Amplified fluorescence response of chemosensors grafted onto silica nanoparticles

In conventional fluorescent chemosensors, the recognition of the target by the receptor unit affects the fluorescence properties of a single covalently coupled fluorescent moiety. Here we show for the first time that when a suitable TSQ derivative is densely grafted onto the surface of preformed silica nanoparticles electronic interactions between the individual chemosensor units enable the free units to recognize the state of the surrounding complexed ones. As a result, the fluorescence transduction is not limited to the local site where binding occurs, but it involves a wider region of the fluorophore network that is able to transfer its excitation energy to the complexed units. Such behavior leads to an amplification of the fluorescence signal. What we report here is the first example of amplification in the case an off-on chemosensor due to its organization onto the surface of silica nanoparticles. We also describe a simple general model to approach amplification in multifluorophoric systems based on the localization of the excited states, which is valid for assemblies such as the supramolecular ones where molecular interactions are weak and do not significantly perturb the individual electronic states. The introduction of an amplification factorf in particular allows for a simple quantitative estimation of the amplification effects

Amplified fluorescence response of chemosensors grafted onto silica nanoparticles

In conventional fluorescent chemosensors, the recognition of the target by the receptor unit affects the fluorescence properties of a single covalently coupled fluorescent moiety. Here we show for the first time that when a suitable TSQ derivative is densely grafted onto the surface of preformed silica nanoparticles electronic interactions between the individual chemosensor units enable the free units to recognize the state of the surrounding complexed ones. As a result, the fluorescence transduction is not limited to the local site where binding occurs, but it involves a wider region of the fluorophore network that is able to transfer its excitation energy to the complexed units. Such behavior leads to an amplification of the fluorescence signal. What we report here is the first example of amplification in the case an off-on chemosensor due to its organization onto the surface of silica nanoparticles. We also describe a simple general model to approach amplification in multifluorophoric systems based on the localization of the excited states, which is valid for assemblies such as the supramolecular ones where molecular interactions are weak and do not significantly perturb the individual electronic states. The introduction of an amplification factorf in particular allows for a simple quantitative estimation of the amplification effects

PANCREATIC CANCER (PaCa)-DERIVED SOLUBLE MEDIATORS INDUCE DENDRITIC CELLS (DC) TO ACQUIRE AN IMMUNESUPPRESSIVE PHENOTYPE BY DOWNREGULATING CTLA4

Objective: An altered function of lymphocytes, DC and immature myeloid cells appears to be an hallmark of tumor-mediated immune suppression and the two inhibitory co-stimulatory receptors PDL-1 and CTLA4 might have a role in this context. The aim of the present in vitro study was to assess whether PaCa cells cross-talk with normal mononuclear circulating cells (PBMC) causing them to acquire an immunesuppressive phenotype and to evaluate whether PDL1 and CTLA4 are involved. Methods: PBMC from blood donors were cultured for 4 days in Control (CTL) and in the PaCa cancer cell line Capan1 conditioned media (CM). Lymphocytes subsets (CD4+, CD8+, CD4+CD25+) and CD33+ immature myeloid cells subsets (CD14+/-; HLA-DR+/-) expressing or not PDL1 and/or CTLA4 were analysed by flow cytometry. To assess immunesuppressive function, myeloid cells were FACS sorted and co-coltured with allogenic total T lymphocytes in 1:20 and 1:40 ratio. Total T lymphocytes proliferation was determined by 3HThymidine uptake. Results: Capan1 CM caused an expansion of CD4+CD25+ (p=0.01) and a reduction of CD33+CD14- HLA-DR+ (p=0.03) cells. In this latter cellular subset, CM caused also an increase of PDL1 (p=0.046) and a decrease of CTLA4 (p=0.05) positive cells. FACS sorted CTL and CM CD33+CD14-HLA-DR+ cells did not significantly affect the proliferation of allogenic total T lymphocytes at 1:20 (p=0,54) or at 1:40 ratio (p=0,81). The CD33+CD14-HLA-DR+ PDL-1+ cells did not significantly modify allogenic T cells proliferation with respect to PDL- cells (p=0,11), while those cells which were CTLA4 negative caused a significant inhibition of T cell proliferation in comparison of CTLA4 positive cells (p=0,008). Conclusions: PaCa-derived soluble factors induce the expansion of the inhibitory lymphocytes subset CD4+CD25+ and a reduction of the immature CD33+CD14-DR+ dendritic cells. The tumor associated reduced expression of the inhibitory molecule CTLA4 in this cell population was demonstrated to characterize an immunosuppressive phenotype and this study suggests to take care in the use of anti-CTLA4 therapies

Sarcoidosis is a Th1/Th17 multisystem disorder

BACKGROUND AND AIMS: Sarcoidosis is characterised by a compartmentalisation of CD4(+) T helper 1 (Th1) lymphocytes and activated macrophages in involved organs, including the lung. Recently, Th17 effector CD4(+) T cells have been claimed to be involved in the pathogenesis of granuloma formation. The objective of this study was to investigate the involvement of Th17 cells in the pathogenesis of sarcoidosis. METHODS: Peripheral and pulmonary Th17 cells were evaluated by flow cytometry, real-time PCR, immunohistochemistry analyses and functional assays in patients with sarcoidosis in different phases of the disease and in control subjects. RESULTS: Th17 cells were detected both in the peripheral blood (4.72 \ub1 2.27% of CD4(+) T cells) and in the bronchoalveolar lavage (BAL) (8.81 \ub1 2.25% of CD4(+) T lymphocytes) of patients with sarcoidosis and T cell alveolitis. Immunohistochemical analysis of lung and lymph node specimens showed that interleukin 17 (IL-17)(+)/CD4(+) T cells infiltrate sarcoid tissues surrounding the central core of the granuloma. IL-17 was expressed by macrophages infiltrating sarcoid tissue and/or forming the granuloma core (7.88 \ub1 2.40% of alveolar macrophages). Analysis of some lung specimens highlighted the persistence of IL-17(+)/CD4(+) T cells in relapsed patients and their absence in the recovered cases. Migratory assays demonstrated the ability of the Th17 cell to respond to the chemotactic stimulus CCL20-that is, the CCR6 ligand (74.8 \ub1 8.5 vs 7.6 \ub1 2.8 migrating BAL lymphocytes/high-powered field, with and without CCL20, respectively). CONCLUSIONS: Th17 cells participate in the alveolitic/granuloma phase and also to the progression towards the fibrotic phase of the disease. The recruitment of this cell subset may be driven by CCL20 chemokine