20 research outputs found
L. Garzón Guillén, "L’immigrazione latinoamericana qualificata in Europa e l’effetto specchio. Note a partire dai casi catalano, italiano e belga"
No full textIl saggio è stato commissionato nel contesto della ricerca di Ateneo dell’Università di Padova sulle migrazioni latinoamericane ideata e coordinata da Valter Zanin.
Luis Garzón Guillén si interroga sui condizionamenti presenti nelle società di partenza e di arrivo i quali favoriscono o inibiscono la migrazione di personale professionalmente altamente qualificato dall’America latina all’Europa, concentrandosi soprattutto sul caso di argentini, ecuadoriani in Italia e in Catalogna, dei quali viene ricostruito il profilo professionale e di cui vengono menzionate problematiche di inserimento nei relativi mercati del lavoro
HHV-8 and EBV are not commonly found in idiopathic pulmonary fibrosis
No full textBACKGROUND AND AIM OF THE WORK: Idiopathic Pulmonary Fibrosis/Usual Interstitial Pneumonia (IPF/UIP) is a diffuse and progressive lung disease whose pathogenesis is incompletely understood. Recently, the presence of Herpesvirus-specific DNA was detected in the large majority of cases of a series of IPF/UIP and other lung interstitial diseases. We have therefore tested our own IPF/UIP series for the presence of HHV-8 and EBV proteins. METHODS: We used a variety of sensitive technologies including immunohistochemistry with NCL-HHV8-LNA and EBV-LMP1 antibodies, as well as of EBV RNA (EBER) by in-situ hybridisation; the presence of HHV-8 and EBV DNA was also investigated by means of PCR and subsequent analysis using a microfluidic apparatus. RESULTS: Despite the good sample quality, immunohistochemical, in-situ hybridisation and PCR results were negative for both EBV and HHV-8. CONCLUSIONS: We conclude that EBV and HHV-8 are not involved in the pathogenesis of IPF/UIP
Snf1 phosphorylates adenylate cyclase and negatively regulates protein kinase A-dependent transcription in saccharomyces cerevisiae
Get PDFIn eukaryotes, nutrient availability and metabolism are coordinated by sensing mechanisms and signaling pathways, which influence a broad set of cellular functions such as transcription and metabolic pathways to match environmental conditions. In yeast, PKA is activated in the presence of high glucose concentrations, favoring fast nutrient utilization, shutting down stress responses, and boosting growth. On the contrary, Snf1/AMPK is activated in the presence of low glucose or alternative carbon sources, thus promoting an energy saving program through transcriptional activation and phosphorylation of metabolic enzymes. The PKA and Snf1/AMPK pathways share common downstream targets. Moreover, PKA has been reported to negatively influence the activation of Snf1/AMPK. We report a new cross-talk mechanism with a Snf1-dependent regulation of the PKA pathway. We show that Snf1 and adenylate cyclase (Cyr1) interact in a nutrient-independent manner. Moreover, we identify Cyr1 as a Snf1 substrate and show that Snf1 activation state influences Cyr1 phosphorylation pattern, cAMP intracellular levels, and PKA-dependent transcription
NS5A inhibitors impair NS5A-phosphatidylinositol 4-kinase III∝ complex formation and cause a decrease of phosphatidylinositol 4-phosphate and cholesterol levels in hepatitis C virus-associated membranes
No full textThe hepatitis C virus (HCV) nonstructural (NS) protein 5A is a multifunctional protein that plays a central role in viral replication and assembly. Antiviral agents directly targeting NS5A are currently in clinical development. Although the elucidation of the mechanism of action (MOA) of NS5A inhibitors has been the focus of intensive research, a detailed understanding of how these agents exert their antiviral effect is still lacking. In this study, we observed that the downregulation of NS5A hyperphosphorylation is associated with the actions of NS5A inhibitors belonging to different chemotypes. NS5A is known to recruit the lipid kinase phosphatidylinositol 4-kinase III∝ (PI4KIII∝) to the HCV-induced membranous web in order to generate phosphatidylinositol 4-phosphate (PI4P) at the sites of replication. We demonstrate that treatment with NS5A inhibitors leads to an impairment in the NS5A-PI4KIII∝ complex formation that is paralleled by a significant reduction in PI4P and cholesterol levels within the endomembrane structures of HCV-replicating cells. A similar decrease in PI4P and cholesterol levels was also obtained upon treatment with a PI4KIII∝-targeting inhibitor. In addition, both the NS5A and PI4KIII∝ classes of inhibitors induced similar subcellular relocalization of the NS5A protein, causing the formation of large cytoplasmic NS5A-containing clusters previously reported to be one of the hallmarks of inhibition of the action of PI4KIII∝. Because of the similarities between the effects induced by treatment with PI4KIII∝ or NS5A inhibitors and the observation that agents targeting NS5A impair NS5A-PI4KIII∝ complex formation, we speculate that NS5A inhibitors act by interfering with the function of the NS5API4KIII∝ complex
Espressione di marcatori di migrazione e invasività nei foci fibroblastici nella fibrosi polmonare idiopatica.
No full textMass spectrometry-based metabolomics : convergence of Snf1/AMPK and methionine metabolism to control mitochondrial respiration
No full textMass spectrometry-based metabolomics has become increasingly popular in biology and molecular medicine. Analysis of the key metabolites in biological fluids has become an important part of improving disease diagnosis. In this context, chromatography coupled with mass spectrometry represents a valid tool to characterize and quantify numerous compounds present in biological matrices. In the present work, we have explored the role of methionine metabolism on cellular functions of a prototrophic yeast strain during exponential growth phase and determining how this affects the metabolic state upon SNF1 deletion.
Experimental
Yeast strains and growth conditions S. cerevisiae strains used in this study are reported in Table 1. Synthetic medium (SD)
contains 2% glucose, 6.7 g/L of Yeast Nitrogen Base (Difco). Methionine was added to the concentrations indicated in figure legends. In these conditions, cells exhibit exponential growth between OD600nm = 0.1 per ml (equivalent to 2*106 cells/ml) and OD600nm = 2.5 per ml (5*107 cells/ml); all experiments were performed in exponential phase of growth (OD600nm/ml 0.5-1) or post-log phase (24 h, 48 h and 72 h after the beginning of exponential phase). Antimycin A was added to a final concentration of 1 μg/ml from 2 mg/ml stock in 100% ethanol; the same volume of ethanol was added in the control culture.
Cells were properly manupulated in order to extract RNA and carried out all the necessary biological investigation in addition to metabolites analysis by GC-MS.
Metabolites analysis
13C-labelling of proteinogenic amino acids was achieved by growth on 20 g/L glucose as a mixture of 80% (w/w) unlabelled and 20% (w/w) uniformly labeled [U-13C]glucose (13C, 99 %; Cambridge Isotope Laboratories, Inc). Cells from an overnight minimal medium culture were washed and used for inoculation below an OD600 of 0.03. 13C-labelled biomass aliquots were harvested by centrifugation during the mid-exponential growth phase at an OD600 of ≤1. The cells (about 0.3 mg of dry biomass) were washed once with sterile water, and hydrolysed in 150 μL 6 M HCl at 105 oC for 6 h. The hydrolysate was dried in a heating block at 80 oC under a constant airflow.
Before the GC/MS analyses all samples were subjected to a derivatization step as follows. Each sample was resuspended in 30 μL of acetonitrile, followed by 30 μL of MBDSTFA (N-methyl-N-ter-butyldimethylsilyl-trifluoroacetamide). The resulting mixture, contained in a closed vial, was stirred for 10 min and centrifuged for 15 sec. Then, the vial was incubated at 85° C. After 1 h, the sample slowly reached room temperature and was analysed by GC-MS (single quadrupole).
A METAFOR (metabolic flux ratio) analysis was perfomed: the mass isotopomer distribution of proteinogenic amino acids was used to calculate the split ratios of key branching points of yeast central metabolism using the software FIAT FLUX [3].
The results show that methionine addition leads to Snf1 and Acc1 phosphorylations as well as a general slow-down of proliferation, boosting mtDNA synthesis, mitochondrial pyruvate uptake and TCA cycle activity. Remarkably, mitochondrial proteomic analysis highlight that methionine upregulates proteins of the pyruvate dehydrogenase complex, TCA cycle and respiratory chain complexes.
Thus, Snf1/AMPK and methionine metabolism converge to control mitochondrial respiration in glucose repressed conditions.
References:
[1] Kushnirov, V. V. (2000). Yeast 16, 857–860. [2] Pessina, S., Tsiarentsyeva, V., Busnelli, S., Vanoni, M., Alberghina, L., and Coccetti, P. (2010). Cell Cycle 9, 2189–2200. [3] Zamboni, N., Fischer, E., and Sauer, U. (2005). BMC Bioinformatics 6, 209
Espressione di marcatori di migrazione e invasività nei foci fibroblastici nella fibrosi polmonare idiopatica
No full textNutritional modulation of CK2 in Saccharomyces cerevisiae: regulating the activity of a constitutive enzyme
No full textCK2 is a highly conserved protein kinase involved in different cellular processes, which shows a higher activity in actively proliferating mammalian cells and in various types of cancer and cancer cell lines. We recently demonstrated that CK2 activity is strongly influenced by growth rate in yeast cells as well. Here, we extend our previous findings and show that, in cells grown in either glucose or ethanol-supplemented media, CK2 presents no alteration in K(m) for both the ATP and the peptide substrate RRRADDSDDDDD, while a significant increase in V (max) is observed. In chemostat-grown cells, no difference of CK2 activity was observed in cells grown at the same dilution rate in media supplemented with either ethanol or glucose, excluding the contribution of carbon metabolism on CK2 activity. By using the eIF2 beta-derived peptide, which can be phosphorylated by the holoenzyme but not by the free catalytic subunits, we show that the holoenzyme activity requires the concurrent presence of both beta and beta' encoding genes. Finally, conditions of nitrogen deprivation leading to a G0-like arrest result in a decrease of total CK2 activity, but have no effect on the activity of the holoenzyme. These findings newly indicate a regulatory role of beta and beta' subunits of CK2 in the nutrient response
Carcinoid crisis induced by receptor radionuclide therapy with 90Y-DOTATOC in a case of liver metastases from bronchial neuroendocrine tumor (atypical carcinoid).
No full textSS receptors are overexpressed in many tumors, mainly of neuroendocrine origin, thus enabling the treatment with SS analogs. The clinical experience of receptor radionuclide therapy with the new analog [90Y-DOTA0-Tyr3 ]-octreotide [90Y-DOTATOC] has been developed over the last decade and is gaining a pivotal role in the therapeutic workout of these tumors. It is well known that some procedures performed in diagnostic and therapeutic management of endocrine tumors, such as agobiopsy and hepatic chemoembolization, can be associated with the occurrence of symptoms related to the release of vasoactive amines and/or hormonal peptides from tumor cell lysis. This is the first report of a severe carcinoid crisis developed after receptor radionuclide therapy with 90Y-DOTATOC administered in a patient affected by liver metastases from bronchial neuroendocrine tumor (atypical carcinoid). Despite protection with H1 receptor antagonists, octreotide and corticosteroids, few days after the therapy the patient complained of persistent flushing of the face and upper trunk, severe labial and periocular oedema, diarrhoea and loss of appetite. These symptoms increased and required new hospitalisation. The patient received iv infusion of octreotide associated with H1 and H2 receptor antagonists and corticosteroid therapy, which induced symptom remission within few days. The case here reported confirms that radionuclide therapy is highly effective in determining early rupture of metastatic tissue and also suggests that pre-medication should be implemented before the radiopeptide administration associated with a close monitoring of the patient in the following days
