Soluble Melanoma Cell Adhesion Molecule (sMCAM/sCD146) Promotes Angiogenic Effects on Endothelial Progenitor Cells through Angiomotin

The melanoma cell adhesion molecule (CD146) contains a circulating proteolytic variant (sCD146), which is involved in inflammation and angiogenesis. Its circulating level is modulated in different pathologies, but its intracellular transduction pathways are still largely unknown. Using peptide pulldown and mass spectrometry, we identified angiomotin as a sCD146-associated protein in endothelial progenitor cells (EPC). Interaction between angiomotin and sCD146 was confirmed by enzyme-linked immunosorbent assay (ELISA), homogeneous time-resolved fluorescence, and binding of sCD146 on both immobilized recombinant angiomotin and angiomotin-transfected cells. Silencing angiomotin in EPC inhibited sCD146 angiogenic effects, i.e. EPC migration, proliferation, and capacity to form capillary-like structures in Matrigel. In addition, sCD146 effects were inhibited by the angiomotin inhibitor angiostatin and competition with recombinant angiomotin. Finally, binding of sCD146 on angiomotin triggered the activation of several transduction pathways that were identified by antibody array. These results delineate a novel signaling pathway where sCD146 binds to angiomotin to stimulate a proangiogenic response. This result is important to find novel target cells of sCD146 and for the development of therapeutic strategies based on EPC in the treatment of ischemic diseases.     

Functional interaction between PARP-1 and PARP-2 in chromosome stability and embryonic development in mouse

The DNA damage-dependent poly(ADP-ribose) polymerases, PARP-1 and PARP-2, homo- and heterodimerize and are both involved in the base excision repair (BER) pathway. Here, we report that mice carrying a targeted disruption of the PARP-2 gene are sensitive to ionizing radiation. Following alkylating agent treatment, parp-2(-/-)-derived mouse embryonic fibroblasts exhibit increased post-replicative genomic instability, G(2)/M accumulation and chromosome mis-segregation accompanying kinetochore defects. Moreover, parp-1(-/-)parp-2(-/-) double mutant mice are not viable and die at the onset of gastrulation, demonstrating that the expression of both PARP-1 and PARP-2 and/or DNA-dependent poly(ADP-ribosyl) ation is essential during early embryogenesis. Interestingly, specific female embryonic lethality is observed in parp-1(+/-)parp-2(-/-) mutants at E9.5. Meta phase analyses of E8.5 embryonic fibroblasts highlight a specific instability of the X chromosome in those females, but not in males. Together, these results support the notion that PARP-1 and PARP-2 possess both overlapping and non-redundant functions in the maintenance of genomic stability.     

Temporal Trends in Treatment Outcomes for HIV-1 and HIV-2-Infected Adults Enrolled in C?te d'Ivoire's National Antiretroviral Therapy Program

Background

In Côte d''Ivoire during 2004–2007, numbers of ART enrollees increased from <5,000 to 36,943. Trends in nationally representative ART program outcomes have not yet been reported.

Methodology/Principal Findings

We conducted a retrospective chart review to assess trends in patient characteristics and attrition [death or loss to follow-up (LTFU)] over time, among a nationally representative sample of 3,682 adults (≥15 years) initiating ART during 2004–2007 at 34 health facilities. Among ART enrollees during 2004–2007, median age was 36, the proportion female was 67%, the proportion HIV-2-infected or dually HIV-1&2 reactive was 5%, and median baseline CD4⁺ T-cell (CD4) count was 135 cells/µL. Comparing cohorts initiating ART in 2004 with cohorts initiating ART in 2007, median baseline weight declined from 55 kg to 52 kg (p = 0.008) and the proportion weighing <45 kg increased from 17% to 22% (p = 0.014). During 2004–2007, pharmacy-based estimates of the percentage of new ART enrollees ≥95% adherent to ART declined from 74% to 60% (p = 0.026), and twelve-month retention declined from 86% to 69%, due to increases in 12-month mortality from 2%–4% and LTFU from 12%–28%. In univariate analysis, year of ART initiation was associated with increasing rates of both LTFU and mortality. Controlling for baseline CD4, weight, adherence, and other risk factors, year of ART initiation was still strongly associated with LTFU but not mortality. In multivariate analysis, weight <45 kg and adherence <95% remained strong predictors of LTFU and mortality.

Conclusions

During 2004–2007, increasing prevalence among ART enrollees of measured mortality risk factors, including weight <45 kg and ART adherence <95%, might explain increases in mortality over time. However, the association between later calendar year and increasing LTFU is not explained by risk factors evaluated in this analysis. Undocumented transfers, political instability, and patient dissatisfaction with crowded facilities might explain increasing LTFU.     

How are sticky chromosomes formed?

Blood lymphocytes in culture were irradiated by gamma-rays 3 h to 30 mn before harvesting. The various induced lesions were analysed, with a particular attention on sticky chromosomes, i.e. radial figures in which chromosomes are not obviously broken, but are linked by a tiny filament. Such anomalies are preferentially induced in mid to late G2-phase. They result from recombinations occurring at nonrandom chromosome regions: junction between hetero- and euchromatin, and telomeric regions. It is proposed that they are formed when double strand breaks are induced while intrachromatidic links have started to be formed in the course of chromosome condensation. If this interpretation is correct, the apparent lack of induced breakage of premitotic chromosomes is artifactual.     

Modelling based method for pharmacokinetic hypotheses test

The aim of this work is to propose methods to test mechanism of synergy of toxic agents in bees. A synergy between prochloraz, an imidazole fungicide, and deltamethrin, a pyrethroid insecticide, was demonstrated experimentally. The hypothesis is that prochloraz modifies the penetration or the metabolism of deltamethrin. This hypothesis is tested using a pharmacokinetic box model. A previous experimental work showed that bee instantaneous mortalities were higher, from the time t ₁ to the time t ₂ after spraying, in groups sprayed with deltamethrin at dose D ₀ in the presence of prochloraz (+P) than in those sprayed with deltamethrin alone at a dose time as high (). We postulate that accrued mortality is proportional to the cumulated internal deltamethrin (ID ₂). ID ₂ of treatment (+P) had to be greater than ID ₂ of treatment () during the period from t ₁ to t ₂ so that the hypothesis would be consistent with the experimental data. The limit, for which the hypothesis is conceivable, is the ID ₂₍₎ = ID _2(+P ) curve. We study, in particular, the asymptotic behaviour of the limit curve when different parameters of the kinetic model tend to 0 or . These limits allow to verify quickly and easily whether a mechanism is conceivable or not As the limits are calculated with algebraic values, the test can be used for other synergies.     

Radiation-induced carcinogenesis: Individual sensitivity and genomic instability

In spite of a well-known relationship between exposure to radiation and increased risk for cancer development, the biological mechanisms involved in radiation-induced carcinogenesis remain poorly documented. Various hypotheses are discussed in this paper. It appears that radiation cannot be directly responsible for the numerous genetic alterations of cancer cells. Most of them occur during tumor progression. Only one or a very limited number of them was induced by radiation many years before tumor growth. This long delay is a major difficulty for experimental research and raises many questions. Recently, it has been shown that a genomic instability occurs after many generations in cells descending from irradiated cells. This instability leads to multiple genetic alterations and, preferentially, affects some chromosome structures, particularly telomeres. This kind of telomeric instability - related to the shortening of telomeric DNA sequences - has also been observed in senescent cells as well as in non-senescent cells from patients predisposed to cancer, and this process may possibly also occur in the progeny of irradiated cells.Invited paper presented at the International Symposium on Heavy Ion Research: Space, Radiation Protection and Therapy, Sophia-Antipolis, France, 21–24 March 1994     

Chemical synthesis and characterization of Pi1, a scorpion toxin from Pandinus imperator active on K+ channels.

Pi1 is a 35-residue toxin cross-linked by four disulfide bridges that has been isolated from the venom of the chactidae scorpion Pandinus imperator. Due to its very low abundance in the venom, we have chemically synthesized this toxin in order to study its biological activity. Enzyme-based proteolytic cleavage of the synthetic Pi1 (sPi1) demonstrates half-cystine pairings between Cys4-Cys25, Cys10-Cys30, Cys14-Cys32 and Cys20-Cys35, which is in agreement with the disulfide bridge organization initially reported on the natural toxin. In vivo, intracerebroventricular injection of sPi1 in mice produces lethal effects with an LD50 of 0.2 microgram per mouse. In vitro, the application of sPi1 induces drastic inhibition of Shaker B (IC50 of 23 nM) and rat Kv1.2 channels (IC50 of 0.44 nM) heterologously expressed in Xenopus laevis oocytes. No effect was observed on rat Kv1.1 and Kv1.3 currents upon synthetic peptide application. Also, sPi1 is able to compete with 125I-labeled apamin for binding onto rat brain synaptosomes with an IC50 of 55 pM. Overall, these results demonstrate that sPi1 displays a large spectrum of activities by blocking both SK- and Kv1-types of K+ channels; a selectivity reminiscent of that of maurotoxin, another structurally related four disulfide-bridged scorpion toxin that exhibits a different half-cystine pairing pattern.     

The rate of chromosome breakage is age dependent in lymphocytes of adult controls

Summary Chromosome breaks and chromatid-type lesions from a prospective study of more than 1000 lymphocyte karyotypes from each of six controls were analysed. These lesions were more frequent in older (75 years old on average) than in younger (29 years old on average) controls, especially after 72h cultures. All controls were found to be carriers of fragile sites. The most frequent were 3p14.3 and 16q23, especially in older controls. At least one fra(X)(q27) mitosis was found in each control. Most deletions occurred after breakage in heterochromatin or in late-replicating euchromatin. As almost all radials were either mitotic chiasmata or triradials (branched chromosomes), it is concluded that chromatid exchanges between non-homologous segments are very rare, and indicate chromosomal instability syndrome or recent exposure to a mutagen.     

Reduction and reoxidation of the neurotoxin II from the scorpion Androctonus australis Hector

Reoxidation of the totally reduced scorpion neurotoxin II from Androctonus australis Hector (four disulfide bridges) has been investigated. The totally reduced toxin was highly insoluble in neutral and alkaline conditions, which prevented the use of the usual air oxidation process for renaturation. We tested a new method in which the reduced molecules were first solubilized in 10% (v/v) acetic acid and then oxidized by air through dialysis against a series of buffers with a slow pH gradient from 2.2 to 7.0 or 8.0. In this system, up to 95% of the protein was recovered in solution. Addition of reduced and oxidized glutathione accelerated refolding and also permitted a better recovery of fully active peptide as measured by both toxicity to mice and ability to displace 125I radiolabeled toxin II from its binding site on rat brain synaptosomal fractions. The reoxidation reaction could also be monitored directly by high pressure liquid chromatography. A strong effect of guanidine hydrochloride concentration as well as the temperature was observed both on the solubility of the reoxidation intermediates and on the refolding pathway. Finally, the method used, i.e. dialysis reoxidation with a pH gradient from 2.2 to 8.0 in 0.1 M sodium phosphate, 0.1 M sodium chloride, 20 mM guanidine hydrochloride, 1 mM oxidized and reduced glutathione allowed regeneration in high yield (70%) of a reoxidized toxin form indistinguishable from the native toxin. A minor stable and inactive molecular species (about 30%) showing a difference in mobility by electrophoresis was also detected.