Resveratrol interrupts the pro-invasive communication between Cancer Associated Fibroblasts and Cholangiocarcinoma cells.
Publication Date: 23/05/2018, on Cancer letters
by Thongchot S, Ferraresi A, Vidoni C, Loilome W, Yongvanit P, Namwat N, Isidoro C
Cholangiocarcinoma (CCA), the cancer arising from the epithelial cells of bile ducts, is a prototype of inflammatory-driven cancer. Cytokines released by cancer associated fibroblasts (CAFs) play a pivotal role in CCA progression, driving the epigenetic Epithelial-to-Mesenchymal transition and the growth and metastasization of CCA cells. Consistently, the conditioned medium from CCA-derived CAFs further stimulated the secretion of IL-6, and to a lesser extent of IL-8, by CCA cells. CCA has a poor prognosis, because of late diagnosis and of high resistance to radio- and chemo-therapy of CCA cells. Targeting the CAFs and their secretion could be an alternative option. We found that while IL-6 indeed promoted the cell migration of invasive CCA cells, the nutraceutical Resveratrol strongly counteracted this effect both in CCA cells and in immortalized cholangiocytes. More importantly, here we show that Resveratrol has the potential to abrogate the secretion of IL-6 by CAFs. While the conditioned medium from CAFs strongly induced IL-6 mediated motility of CCA cells, the conditioned medium from CAFs pre-treated with Resveratrol completely halted cancer cell motility and reverted the N-to E-cadherin switch in migrating cells. This effect was associated with stimulation of autophagy in the cancer cells. This is the first demonstration that CAFs secretory products directly affect the regulation of autophagy and consequently the behavior of CCA cells, and that a nutraceutical may revert the malignant phenotype of cancer cells by acting on CAFs metabolism and secretion.
High-level production of single chain monellin mutants with enhanced sweetness and stability in tobacco chloroplasts.
Publication Date: 18/05/2018, on Planta
by Castiglia D, Leone S, Tamburino R, Sannino L, Fonderico J, Melchiorre C, Carpentieri A, Grillo S, Picone D, Scotti N
Plastid-based MNEI protein mutants retain the structure, stability and sweetness of their bacterial counterparts, confirming the attractiveness of the plastid transformation technology for high-yield production of recombinant proteins. The prevalence of obesity and diabetes has dramatically increased the industrial demand for the development and use of alternatives to sugar and traditional sweeteners. Sweet proteins, such as MNEI, a single chain derivative of monellin, are the most promising candidates for industrial applications. In this work, we describe the use of tobacco chloroplasts as a stable plant expression platform to produce three MNEI protein mutants with improved taste profile and stability. All plant-based proteins were correctly expressed in tobacco chloroplasts, purified and subjected to in-depth chemical and sensory analyses. Recombinant MNEI mutants showed a protein yield ranging from 5% to more than 50% of total soluble proteins, which, to date, represents the highest accumulation level of MNEI mutants in plants. Comparative analyses demonstrated the high similarity, in terms of structure, stability and function, of the proteins produced in plant chloroplasts and bacteria. The high yield and the extreme sweetness perceived for the plant-derived proteins prove that plastid transformation technology is a safe, stable and cost-effective production platform for low-calorie sweeteners, with an estimated production of up to 25-30 mg of pure protein/plant.
Psychometric properties of the Italian version of the Cognitive Reserve Scale (I-CRS).
Publication Date: 04/05/2018, on Neurological sciences : official journal of the Italian Neurological Society and of the Italian Society of Clinical Neurophysiology
by Altieri M, Siciliano M, Pappacena S, Roldán-Tapia MD, Trojano L, Santangelo G
The original definition of cognitive reserve (CR) refers to the individual differences in cognitive performance after a brain damage or pathology. Several proxies were proposed to evaluate CR (education, occupational attainment, premorbid IQ, leisure activities). Recently, some scales were developed to measure CR taking into account several cognitively stimulating activities. The aim of this study is to adapt the Cognitive Reserve Scale (I-CRS) for the Italian population and to explore its psychometric properties. I-CRS was administered to 547 healthy participants, ranging from 18 to 89 years old, along with neuropsychological and behavioral scales to evaluate cognitive functioning, depressive symptoms, and apathy. Cronbach's α, corrected item-total correlations, and the inter-item correlation matrix were calculated to evaluate the psychometric properties of the scale. Linear regression analysis was performed to build a correction grid of the I-CRS according to demographic variables. Correlational analyses were performed to explore the relationships between I-CRS and neuropsychological and behavioral scales. We found that age, sex, and education influenced the I-CRS score. Young adults and adults obtained higher I-CRS scores than elderly adults; women and participants with high educational attainment scored higher on I-CRS than men and participants with low education. I-CRS score correlated poorly with cognitive and depression scale scores, but moderately with apathy scale scores. I-CRS showed good psychometric properties and seemed to be a useful tool to assess CR in every adult life stage. Moreover, our findings suggest that apathy rather than depressive symptoms may interfere with the building of CR across the lifespan.
Targeted therapy of human glioblastoma via delivery of a toxin through a peptide directed to cell surface nucleolin.
Publication Date: 01/05/2018, on Journal of cellular physiology
by Dhez AC, Benedetti E, Antonosante A, Panella G, Ranieri B, Florio TM, Cristiano L, Angelucci F, Giansanti F, Di Leandro L, d'Angelo M, Melone M, De Cola A, Federici L, Galzio R, Cascone I, Raineri F, Cimini A, Courty J, Giordano A, Ippoliti R
Targeted anticancer therapies demand discovery of new cellular targets to be exploited for the delivery of toxic molecules and drugs. In this perspective, in the last few years, nucleolin has been identified as an interesting surface marker to be used for the therapy of glioblastoma. In this study, we investigated whether a synthetic antagonist of cell-surface nucleolin known as N6L, previously reported to decrease both tumor growth and tumor angiogenesis in several cancer cell lines, including glioblastoma cells, as well as endothelial cells proliferation, could be exploited to deliver a protein toxin (saporin) to glioblastoma cells. The pseudopeptide N6L cross-linked to saporin-S6 induced internalization of the toxin inside glioblastoma cancer cells. Our results in vitro demonstrated the effectiveness of this conjugate in inducing cell death, with an ID four orders of magnitude lower than that observed for free N6L. Furthermore, the preliminary in vivo study demonstrated efficiency in reducing the tumor mass in an orthotopic mouse model of glioblastoma.
Multiple Reaction Monitoring Tandem Mass Spectrometry Approach for the Identification of Biological Fluids at Crime Scene Investigations.
Publication Date: 01/05/2018, on Analytical chemistry
by Illiano A, Arpino V, Pinto G, Berti A, Verdoliva V, Peluso G, Pucci P, Amoresano A
Knowledge of the nature of biofluids at a crime scene is just as important as DNA test to link the nature of the biofluid, the criminal act, and the dynamics of the crime. Identification of methods currently used for each biological fluid (blood, semen, saliva, urine) suffer from several limitations including instability of assayed biomolecules, and low selectivity and specificity; as an example of the latter issue, it is not possible to discriminate between alpha-amylase 1 (present in saliva) and alpha-amylase 2 (present in semen and vaginal secretion. In this context, the aim of the work has been to provide a predictive protein signature characteristic of each biofluid by the recognition of specific peptides unique for each protein in a single analysis. A panel of four protein biomarkers for blood, four for saliva, five for semen, and two for urine has been monitored has been monitored by using a single multiple reaction monitoring (MRM)-based method targeting concomitantly 46 different peptides. Then, The optimized method allows four biological matrices to be identified when present on their own or in 50:50 mixture with another biofluid. Finally, a valid strategy combining both DNA analysis and liquid chromatographic-tandem mass spectrometric multiple reaction monitoring (LC-MS-MRM) identification of biofluids on the same sample has been demonstrated to be particularly effective in forensic investigation of real trace evidence collected at a crime scene.
Aquaporin 4 expression in human skeletal muscle fiber types.
Publication Date: 01/05/2018, on Muscle & nerve
by Vizzaccaro E, Terracciano C, Rastelli E, Massa R
Aquaporins (AQPs) are a family of transmembrane proteins involved in the maintenance of osmotic gradients. AQP4 is abundant in skeletal muscle, where it seems to be associated with glycolytic metabolism. We investigated the pattern of expression of AQP4 in normal human myofibers relative to the main forms of myosin heavy chain (MHC).
Caged noble metals: Encapsulation of a cytotoxic platinum(II)-gold(I) compound within the ferritin nanocage.
Publication Date: 30/04/2018, on International journal of biological macromolecules
by Ferraro G, Petruk G, Maiore L, Pane F, Amoresano A, Cinellu MA, Monti DM, Merlino A
The encapsulation of Pt and Au-based anticancer agents within a protein cage is a promising way to enhance the selectivity of these potential drugs. Here a cytotoxic organometallic compound containing platinum(II) and gold(I) has been encapsulated within a ferritin nanocage (AFt). Inductively plasma coupled mass spectrometry data, collected to evaluate the amount of Pt and Au within the cage, indicate disruption of the starting heterobimetallic complex upon encapsulation within the nanocage. The drug-loaded protein (Pt(II)/Au(I)-AFt) has been characterized by UV-Vis spectroscopy, circular dichroism and X-ray diffraction analysis. Data indicate that the protein maintains its fold upon encapsulation of the metallodrug and that Au(I) and Pt(II)-containing fragments are encapsulated within the AFt cage, with Au(I) ion that binds the side chain of Cys126 and Pt(II) in the bulk, respectively. The in vitro cytotoxicity of Pt(II)Au(I)-AFt, as well as that of the free heterobimetallic complex, has been comparatively evaluated on human cervix and breast cancer cells and against cardiomyoblasts and keratinocytes non-tumorigenic cells. Our data demonstrate that it is possible to obtain a protein nanocarrier containing both Pt and Au atoms starting from a bimetallic compound, opening the way for the design and development of new potential drugs based on protein nanocarriers.
New insights on the functional role of URG7 in the cellular response to ER stress.
Publication Date: 28/04/2018, on Biology of the cell
by Armentano MF, Caterino M, Miglionico R, Ostuni A, Pace MC, Cozzolino F, Monti M, Milella L, Carmosino M, Pucci P, Bisaccia F
Up-regulated Gene clone 7 (URG7) is an ER resident protein, whose expression is upregulated in the presence of hepatitis B virus X antigen (HBxAg) during HBV infection. In virus-infected hepatocytes, URG7 shows an anti-apoptotic activity due to the PI3K/AKT signaling activation, does not seem to have tumorigenic properties, but it appears to promote the development and progression of fibrosis. However, the molecular mechanisms underlying URG7 activity remain largely unknown.
Microglial Phagocytosis and Its Regulation: A Therapeutic Target in Parkinson's Disease?
Publication Date: 27/04/2018, on Frontiers in molecular neuroscience
by Janda E, Boi L, Carta AR
The role of phagocytosis in the neuroprotective function of microglia has been appreciated for a long time, but only more recently a dysregulation of this process has been recognized in Parkinson's disease (PD). Indeed, microglia play several critical roles in central nervous system (CNS), such as clearance of dying neurons and pathogens as well as immunomodulation, and to fulfill these complex tasks they engage distinct phenotypes. Regulation of phenotypic plasticity and phagocytosis in microglia can be impaired by defects in molecular machinery regulating critical homeostatic mechanisms, including autophagy. Here, we briefly summarize current knowledge on molecular mechanisms of microglia phagocytosis, and the neuro-pathological role of microglia in PD. Then we focus more in detail on the possible functional role of microglial phagocytosis in the pathogenesis and progression of PD. Evidence in support of either a beneficial or deleterious role of phagocytosis in dopaminergic degeneration is reported. Altered expression of target-recognizing receptors and lysosomal receptor CD68, as well as the emerging determinant role of α-synuclein (α-SYN) in phagocytic function is discussed. We finally discuss the rationale to consider phagocytic processes as a therapeutic target to prevent or slow down dopaminergic degeneration.
Analysis of proautophagic activities of Citrus flavonoids in liver cells reveals the superiority of a natural polyphenol mixture over pure flavones.
Publication Date: 26/04/2018, on The Journal of nutritional biochemistry
by Lascala A, Martino C, Parafati M, Salerno R, Oliverio M, Pellegrino D, Mollace V, Janda E
Autophagy dysfunction has been implicated in the pathogenesis of nonalcoholic fatty liver disease (NAFLD). Natural compounds present in bergamot polyphenol fraction (BPF) prevent NAFLD and induce autophagy in rat livers. Here, we employed HepG2 cells expressing DsRed-LC3-GFP, a highly sensitive model system to screen for proautophagic compounds present in BPF. BPF induced autophagy in a time- and dose-dependent fashion and the effect was amplified in cells loaded with palmitic acid. Autophagy was mediated by the hydrophobic fraction of acid-hydrolyzed BPF (A-BPF), containing six flavanone and flavone aglycones as identified by liquid chromatography-high-resolution mass spectrometry. Among them, naringenin, hesperitin, eriodictyol and diosmetin were weak inducers of autophagy. Apigenin showed the strongest and dose-dependent proautophagic activity at early time points (6 h). Luteolin induced a biphasic autophagic response, strong at low doses and inhibitory at higher doses. Both flavones were toxic in HepG2 cells and in differentiated human liver progenitors HepaRG upon longer treatments (24 h). In contrast, BPF and A-BPF did not show any toxicity, but induced a persistent increase in autophagic flux. A mixture of six synthetic aglycones mimicking A-BPF was sufficient to induce a similar autophagic response, but it was mildly cytotoxic. Thus, while six main BPF flavonoids fully account for its proautophagic activity, their combined effect is not sufficient to abrogate cytotoxicity of individual compounds. This suggests that a natural polyphenol phytocomplex, such as BPF, is a safer and more effective strategy for the treatment of NAFLD than the use of pure flavonoids.
DNA in Squid Synaptosomes.
Publication Date: 19/04/2018, on Molecular neurobiology
by Cefaliello C, Prisco M, Crispino M, Giuditta A
The synthesis of brain metabolic DNA (BMD) is modulated by learning and circadian oscillations and is not involved in cell division or DNA repair. Data from rats have highlighted its prevalent association with the mitochondrial fraction and its lack of identity with mtDNA. These features suggested that BMD could be localized in synaptosomes that are the major contaminants of brain mitochondrial fractions. The hypothesis has been examined by immunochemical analyses of the large synaptosomes of squid optic lobes that are readily prepared and identified. Optic lobe slices were incubated with 5-bromo-2-deoxyuridine (BrdU) and the isolated synaptosomal fraction was exposed to the green fluorescent anti-BrdU antibody. This procedure revealed that newly synthesized BrdU-labeled BMD is present in a significant percent of the large synaptosomes derived from the nerve terminals of retinal photoreceptor neurons and in synaptosomal bodies of smaller size. Synaptosomal BMD synthesis was strongly inhibited by actinomycin D. In addition, treatment of the synaptosomal fraction with Hoechst 33258, a blue fluorescent dye specific for dsDNA, indicated that native DNA was present in all synaptosomes. The possible role of synaptic BMD is briefly discussed.
Cardiotoxic effects of the novel approved anti-ErbB2 agents and reverse cardioprotective effects of ranolazine.
Publication Date: 19/04/2018, on OncoTargets and therapy
by De Lorenzo C, Paciello R, Riccio G, Rea D, Barbieri A, Coppola C, Maurea N
Pertuzumab, a novel anti-epidermal growth factor receptor 2 humanized monoclonal antibody, and trastuzumab-emtansine (TDM1), a novel antibody-drug conjugate made up of trastuzumab covalently linked to the highly potent microtubule inhibitory agent DM1, have been recently approved by the US Food and Drug Administration for increasing the efficiency and safety of breast cancer therapy with trastuzumab. We investigated for the first time the potential cardiotoxic effects of pertuzumab and TDM1, which are not yet fully elucidated, and we tested whether ranolazine could blunt their cardiotoxicity.
The Effect of Low-Dose Ionizing Radiation on Stem Cell Biology: A Contribution to Radiation Risk.
Publication Date: 17/04/2018, on Stem cells (Dayton, Ohio)
by Squillaro T, Galano G, De Rosa R, Peluso G, Galderisi U
Exposure to high levels of ionizing radiation (IR) (>0.5Gy), negatively affect health. but, less is known about the effects of low dose IR (LDIR) but recent, evidence suggests that it may have profound effects on cellular functions. We are commonly exposed to LDIR over natural background levels from numerous sources: people may be exposed to low dose IR for medical diagnosis and therapy, air travel, illegal IR waste dumpsites or by occupational exposures in the nuclear and medical sectors. Stem cells reside for long periods of time in our bodies, and this increases the possibility that they may be accumulate genotoxic damage derived from extrinsic LDIR or intrinsic sources (such as DNA replication). In this review we provide an overview of LDIR effects on biology of stem cell compartments. The principal findings and issues reported in the scientific literature are discussed in order to present the current understanding of the LDIR exposure risk, and assess whether it may impact human health. We first consider the general biological consequences of LDIR exposure. Following this, we discuss the effects of LDIR on stem cells as discovered through in vitro and in vivo studies. This article is protected by copyright. All rights reserved.
LPA Induces Metabolic Reprogramming in Ovarian Cancer via a Pseudohypoxic Response.
Publication Date: 15/04/2018, on Cancer research
by Ha JH, Radhakrishnan R, Jayaraman M, Yan M, Ward JD, Fung KM, Moxley K, Sood AK, Isidoro C, Mukherjee P, Song YS, Dhanasekaran DN
Although hypoxia has been shown to reprogram cancer cells toward glycolytic shift, the identity of extrinsic stimuli that induce metabolic reprogramming independent of hypoxia, especially in ovarian cancer, is largely unknown. In this study, we use patient-derived ovarian cancer cells and high-grade serous ovarian cancer cell lines to demonstrate that lysophosphatidic acid (LPA), a lipid growth factor and GPCR ligand whose levels are substantially increased in ovarian cancer patients, triggers glycolytic shift in ovarian cancer cells. Inhibition of the G protein α-subunit Gαi2 disrupted LPA-stimulated aerobic glycolysis. LPA stimulated a pseudohypoxic response via Rac-mediated activation of NADPH oxidase and generation of reactive oxygen species, resulting in activation of HIF1α. HIF1α in turn induced expression of glucose transporter-1 and the glycolytic enzyme hexokinase-2 (HKII). Treatment of mice bearing ovarian cancer xenografts with an HKII inhibitor, 3-bromopyruvate, attenuated tumor growth and conferred a concomitant survival advantage. These studies reveal a critical role for LPA in metabolic reprogramming of ovarian cancer cells and identify this node as a promising therapeutic target in ovarian cancer. These findings establish LPA as a potential therapeutic target in ovarian cancer, revealing its role in the activation of HIF1α-mediated metabolic reprogramming in this disease. .
Pro-inflammatory cytokines activate hypoxia-inducible factor 3α via epigenetic changes in mesenchymal stromal/stem cells.
Publication Date: 11/04/2018, on Scientific reports
by Cuomo F, Coppola A, Botti C, Maione C, Forte A, Scisciola L, Liguori G, Caiafa I, Ursini MV, Galderisi U, Cipollaro M, Altucci L, Cobellis G
Human mesenchymal stromal/stem cells (hMSCs) emerged as a promising therapeutic tool for ischemic disorders, due to their ability to regenerate damaged tissues, promote angiogenesis and reduce inflammation, leading to encouraging, but still limited results. The outcomes in clinical trials exploring hMSC therapy are influenced by low cell retention and survival in affected tissues, partially influenced by lesion's microenvironment, where low oxygen conditions (i.e. hypoxia) and inflammation coexist. Hypoxia and inflammation are pathophysiological stresses, sharing common activators, such as hypoxia-inducible factors (HIFs) and NF-κB. HIF1α and HIF2α respond essentially to hypoxia, activating pathways involved in tissue repair. Little is known about the regulation of HIF3α. Here we investigated the role of HIF3α in vitro and in vivo. Human MSCs expressed HIF3α, differentially regulated by pro-inflammatory cytokines in an oxygen-independent manner, a novel and still uncharacterized mechanism, where NF-κB is critical for its expression. We investigated if epigenetic modifications are involved in HIF3α expression by methylation-specific PCR and histone modifications. Robust hypermethylation of histone H3 was observed across HIF3A locus driven by pro-inflammatory cytokines. Experiments in a murine model of arteriotomy highlighted the activation of Hif3α expression in infiltrated inflammatory cells, suggesting a new role for Hif3α in inflammation in vivo.