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Find video protocols related to scientific articles indexed in Pubmed.
In Vivo Efficacy Study of Milk Thistle Extract (ETHIS-094™) in STAM™ Model of Nonalcoholic Steatohepatitis.
Drugs R D
PUBLISHED: 11-19-2014
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A subcategory of nonalcoholic fatty liver disease (NAFLD), nonalcoholic steatohepatitis (NASH) is characterized by accumulation of fat accompanied by inflammatory infiltration and hepatocellular damage. The active complex of milk thistle is a lipophilic extract from its seeds, comprising three isomers, collectively known as silymarin. Silymarin has demonstrated antioxidant, anti-inflammatory, and antifibrotic properties, and has been extensively studied in the treatment of liver diseases. The majority of published clinical research on silymarin has used Legalon(®) (Rottapharm/Madaus), containing the patented extract of milk thistle ETHIS-094™ (Euromed). The current study was undertaken to examine the effects of ETHIS-094™ in the Stelic Animal Model (STAM™), a validated and widely used animal model for NASH.
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Co-Culture System of human salivary gland epithelial cells and immune cells from primary Sjögren's syndrome patients: an in vitro approach to study the Rituximab effects on the Raf-1/ERK1/2 pathway activation.
Int. Immunol.
PUBLISHED: 11-10-2014
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Primary Sjögren's syndrome (pSS) is a chronic autoimmune disorder of the exocrine glands with associated lymphocytic infiltrates of the affected glands. Dryness of the mouth and eyes results from involvement of the salivary and lacrimal glands. Efficacy of Rituximab (RTX) in pSS is still open to debate. This study delineates the signalling pathway involved in RTX-mediated pro-inflammatory factors down-regulation in a co-culture system of pSS salivary gland epithelial cells (SGEC) with syngeneic pSS B-lymphocytes. In addition, the effects of RTX on the Raf-1/ERK1/2 pathway activation in pSS SGEC co-cultured with syngeneic pSS T-lymphocytes were also investigated. This study demonstrated that RTX may interfere with the ERK1/2 pathway in a syngeneic co-culture of pSS SGEC with pSS B lymphocytes, leading to decreased cytokines production by SGEC. These novel findings reveal that syngeneic co-culture of pSS SGEC with pSS B lymphocytes leads to a down-regulation of Raf-1 in epithelial cells that adversely regulates the activity of the ERK1/2 pathway and determines a subsequent reduction of the pro-inflammatory factors release.
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Skin/nail infections with the addition of pertuzumab to trastuzumab-based chemotherapy.
Breast Cancer Res. Treat.
PUBLISHED: 10-30-2014
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We report a series of breast cancer patients with invasive skin and nail infections with Staphylococcus species that we attribute to the addition of pertuzumab to trastuzumab-based therapy. With the suspicion of an increased incidence of cutaneous infection in patients treated with pertuzumab and trastuzumab-based chemotherapy, treating medical oncologists identified patients receiving therapy who experienced infection. Between March and October 2014, 18 patients treated with pertuzumab and trastuzumab-based chemotherapy were found to have 21 separate skin/nail infections. Treatment was administered as neoadjuvant therapy in 12 (67 %) patients, adjuvant therapy in four (22 %) patients, and for metastatic disease in two (11 %) patients. Granulocyte growth factors were administered in 11 (61 %) patients and no patients were documented to be neutropenic. New skin and nail lesions developed as early as cycle 1 and as late as 8 months from initial therapy. The 21 separate infections documented were folliculitis and "bite-like" lesions (10), abscess (6), paronychia (3), and cellulitis (2). The appearance of these lesions was distinct from typical EGFR-associated skin changes. When cultures were obtained, Staphylococcus species were isolated. Quantitative immunoglobulins were assessed in 14 (78 %) patients and were abnormally low in six (43 %) of these patients. The skin infections resulted in treatment delay in two (11 %) patients and premature discontinuation of therapy in one patient. We believe that the skin/nail infections reported here in patients treated with the combination of pertuzumab and trastuzumab represent a previously unrecognized toxicity of adding pertuzumab to trastuzumab-based therapies.
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Impact of treatment site in adolescents and young adults with central nervous system tumors.
J. Natl. Cancer Inst.
PUBLISHED: 09-01-2014
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Adolescents and young adults (AYAs; aged 15-39 years) have inferior survival in comparison with younger (aged 0-14 years) cancer patients. Impact of care at specialized centers such as National Cancer Institute-designated Comprehensive Cancer Centers (NCICCC) for AYAs of all ages or the Children's Oncology Group (COG) for AYAs aged 15 to 21 years with central nervous system (CNS) tumors remains unstudied.
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ADAM17 at the interface between inflammation and autoimmunity.
Immunol. Lett.
PUBLISHED: 08-27-2014
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The discovery of the disintegrin and metalloproteinase 17 (ADAM17), originally identified as tumor necrosis factor-a converting enzyme (TACE) for its ability as sheddase of TNF-? inspired scientists to attempt to elucidate the molecular mechanisms underlying ADAM17 implication in diseased conditions. In recent years, it has become evident that this protease can modify many non matrix substrates, such as cytokines (e.g. TNF-?), cytokine receptors (e.g. IL-6R and TNF-R), ligands of ErbB (e.g. TGF-? and amphiregulin) and adhesion proteins (e.g. Lselectin and ICAM-1). Several recent studies have described experimental model system to better understand the role of specific signaling molecules, the interplay of different signals and tissue interactions in regulating ADAM17-dependent cleavage of most relevant substrates in inflammatory diseases. The central question is whether ADAM17 can influence the outcome of inflammation and if so, how it performs this regulation in autoimmunity, since inflammatory autoimmune diseases are often characterized by deregulated metalloproteinase activities. This review will explore the latest research on the influence of ADAM17 on the progression of inflammatory processes linked to autoimmunity and its role as modulator of inflammation.
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Sustained virological response with intravenous silibinin: individualized IFN-free therapy via real-time modelling of HCV kinetics.
Liver Int.
PUBLISHED: 07-31-2014
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Intravenous silibinin (SIL) is a potent antiviral agent against hepatitis C virus (HCV) genotype-1. In this proof of concept case-study we tested: (i) whether interferon-alfa (IFN)-free treatment with SIL plus ribavirin (RBV) can achieve sustained virological response (SVR); (ii) whether SIL is safe and feasible for prolonged duration of treatment and (iii) whether mathematical modelling of early on-treatment HCV kinetics can guide duration of therapy to achieve SVR.
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New Approach Based on Compressive Sampling for Sample Rate Enhancement in DASs for Low-Cost Sensing Nodes.
Sensors (Basel)
PUBLISHED: 07-03-2014
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The paper deals with the problem of improving the maximum sample rate of analog-to-digital converters (ADCs) included in low cost wireless sensing nodes. To this aim, the authors propose an efficient acquisition strategy based on the combined use of high-resolution time-basis and compressive sampling. In particular, the high-resolution time-basis is adopted to provide a proper sequence of random sampling instants, and a suitable software procedure, based on compressive sampling approach, is exploited to reconstruct the signal of interest from the acquired samples. Thanks to the proposed strategy, the effective sample rate of the reconstructed signal can be as high as the frequency of the considered time-basis, thus significantly improving the inherent ADC sample rate. Several tests are carried out in simulated and real conditions to assess the performance of the proposed acquisition strategy in terms of reconstruction error. In particular, the results obtained in experimental tests with ADC included in actual 8- and 32-bits microcontrollers highlight the possibility of achieving effective sample rate up to 50 times higher than that of the original ADC sample rate.
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NAMI-A is highly cytotoxic toward leukaemia cell lines: evidence of inhibition of KCa 3.1 channels.
Dalton Trans
PUBLISHED: 07-01-2014
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We report here that the established anticancer ruthenium(iii) complex NAMI-A induces potent and selective cytotoxic effects in a few leukaemia cell lines. These results sound very surprising after 20 years of intense studies on NAMI-A, commonly considered as a "non-cytotoxic" antimetastatic agent. In addition, evidence is given for selective inhibition of KCa 3.1 channels. The implications of these findings are discussed.
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Universal scaling effects of a temperature gradient at first-order transitions.
Phys Rev E Stat Nonlin Soft Matter Phys
PUBLISHED: 06-25-2014
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We study the effects of smooth inhomogeneities at first-order transitions. We show that a temperature gradient at a thermally driven first-order transition gives rise to nontrivial universal scaling behaviors with respect to the length scale l_{t} of the variation of the local temperature T_{x}. We propose a scaling ansatz to describe the crossover region at the surface where T_{x}=T_{c}, where the typical discontinuities of a first-order transition are smoothed out. The predictions of this scaling theory are checked, and get strongly supported, by numerical results for the two-dimensional (2D) Potts models, for a sufficiently large number of states to have first-order transitions. Comparing with analogous results at the 2D Ising transition, we note that the scaling behaviors induced by a smooth inhomogeneity appear quite similar in first-order and continuous transitions.
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ER Reorganization is Remarkably Induced in COS-7 Cells Accumulating Transmembrane Protein Receptors Not Competent for Export from the Endoplasmic Reticulum.
J. Membr. Biol.
PUBLISHED: 05-27-2014
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The newly synthesized mutant L501fsX533 Frizzled-4 form and the alpha3beta4 nicotinic acetylcholine receptor expressed in the absence of nicotine accumulate in the endoplasmic reticulum of COS-7 cells and induce the formation of large areas of smooth and highly convoluted cisternae. This results in a generalized block of the transport to the Golgi complex of newly synthesized proteins. Intriguingly, both effects happen peculiarly in COS-7 cells; HeLa, Huh-7, and HEK293 cells expressing the two receptors at similar level than COS-7 cells show normal ER and normal transport toward the plasma membrane. These results question the conclusion that a dominant-negative mechanism would explain the dominance of the mutant L501fsX533 Fz4 allele in the transmission of a form of Familial exudative vitreoretinopathy. Moreover, they indicate that the coordination of endoplasmic reticulum homeostasis in COS-7 cells is particularly error prone. This finding suggests that COS-7 cells may be extremely useful to study the molecular mechanisms regulating endoplasmic reticulum size and architecture.
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Severity of liver disease affects HCV kinetics in patients treated with intravenous silibinin monotherapy.
Antivir. Ther. (Lond.)
PUBLISHED: 05-21-2014
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HCV kinetic analysis and modeling during antiviral therapy have not been performed in decompensated cirrhotic patients awaiting liver transplantation. Here, viral and host parameters were compared in patients treated with daily intravenous silibinin (SIL) monotherapy for 7 days according to the severity of their liver disease.
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Neovascularization is prominent in the chronic inflammatory lesions of Sjögren's syndrome.
Int J Exp Pathol
PUBLISHED: 04-29-2014
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Angiogenesis is a common finding in chronic inflammatory diseases; however, its role in Sjögren's syndrome (SS) remains to be elucidated. Previous SS studies have demonstrated an increase in VEGF-A/VEGFR-2 system expression in minor salivary gland (MSG) biopsies from patients with SS, but differences in the new blood vessel formation between the different grades of disease severity have not been reported. Therefore, experiments were performed to demonstrate angiogenesis during different phases of primary SS (pSS) and to define the relationship between the microvessel density (MVD), macrophage infiltration and histiocyte distribution in SS MSG inflammatory lesions. In this series of experiments, immunohistochemistry was used to examine angiogenesis in serial sections of pSS MSG. Patients with pSS were classified accordingly with the grade of inflammatory lesions as I = low-grade (low focus score of 1 or 2), II = intermediate-grade (focus score of 3–6) and III = extensive inflammation in the MSG (high focus score of 12). Histological examination demonstrated that the MVD increased with the severity of the inflammatory lesions, and in addition, we found an increased infiltration of inflammatory and pro-angiogenic cells.These findings reveal that angiogenesis is intimately involved in the progression of pSS, may be central to the propagation of the chronic immune response observed in pSS and could represent a novel potential biomarker of pSS disease activity.
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Mapping glucose-mediated gut-to-brain signalling pathways in humans.
Neuroimage
PUBLISHED: 02-17-2014
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Previous fMRI studies have demonstrated that glucose decreases the hypothalamic BOLD response in humans. However, the mechanisms underlying the CNS response to glucose have not been defined. We recently demonstrated that the slowing of gastric emptying by glucose is dependent on activation of the gut peptide cholecystokinin (CCK1) receptor. Using physiological functional magnetic resonance imaging this study aimed to determine the whole brain response to glucose, and whether CCK plays a central role.
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HDAC-inhibitor (S)-8 disrupts HDAC6-PP1 complex prompting A375 melanoma cell growth arrest and apoptosis.
J. Cell. Mol. Med.
PUBLISHED: 02-13-2014
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Histone deacetylase inhibitors (HDACi) are agents capable of inducing growth arrest and apoptosis in different tumour cell types. Previously, we reported a series of novel HDACi obtained by hybridizing SAHA or oxamflatin with 1,4-benzodiazepines. Some of these hybrids proved effective against haematological and solid cancer cells and, above all, compound (S)-8 has emerged for its activities in various biological systems. Here, we describe the effectiveness of (S)-8 against highly metastatic human A375 melanoma cells by using normal PIG1 melanocytes as control. (S)-8 prompted: acetylation of histones H3/H4 and ?-tubulin; G0 /G1 and G2 /M cell cycle arrest by rising p21 and hypophos-phorylated RB levels; apoptosis involving the cleavage of PARP and caspase 9, BAD protein augmentation and cytochrome c release; decrease in cell motility, invasiveness and pro-angiogenic potential as shown by results of wound-healing assay, down-regulation of MMP-2 and VEGF-A/VEGF-R2, besides TIMP-1/TIMP-2 up-regulation; and also intracellular accumulation of melanin and neutral lipids. The pan-caspase inhibitor Z-VAD-fmk, but not the antioxidant N-acetyl-cysteine, contrasted these events. Mechanistically, (S)-8 allows the disruption of cytoplasmic HDAC6-protein phosphatase 1 (PP1) complex in A375 cells thus releasing the active PP1 that dephosphorylates AKT and blocks its downstream pro-survival signalling. This view is consistent with results obtained by: inhibiting PP1 with Calyculin A; using PPP1R2-transfected cells with impaired PP1 activity; monitoring drug-induced HDAC6-PP1 complex re-shuffling; and, abrogating HDAC6 expression with specific siRNA. Altogether, (S)-8 proved very effective against melanoma A375 cells, but not normal melanocytes, and safe to normal mice thus offering attractive clinical prospects for treating this aggressive malignancy.
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Chronic inflammation enhances NGF-?/TrkA system expression via EGFR/MEK/ERK pathway activation in Sjögren's syndrome.
J. Mol. Med.
PUBLISHED: 01-28-2014
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Primary Sjögren's syndrome (pSS) is a chronic autoimmune exocrine disease associated with variable lymphocytic infiltration of the affected organs (primarily salivary and lachrymal glands). To investigate the potential implication of nerve growth factor-? (NGF-?) and its high affinity receptor tyrosine kinase receptor A (TrkA) in the regulation of pSS inflammatory responses, we studied their expression in the human salivary gland epithelial cells (SGEC) cultures from pSS minor salivary glands (MSG) biopsies and their relationship with histopathological disease parameters. Here, we demonstrated an increased expression of the NGF-?/TrkA system in pSS SGEC, correlated with the MSG inflammation grade. The results demonstrate that the pro-inflammatory cytokines TNF-? and IL-6 enhance NGF-? production; on the contrary, NGF-? production was reduced in the presence of both Raf-1 kinase and MEK inhibitors. Furthermore, TNF-?/IL-6 treatment increased ERK1/2 phosphorylation. Inhibition of the EGF/EGFR system also decreased NGF-? release by pSS SGEC, indicating that the chronic inflammatory condition characteristic of pSS enhances NGF-? production via EGFR/Raf-1/MEK/ERK pathway activation.
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hERG1 channels regulate VEGF-A secretion in human gastric cancer: clinicopathological correlations and therapeutical implications.
Clin. Cancer Res.
PUBLISHED: 01-21-2014
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hERG1 channels are aberrantly expressed in several types of human cancers, where they affect different aspects of cancer cell behavior. A thorough analysis of the functional role and clinical significance of hERG1 channels in gastric cancer is still lacking.
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Rituximab-mediated Raf kinase inhibitor protein induction modulates NF-?B in Sjögren syndrome.
Immunology
PUBLISHED: 01-15-2014
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Primary Sjögren syndrome (pSS) is an autoimmune disorder characterized by an epithelial injury surrounded by dense lymphocytic infiltrates. The conditions for the long-term maintenance of human salivary gland epithelial cells from pSS patients and a co-culture system with pSS lymphocytes were used to assess the effect of Rituximab (RTX) on the inflammatory condition and progression in pSS. Quantitative real-time PCR, genes and protein array analysis, Western blot, flow cytometry, small interfering RNA transfection and nuclear factor-?B (NF-?B) DNA binding assays were used as methods. Supporting the benefits of RTX, this study demonstrates that RTX decreases NF-?B activity and interrupts the NF-?B signalling pathway through the up-regulation of the Raf-1 kinase inhibitor protein (RKIP). Over-expression of RKIP down-regulates interleukins, their receptors and the expression of genes encodes proteins that attracted lymphocytes. Silencing of the RKIP gene leads to significantly increased expression and release of pro-inflammatory mediators supporting that RKIP expression could be involved in the suppression of NF-?B activation in pSS salivary gland epithelial cells.
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Antiviral activity and safety profile of silibinin in HCV patients with advanced fibrosis after liver transplantation: a randomized clinical trial.
Transpl. Int.
PUBLISHED: 01-14-2014
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Response to interferon-based therapies in HCV recurrence after liver transplantation (LT) is unsatisfactory, and major safety issues aroused in preliminary experience with boceprevir and telaprevir. As transplant community identified HCV viral clearance as a critical matter, efficacious and safe anti-HCV therapies are awaited. The aim of this study was to assess efficacy and safety of intravenous silibinin monotherapy in patients with established HCV recurrence after LT, nonresponders to pegylated interferon and ribavirin. This is a single center, prospective, randomized, parallel-group, double-blind, placebo-controlled, phase 2 trial including 20 patients randomly assigned (3:1) to receive daily 20 mg/kg of intravenous silibinin or saline as placebo, for 14 consecutive days. On day 14 of treatment, viral load decreased by 2.30 ± 1.32 in silibinin group versus no change in the placebo group (P = 0.0002). Sixteen days after the end of the treatment, viral load mean values were similar to baseline. Treatment resulted well tolerated apart from a transient and reversible increase in bilirubin. Neither changes in immunosuppressant through levels nor dosage adjustments were necessary. Silibinin monotherapy has a significant antiviral activity in patients with established HCV recurrence on the graft not responding to standard therapy and confirms safety and tolerability without interaction with immunosuppressive drugs (ClinicalTrials.gov number: NCT01518933).
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The metalloproteinase ADAM17 and the epidermal growth factor receptor (EGFR) signaling drive the inflammatory epithelial response in Sjögren's syndrome.
Clin. Exp. Med.
PUBLISHED: 01-14-2014
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Primary Sjögren's syndrome (pSS) is a chronic autoimmune disorder that particularly compromises the function of exocrine glands. The pathogenetic mechanisms of this autoimmune exocrinopathy have not been fully elucidated. Since increasing evidence actually suggests that the epidermal growth factor receptor (EGFR) pathway has a major impact on the inflammatory/immune reactions of the epithelial cells, in the apparent effort of enhancing innate immune defense while opposing overactivation of pro-inflammatory functions, the focus of the work presented here is clarify whether the EGFR-extracellular-signal-regulated kinase (ERK) pathway plays a role in the pro-inflammatory responses mounted by pSS salivary gland epithelial cells (SGEC). Investigations revealed that the EGFR-mediated activation of the downstream effectors ERK1/2 in pSS SGEC appeared to require ADAM17-dependent release of the endogenous EGFR ligand amphiregulin and transactivation of the EGFR. Moreover, blockade of amphiregulin bioactivity using a neutralizing Ab significantly reduced EGFR transactivation and ERK1/2 phosphorylation. In addition, pSS SGEC treated with the specific ADAM17 inhibitor TAPI-1 and with the EGFR inhibitor AG1478 exhibited deactivated AREG/EGFR/ERK signaling pathway and reduced pro-inflammatory cytokines released.
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Hypoxia-resistant profile implies vulnerability of cancer stem cells to physiological agents, which suggests new therapeutic targets.
Cell Cycle
PUBLISHED: 11-05-2013
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We have previously shown that peculiar metabolic features of cell adaptation and survival in hypoxia imply growth restriction points that are typical of embryonic stem cells and disappear with differentiation. Here we provide evidence that such restrictions can be exploited as specific antiblastic targets by physiological factors such as pyruvate, tetrahydrofolate, and glutamine. These metabolites act as powerful cytotoxic agents on cancer stem cells (CSCs) when supplied at doses that perturb the biochemical network, sustaining the resumption of aerobic growth after the hypoxic dormant state. Experiments were performed in vivo and in vitro using CSCs obtained from various anaplastic tumors: human melanoma, leukemia, and rat hepatoma cells. Pretreatment of melanoma CSCs with pyruvate significantly reduces their self-renewal in vitro and tumorigenicity in vivo. The metabolic network underlying the cytotoxic effect of the physiological factors was thoroughly defined, principally using AH130 hepatoma, a tumor spontaneously reprogrammed to the embryonic stem stage. This network, based on a tight integration of aerobic glycolysis, cellular redox state, and folate metabolism, is centered on the cellular NADP/NADPH ratio that controls the redox pathway of folate utilization in purine synthesis. On the whole, this study indicates that pyruvate, FH 4, and glutamine display anticancer activity, because CSCs are committed to survive and maintain their stemness in hypoxia. When CSC need to differentiate and proliferate, they shift from anaerobic to aerobic status, and the few mitochondria available makes them susceptible to the injury of the above physiological factors. This vulnerability might be exploited for novel therapeutic treatments.
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Magnetic resonance imaging tracking of ferumoxytol-labeled human neural stem cells: studies leading to clinical use.
Stem Cells Transl Med
PUBLISHED: 09-06-2013
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Numerous stem cell-based therapies are currently under clinical investigation, including the use of neural stem cells (NSCs) as delivery vehicles to target therapeutic agents to invasive brain tumors. The ability to monitor the time course, migration, and distribution of stem cells following transplantation into patients would provide critical information for optimizing treatment regimens. No effective cell-tracking methodology has yet garnered clinical acceptance. A highly promising noninvasive method for monitoring NSCs and potentially other cell types in vivo involves preloading them with ultrasmall superparamagnetic iron oxide nanoparticles (USPIOs) to enable cell tracking using magnetic resonance imaging (MRI). We report here the preclinical studies that led to U.S. Food and Drug Administration approval for first-in-human investigational use of ferumoxytol to label NSCs prior to transplantation into brain tumor patients, followed by surveillance serial MRI. A combination of heparin, protamine sulfate, and ferumoxytol (HPF) was used to label the NSCs. HPF labeling did not affect cell viability, growth kinetics, or tumor tropism in vitro, and it enabled MRI visualization of NSC distribution within orthotopic glioma xenografts. MRI revealed dynamic in vivo NSC distribution at multiple time points following intracerebral or intravenous injection into glioma-bearing mice that correlated with histological analysis. Preclinical safety/toxicity studies of intracerebrally administered HPF-labeled NSCs in mice were also performed, and they showed no significant clinical or behavioral changes, no neuronal or systemic toxicities, and no abnormal accumulation of iron in the liver or spleen. These studies support the clinical use of ferumoxytol labeling of cells for post-transplant MRI visualization and tracking.
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Magnetic susceptibility of strongly interacting matter across the deconfinement transition.
Phys. Rev. Lett.
PUBLISHED: 07-31-2013
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We propose a method to determine the total magnetic susceptibility of strongly interacting matter by lattice QCD simulations and present numerical results for the theory with two light flavors, which suggest a weak magnetic activity in the confined phase and the emergence of strong paramagnetism in the deconfined, quark-gluon plasma phase.
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The cytosolic chaperone ?-crystallin B rescues folding and compartmentalization of misfolded multispan transmembrane proteins.
J. Cell. Sci.
PUBLISHED: 07-10-2013
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The ?-crystallin B chain (CRYAB or HspB5) is a cytosolic chaperone belonging to the small heat shock protein family, which is known to help in the folding of cytosolic proteins. Here we show that CRYAB binds the mutant form of at least two multispan transmembrane proteins (TMPs), exerting an anti-aggregation activity. It rescues the folding of mutant Frizzled4, which is responsible for a rare autosomal dominant form of familial exudative vitreoretinopathy (Fz4-FEVR), and the mutant ATP7B Cu transporter (ATP7B-H1069Q) associated with a common form of Wilsons disease. In the case of Fz4-FEVR, CRYAB prevents the formation of inter-chain disulfide bridges between the lumenal ectodomains of the aggregated mutant chains, which enables correct folding and promotes appropriate compartmentalization on the plasma membrane. ATP7B-H1069Q, with help from CRYAB, folds into the proper conformation, moves to the Golgi complex, and responds to copper overload in the same manner as wild-type ATP7B. These findings strongly suggest that CRYAB plays a pivotal role, previously undetected, in the folding of multispan TMPs and, from the cytosol, is able to orchestrate folding events that take place in the lumen of the ER. Our results contribute to the explanation of the complex scenario behind multispan TMP folding; additionally, they serve to expose interesting avenues for novel therapeutic approaches.
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Familial Exudative Vitreoretinopathy caused by a Homozygous Mutation in TSPAN12 in a Cystic Fibrosis Infant.
Ophthalmic Genet.
PUBLISHED: 07-08-2013
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Abstract Familial exudative vitreoretinopathy (FEVR) is a genetic disease affecting the vascularization of the peripheral retina. The clinical manifestations are very heterogeneous, ranging from mildly affected patients, who could present no visual defects, to severe conditions which can also cause complete blindness at birth or in the first decade. FEVR can be inherited in all the three genetic forms: dominant, recessive and X-linked. To date, four genes have been associated with the condition: TSPAN12. NDP. FDZ4 and LRP5. Interestingly, mutations in TSPAN12 have been considered causative of both a dominant and recessive inheritance and a FEVR phenotype sensitive to the number of TSPAN12 mutations has been supposed. Here we describe a case of a female infant affected by cystic fibrosis and by a severe form of exudative vitreoretinopathy. In particular, we have detected the homozygous missense mutation c.668 T?>?C in TSPAN12. Neither of the heterozygous parents has ocular manifestations of the disease, suggesting a classic recessive mendelian pattern of inheritance.
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hERG1 channels modulate integrin signaling to trigger angiogenesis and tumor progression in colorectal cancer.
Sci Rep
PUBLISHED: 06-07-2013
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Angiogenesis is a potential target for cancer therapy. We identified a novel signaling pathway that sustains angiogenesis and progression in colorectal cancer (CRC). This pathway is triggered by ?1 integrin-mediated adhesion and leads to VEGF-A secretion. The effect is modulated by the human ether-à-go-go related gene 1 (hERG1) K(+) channel. hERG1 recruits and activates PI3K and Akt. This in turn increases the Hypoxia Inducible Factor (HIF)-dependent transcription of VEGF-A and other tumour progression genes. This signaling pathway has novel features in that the integrin- and hERG1-dependent activation of HIF (i) is triggered in normoxia, especially after CRC cells have experienced a hypoxic stage, (ii) involves NF-kB and (iii) is counteracted by an active p53. Blocking hERG1 switches this pathway off also in vivo, by inhibiting cell growth, angiogenesis and metastatic spread. This suggests that non-cardiotoxic anti-hERG1 drugs might be a fruitful therapeutic strategy to prevent the failure of anti-VEGF therapy.
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Potassium channels: novel emerging biomarkers and targets for therapy in cancer.
Recent Pat Anticancer Drug Discov
PUBLISHED: 06-05-2013
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K+ Channels form the largest family among ion channels. Besides regulating many physiological functions, K+ channels, being aberrantly expressed in different types of tumors, affect several hallmarks of cancer. In cancer cells, K+ channel activity regulates cell proliferation, resistance to apoptotic cell death, tumor angiogenesis, invasiveness and metastatic spread. Moreover, being expressed in cells of the tumor microenvironment, K+ channels can also modulate the immune/inflammatory response which contributes to drive cancer establishment and progression. After almost 30 years of studies, some K+ channels are emerging as novel cancer biomarkers, to be employed to stratify patients for either prognostic or predictive purposes. Moreover, it is not remote the time in which it will be possible to target specific K+ channels in cancer for therapeutic purposes. One hindrance for applying a K+ channel-based therapy to cancer is the fact that K+ channel blockers can cause side effects, which often overlap with, but can mask, benefits. We here show some strategies to overcome harmful side effects caused by blocking K+ channels. Once taken into account these strategies, K+ channels may represent suitable and easily accessible cancer biomarkers and targets for therapy. The relevant patents related to K+ channels and cancer are discussed.
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A disorder-to-order structural transition in the COOH-tail of Fz4 determines misfolding of the L501fsX533-Fz4 mutant.
Sci Rep
PUBLISHED: 05-28-2013
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Frizzled 4 belongs to the superfamily of G protein coupled receptors. The unstructured cytosolic tail of the receptor is essential for its activity. The mutation L501fsX533 in the fz4 gene results in a new COOH-tail of the receptor and causes a form of Familial exudative vitreoretinopathy. Here we show that the mutated tail is structured. Two amphipathic helices, displaying affinity for membranes and resembling the structure of Influenza Hemagglutinin fusion peptide, constitute the new fold. This tail induces the aggregation of the receptor in the Endoplasmic Reticulum and it is sufficient to block the export to the Golgi of a chimeric VSVG protein containing the mutated tail. Affecting the tails structure, net charge or amphipathicity relocates the mutated Fz4 receptor to the Plasma Membrane. Such disorder-to-order structural transition was never described in GPCRs and opens a new scenario on the possible effect of mutations on unstructured regions of proteins.
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Neural stem cell-mediated enzyme/prodrug therapy for glioma: preclinical studies.
Sci Transl Med
PUBLISHED: 05-10-2013
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High-grade gliomas are extremely difficult to treat because they are invasive and therefore not curable by surgical resection; the toxicity of current chemo- and radiation therapies limits the doses that can be used. Neural stem cells (NSCs) have inherent tumor-tropic properties that enable their use as delivery vehicles to target enzyme/prodrug therapy selectively to tumors. We used a cytosine deaminase (CD)-expressing clonal human NSC line, HB1.F3.CD, to home to gliomas in mice and locally convert the prodrug 5-fluorocytosine to the active chemotherapeutic 5-fluorouracil. In vitro studies confirmed that the NSCs have normal karyotype, tumor tropism, and CD expression, and are genetically and functionally stable. In vivo biodistribution studies demonstrated NSC retention of tumor tropism, even in mice pretreated with radiation or dexamethasone to mimic clinically relevant adjuvant therapies. We evaluated safety and toxicity after intracerebral administration of the NSCs in non-tumor-bearing and orthotopic glioma-bearing immunocompetent and immunodeficient mice. We detected no difference in toxicity associated with conversion of 5-fluorocytosine to 5-fluorouracil, no NSCs outside the brain, and no histological evidence of pathology or tumorigenesis attributable to the NSCs. The average tumor volume in mice that received HB1.F3.CD NSCs and 5-fluorocytosine was about one-third that of the average volume in control mice. On the basis of these results, we conclude that combination therapy with HB1.F3.CD NSCs and 5-fluorocytosine is safe, nontoxic, and effective in mice. These data have led to approval of a first-in-human study of an allogeneic NSC-mediated enzyme/prodrug-targeted cancer therapy in patients with recurrent high-grade glioma.
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Growth, photosynthetic efficiency, and biochemical composition of Tetraselmis suecica F&M-M33 grown with LEDs of different colors.
Biotechnol. Bioeng.
PUBLISHED: 05-03-2013
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The effect of light quality on cell size and cell cycle, growth rate, productivity, photosynthetic efficiency and biomass composition of the marine prasinophyte Tetraselmis suecica F&M-M33 grown in 2-L flat panel photobioreactors illuminated with light emitting diodes (LEDs) of different colors was investigated. Biomass productivity and photosynthetic efficiency were comparable between white and red light, while under blue and green light productivity decreased to less than half and photosynthetic efficiency to about one third. Differences in cell size and number correlated with the cell cycle phase. Under red light cells were smaller and more motile. Chlorophyll content was strongly reduced with red and enhanced with blue light, while carotenoids and gross biomass composition were not affected by light quality. The eicosapentaenoic acid content increased under red light. Red light can substitute white light without affecting productivity of T. suecica F&M-M33, leading to smaller and more motile cells and increased eicosapentaenoic acid content. Red LEDs can thus be profitably used for the production of this microalga for aquaculture. Biotechnol. Bioeng. 2013;9999: 1-9. © 2013 Wiley Periodicals, Inc.
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Massive intra-abdominal imatinib-resistant gastrointestinal stromal tumor in a 21-year-old male.
Case Rep Med
PUBLISHED: 04-09-2013
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Gastrointestinal stromal tumors (GISTs) in adolescence are far less common than adult GISTs and have varied GIST genotypes that present diagnostic and therapeutic challenges. Here, we discuss a 21-year-old male with diagnosis of unresectable, imatinib-resistant GIST. At initial evaluation, a neoadjuvant treatment approach was recommended. As such, the patient received imatinib over the course of one year. Unfortunately, the GIST increased in size, and a subsequent attempt at surgical resection was aborted fearing infiltration of major vascular structures. The patient was then referred to our institution, at which time imatinib therapy was discontinued. Surgical intervention was again considered and the patient underwent successful resection of massive intra-abdominal GIST with total gastrectomy and Roux-en-Y esophagojejunostomy. Since pediatric GISTs are typically resistant to imatinib, we performed genotype analysis of the operative specimen that revealed KIT mutations associated with imatinib sensitivity and resistance. Given the sequencing data and operative findings, the patient was started postoperatively on sunitinib. This case illustrates the importance of understanding both adult and pediatric GISTs when implementing appropriate treatment regimens. Since the genotype of GISTs dictates phenotypic behavior, mutational analysis is an important component of care especially for adolescents whose disease may mirror the pediatric or adult population.
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[Pagets disease of bone: new therapeutic strategies].
Recenti Prog Med
PUBLISHED: 04-04-2013
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Pagets disease of bone is a chronic disorder of unknown etiology that can result in enlarged and misshapen bones. The excessive breakdown and formation of bone tissue cause affected bones to weaken, resulting in pain, misshapen bones, fractures, and arthritis in the joints. In most cases the diagnosis is achieved casually, as only 5% of patients develop burning pain at the level of affected bones. As regards therapy, the use of anti-reabsorbing drugs, such as bisphosphonates and calcitonin, appears reasonable. Given the disease pathogenesis, the administration of denosumab and tocilizumab may be a valuable alternative to inhibit RANK expression, and thus osteoclast formation, and interleukin-6 production.
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Susceptibility of the QCD vacuum to CP-odd electromagnetic background fields.
Phys. Rev. Lett.
PUBLISHED: 02-21-2013
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We investigate two flavor quantum chromodynamics (QCD) in the presence of CP-odd electromagnetic background fields and determine, by means of lattice QCD simulations, the induced effective ? term to first order in E[over ?] · B[over ?]. We employ a rooted staggered discretization and study lattice spacings down to 0.1 fm and Goldstone pion masses around 480 MeV. In order to deal with a positive measure, we consider purely imaginary electric fields and real magnetic fields, and then exploit the analytic continuation. Our results are relevant to a description of the effective pseudoscalar quantum electrodynamics-QCD interactions.
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GRO-?/CXCR2 system and ADAM17 correlated expression in Sjögrens syndrome.
Inflammation
PUBLISHED: 02-08-2013
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The chemokine GRO-? and its receptor CXCR2 are associated with the chronic inflammation in Sjögrens syndrome (SS). To better understand the molecular mechanisms by which the GRO-?/CXCR2 system is involved in the SS inflammatory condition, our studies were designed to clarify the role of ADAM17 activation in the modulation of the GRO-?/CXCR2 chemokine system in epithelial cells (SGEC) from SS salivary glands. The CXCR2 overexpression observed in SS SGEC was dramatically decreased by ADAM17 inhibitor TAPI-1. In addition, comparing the expression levels of ADAM17 in healthy SGEC in presence or not of GRO-? treatment, we observed that GRO-? dose-dependently influences ADAM17 activation, an effect that was inhibited by blocking the interaction of GRO-? with its CXCR2 receptor. Our data show for the first time that ADAM17 has an important role in GRO-?/CXCR2 system activity regulation, suggesting that regulating CXCR2/ADAM17 interaction could be an attractive therapeutic target in SS.
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Change of ? dependence in 4D SU(N) gauge theories across the deconfinement transition.
Phys. Rev. Lett.
PUBLISHED: 02-06-2013
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We study the dependence of 4D SU(N) gauge theories on the topological ? term at finite temperature T. We exploit the lattice formulation of the theory, presenting numerical results for the expansion of the free energy up to O(?6), for N=3 and N=6. Our analysis shows that the ? dependence drastically changes across the deconfinement transition: the low-T phase is characterized by a large-N scaling with ?/N as relevant variable, while in the high-T phase the scaling variable is just ? and the free energy is essentially determined by the instanton-gas approximation.
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Salivary gland expression level of I?B? regulatory protein in Sjögrens syndrome.
J. Mol. Histol.
PUBLISHED: 01-28-2013
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Diagnosis and therapeutic strategies in Sjögrens syndrome (SS) might greatly benefit of the present multidisciplinary approach to studying the molecular pathogenesis of the disease. A deregulated inflammatory response has been described in the SS. The research in the last years sheds light on the importance of the NF-?B pathway regulating the pro-inflammatory cytokine production and leukocyte recruitment. These are important contributors to the inflammatory response during the development of SS. In this study we examine the expression of the NF-?B inhibitory protein termed I?B? in salivary glands epithelial cells (SGEC) comparing it with SGEC from healthy controls, to test the hypothesis that an altered expression of I?B? occurs in SGEC from SS biopsies. Real-Time PCR, western blot and immunohistochemistry demonstrated that the expression level of I?B? was significantly lower in SS with respect to healthy controls leading to an increased NF-?B activity. Our results suggest that the analysis of I?B? expression at salivary gland epithelial cell level could be a potential new hallmark of SS progression and sustain a rationale to more deeply investigate the therapeutic potential of specific NF-?B inhibitors in SS.
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COUP-TFII in pancreatic adenocarcinoma: Clinical implication for patient survival and tumor progression.
Int. J. Cancer
PUBLISHED: 01-23-2013
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Despite the accumulating knowledge of alterations in pancreatic cancer molecular pathways, no substantial improvements in the clinical prognosis have been made and this malignancy continues to be a leading cause of cancer death in the Western World. The orphan nuclear receptor COUP-TFII is a regulator of a wide range of biological processes and it may exert a pro-oncogenic role in cancer cells; interestingly, indirect evidences suggest that the receptor could be involved in pancreatic cancer. The aim of this study was to evaluate the expression of COUP-TFII in human pancreatic tumors and to unveil its role in the regulation of pancreatic tumor growth. We evaluated COUP-TFII expression by immunohistochemistry on primary samples. We analyzed the effect of the nuclear receptor silencing in human pancreatic cancer cells by means of shRNA expressing cell lines. We finally confirmed the in vitro results by in vivo experiments on nude mice. COUP-TFII is expressed in 69% of tested primary samples and correlates with the N1 and M1 status and clinical stage; Kaplan-Meier and Cox regression analysis show that it may be an independent prognostic factor of worst outcome. In vitro silencing of COUP-TFII reduces the cell growth and invasiveness and it strongly inhibits angiogenesis, an effect mediated by the regulation of VEGF-C. In nude mice, COUP-TFII silencing reduces tumor growth by 40%. Our results suggest that COUP-TFII might be an important regulator of the behavior of pancreatic adenocarcinoma, thus representing a possible new target for pancreatic cancer therapy.
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Emerging avenues linking inflammation, angiogenesis and Sjögrens syndrome.
Cytokine
PUBLISHED: 01-20-2013
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Sjögrens syndrome (SS) is an autoimmune disease characterized by an inflammatory mononuclear infiltration and the destruction of epithelial cells of the lachrymal and salivary glands. The aetiology is unknown. The expression "autoimmune epithelitis" has been proposed as an alternative to SS, in view of the emerging central role of the epithelial cells in the disease pathogenesis. At the biomolecular level, the epithelial cells play an important role in triggering the autoimmune condition via antigen presentation, apoptosis, and chemokine and cytokines release. Inflammation and angiogenesis are frequently coupled in the pathological conditions associated to autoimmune diseases, and an angiogenic imbalance contributes to the pathogenesis of a number of inflammatory disorders. This work reviews the current knowledge of the molecular and cellular mechanisms underlying the pathogenesis of the inflammatory reactions that characterize SS. The literature and our data on the role of angiogenesis in the pathophysiology of the disease are discussed.
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?3-PUFAs exert anti-inflammatory activity in visceral adipocytes from colorectal cancer patients.
PLoS ONE
PUBLISHED: 01-01-2013
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The aim of this study was to correlate specific fatty acid profiles of visceral white adipose tissue (WAT) with inflammatory signatures potentially associated with colorectal cancer (CRC).
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Design and assessment of a low-cost, electromyographically controlled, prosthetic hand.
Med Devices (Auckl)
PUBLISHED: 01-01-2013
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The study reported here explored the design and realization of a low-cost, electromyographically controlled hand prosthesis for amputees living in developing countries. The developed prosthesis is composed of a light aluminum structure with opposing fingers connected to a DC motor that imparts only the movement of grasp. Problems associated with surface electromyographic signal acquisition and processing, motor control, and evaluation of grasp force were addressed, with the goal of minimizing cost and ensuring easy assembly. Simple analog front ends amplify and condition the electromyographic signals registered from two antagonist muscles by surface electrodes. Analog signals are sampled at 1 kHz and processed by a microcontroller that drives the motor with a supply voltage proportional to the muscular contraction, performing the opening and closing of the opposing fingers. Reliable measurements of the level of muscle contractions were obtained by specific digital processing: real-time operators implementing the root mean square value, mean absolute value, standard deviation, and mean absolute differential value were compared in terms of efficiency to estimate the EMG envelope, computational load, and time delay. The mean absolute value operator was adopted at a time window of 64 milliseconds. A suitable calibration procedure was proposed to overcome problems associated with the wide variation of electromyograph amplitude and background noise depending on the specific patients muscles selected. A pulse-width modulated signal drives the DC motor, allowing closing and opening of the prosthesis. The relationship between the motor-driver signal and the actual hand-grasp force developed by the prosthesis was measured using a hand-held grip dynamometer. The resulting force was proportional to current for moderate values of current and then saturated. The motor torque, and, in turn, the force elicited, can be measured by sensing the current absorbed by the motor. Therefore, the grasp force can be opportunely limited or controlled. The cost of the only electronic and mechanical components of the electromyographically controlled hand was about US$50; other costs, such as the cost of labor to assemble the prosthesis and the production of adapters for patients, were not estimated.
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A neuropharmacokinetic assessment of bafetinib, a second generation dual BCR-Abl/Lyn tyrosine kinase inhibitor, in patients with recurrent high-grade gliomas.
Eur. J. Cancer
PUBLISHED: 01-01-2013
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The primary objective of this study was to use intracerebral microdialysis (ICMD) to determine the neuropharmacokinetics of bafetinib, a dual BCR-Abl/Lyn tyrosine kinase inhibitor that may have activity against gliomas.
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Sjögrens syndrome autoantibodies provoke changes in gene expression profiles of inflammatory cytokines triggering a pathway involving TACE/NF-?B.
Lab. Invest.
PUBLISHED: 12-12-2011
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We explore the association of the inflammatory gene expression profile observed in the chronic inflammatory autoimmune disorder Sjögrens syndrome (SS) with changes in TNF-? converting enzyme (TACE), tumor necrosis factor (TNF)-? and nuclear factor (NF)-?B levels showing that pathways that include TNF-? signaling converge on NF-?B contributing to exacerbate the diseases. The treatment of human salivary gland epithelial cells (SGECs) with SS anti-Ro/SSA autoantibodies (Abs) result in a progressive increase in NF-?B-DNA binding, that includes a marked enhancement in NF-?B subunit p65 protein-DNA binding. A human cytokine multi-analyte array demonstrated that the NF-?B proinflammatory target genes, increased by anti-Ro/SSA Abs treatment, includes CXC chemokines (CXCL1, CXCL6 and CXCL9), CC chemokines (CCL2, CCL13 and CCL20), interleukins (IL-1?, IL-1?, IL-1F8, IL-6, IL-8, IL-9, IL-13, IL-17 and IL-22) and their receptors (IL-1RN, IL-10R?, IL-13R?, CCR1, CCR2, CCR3, CCR4 and CXCR1). Blockade of TACE through the use of the specific inhibitor TAPI-1 regulates proinflammatory cytokines production in SGEC treated with anti-Ro/SSA Abs inhibiting NF-?B nuclear translocation and activation. To further investigate the role of NF-?B on anti-Ro/SSA Abs-determined proinflammatory gene expression, we used the inhibitory protein I?B-? dominant negative super-repressor as inhibitor of NF-?B-DNA binding, demonstrating that transfection with dominant-negative I?B-? in anti-Ro/SSA-treated SGEC determined a marked reduction of proinflammatory cytokines gene expression. Although further studies are needed to clarify the mechanisms underlying SS, our results demonstrate that SS Abs exert their pathogenic effects via triggering the TACE/TNF-?/NF-?B axis.
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Ligand of Numb proteins LNX1p80 and LNX2 interact with the human glycoprotein CD8? and promote its ubiquitylation and endocytosis.
J. Cell. Sci.
PUBLISHED: 11-01-2011
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E3 ubiquitin ligases give specificity to the ubiquitylation process by selectively binding substrates. Recently, their function has emerged as a crucial modulator of T-cell tolerance and immunity. However, substrates, partners and mechanism of action for most E3 ligases remain largely unknown. In this study, we identified the human T-cell co-receptor CD8 ?-chain as binding partner of the ligand of Numb proteins X1 (LNX1p80 isoform) and X2 (LNX2). Both LNX mRNAs were found expressed in T cells purified from human blood, and both proteins interacted with CD8? in human HPB-ALL T cells. By using an in vitro assay and a heterologous expression system we showed that the interaction is mediated by the PDZ (PSD95-DlgA-ZO-1) domains of LNX proteins and the cytosolic C-terminal valine motif of CD8?. Moreover, CD8? redistributed LNX1 or LNX2 from the cytosol to the plasma membrane, whereas, remarkably, LNX1 or LNX2 promoted CD8? ubiquitylation, downregulation from the plasma membrane, transport to the lysosomes, and degradation. Our findings highlight the function of LNX proteins as E3 ligases and suggest a mechanism of regulation for CD8? localization at the plasma membrane by ubiquitylation and endocytosis.
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Polyphenols and human health: a prospectus.
Crit Rev Food Sci Nutr
PUBLISHED: 09-21-2011
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The lay press often heralds polyphenols as panacea for all sorts of diseases. The rationale is that their antioxidant activity would prevent free radical damage to macromolecules. However, basic and clinical science is showing that the reality is much more complex than this and that several issues, notably content in foodstuff, bioavailability, or in vivo antioxidant activity are yet to be resolved. We summarize the recent findings concerning the effects of polyphenols on human health, analyze the current limitations at pitfalls, and propose future directions for research.
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Cyanidin-3-O-?-glucoside and protocatechuic acid exert insulin-like effects by upregulating PPAR? activity in human omental adipocytes.
Diabetes
PUBLISHED: 07-25-2011
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Insulin resistance (IR) represents an independent risk factor for metabolic, cardiovascular, and neoplastic disorders. Preventing/attenuating IR is a major objective to be reached to preserve population health. Because many insulin-sensitizing drugs have shown unwanted side effects, active harmless compounds are sought after. Dietary anthocyanins have been demonstrated to ameliorate hyperglycemia and insulin sensitivity. This study aimed at investigating whether cyanidin-3-O-?-glucoside (C3G) and its metabolite protocatechuic acid (PCA) might have a role in glucose transport activation in human omental adipocytes and 3T3-L1 cells.
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Tethered bilayer lipid micromembranes for single-channel recording: the role of adsorbed and partially fused lipid vesicles.
Phys Chem Chem Phys
PUBLISHED: 06-24-2011
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A mercury-supported bilayer lipid micromembrane was prepared by anchoring a thiolipid monolayer to a mercury cap electrodeposited on a platinum microdisc about 20 ?m in diameter; a lipid monolayer was then self-assembled on top of the thiolipid monolayer either by vesicle fusion or by spilling a few drops of a lipid solution in chloroform on the cap and allowing the solvent to evaporate. Single-channel recording following incorporation of the alamethicin channel-forming peptide exhibits quite different features, depending on the procedure followed to form the distal lipid monolayer. The "spilling" procedure, which avoids the formation of adsorbed or partially fused vesicles, yields very sharp single-channel currents lasting only one or two milliseconds. These are ascribed to ionic flux into the hydrophilic spacer moiety of the thiolipid. Conversely, the vesicle-fusion procedure yields much longer single-channel openings analogous to those obtained with conventional bilayer lipid membranes, albeit smaller. This difference in behavior is explained by ascribing the latter single-channel currents to ionic flux into vesicles adsorbed and/or partially fused onto the tethered lipid bilayer, via capacitive coupling.
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Predominant role of obesity/insulin resistance in oxidative stress development.
Eur. J. Clin. Invest.
PUBLISHED: 06-16-2011
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? Hyperlipidaemia, hyperglycaemia and hyperinsulinaemia, hallmarks of the postprandial state, have been also associated with increased oxidative stress and lipoprotein oxidation contributing to vascular injury and atherosclerosis. However, the specific links among metabolic disorders, postprandial state, insulin resistance and oxidative stress are still to be clarified. This study aimed at investigating the individual role played by obesity, insulin resistance and type 2 diabetes in the occurrence of fasting and postprandial oxidative stress.
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A failure of TNFAIP3 negative regulation maintains sustained NF-?B activation in Sjögrens syndrome.
Histochem. Cell Biol.
PUBLISHED: 05-12-2011
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Sjögrens syndrome (SS) is characterized by the features of systemic autoimmunity and exocrine gland dysfunction and inflammation. Deregulated cytokine production is known to contribute to the etiology of SS but the underlying molecular mechanism is still remains to be unclear. TNF-?-induced protein 3 or TNFAIP3 is involved in the negative feedback regulation of nuclear factor-?B (NF-?B) signaling in response to specific pro-inflammatory stimuli in different cell types. To define the contribution of TNFAIP3 to SS, the levels of TNFAIP3 expression in human salivary gland epithelial cells (SGEC) derived from active primary SS patients were analyzed. Histological analysis was performed on paraffin-embedded human Sjögrens samples and healthy tissues. In separate experiments, immunofluorescence staining, western blot analysis and quantitative real-time PCR for TNFAIP3 was conducted in SGEC from SS and healthy subjects. Our findings clearly demonstrate changes in levels of the protein and gene expression between healthy controls and SS patients, depicting a very weak positivity for TNFAIP3 in SS samples. TNFAIP3 was found down-regulated in SGECs derived from SS patients in comparison with controls, and the cells with down-regulated TNFAIP3 expression exhibited enhanced NF-?B activities. In addition, to investigate the role of TNFAIP3 in the activation of NF-?B, we depleted TNFAIP3 expression by siRNA in healthy SGEC after treatment with or without TNF-?. Intriguingly, the silencing of TNFAIP3 by its siRNA in healthy SGEC increased NF-?B activation that could explain the deregulated cytokines production observed in SS.
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CCAAT/enhancer-binding protein-? participates in oxidized LDL-enhanced proliferation in 3T3-L1 cells.
Biochimie
PUBLISHED: 05-10-2011
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Increased circulating oxidized LDL (oxLDL) have been found in obese subjects. Obesity is characterized by an excess of fat mass resulting from an increase in adipocyte number and size. The generation of new adipocytes is a tightly controlled process where multiple factors acting in a signaling cascade follow a precise temporal expression pattern; oxLDL appear to have a role in the impairment of this process. The purpose of this study was to examine the effects of oxLDL on the mechanisms involved in the proliferative stage of the differentiation process in 3T3-L1 cells. After hormonal induction, 3T3-L1 cells undergo approximately two rounds of mitotic clonal expansion (MCE), a process required for adipogenesis. CCAAT/enhancer-binding protein ? (C/EBP?) is immediately expressed after induction, and plays a crucial role in MCE, but its expression must decrease to allow preadipocytes to mature into adipocytes. We found that, in the presence of stimuli to differentiate, oxLDL induced a higher proliferation rate in this cell line, associated with a sustained up-regulation of C/EBP?, which remained activated inside the nucleus for several days. RNAi-mediated knockdown of C/EBP? 24 h after oxLDL treatment counteracted the increase in proliferation rate. Both C/EBP? expression and proliferation processes appear to be influenced by cAMP/protein kinase A (PKA) and extracellular signal-regulated kinases1/2 (ERK1/2) pathways. OxLDL treatment led to increased levels of cAMP, and to a strong, prolonged phosphorylation of ERK1/2 and C/EBP?. The addition of cAMP and PKA inhibitors, SQ22536 and H-89, respectively, reduced proliferation only in oxLDL-treated cells, whereas the addition of ERK1/2 inhibitor U0126 blocked proliferation in both control and oxLDL-treated cells. C/EBP? nuclear expression and DNA-binding activity were reduced by U0126, under all tested conditions. These findings show that the altered expression pattern of C/EBP? is involved in the increase in the number of proliferating cells induced by oxLDL, in hormone-stimulated 3T3-L1 cells.
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[The treatment of the glucocorticoids-induced osteoporosis: recent knowledge].
Recenti Prog Med
PUBLISHED: 12-08-2010
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Osteoporosis is a skeletal disorder characterized by compromised bone strength predisposing to an increased risk of fractures that affects both cortical bone and trabecular bone. Osteoporosis may be either primary or secondary. Among the secondary forms, the glucocorticoid-induced osteoporosis is most common form that occurs, regardless of age, sex, and even with low doses of glucocorticoid.
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Chemotherapy resistance in acute lymphoblastic leukemia requires hERG1 channels and is overcome by hERG1 blockers.
Blood
PUBLISHED: 11-03-2010
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Bone marrow mesenchymal cells (MSCs) can protect leukemic cells from chemotherapy, thus increasing their survival rate. We studied the potential molecular mechanisms underlying this effect in acute lymphoblastic leukemia (ALL) cells. Coculture of ALL cells with MSCs induced on the lymphoblast plasma membrane the expression of a signaling complex formed by hERG1 (human ether-à-go-go-related gene 1) channels, the ?(1)-integrin subunit, and the chemokine receptor CXC chemokine receptor-4. The assembly of such a protein complex activated both the extracellular signal-related kinase 1/2 (ERK1/2) and the phosphoinositide 3-kinase (PI3K)/Akt prosurvival signaling pathways. At the same time, ALL cells became markedly resistant to chemotherapy-induced apoptosis. hERG1 channel function appeared to be important for both the initiation of prosurvival signals and the development of drug resistance, because specific channel blockers decreased the protective effect of MSCs. NOD/SCID mice engrafted with ALL cells and treated with channel blockers showed reduced leukemic infiltration and had higher survival rates. Moreover, hERG1 blockade enhanced the therapeutic effect produced by corticosteroids. Our findings provide a rationale for clinical testing of hERG1 blockers in the context of antileukemic therapy for patients with ALL.
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Blockade of TNF-? signaling suppresses the AREG-mediated IL-6 and IL-8 cytokines secretion induced by anti-Ro/SSA autoantibodies.
Lab. Invest.
PUBLISHED: 09-20-2010
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The aim of this study was to analyze the Furin-TNF-?-converting enzyme (TACE)-amphiregulin (AREG)-IL-6/IL-8 secretion pathway in non-neoplastic human salivary gland epithelial cells (SGECs) stimulated with anti-Ro/SSA autoantibodies (Abs). We examined whether anti-Ro/SSA Abs-mediated TACE activation is responsible for AREG activation. As recent studies have demonstrated that AREG could induce proinflammatory cytokines secretion in epithelial cells, we discuss how TACE-mediated AREG shedding, caused by anti-Ro/SSA Abs treatment, could have a critical role in TNF-?-induced IL-6 and IL-8 secretion by SGEC. Furthermore, the effects of TNF-? blockade on AREG expression and TNF-?-AREG-mediated IL-6 and IL-8 secretion were evaluated. We have discovered that the upregulation of AREG occurs through TNF-? produced after anti-Ro/SSA Abs uptake via Fc? receptors. Biological drug adalimumab and the gene silencing technique were used to study the AREG-IL-6/IL-8 secretion pathway, demonstrating that (i) adalimumab-mediated TNF-? blocking and TNF-? gene silencing provoke a significant decrease of proinflammatory cytokines production and AREG expression in anti-Ro/SSA Abs-treated SGEC; (ii) AREG gene silencing has a potent inhibitory effect on TNF-?-induced IL-6 and IL-8 secretion in SGEC treated with anti-Ro/SSA Abs; (iii) an inspection of the kinetics of cytokine production after exogeni TNF-? and AREG addition, and the use of cycloheximide in the presence of exogenous TNF-? as stimulant, clarified that TNF-? induces IL-6 and IL-8 secretion through AREG.Laboratory Investigation advance online publication, 20 September 2010; doi:10.1038/labinvest.2010.168.
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Expression of pro-inflammatory TACE-TNF-?-amphiregulin axis in Sjögrens syndrome salivary glands.
Histochem. Cell Biol.
PUBLISHED: 08-12-2010
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The tumor-necrosis-factor-converting-enzyme (TACE)-TNF-?-Amphiregulin (AREG) axis plays an important pathogenic role in inflammatory and autoimmune disorders. However, the pathological roles of these proteins in the chronic autoimmune disease Sjögrens syndrome (SS) remain to be elucidated. It is known that the TACE-AREG axis is clearly part of a larger cascade of signals that starts with the activation of Furin, responsible for maturation of TACE that, in turn, determines the production of active TNF-?, directly involved in the up-regulation of AREG expression. This study showed that Furin, TACE, TNF-?, and AREG proteins, detected in acinar and ductal cells of human salivary glands from SS patients, increased remarkably in comparison with biopsies of labial salivary glands from healthy controls. The changes in Furin, TACE, TNF- ?, and AREG proteins level detected in salivary glands biopsies of SS patients could be responsible for pro-inflammatory cytokines overexpression characterizing Sjögrens syndrome.
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The Kamil Crater in Egypt.
Science
PUBLISHED: 07-22-2010
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We report on the detection in southern Egypt of an impact crater 45 meters in diameter with a pristine rayed structure. Such pristine structures are typically observed on atmosphereless rocky or icy planetary bodies in the solar system. This feature and the association with an iron meteorite impactor and shock metamorphism provides a unique picture of small-scale hypervelocity impacts on Earths crust. Contrary to current geophysical models, ground data indicate that iron meteorites with masses of the order of tens of tons can penetrate the atmosphere without substantial fragmentation.
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PCNL in Italy.
Arch Ital Urol Androl
PUBLISHED: 07-03-2010
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The first Italian meeting on percutaneous nephrolithotomy (PCNL) was held in Milan in 1984. Since then PCNO has been practised in many centres but is diffusion has not been fast.
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Choosing the nephrostomy size after percutaneous nephrolithotomy.
World J Urol
PUBLISHED: 05-24-2010
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To evaluate the effect of nephrostomy tube size on perioperative outcomes of percutaneous nephrolithtotmy (PCNL).
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Pro-inflammatory role of Anti-Ro/SSA autoantibodies through the activation of Furin-TACE-amphiregulin axis.
J. Autoimmun.
PUBLISHED: 05-04-2010
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Prolonged inflammation can be detrimental because it may cause host toxicity and tissue damage. Indeed, excessive production of inflammatory cytokines is often associated with many autoimmune diseases. In this study we demonstrate that the anti-Ro/SSA autoantibodies (Abs) stimulate the production of pro-inflammatory cytokines IL-6 and IL-8 by human healthy salivary gland epithelial cells (healthy SGEC). The secretion of these cytokines is due to amphiregulin (AREG) that is overexpressed in healthy SGEC treated with anti-Ro/SSA Abs and in Sjögrens syndrome. We have discovered that the up-regulation of AREG occurs through TNF-alpha produced following anti-Ro/SSA Abs treatment. The gene silencing technique was used to study the AREG-TNF-alpha-IL-6/IL-8 secretion pathway, demonstrating that: (i) TNF-alpha gene silencing provokes a significant decrease of proinflammatory cytokines production and AREG expression in anti-Ro/SSA Abs-treated healthy SGEC; (ii) AREG gene silencing has a potent inhibitory effect on TNF-alpha-induced IL-6 and IL-8 secretion in healthy SGEC treated with anti-Ro/SSA Abs. These findings indicate that TACE-mediated AREG shedding plays a critical role in TNF-alpha-induced IL-6 and IL-8 secretion by the human healthy salivary gland epithelial cells, suggesting that this may be one of the possible intracellular mechanisms involved in the salivary glands inflammatory response in Sjögrens syndrome.
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Proposing a relationship between Mycobacterium avium subspecies paratuberculosis infection and Hashimotos thyroiditis.
Scand. J. Infect. Dis.
PUBLISHED: 05-01-2010
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Humans are widely exposed to Mycobacterium avium subspecies paratuberculosis (MAP), a proven multi-host chronic enteric pathogen that has recently been linked to autoimmune diabetes. In the present study we used a MAP species-specific polymerase chain reaction with the insertion element IS900-specific probe to detect MAP infection in members of the same family suffering from Hashimotos thyroiditis.
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Acycloguanosyl 5-thymidyltriphosphate, a thymidine analogue prodrug activated by telomerase, reduces pancreatic tumor growth in mice.
Gastroenterology
PUBLISHED: 02-16-2010
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Gemcitabine is the standard of care for metastatic and nonresectable pancreatic tumors. Phase II and III trials have not demonstrated efficacy of recently developed reagents, compared with gemcitabine alone; new chemotherapic agents are needed. Ninety percent of pancreatic tumors have telomerase activity, and expression correlates with tumor stage. We developed a thymidine analogue prodrug, acycloguanosyl 5-thymidyltriphosphate (ACV-TP-T), that is metabolized by telomerase and releases the active form of acyclovir. We investigated the antitumor efficacy of ACV-TP-T in vitro and in vivo.
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Immediate rehabilitation of the extremely atrophic mandible with fixed full-prosthesis supported by four implants.
Clin Implant Dent Relat Res
PUBLISHED: 02-11-2010
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To prospectively assess the outcome of immediate rehabilitation of extremely atrophic mandibles by a full-arch fixed bridge anchored to four implants.
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Advances in the understanding of the Fc gamma receptors-mediated autoantibodies uptake.
Clin. Exp. Med.
PUBLISHED: 02-04-2010
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Receptors for the Fc fragment of immunoglobulin G (Fc?Rs) are important molecules not only to mediate and control the effectors functions of IgG antibodies, but they also control the autoimmunity-tolerance balance in the periphery. In humans, three different types of Fc?Rs, belonging to the Ig gene superfamily, have been identified; Fc?RI (cluster of differentiation (CD64), Fc?RII (CD32) and Fc?RIII (CD16). A wide range of inflammatory and autoimmune diseases, such as vasculitis, glomerulonephritis, and autoimmune hemolytic anemia, seems to be mediated, in part, by Fc?Rs. Recent findings supposed that, under certain conditions, Fc?Rs are involved in the penetration of antibodies into cells and Fc?Rs constitute one of the main effector mechanisms through which autoantibodies exert their action. In this review, we concentrate on the role of human Fc?Rs in autoantibodies penetration and summarize the current knowledge on the structure, ligand binding capacity and their role in autoimmunity and pathogenic effect of autoantibodies.
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Bioavailability of the polyphenols: status and controversies.
Int J Mol Sci
PUBLISHED: 01-27-2010
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The current interest in polyphenols has been driven primarily by epidemiological studies. However, to establish conclusive evidence for the effectiveness of dietary polyphenols in disease prevention, it is useful to better define the bioavailability of the polyphenols, so that their biological activity can be evaluated. The bioavailability appears to differ greatly among the various phenolic compounds, and the most abundant ones in our diet are not necessarily those that have the best bioavailability profile. In the present review, we focus on the factors influencing the bioavailability of the polyphenols. Moreover, a critical overview on the difficulties and the controversies of the studies on the bioavailability is discussed.
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Defining the role of cholecystokinin in the lipid-induced human brain activation matrix.
Gastroenterology
PUBLISHED: 01-18-2010
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In human beings, as in most vertebrates, the release of the intestinal peptide cholecystokinin (CCK) by ingested food plays a major role both in digestion and the regulation of further food intake, but the changes in brain function and their underlying activation mechanisms remain unknown. Our aim was to explore, using a novel physiologic magnetic resonance imaging approach, the temporospatial brain activation matrix, in response to ingestion of a lipid meal and, by use of a CCK-1 receptor antagonist, to define the role of CCK in this activation.
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Recognition and killing of brain tumor stem-like initiating cells by CD8+ cytolytic T cells.
Cancer Res.
PUBLISHED: 11-10-2009
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Solid tumors contain a subset of stem-like cells that are resistant to the cytotoxic effects of chemotherapy/radiotherapy, but their susceptibility to cytolytic T lymphocyte (CTL) effector mechanisms has not been well characterized. Using a panel of early-passage human brain tumor stem/initiating cell (BTSC) lines derived from high-grade gliomas, we show that BTSCs are subject to immunologic recognition and elimination by CD8(+) CTLs. Compared with serum-differentiated CD133(low) tumor cells and established glioma cell lines, BTSCs are equivalent with respect to expression levels of HLA class I and ICAM-1, similar in their ability to trigger degranulation and cytokine synthesis by antigen-specific CTLs, and equally susceptible to perforin-dependent CTL-mediated cytolysis. BTSCs are also competent in the processing and presentation of antigens as evidenced by the killing of these cells by CTL when antigen is endogenously expressed. Moreover, we show that CTLs can eliminate all BTSCs with tumor-initiating activity in an antigen-specific manner in vivo. Current models predict that curative therapies for many cancers will require the elimination of the stem/initiating population, and these studies lay the foundation for developing immunotherapeutic approaches to eradicate this tumor population.
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[New and old strategies against osteoporosis].
Recenti Prog Med
PUBLISHED: 11-05-2009
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Osteoporosis is a skeletal disorder characterized by compromised bone strength predisposing to an increased risk of fracture that affects both cortical bone and trabecular bone. Osteoporosis may be either primary or secondary. Primary osteoporosis can be distinguished into two types: diffuse or local. The treatment can be managed through drugs, exercise and lifestyle. The aim of this paper is to highlight new and old therapeutic strategies.
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Induction of TNF-alpha-converting enzyme-ectodomain shedding by pathogenic autoantibodies.
Int. Immunol.
PUBLISHED: 10-23-2009
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The release of the soluble form of tumor necrosis factor (TNF)-alpha from the plasma membrane occurs through the activation of the secretase tumor necrosis factor-alpha-converting enzyme (TACE). The current study was designed to examine whether the anti-Ro/SSA autoantibodies (Abs) are capable to regulate TACE expression in non-neoplastic human salivary gland epithelial cells (SGEC) cultures. We investigated the effect of anti-Ro/SSA Abs on the localization and abundance of cell-surface TACE and on TACE pro-domain-shedding and activation. In addition, the potential physiological consequences of TNF-alpha blockage by the biological agent Adalimumab on post-translational regulation of TACE are discussed. Anti-Ro/SSA Abs were purified from IgG fractions of patients with primary Sjögrens syndrome, using Sepharose 4B-Ro/SSA affinity columns. Flow cytometry, reverse transcription-PCR, western blot and immunohistochemistry were used to study TACE expression on SGEC and TACE regulation by Abs. Our study demonstrated a dose-dependent increase of TACE messenger RNA (mRNA) expression in anti-Ro/SSA Abs-treated SGEC, followed by internalization, pro-domain shedding and activation of TACE protein, suggesting that increased TACE activity is necessary for the release of TNF-alpha observed in anti-Ro/SSA Abs-stimulated SGEC. Adalimumab treatment brought TACE mRNA and surface TACE expression to levels than those observed in untreated SGEC. These data suggest that the effect of anti-Ro/SSA Abs on TACE expression and intracellular distribution is exerted by TNF-alpha production.
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Role of fat hydrolysis in regulating glucagon-like Peptide-1 secretion.
J. Clin. Endocrinol. Metab.
PUBLISHED: 10-16-2009
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Context: Glucagon-like peptide-1 (GLP-1) is produced by specialized cells in the gut and secreted in response to carbohydrates and lipids. The mechanisms regulating fat-stimulated GLP-1 release have, however, not been clarified in detail. Aim: We aimed to investigate the effect of intraduodenal (ID) fat hydrolysis on GLP-1 release and test whether the signal is mediated through cholecystokinin (CCK)-1 receptors. Design and Setting: Thirty-four healthy, male ambulatory volunteers were studied in three consecutive, randomized, double blind, crossover studies. Intervention: There were three interventions: 1) 12 subjects received an ID fat infusion with or without orlistat, an irreversible inhibitor of gastrointestinal lipases, in comparison with vehicle; 2) 12 subjects received ID sodium oleate (C18:1), ID sodium caprylate (C8:0), or ID vehicle; and 3) 10 subjects received ID sodium oleate with and without the CCK-1 receptor antagonist dexloxiglumide or ID vehicle plus iv saline (placebo). The effect of these treatments on GLP-1 concentrations and CCK release was quantified. Results: The following results were reached: 1) ID fat induced significant increase in GLP-1 concentrations (P < 0.004), and inhibition of fat hydrolysis by orlistat abolished this effect; 2) sodium oleate significantly stimulated GLP-1 release (P < 0.008), whereas sodium caprylate was ineffective compared with controls; and 3) dexloxiglumide administration abolished the effect of sodium oleate on GLP-1. ID fat or sodium oleate significantly stimulated plasma CCK (P < 0.006 and P < 0.004) compared with saline, whereas sodium caprylate did not. Conclusion: Generation of long-chain fatty acids through hydrolysis of fat is a critical step for fat-induced stimulation of GLP-1 in humans; the signal is mediated via CCK release and CCK-1 receptors.
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TNF blocker drugs modulate human TNF-?-converting enzyme pro-domain shedding induced by autoantibodies.
Immunobiology
PUBLISHED: 10-06-2009
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Novel biologic therapies targeted against specific components of the immune system, including blockade of TNF-? have revolutionized therapeutic approaches to inflammatory conditions and systemic inhibitors of TNF-? have been approved for the treatment of a wide variety of autoimmune diseases. No studies aimed to elucidate the effects of anti-TNF-? blockers on tumour necrosis factor-? convertase (TACE) expression and activation have yet been published. TACE is the principal protease involved in the activation of pro-TNF-? and is a target for anti-TNF-? therapy. Here we focused on regulation of TACE expression in human salivary gland epithelial cells (SGEC) treated by anti-Ro/SSA autoantibodies (autoAbs), characterizing primary Sjögrens syndrome and on the effect of anti-Ro/SSA autoAbs on TACE pro-domain shedding and activation. To test the hypothesis that anti-TNF-? blocker drugs affect TACE expression, we used Adalimumab and Etanercept to block TNF-? and evaluate the effects of these biological agents on post-translational regulation of TACE. Anti-Ro/SSA autoAbs determines TACE pro-domain shedding suggesting that TACE activity is necessary for the release of TNF-? observed in anti-Ro/SSA autoAbs-stimulated cells. The comparative efficacy analysis of the regulation of TACE activity by Adalimumab and Etanercept revealed that Adalimumab appear to be significantly more efficacious than Etanercept in preventing TACE activation caused by anti-Ro/SSA autoAbs. It is intriguing to consider that regulation of TACE may participate in the pathogenic role of autoantibodies and the modulation of TACE expression by TNF-? antagonists might contribute to the beneficial effect of these drugs in inflammatory and autoimmune diseases.
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Regulation of mRNA caspase-8 levels by anti-nuclear autoantibodies.
Clin. Exp. Med.
PUBLISHED: 09-29-2009
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Apoptosis of the acinar and ductal epithelial cells of the salivary glands has been proposed as a mechanism possibly responsible for the impairment of the secretory function in Sjögrens syndrome, an organ-specific autoimmune disorder characterized by destruction of these glandular structures. The presence of serum autoantibodies (Abs) directed against the ribonucleoproteic antigens Ro and La is one of the classification criteria used to identify Sjögren patients, and there is increasing evidence of the direct involvement of Abs in tissue pathogenesis. Our recent report demonstrated that anti-Ro and anti-La Abs are able to trigger the extrinsic pathway of apoptosis in the human salivary gland cells. To better understand how the anti-Ro and anti-La Abs exert their apoptotic effect, human caspase-8 gene expression was examined in primary human salivary gland epithelial cell (SGEC) cultures established from biopsies of labial minor salivary glands. To measure mRNA expression changes of initiating caspase-8, the real-time polymerase chain reaction was employed. This was combined with western blot to study the activation of caspase-8 detecting the cleaved form of caspase-8 and the cleaved poly (ADP-ribose) polymerase, downstream consequences of caspases activation. Data obtained suggest that the anti-Ro and anti-La Abs determine a transcriptional up-regulation and activation of caspase-8. Study of the mRNA in SGEC experimental model may provide insight into the signal transduction pathway stimulated by anti-nuclear autoantibodies.
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Tumor necrosis factor inhibitors block apoptosis of human epithelial cells of the salivary glands.
Ann. N. Y. Acad. Sci.
PUBLISHED: 09-03-2009
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Inhibition of tumor necrosis factor-alpha (TNF-alpha) in organ-specific autoimmune disease is proving efficacious for a large number of patients. A wide array of biological agents has been designed to inhibit TNF-alpha, such as adalimumab (fully humanized) and etanercept (soluble TNF-alpha receptor fusion constructs p75 subunit). Recently, we suggested that anti-Ro and anti-La autoantibodies (Abs) isolated from patients with Sjögrens syndrome, an autoimmune rheumatic disease, are able to trigger cell death through extrinsic apoptotic mechanisms in human salivary gland epithelial cells (SGEC). We analyzed if primary human SGEC cultures, established from biopsy of labial minor salivary glands, are able to produce TNF-alpha, an inductor of the extrinsic apoptotic pathway, when treated with anti-Ro autoantibodies. A comparative study was performed to test the efficacy of adalimumab and etanercept to block TNF-alpha-mediated apoptosis. ELISA assay and RT-PCR were employed to visualize TNF-alpha production, and apoptosis was evaluated by DNA ladder and flow cytometry. We found that cell treatment with anti-Ro autoantibodies determines TNF-alpha production that reaches a maximum at 16 h and is decreased (P < 0.05) at 24 and 48 h. Adalimumab seems to be more efficacious than etanercept in blocking TNF-alpha-mediated apoptosis. The YOPRO-1 (+) and propidium iodide (-) method revealed 60% of apoptotic cells after 24 h of incubation with anti-Ro compared with 15% of apoptotic cells treated with anti-Ro plus adalimumab and 25% of apoptotic cells treated with anti-Ro plus etanercept. The antiapoptotic effect of adalimumab and etanercept was supported by inhibition of DNA laddering induced by anti-Ro Abs. These data validate the therapeutic efficacy of the anti-TNF reagents in the treatment of autoimmune disorders.
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