1,720,979 research outputs found
GENETIC POLYMORPHISMS OF THE FOLATE METABOLIC PATHWAY IN CHILDHOOD ACUTE LYMPHOBLASTIC LEUKEMIA. A MOLECULAR STUDY AND A PROPOSAL FOR AN INTERPRETATIVE MODEL.
No full textAcute lymphoblastic leukemia (ALL) is the most common pediatric cancer accounting for 80% of childhood leukemia. The uncontrolled proliferation of lymphoid progenitors in the bone marrow and the accumulation of malignant lymphoblasts in peripheral blood characterize the disease. The molecular analysis of common genetic alterations in lymphoblastic cells has strongly contributed to the comprehension of ALL pathogenesis. Different gene polymorphisms (most of them SNPs) play an important role in the susceptibility to childhood ALL which probably derives from a combination and relation of genetic and environmental factors. Folic acid and the pool of folate of the one carbon-metabolic-pathway are key elements involved in several processes including DNA synthesis and methylation. Polymorphisms in genes coding for enzymes of the folate metabolic pathway can alter the intracellular folate status or distribution and sub-optimal/anomalous folate levels/distribution increase the risk of developing several neoplasias. The two main enzymes involved in cyclization of folate isoforms are DHFR and MTHFR. The first one is responsible for the conversion of dihydrofolate to tetrahydrofolate whilst the second one catalyzes the reduction of 5,10-methylene-THF to 5-methyl-THF. Studies have demonstrated that subjects with the homozygous DD-genotype have higher DHFR mRNA levels that may be responsible for storage of THF and other isoforms within the cell; reduced enzyme activity instead, has been associated to MTHFR 677TT homozygotes. This condition is responsible for an underutilization of methylene-THF in the cell with storage of folate reduced isoforms. Several studies have demonstrated that DHFR and MTHFR polymorphisms may be protective against hematological cancers such as ALL. It is to note that current treatment regimens achieve levels about 80% in overall survival (OS). Unfortunately, the side effects derived from the chemotherapeutic agents used can be severe, especially for high-risk patients. Therefore, the identification of additional markers which can improve risk stratification and individual tailored therapy regimens would be a great goal, in order to avoid over-treatment which can increase long-term adverse side effects.
The aim of the present study was, therefore, to investigate whether common polymorphisms (i.e. MTHFR C677T and A1298C in addition to DHFR 19 bp INS/DEL and Bcl-2 -938 C>A) might influence the risk of childhood ALL. After a single analysis we can ascribe to MTHFR C677T gene polymorphism a protective significant role against childhood ALL (P=0.046), whilst Bcl-2 -938 C>A gene polymorphism seems to be a risk factor for the susceptibility to the disease (P=0.049). Then, considering parameters such as disease onset and therapy duration we can observe a significant higher mean age onset disease for homozygotes MTHFR 1298-CC (P=0.05). From the analysis of the therapy duration we found a significant association for MTHFR A1298C and Bcl-2 -938 C>A polymorphisms: MTHFR 1298CC homozygotes showed a slight higher mean therapy duration (P=0.05), as well as Bcl-2 -938AA homozygotes (P=0.03). Finally, in an exploratory way, to validate the proposed model we evaluated among healthy PBL cells harvested from subjects with opposite genotype condition considering DFHR and MTHFR genes (respectively, WW/CC and DD/TT), possible differences in base-line cellular viability and under MTX treatment. The pharmacological induced restriction in folate availability (MTX) yields to results in favor of WW/CC, whilst the base-line cell viability yields comparable results among genotypes. This is in line with the hypothesis that a higher MTX level could be present in DD/TT cells, prone to storage either natural folate isoforms (useful for the cell viability) or synthetic toxic analogue (MTX). This observation argues us into hypothesizing that also MTX being itself a synthetic folate analogue follows the same handling process. Now, inside the cell it should result in an elevated toxicity level, being responsible for an elevated death
Surdité brusque (SB): recherche de nouveaux marqueurs moléculaires.
No full textThis study is the first recognition of genetic implications (i.e. SNPs) in the field of sudden sensorineural hearing loss. In detail, SNPs of iron homeostasis genes seem to have a role in sudden sensorineural hearing loss establishment. Pharmacogenetics aspects have been discussed
Sudden sensorineural hearing loss and polymorphisms in iron homeostasis genes: New insights from a case-control study
Full text linkBackground. Even if various pathophysiological events have been proposed as explanations, the putative cause of sudden hearing loss remains unclear. Objectives. To investigate and to reveal associations (if any) between the main iron-related gene variants and idiopathic sudden sensorineural hearing loss. Study Design. Case-control study. Materials and Methods. A total of 200 sudden sensorineural hearing loss patients (median age 63.65 years; range 10-92) were compared with 400 healthy control subjects. The following genetic variants were investigated: the polymorphism c.-8CG in the promoter of the ferroportin gene (FPN1; SLC40A1), the two isoforms C1 and C2 (p.P570S) of the transferrin protein (TF), the amino acidic substitutions p.H63D and p.C282Y in the hereditary hemochromatosis protein (HFE), and the polymorphism c.-582AG in the promoter of the HEPC gene, which encodes the protein hepcidin (HAMP). Results. The homozygous genotype c.-8GG of the SLC40A1 gene revealed an OR for ISSNHL risk of 4.27 (CI 95%, 2.65-6.89; P = 0.001), being overrepresented among cases. Conclusions. Our study indicates that the homozygous genotype FPN1 -8GG was significantly associated with increased risk of developing sudden hearing loss. These findings suggest new research should be conducted in the field of iron homeostasis in the inner ear
P2X7 Receptor Orchestrates Multiple Signalling Pathways Triggering Inflammation, Autophagy and Metabolic/Trophic Responses
No full textP2X7 receptor is an ion channel activated by extracellular adenosine trisphosphate (eATP) that attracted increasing attention for its role in immune reactions, neurobiology and oncology. As receptor for an extracellular ligand, P2X7 activates a series of intracellular signalling pathways mainly via alterations of the ion permeability, but also through formation of a large unselective pore and direct interaction with other proteins. Here we wish to give an overview on the main biochemical paths initiated by P2X7 activation by revising recent and established literature on P2X7-triggered signalling cascades leading to cell death, inflammatory and immune response activation, proliferation and metabolism modulation. We will focus on the well-known P2X7 inflammasome/NF-kB and pro-apoptotic networks but also cover P2X7-activated emerging autophagic, pyroptotic and proliferative-oncogenic pathways, like beclin-1/LC3-II, caspase-11, Akt and VEGF axes
P2X7 Receptor as a Therapeutic Target
No full textP2X7 receptor is an ATP-gated cation channel that upon agonist interaction leads to cellular influx of Na+ and Ca2 + and efflux of K+. P2X7 is expressed by a wide variety of cells and its activation mediates a large number of biological processes like inflammation, neuromodulation, cell death or cell proliferation and it has been associated to related pathological conditions including infectious, inflammatory, autoimmune, neurological, and musculoskeletal disorders and, in the last years, to cancer. This chapter describes structural features of P2X7, chemical properties of its agonist, antagonist, and allosteric modulators and summarizes recent advances on P2X7 receptor as therapeutic target in the aforementioned diseases. We also give an overview on recent literature suggesting that P2X7 single-nucleotide polymorphisms could be exploited as diagnostic biomarkers for the development of tailored therapies
Acute myeloid leukaemia:analysis of the expression of P2X7a and P2X7B isoforms
No full textAcute Myeloid Leukaemia (AML) is a clonal disorder originating from the accumulation of myeloid progenitor cells in the bone marrow. An increasing number of reports correlate P2X7 with tumor growth and progression (Adinolfi et al. 2015). Haemopoietic lympho-proliferative disorders, among which AML (Salvestrini et al. 2012) were the first neoplastic models where P2X7 oncopromoting activity was demonstrated (Adinolfi et al., 2002; Chong et al., 2010).The P2X7 receptor for extracellular ATP is a ionotropic, ligand-gated, cation channel and it is expressed by different cell types such as neurons, macrophages, dendritic and microglial cells, fibroblasts, endothelial cells and lymphocytes. Nine different P2X7 splice variants have been detected in humans, among those just two, P2X7A and P2X7B are functional ion channels (Cheewatrakoolpong et al., 2005) both showing growth promoting activity (Adinolfi et al., 2010; Giuliani et al., 2014).
Isoform B is a naturally truncated P2X7 splice variant, widely expressed in several human tissues especially in lymphoid tissue and lymphocytes themselves, often to a much higher level than P2X7A. Considering that a wide characterization of P2X7B is still lacking, we analyzed the differential expression of P2X7A and P2X7B in a cohort of AML patients by Real Time PCR. We obtained a significant correlation between high P2X7B expression and relapse as compared to onset of disease. Moreover, low P2X7A expression associates with favorable karyotype and, although not significant, there is a similar tendency for P2X7B. Futhermore, preliminary data suggest a correlation between age of onset (>65 years) and high P2X7B expression. Although a wider cohort analysis will be required, these data prompt us to postulate that different P2X7A and B expression levels could be involved in AML development, suggesting that P2X7 could be a valuable prognostic marker and therapeutic target for the disease
Dihydrofolate reductase (DHFR) 19-bp ins/del polymorphism and methylenetetrahydrofolate reductase (MTHFR) C677T in coronary heart disease patients: potential intracellular folate unbalancing.
No full textElevated homocysteine (Hcy) due to common gene variants is correlated with coronary heart disease (CHD), but definite causality remains uncertain. Hcy increases due to genetic and environment interactions. Assuming no direct effects of folate on CHD except via increased Hcy, and that folate drives Hcy metabolism, investigating on how genes influence folate balancing is strongly useful. DHFR and MTHFR fully activate diet folate. Common polymorphisms influence activity/level of enzymes. MTHFR677TT reduces enzyme efficiency yielding ~20% higher Hcy. Less definitive is the role ascribed to DHFR 19bp ins/del; some authors reported ~50% higher mRNA in del/del genotype. We selected 14 CHD patients from our Center carrying opposite DHFR/MTHFR genotypes. We hypothesized that patients (n=7) carrying both polymorphisms in homozygous condition (DHFRdel/del and MTHFR677TT) might have different folate status and respond differently, than double wildtypes (n=7), under controlled in vitro folate levels. So, we put whole blood in culture (t0, t48, t96-120 hours) containing 10ng/ml folate. DD/TT patients had significant lower basal folate levels (plasma and RBC) versus wildtypes. Interestingly, in vitro culture showed DD/TT genotype rapidly gained folate stores, to completely cancel the gap at ~96h. These results are partially in contrast with previous reports in which DHFR properties were tested without consider MTHFR, and strongly advice to analyze these two common variants before ascribing disease risk and public health implications
Una "rete" per proteggere il cuore. (FAR 2013)
No full textBackground: Dopo un infarto del miocardio (MI), il tessuto danneggiato del cuore deve essere riparato e sostituito da nuovo tessuto. Nella maggior parte dei pazienti con MI, i meccanismi di riparazione inducono profondi cambiamenti strutturali e funzionali. Queste modificazioni purtroppo non sono limitate esclusivamente alla zona infartuata, ma si estendono anche alle zone non compromesse spesso determinando cambiamenti disfunzionali.
Nelle fasi iniziali dopo un MI, si verifica un assottigliamento ed una estensione della zona colpita, e una patologica ipertrofia dei cardiomiociti, con apoptosi e rimodellamento della matrice extracellulare (ECM) della zona cosi detta “remota” cioè non direttamente colpita dall’infarto. Questi ultimi processi possono estendersi nel tempo determinando alterazioni della complessa geometria del ventricolo sinistro (LV), mediante cambi di forma, massa, volume e funzione del LV (Nahrendorf et al; Eur Heart J. 2008).
Sebbene alcuni di questi cambiamenti possono essere fisiologici e di natura adattativa, come risposta compensatoria a breve termine a causa della improvvisa perdita di funzione contrattile del cuore nell’area infartuata, nel lungo periodo possono causare scompenso cardiaco e morte cardiovascolare. I principali determinanti di un rimodellamento anomalo post-infarto comprendono il grado di estensione dell’infarto e le condizioni di carico del LV.
Fino ad oggi numerosi sforzi sono stati fatti per ridurre l’estensione dell’infarto mediante una tempestiva riperfusione (con drastica diminuzione delle morti in fase acuta) e per ridurre farmacologicamente il carico di lavoro del LV. A fronte di una significativo calo delle morti, si verifica oggi un altrettanto drastico aumento di patologie cardiache post-MI a causa della mancanza di trattamenti efficaci per trattare a lungo termine i sopravvissuti ad un MI. Il frequente anomalo rimodellamento cardiaco post-MI è un complesso ed articolato processo mediato da cellule e fattori che solo ultimamente stanno emergendo come strategie alternative di trattamento (Nahrendorf et al; Circulation 2010).
L’healing cardiaco post-MI può essere schematizzato in tre fasi:
-Fase infiammatoria: reclutamento di monociti e neutrofili, mediatori chimici (complemento e chemokine ligand) e attivazione delle MMPs, per rimuovere cellule morte e degradare la matrice extracellulare (ECM).
-Fase proliferativa: monociti/macrofagi producono citokine e fattori d crescita per reprimere la risposta infiammatoria e regolare la formazione del tessuto di granulazione. In questa fase angiogenesi e deposizione di nuova ECM da parte dei fibroblasti sono processi determinanti.
-Fase di maturazione: rimodellamento della ECM, apoptosi cellulare e formazione di una cicatrice matura di collagene con apprezzabili proprietà elastiche.
Un ottimale healing cardiaco necessita di una riposta infiammatoria bilanciata. La fase infiammatoria è necessaria, ma non deve essere né eccessiva né limitata (nel tempo o nell’intensità), così come le fasi proliferativa e di maturazione che se non ottimali determinano cicatrici fibrose e poco elastiche. I processi di healing cardiaco post-MI, a differenza di altri distretti, avvengono in un organo in continuo movimento. La lesione deve essere riparata in condizioni di stress meccanico dovuto ai cicli della contrazione cardiaca e alla pressione intraventricolare. In queste condizioni estreme si assiste alla formazione di nuove mini lesioni e/o alla estensione della lesione primaria con conseguente sfasamento ed accavallamento di nuovi processi di riparazione e grave sbilanciamento del timing della fase ripartiva. Questo porta ad un insufficiente processo ripartivo con sostituzione di tessuto cardiaco fibrotico scarsamente performante le funzioni e le performance cardiache divenendo ipocinetico/discinetico. A differenza della riparazione in altri distretti (cute, ossa, etc.) in cui la presenza di scaffold rigidi favorisce la riparazione, il cuore necessiterebbe di scaffold compiacenti che assecondino tutti i suoi movimenti senza danneggiarsi. La deposizione di strutture pseudo-rigide (polimeri, reti composte da fili da sutura) intorno al cuore in un modello ripartivo post-MI animale ha fornito dati incoraggianti i cui risultati convergono nel dimostrare che la “costrizione” e circoscrizione della zona infartuata riduce significativamente espansione e rimodellamento anomalo del LV scongiurando l’instaurazione di condizioni di scompenso cardiaco (Mukherjee et al; Criculation 2011).
Proposta del progetto: Il nostro gruppo recentemente ha dimostrato come i livelli circolanti di un fattore polimerizzante la Fibrina (FXIII) si riducono significativamente durante le prime giornate post-MI per raggiungere valori medi molto bassi (20-30% del normale) intorno alla 4°-5° giornata (Gemmati et al, Mol Medicine 2007). La riduzione dei livelli è associata significativamente a prognosi negativa post-MI in particolare a scompenso cardiaco. Altri gruppi hanno dimostrato che l’assenza di FXIII in topi KO -- / +- per il gene, porta a morte post-MI entro la 5° gg a causa della rottura del ventricolo sn., e che la reinfusione di FXIII nei topi KO elimina completamente questa severa complicanza (Nahrendorf et al; Circulation 2006). Il FXIII è recentemente considerato un fattore che influenza positivamente tutte le tre fasi di riparazione tessutale con spiccate capacità neo-angiogeniche, intersecando coagulazione, fibrinolisi, infiammazione e riparazione tessutale (Ichinose A; Int J Hematol 2012).
La nostra proposta di ricerca si basa sull’ipotesi che una rete biologica con aumentate caratteristiche elastiche ed estensibili possa essere costituita dalla fibrina cross-linkata dal FXIII, e che questa rete si vada a posizionare spontaneamente intorno alla lesione cardiaca post-MI per la naturale affinità esistente tra strutture lese-Fibrina-FXIII presente in tutti i processi emostatici/coagulativi/riparativi che seguono l’instaurarsi di una lesione.
-Nei laboratori del Centro Emostasi e Trombosi si allestiranno in vitro reti di fibrina cross-linkata a differenti concentrazioni di FXIII a tre livelli di concentrazione di Fibrinogeno (low, medium, high). Queste tre differenti concentrazioni di fibrinogeno sono state scelte per simulare le differenti condizioni che si presentano in vivo durante la riparazione tessutale a seconda che essa sia precoce o ritardata in presenza di differenti condizioni di infiammazione
-Varianti geniche del FXIII, espresse in vitro in collaborazione con il Dipartimento di Biochimica e Biologia Molecolare, note per possedere attività funzionali differenti, saranno utilizzate per ottenere reti di fibrina con potenziali differenze strutturali.
-La composizione qualitativa della rete sarà analizzata dopo riduzione chimica ed analisi elettroforetica al Western Blotting. Questo rivelerà importanti informazioni sul timing di polimerizzazione e sul grado di legami covalenti eseguiti da parte del FXIII sulla maglia di Fibrina. Sono queste le variabili che influenzano elasticità ed estensibilità della maglia di fibrina (Li uet al, Science 2006).
-In collaborazione con il Dipartimento di Fisica dell’Università di Ferrara e dell’Università di Messina si analizzeranno al microscopio elettronico (ME) la struttura tridimensionale della maglia di fibrina, e numerose proprietà fisiche (in particolare elasticità ed estensibilità) con differenti approcci sperimentali: microscopia a forza atomica (AFM) e sollecitazione meccanica (Univ. di Messina); calcolo del ritardo di risposta dopo sollecitazione con ultrasuoni (Univ. di Ferrara).
Ricadute traslazionali:
Alla luce delle numerose proprietà biologiche e fisiche favorenti la riparazione tessutale recentemente riconosciute al FXIII da parte di numerosi gruppi di ricerca indipendenti, proponiamo una accurata caratterizzazione di molecole ricombinanti di FXIII e successiva valutazione degli effetti biologici che esse possono esercitare sulla maglia di Fibrina. La successiva selezione di combinazioni geniche con potenziale ruolo pro-healing è la premessa essenziale per le successive fasi di ricerca traslazionale. Trattamenti personalizzati sulla base dei principi della farmacogenetica, restituirebbero risultati significativi nel campo della riparazione tessutale e nello specifico per contrastare ed interrompere efficacemente l’anomalo rimodellamento post-MI
Role of the P2X7 receptor in tumor-associated inflammation
No full textInflammation is constantly associated to cancer. Malignant tumors often develop at sites of chronic inflammation, and inflammation promotes tumor progression. But, at the same time, inflammation is crucial for anti-tumor immune response. Many factors are responsible for this ‘Dr Jekyll/Mr Hyde’ roles of inflammation, among which one that is attracting increasing attention is the P2X7 receptor (P2X7R). This receptor is expressed by most malignant tumors and widely diffused in innate and adaptive immune cells, where it supports proliferation, chemotaxis, growth factor, and cytokine release. P2X7R-targeting may offer novel avenues for anti-cancer therapeutic intervention, but might also impair host anti-tumor responses. This short review highlights recent findings on the dual role of the P2X7R in cancer-associated inflammation
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