays of spots containing target genetic material to which fluorescently labeled samples are hybridized. Whereas there are a number of methods in use for the quantification of images, many of the software systems in wide use either encourage or require extensive human interaction at the level of individual spots on arrays. We present a fully automatic system for microarray image quantification. The system automatically locates both subarray grids and individual spots, requiring no user identification of any image coordinates. Ratios are computed based on explicit segmentation of each spot. On a typical image of 6000 spots, the entire process takes less than 20 sec. We present a quantitative assessment of performance on multiple replicates of genome-wide array-based comparative genomic hybridization experiments. By explicitly identifying the pixels in each spot, the system yields more accurate estimates of ratios than systems assuming spot circularity. The software, called, runs on Windows platforms and is available free of charge for academic use.     

Transport of Proteins Across the Human Placenta

PROBLEM: The transport of various proteins across the human placenta was investigated by comparing maternal and fetal concentrations of tetanus antigen (TT-AG), anti-tetanus (TT)-immunoglobulin G (IgG) (following maternal vaccination), IgA, human chorionic gonadotropin (hCG), human placental lactogen (hPL), and alpha-fetoprotein (AFP) at term. METHOD OF STUDY: The concentrations of the six proteins were determined using enzyme-linked immunosorbent assay in serum of maternal venous and umbilical (fetal) vein samples obtained at delivery from uncomplicated term pregnancies (n = 16). RESULTS: The ratios (mean ± standard deviation) of fetal (umbilical) to maternal level were 1.41 ± 0.33 (anti-TT-IgG), 0.91 ± 0.37 (TT-AG), 0.002 ± 0.001 (IgA), 0.003 ± 0.001 (hCG), and 0.008 ± 0.004 (hPL), while the maternal:fetal concentration ratio of AFP was 0.002 ± 0.002. IgA, hCG, hPL, and AFP showed a close correlation between maternal and fetal levels varying between r² = 0.47 to 0.73 (P < 0.004–0.0001). Because AFP is produced by the fetus while IgA originates in the mother, the appearance of small amounts of these two proteins in the maternal or fetal compartment, respectively, suggests a slow rate of diffusion following a high concentration gradient. The detection of hCG and hPL in fetal serum is also interpreted as diffusion from the maternal into the fetal blood. Anti-TT-IgG has a significantly higher concentration in the fetal as compared with the maternal serum, which is in line with the well-documented active transfer of IgG. Fetal TT-antigen levels were similar to maternal concentrations, showing a close correlation (r² = 0.74, P < 0.0001) between the two proteins. CONCLUSIONS: The correlation between maternal and fetal concentrations of various proteins like IgA (150,000 Da), hCG (42,000 Da), and hPL (21,000 Da) suggests passive diffusion of these macromolecules across the placenta from the maternal to the fetal side, albeit at a slow rate. A similar process is postulated for AFP (70,000 Da) diffusing in the opposite direction from the fetus to the mother. There was no significant difference between the transplacental fetomaternal gradient of IgA and hCG and the maternal-fetal gradient of AFP. In view of the substantially larger volume of circulating maternal as compared with fetal blood, a significantly higher rate of crossing of AFP as compared with the other proteins must be assumed. It is uncertain whether a difference in the rate of transplacental transfer in the two directions or an additional source of AFP production in the maternal compartment explains the high maternal level. Anti-TT-IgG concentration is significantly higher in fetal than in maternal serum suggesting active transfer from the mother to the fetus. Furthermore, there is considerable transfer of TT-AG and a close correlation of fetal:maternal ratios of anti-TT-IgG (150,000 Da) and TT-AG (150,000 Da) could be an indication for a specific transfer of the antigen antibody complex.     

The anti-apoptotic factor Bcl-2 can functionally substitute for the B cell survival but not for the marginal zone B cell differentiation activity of BAFF

The TNF family ligand B cell-activating factor (BAFF, BLyS, TALL-1) is an essential factor for B cell development. BAFF binds to three receptors, BAFF-R, transmembrane activator and CAML interactor (TACI), and B cell maturation antigen (BCMA), but only BAFF-R is required for successful survival and maturation of splenic B cells. To test whether the effect of BAFF is due to the up-regulation of anti-apoptotic factors, TACI-Ig-transgenic mice, in which BAFF function is inhibited, were crossed with transgenic mice expressing FLICE-inhibitory protein (FLIP) or Bcl-2 in the B cell compartment. FLIP expression did not rescue B cells, while enforced Bcl-2 expression restored peripheral B cells and the ability to mount T-dependent antibody responses. However, many B cells retained immaturity markers and failed to express normal amounts of CD21. Marginal zone B cells were not restored and the T-independent IgG3, but not IgM, response was impaired in the TACI-IgxBcl-2 mice. These results suggest that BAFF is required not only to inhibit apoptosis of maturating B cells, but also to promote differentiation events, in particular those leading to the generation of marginal zone B cells.