Platelet-activating factor secreted by DDAVP-treated monocytes mediates von willebrand factor release from endothelial cells

We have previously shown that although DDAVP (1-deamino-8-D-arginine vasopressin), a synthetic analogue of the natural hormone arginine vasopressin, does not directly promote release of vWf from human umbilical vein endothelial cells (ECs), enhanced release does occur when ECs were exposed to either monocytes or to supernatants recovered from DDAVP-treated monocytes. In the present study, we have found that exposure of monocytes to DDAVP did not increase secretion of interleukins (IL)-1β, IL-6, IL-8, tumor necrosis factor (TNF-α), growth factors G-CSF (granulocyte-), GM-CSF (granulocyte, monocyte-colony stimulating factor), prostaglandins (PG) E₂, PGF_2α, or PGI₂ or purine nucleotides such as ATP and ADP. However, increased levels of platelet-activating factor (PAF) were secreted by DDAVP-treated monocytes in a time- and dose-dependent manner that positively correlated with the enhancement in vWf release from ECs. Moreover, this effect could also be elicited when lipid extracts of these supernatants or purified PAF were added directly to ECs. This response could be inhibited with (±)-trans-2,5-Bis(3,4,5-trimethoxyphenyl)-1,3-dioxolane, a specific PAF receptor antagonist, when the ECs were exposed to supernatants from DDAVP-treated monocytes or to pure PAF. The present data indicate that enhanced secretion of PAF from monocytes is one mechanism whereby DDAVP can provoke release of vWf from ECs. © 1993 Wiley-Liss, Inc.     

A comparative study on immunosuppressive effects of cyclosporin A and FK 506 on peripheral blood lymphocytes in dogs

Immunosuppressive effects of cyclosporin A (CsA) and FK 506 (FK) on peripheral blood lymphocytes were studied in dogs in respect to mixed lymphocyte reaction, proliferative responses to recombinant interleukin-2 (rIL-2), phytohemagglutinin (PHA) and concanavalin-A (Con-A); phenotypes of OKIa1, CD3, CD8 and surface IgM; cytotoxic activity against xenogeneic tumor cells. CsA (2.0 or 5.0 mg/kg, intravenously) or FK (0.16 mg/kg, intramuscularly) was given to mongrel dogs every morning for serial 21 days. The blood concentrations of CsA, measured as trough levels by fluorescence polarization method, ranged from 37 to 350 ng/ml in dogs administered at 2.0 mg/kg and from 170 to 894 ng/ml in dogs administered at 5.0 mg/kg during treatment, respectively. In dogs treated with FK at a dose of 0.16 mg/kg, the drug concentrations in the plasma during treatment ranged from 0.16 to 1.8 ng/ml. Mixed lymphocyte reaction and proliferative responses to rIL-2, PHA and Con-A, which were declined by CsA, were not affected by FK. In contrast, the proportion of OKIa1+ cells was not affected by CsA, whereas FK decreased the proportion of OKIa1+ cells progressively during the course of treatment. Cytotoxic activity was suppressed by both CsA and FK. These results possibly indicate that CsA and FK exert their immunosuppressive effects via different mechanisms.     

Meiotic recombination in an Irish family with beta-thalassaemia

Using the technique of allele-specific priming of the polymerase chain reaction (PCR), the C-T substitution in codon 39 was identified as the cause of -thalassaemia in an Irish family. Analysis of the restriction fragment length polymorphisms (RFLPs) in the -globin gene cluster established linkage of the -thalassaemia mutation to a particular -haplotype but indicated that a recombinational event had occurred in the paternal chromosome in the younger of two affected children. Non-paternity was excluded by DNA fingerprinting analysis with hypervariable minisatellite probes. This is the fourth case of recombination in the -globin gene cluster to be reported. The event has occurred 5 of the polymorphic RsaI site at position-550 bp upstream of the -globin gene mRNA Cap site, within the 9.1-kb region that has been shown to be a hot spot for recombination in the -globin gene cluster.     

Erythroid and granulocytic colony growth in cultures supplemented with human serum lipoproteins.

The ability of human serum to support erythroid and granulocytic colony formation has been investigated. It was found that normal human serum could replace fetal calf serum in the cultures and was able to support the growth of these hemopoietic colonies. Serum fractions enriched for low density lipoproteins, either by precipitation with Heparin-Mn++ or by ultracentrifugation, was found to contain this growth supporting activity of human serum.     

Over‐Expression of RNA Processing,Heat Shock,and DNA Repair Proteins in Breast Tumor Compared to Normal Tissue

This study identifies the main changes in protein expression in human breast tumors compared to normal breast tissue. Malignant tumors (32) and normal breast tissue samples (23), from formaldehyde‐fixed, paraffin‐embedded specimens are subjected to discovery proteomics using liquid chromatography/tandem mass spectrometry, with spectral counts for quantitation. The dataset contains 1406 proteins. Differential expression is measured using a method that takes advantage of estimates of the percentage of tumor on a slide. This analysis shows that the major classes of proteins over‐expressed by tumors are RNA‐binding, heat shock and DNA repair proteins. RNA‐binding proteins, including heterogeneous nuclear ribonucleoproteins (HNRNPs), SR splice factors (SRSF) and elongation factors form the largest group. Comparison with results from another study demonstrates that the RNA‐binding proteins are associated specifically with malignant transformation, rather than with cell proliferation. HNRNP and SRSF proteins help define splice sites in normal cells. Their over‐expression may dysregulate splicing, which in turn has the potential to promote malignant transformation.     

Genotoxic and mutagenic effects of lipid-coated CdSe/ZnS quantum dots

We proposed to evaluate the genotoxicity and mutagenicity of a new quantum dots (QDs) nanoplatform (QDsN), consisting of CdSe/ZnS core–shell QDs encapsulated by a natural fusogenic lipid (1,2-di-oleoyl-sn-glycero-3-phosphocholine (DOPC)) and functionalized by a nucleolipid N-[5′-(2′,3′-di-oleoyl) uridine]-N′,N′,N′-trimethylammoniumtosylate (DOTAU). This QDs nanoplatform may represent a new therapeutic tool for the diagnosis and treatment of human cancers. The genotoxic, mutagenic and clastogenic effects of QDsN were compared to those of cadmium chloride (CdCl₂). Three assays were used: (1) the Salmonella/microsome assay with four tester strains, (2) the comet assay and (3) the micronucleus test on CHO cells. The contribution of simulated sunlight was studied in the three assays while oxidative events were only explored in the comet assay in aliquots pretreated with the antioxidant l-ergothioneine. We found that QDsN could enter CHO-K1 cells and accumulate in cytoplasmic vesicles. It was not mutagenic in the Salmonella/mutagenicity test whereas CdCl₂ was weakly positive. In the dark, both the QDsN and CdCl₂ similarly induced dose-dependent increases in single-strand breaks and micronuclei. Exposure to simulated sunlight significantly potentiated the genotoxic activities of both QDsN and CdCl₂, but did not significantly increase micronucleus frequencies. l-Ergothioneine significantly reduced but did not completely suppress the DNA-damaging activity of QDsN and CdCl₂. The present results clearly point to the genotoxic properties and the risk of long-term adverse effects of such a nanoplatform if used for human anticancer therapy and diagnosis in the future.     

Modification of plant bait technique for direct diagnosis for Rhizoctonia rice pathogens from Myanmar paddy field soils

A modified baiting technique was conducted for selective isolation, fungal DNA diagnosis and fungal cell lipid assay derived from Myanmar isolates of Rhizoctonia spp., causal agents of rice sheath diseases by trapping selective plant stem segments. Bait plant materials of rice, mat rush and cotton were successfully used to isolate R. solani AG1-IA, R. oryzae and R. oryzae-sativae. Moreover, the three plant materials were also effectively used to detect genomic DNA derived from all Rhizoctonia spp. obtained from Myanmar. Rice segment was the most successful materials for detection of fungal cell lipids including palmitic, stearic and linoleic acids. The results of this experiment demonstrate that bait plant materials of rice, mat rush and cotton were the best useful tools for not only direct isolation, but also fungal DNA diagnosis and cell lipid assay of Myanmar soil environmental conditions.     

Focused Examination of the Intestinal Epithelium Reveals Transcriptional Signatures Consistent with Disturbances in Enterocyte Maturation and Differentiation during the Course of SIV Infection

The Gastrointestinal (GI) tract plays a pivotal role in AIDS pathogenesis as it is the primary site for viral transmission, replication and CD4⁺ T cell destruction. Accordingly, GI disease (enteropathy) has become a well-known complication and a driver of AIDS progression. To better understand the molecular mechanisms underlying GI disease we analyzed global gene expression profiles sequentially in the intestinal epithelium of the same animals before SIV infection and at 21 and 90 days post infection (DPI). More importantly we obtained sequential excisional intestinal biopsies and examined distinct mucosal components (epithelium. intraepithelial lymphocytes, lamina propria lymphocytes, fibrovascular stroma) separately. Here we report data pertaining to the epithelium. Overall genes associated with epithelial cell renewal/proliferation/differentiation, permeability and adhesion were significantly down regulated (<1.5–7 fold) at 21 and 90DPI. Genes regulating focal adhesions (n = 6), gap junctions (n = 3), ErbB (n = 3) and Wnt signaling (n = 4) were markedly down at 21DPI and the number of genes in each of these groups that were down regulated doubled between 21 and 90DPI. Notable genes included FAK, ITGA6, PDGF, TGFβ3, Ezrin, FZD6, WNT10A, and TCF7L2. In addition, at 90DPI genes regulating ECM-receptor interactions (laminins and ITGB1), epithelial cell gene expression (PDX1, KLF6), polarity/tight junction formation (PARD3B&6B) and histone demethylase (JMJD3) were also down regulated. In contrast, expression of NOTCH3, notch target genes (HES4, HES7) and EZH2 (histone methyltransferase) were significantly increased at 90DPI. The altered expression of genes linked to Wnt signaling together with decreased expression of PDX1, PARD3B, PARD6B and SDK1 suggests marked perturbations in intestinal epithelial function and homeostasis leading to breakdown of the mucosal barrier. More importantly, the divergent expression patterns of EZH2 and JMJD3 suggests that an epigenetic mechanism involving histone modifications may contribute to the massive decrease in gene expression at 90DPI leading to defects in enterocyte maturation and differentiation.