A novel approach to evaluate the lung cancer risk of airborne particles emitted in a city.

Air quality still represents a main threat to human health in cities. Even in developed countries, decades of air pollution control not yet allowed to reduce pollutant concentrations in urban areas adequately. Indeed, high airborne particle concentrations are measured in several European cities; this is a main issue since particles represent a carrier for carcinogenic compounds. Numerous researches measuring the exposure to the different aerosol metrics in urban areas were recently performed, nonetheless, few data on the lung cancer risk in such environments are available. In the present paper a novel approach to evaluate the lung cancer risk related to the airborne particles emitted by the different sources located in a city is proposed and applied to a pilot case-study (i.e. an Italian city). In particular, an existing lung cancer risk model was modified and applied to assess the particle-related lung cancer "emitted" by the different sources of the city using pollutant emission factors provided by accredited emission inventory databases. Therefore, the average toxicity of the particles emitted by the city (i.e. lung cancer slope factor) and the lung cancer risk globally emitted by the city, expressed as new cases of lung cancer, were evaluated. The proposed emission inventory also allowed to identify and localize the main contributors to the overall risk emitted in a city. As an example, for the city under investigation, the research revealed that the main contributor, amongst the sources considered, is the vehicular traffic which is characterized by a lower mass fraction of carcinogenic compounds but a much higher sub-micron particle emission with respect to the other sources.

Method for measuring settling phenomena by means of frequency domain instrumentation.

The paper deals with the analysis of settling phenomena that characterize the step response of digital to analog converters, amplifiers, and several other devices. Settling is described by means of a minimal second order model that is suitable to account for the distortion terms recognized in the signal spectrum. An alternative method for dynamic performance assessment of systems characterized by poor settling performance is then proposed. Thanks to the use of high bandwidth spectrum analyzers, the proposed method overtakes the limits characterizing the measurement approaches based on the use of time-domain instruments in the presence of modern ultra-wideband systems.

Neural stem cell-mediated CE/CPT-11 enzyme/prodrug therapy in transgenic mouse model of intracerebellar medulloblastoma.

Medulloblastoma is a heterogeneous diffuse neoplasm that can be highly disseminated, and is the most common malignant childhood brain tumor. Although multimodal treatments have improved survival rates for patients with medulloblastoma, these tumors are associated with high morbidity and mortality. New treatment strategies are urgently needed to improve cure rates and, importantly, to spare normal brain tissue from neurotoxicity and patients from life-long cognitive and functional deficits associated with current therapies. In numerous preclinical brain tumor models, neural stem cells (NSCs) have shown great promise as delivery vehicles for therapeutic genes. Here, we have used an established, genetically modified human NSC line (HB1.F3.CD) to deliver carboxylesterase (CE) to cerebellar tumor foci and locally activate the prodrug camptothecin-11 (CPT-11) (Irinotecan) to the potent topoisomerase I inhibitor SN-38. HB1.F3.CD NSC tumor tropism, intratumoral distribution and therapeutic efficacy were investigated in clinically relevant experimental models. Magnetic resonance imaging was used for in vivo tracking of iron nanoparticle-labeled NSCs, and to assess the therapeutic efficacy of CE-expressing HB1.F3.CD cells. As compared with controls, a significant decrease in tumor growth rate was seen in mice that received both NSCs and CPT-11 as their treatment regimen. Thus, this study provides proof-of-concept for NSC-mediated CE/CPT-11 treatment of medulloblastoma, and serves as a foundation for further studies toward potential clinical application.

Successful allogeneic hematopoietic stem cell transplantation for acute myelogenous leukemia in a patient with hepatic echinococcal cyst managed by delayed hepatectomy.

We report the case of a patient who underwent cytotoxic chemotherapy and allogeneic hematopoietic stem cell transplantation (HSCT) for acute myelogenous leukemia in the presence of hepatic cystic echinococcal infection. The presence of Echinococcus granulosus infection in an immunocompromised host is extremely rare, with lack of established data regarding optimal management. Successful management of the patient's disease processes required a multidisciplinary approach, which included systemic chemotherapy, HSCT, treatment of chronic graft-versus-host-disease, and elective en bloc resection of the hepatic cyst.

Abundant GFP expression and LTP in hippocampal acute slices by in vivo injection of sindbis virus.

Virus-mediated gene transfer into neurons is a powerful tool for the analysis of neuronal structure and function. Recombinant sindbis virus has been previously used to study protein function in hippocampal neuron cultures as well as in hippocampal organotypic slice cultures. Nevertheless, some concern still exists about the physiological relevance of these cultured preparations. Acute hippocampal slices are a widely used preparation for the study of synaptic transmission, but currently recombinant gene delivery is usually achieved only through time-consuming transgenic techniques. In this study, we show that a subregion of the CA1 area in acute hippocampal slices can be specifically altered to express a gene of interest. A sindbis virus vector carrying an enhanced green fluorescent protein (EGFP) reporter was injected in vivo into the hippocampus of adult rats. After 18 h, rats were killed, and acute hippocampal slices, infected in the CA1 field, were analyzed morphologically and electrophysiologically. Infected slices showed healthy and stable electrophysiological responses as well as long-term potentiation. In addition, infected pyramidal cells were readily recognized in living slices by two-photon imaging. Specifically, the introduction of an EGFP-Actin fusion protein greatly enhanced the detection of fine processes and dendritic spines. We propose this technique as an efficient tool for studying gene function in adult hippocampal neurons.

TYMSTR, a putative chemokine receptor selectively expressed in activated T cells, exhibits HIV-1 coreceptor function.

BACKGROUND: Chemokines bind to specific receptors and mediate leukocyte migration to sites of inflammation. Recently, some chemokine receptors, notably CXCR4 and CCR5, have been shown to be essential fusion factors on target cells for infection by human immunodeficiency virus (HIV); the chemokines bound by these receptors have also been shown to act as potent inhibitors of HIV infection. Here, we describe the isolation of a novel, putative chemokine receptor. RESULTS: We have isolated the cDNA for a putative human chemokine receptor, which we have termed TYMSTR (T-lymphocyte-expressed seven-transmembrane domain receptor). The TYMSTR gene is localized to human chromosome 3 and encodes a protein that has a high level of identity with chemokine receptors. TYMSTR mRNA was selectively expressed in interleukin-2-stimulated T lymphocytes but not in freshly isolated lymphocytes and leukocytes or related cell lines. The natural ligand for TYMSTR was not identified among 32 human chemokines and other potential ligands. Cells co-expressing TYMSTR and human CD4 fused with cells expressing envelope glycoproteins of macrophage (M)-tropic HIV-1 as well as T-cell line (T)-tropic HIV-1 isolates. Addition of infectious, T-tropic HIV-1 particles to TYMSTR/CD4-expressing cells resulted in viral entry and proviral DNA formation. CONCLUSIONS: Our findings demonstrate that TYMSTR, in combination with CD4, mediates HIV-1 fusion and entry. The high-level expression of TYMSTR in CD4(+) T lymphocytes and the selectivity of this receptor for T-tropic and M-tropic HIV-1 strains indicates that TYMSTR might function as HIV coreceptor at both early and late stages of infection.

The chemokine SDF-1, stromal cell-derived factor 1, attracts early stage B cell precursors via the chemokine receptor CXCR4.

In the bone marrow, progenitor (pro-) and precursor (pre-) B cells depend on close contact with stromal cells for growth and maturation. Stromal cell-derived factor 1 (SDF-1), also known as pre-B cell growth-stimulating factor, is produced by bone marrow stromal cells and was reported to act together with interleukin-7 as co-mitogen for pre-B cells. SDF-1 was recently shown to be a chemokine which is chemotactic for different types of leukocytes and acts via the chemokine receptor CXCR4. Using sorted B220+ bone marrow cells and several B cell lines characteristic for different stages of B lymphopoiesis, we now show that SDF-1 is a potent attractant for pro- and pre-B cells, but is inactive on B cells at later stages of development. In early B cell precursors, SDF-1 induced intracellular Ca2+ mobilization and in vitro migration with a potency and efficacy similar to that observed for chemokines acting on blood leukocytes. These responses were mediated via CXCR4 as they could be inhibited by an antireceptor antibody. SDF-1 is the first chemokine shown to act on early-stage B cell precursors. Mice lacking SDF-1 die perinatally and show a severe deficiency in B lymphopoiesis. We propose that SDF-1 released from the stromal cells exerts its critical hematopoietic function by selectively attracting and confining early B cell precursors within the bone marrow microenvironment that provides the necessary factors for growth and differentiation.

Cloning of ATAC, an activation-induced, chemokine-related molecule exclusively expressed in CD8+ T lymphocytes.

A cDNA clone, designated ATAC, was isolated from a collection of human T cell activation genes. Analysis of tissue distribution determined that ATAC mRNA of approximately 0.9 kb is exclusively expressed in activated CD8+ T cells. Induction of the ATAC gene requires stimulation by both phorbol 12-myristate 13-acetate and Ca2+ ionophore A23187 ("two-signal gene") and is fully abrogated by the immunosuppressive agent cyclosporin A. Upon stimulation, ATAC mRNA is detectable within 30 min, maximal expression is seen after 4 h. The polypeptide encoded by the open reading frame of ATAC mRNA is 114 amino acids long with a calculated M(r) of 12.52 kDa. The structural features predict the cleavage and secretion of a mature ATAC protein of approximately 10 kDa from the 12.52-kDa precursor. ATAC is highly similar to a very recently identified murine molecule designated lymphotactin both at the cDNA (73.8% identity) and the protein (61.4% identity) levels, and related to members of the C-C and C-X-C chemokine families. Two variants of the ATAC protein were expressed and tested for chemotaxis and Ca2+ release on a variety of target cells. The ATAC gene was located to chromosome 1q23.

Actions of the chemotactic cytokines MCP-1, MCP-2, MCP-3, RANTES, MIP-1 alpha and MIP-1 beta on human monocytes.

The activities of six synthetic CC chemokines, MCP-1, MCP-2, MCP-3, RANTES, MIP-1 alpha and MIP-1 beta on human blood monocytes were studied. All CC chemokines elicited a bimodal migration response in vitro. Highest numbers of migrating cells were obtained with the monocyte chemotactic proteins (MCP) and RANTES, somewhat lower numbers with MIP-1 alpha, and only weak migration with MIP-1 beta. The most potent attractants were MCP-1 and MIP-1 alpha which reached maximum efficacy at 0.1 to 1 nM. All CC chemokines also induced the release of N-acetyl-beta-D-glucosaminidase from cytochalasin B-pretreated monocytes. The MCP were most effective (MCP-1 > MCP-3 > MCP-2), RANTES and MIP-1 alpha showed moderate (1/3 of MCP-1 activity), and MIP-1 beta only minimal activity. Cytosolic free Ca2+ changes and exocytosis were used to monitor receptor desensitization. Marked cross-desensitization was observed among MCP-1, MCP-2 and MCP-3 on the one hand, and RANTES, MIP-1 alpha and MIP-1 beta on the other, indicating receptor sharing within these two subgroups of CC chemokines. The responses to RANTES, MIP-1 alpha and MIP-1 beta were also moderately to markedly desensitized by pretreatment with MCP-1, MCP-2 or MCP-3, while the responses to the MCP were virtually unaffected by pretreatment with RANTES, MIP-1 alpha and MIP-1 beta. These results suggest that the MCP also interact with receptors recognized by RANTES, MIP-1 alpha and MIP-1 beta, but not vice versa. Binding studies were performed with radiolabeled MCP-1 or MIP-1 alpha. All MCP competed readily for labeled MCP-1 yielding a concentration-dependent sigmoidal displacement curve. Displacement with RANTES, MIP-1 alpha and MIP-1 beta was observed at higher concentrations, but was not complete. Radiolabeled MIP-1 alpha was displaced efficiently by MIP-1 alpha or MIP-1 beta, but only partially by RANTES. Of the MCP, only MC-3 completely displaced MIP-1 alpha, while only partial displacement was observed with MCP-1 and MCP-2.

A cell surface ELISA for the screening of monoclonal antibodies to antigens on viable cells in suspension.

To simplify the screening of monoclonal antibodies to different human T cell surface molecules a live cell enzyme-linked immunosorbent assay (cell ELISA) has been established and optimized. The assay was performed in 96-well plates. By using living human T lymphocytes in suspension surface modification by fixation or insolubilization of the cells was avoided. Several parameters influencing sensitivity and specificity were studied. About 150 ng/ml of mouse monoclonal antibodies to cell surface antigens could be detected when using 5 x 10(4) cells per well and a 1/1000 dilution of the anti-mouse IgG-alkaline phosphatase conjugate. This sensitivity permitted the primary screening of cell specific antibodies from hybridoma supernatants. The same detection limit was obtained in flow cytometric analysis. If required, the sensitivity of the cell ELISA could be increased using higher cell numbers and conjugate concentration. When analysing different cell lines with selected antibodies the cell ELISA was found to be as sensitive and specific as the fluorescence assay. The assay was applied to the screening of supernatants from hybridomas developed against human T helper cell clones and the detection of V beta specificities of T cell clones.