Homeostasis of naive and memory CD4+ T cells: IL-2 and IL-7 differentially regulate the balance between proliferation and Fas-mediated apoptosis

Cytokines play a crucial role in the maintenance of polyclonal naive and memory T cell populations. It has previously been shown that ex vivo, the IL-7 cytokine induces the proliferation of naive recent thymic emigrants (RTE) isolated from umbilical cord blood but not mature adult-derived naive and memory human CD4(+) T cells. We find that the combination of IL-2 and IL-7 strongly promotes the proliferation of RTE, whereas adult CD4(+) T cells remain relatively unresponsive. Immunological activity is controlled by a balance between proliferation and apoptotic cell death. However, the relative contributions of IL-2 and IL-7 in regulating these processes in the absence of MHC/peptide signals are not known. Following exposure to either IL-2 or IL-7 alone, RTE, as well as mature naive and memory CD4(+) T cells, are rendered only minimally sensitive to Fas-mediated cell death. However, in the presence of the two cytokines, Fas engagement results in a high level of caspase-dependent apoptosis in both RTE as well as naive adult CD4(+) T cells. In contrast, equivalently treated memory CD4(+) T cells are significantly less sensitive to Fas-induced cell death. The increased susceptibility of RTE and naive CD4(+) T cells to Fas-induced apoptosis correlates with a significantly higher IL-2/IL-7-induced Fas expression on these T cell subsets than on memory CD4(+) T cells. Thus, IL-2 and IL-7 regulate homeostasis by modulating the equilibrium between proliferation and apoptotic cell death in RTE and mature naive and memory T cell subsets.     

Functional polymorphisms in the TERT promoter are associated with risk of serous epithelial ovarian and breast cancers

Genetic variation at the TERT-CLPTM1L locus at 5p15.33 is associated with susceptibility to several cancers, including epithelial ovarian cancer (EOC). We have carried out fine-mapping of this region in EOC which implicates an association with a single nucleotide polymorphism (SNP) within the TERT promoter. We demonstrate that the minor alleles at rs2736109, and at an additional TERT promoter SNP, rs2736108, are associated with decreased breast cancer risk, and that the combination of both SNPs substantially reduces TERT promoter activity.     

Cell surface expression of the bovine leukemia virus-binding receptor on B and T lymphocytes is induced by receptor engagement

Bovine leukemia virus (BLV), one of the most common infectious viruses of cattle, is endemic in many herds. Approximately 30-40% of adult cows in the United States are infected by this oncogenic C-type retrovirus and 1-5% of animals will eventually develop a malignant lymphoma. BLV, like the human and simian T cell leukemia viruses, is a deltaretrovirus but, in contrast with the latter, the BLV receptor remains unidentified. In this study, we demonstrate that the amino-terminal 182 residues of the BLV envelope glycoprotein surface unit encompasses the receptor-binding domain. A bona fide interaction of this receptor-binding domain with the BLV receptor was demonstrated by specific interference with BLV, but not human T cell leukemia virus, envelope glycoprotein-mediated binding. We generated a rabbit Ig Fc-tagged BLV receptor-binding domain construct and ascertained that the ligand binds the BLV receptor on target cells from multiple species. Using this tool, we determined that the BLV-binding receptor is expressed on differentiating pro/pre-B cells in mouse bone marrow. However, the receptor was not detected on mature/quiescent B cells but was induced upon B cell activation. Activation of human B and T lymphocytes also induced surface BLV-binding receptor expression and required de novo protein synthesis. Receptor levels were down-regulated as activated lymphocytes returned to quiescence. In the human thymus, BLV-binding receptor expression was specifically detected on thymocytes responding to the IL-7 cytokine. Thus, expression of the BLV-binding receptor is a marker of enhanced metabolic activity in B cells, T cells, and thymocytes.     

Galactodendritic Phthalocyanine Targets Carbohydrate-Binding Proteins Enhancing Photodynamic Therapy

Photosensitizers (PSs) are of crucial importance in the effectiveness of photodynamic therapy (PDT) for cancer. Due to their high reactive oxygen species production and strong absorption in the wavelength range between 650 and 850 nm, where tissue light penetration is rather high, phthalocyanines (Pcs) have been studied as PSs of excellence. In this work, we report the evaluation of a phthalocyanine surrounded by a carbohydrate shell of sixteen galactose units distributed in a dendritic manner (PcGal₁₆) as a new and efficient third generation PSs for PDT against two bladder cancer cell lines, HT-1376 and UM-UC-3. Here, we define the role of galacto-dendritic units in promoting the uptake of a Pc through interaction with GLUT1 and galectin-1. The photoactivation of PcGal₁₆ induces cell death by generating oxidative stress. Although PDT with PcGal₁₆ induces an increase on the activity of antioxidant enzymes immediately after PDT, bladder cancer cells are unable to recover from the PDT-induced damage effects for at least 72 h after treatment. PcGal₁₆ co-localization with galectin-1 and GLUT1 and/or generation of oxidative stress after PcGal₁₆ photoactivation induces changes in the levels of these proteins. Knockdown of galectin-1 and GLUT1, via small interfering RNA (siRNA), in bladder cancer cells decreases intracellular uptake and phototoxicity of PcGal₁₆. The results reported herein show PcGal₁₆ as a promising therapeutic agent for the treatment of bladder cancer, which is the fifth most common type of cancer with the highest rate of recurrence of any cancer.     

Flow cytometric analysis of recombinant murine GM-CSF (rmuGM-CSF) induced changes in the distribution of specific cell populations in vivo

The changes in the distribution of granulocytes, monocytes, and lymphocytes in various tissue compartments following subcutaneous (SC) administration of recombinant murine GM-CSF (rmuGM-CSF) in vivo was determined by flow cytometry in time course studies. Balb/c mice were given single, daily SC injections of 1 or 4 micrograms of rmuGM-CSF for 10 days. Flow cytometric analysis was performed on bone marrow (BMC), peritoneal exudate (PEC), and peripheral blood (PBC) cell preparations from mice treated for 1, 3, and 10 days. Dual fluorescence was employed to gate on leukocytes (T200+) and analyze for Ig+, Thy 1.2+, MAC+, and 8C5+ (granulocytes) cells. The analyses indicated that SC-rmuGM-CSF increased the percentage of 8C5+ cells in PBC after 1 day of treatment. However, significant changes in the cell composition of PEC and BMC were not observed until day 10 of treatment and included increases in 8C5+ cells and the myeloid cell population, respectively. Side scatter analysis (cell density) of PBC and PEC indicated that the percentage of the granulocytic cell population increased significantly in rmuGM-CSF treated mice. The changes observed in PEC and BMC appeared to be dose-related whereas those observed in PBC were not. These data clearly demonstrate the utility of flow cytometric analyses for detecting selective effects of cytokines on cell populations that are involved in host defense mechanisms.     

Characterization of a novel type of HIV-1 particle assembly inhibitor using a quantitative luciferase-Vpr packaging-based assay

The HIV-1 auxiliary protein Vpr and Vpr-fusion proteins can be copackaged with Gag precursor (Pr55Gag) into virions or membrane-enveloped virus-like particles (VLP). Taking advantage of this property, we developed a simple and sensitive method to evaluate potential inhibitors of HIV-1 assembly in a living cell system. Two proteins were coexpressed in recombinant baculovirus-infected Sf9 cells, Pr55Gag, which formed the VLP backbone, and luciferase fused to the N-terminus of Vpr (LucVpr). VLP-encapsidated LucVpr retained the enzymatic activity of free luciferase. The levels of luciferase activity present in the pelletable fraction recovered from the culture medium correlated with the amounts of extracellular VLP released by Sf9 cells assayed by conventional immunological methods. Our luciferase-based assay was then applied to the characterization of betulinic acid (BA) derivatives that differed from the leader compound PA-457 (or DSB) by their substituant on carbon-28. The beta-alanine-conjugated and lysine-conjugated DSB could not be evaluated for their antiviral potentials due to their high cytotoxicity, whereas two other compounds with a lesser cytotoxicity, glycine-conjugated and ε-NH-Boc-lysine-conjugated DSB, exerted a dose-dependent negative effect on VLP assembly and budding. A fifth compound with a low cytotoxicity, EP-39 (ethylene diamine-conjugated DSB), showed a novel type of antiviral effect. EP-39 provoked an aberrant assembly of VLP, resulting in nonenveloped, morula-like particles of 100-nm in diameter. Each morula was composed of nanoparticle subunits of 20-nm in diameter, which possibly mimicked transient intermediates of the HIV-1 Gag assembly process. Chemical cross-linking in situ suggested that EP-39 favored the formation or/and persistence of Pr55Gag trimers over other oligomeric species. EP-39 showed a novel type of negative effect on HIV-1 assembly, targeting the Pr55Gag oligomerisation. The biological effect of EP-39 underlined the critical role of the nature of the side chain at position 28 of BA derivatives in their anti-HIV-1 activity.     

The ubiquitous glucose transporter GLUT-1 is a receptor for HTLV

The human T cell leukemia virus (HTLV) is associated with leukemia and neurological syndromes. The physiopathological effects of HTLV envelopes are unclear and the identity of the receptor, present on all vertebrate cell lines, has been elusive. We show that the receptor binding domains of both HTLV-1 and -2 envelope glycoproteins inhibit glucose transport by interacting with GLUT-1, the ubiquitous vertebrate glucose transporter. Receptor binding and HTLV envelope-driven infection are selectively inhibited when glucose transport or GLUT-1 expression are blocked by cytochalasin B or siRNAs, respectively. Furthermore, ectopic expression of GLUT-1, but not the related transporter GLUT-3, restores HTLV infection abrogated by either GLUT-1 siRNAs or interfering HTLV envelope glycoproteins. Therefore, GLUT-1 is a receptor for HTLV. Perturbations in glucose metabolism resulting from interactions of HTLV envelope glycoproteins with GLUT-1 are likely to contribute to HTLV-associated disorders.     

Chemical composition and industrial applications of Maritime pine (Pinus pinaster Ait.) bark and other non-wood parts

Reviews in Environmental Science and Bio/Technology - Maritime pine (Pinus pinaster Ait.) is native to the western Mediterranean basin and has an economic importance in southwestern Europe. This...