Caratterizzazione ed isolamento di intermedi delle reazioni di sostituzione elettrofila e nucleofila in serie aromatica

1,3,5–Tris(N,N-dialkylamino)benzene derivatives are strongly activated neutral carbon nucleophiles able to stress some reactivity aspects toward more or less activated electrophilic substrates. These very interesting electron-rich benzenes have been firstly synthesized in 1967 and extensively studied. Their supernucleophilic character permits to perform reactions in particularly mild conditions, and make them suitable for mechanistic investigations. In many reactions they permit to isolate –complexes in electrophilic aromatic reactions. The possibility to form moderately stable Wheland intermediates depends both, on the activation of the reagents and on the experimental conditions which makes slow the proton elimination in the re-aromatization process. In presence of a carbon super electrophile reagent as 4,6-dinitrobenzofuroxan or 4,6-dinitrotetrazolepiridine, 1,3,5–tris(N,N-dialkylamino)benzene derivatives afford C–C coupling products which are “double σ complexes”, Wheland–like on the 1,3,5-tris(N,N-dialkylamino)benzene moiety, and Meisenheimer–like on the electrophile moiety. We named these complexes as Wheland–Meisenheimer (W-M) complexes. These complexes are moderately stable at low temperature and they were characterized by NMR spectroscopy methods. Others nucleophile reagents as 2-aminothiazole derivatives give a Wheland-Meisenheimer complex with 4,6-dinitrobenzofuroxan.

Genetica molecolare dell'autismo: studi di associazione ed analisi di geni candidati

Autism is a neurodevelpmental disorder characterized by impaired verbal communication, limited reciprocal social interaction, restricted interests and repetitive behaviours. Twin and family studies indicate a large genetic contribution to ASDs (Autism Spectrum Disorders). During my Ph.D. I have been involved in several projects in which I used different genetic approaches in order to identify susceptibility genes in autism on chromosomes 2, 7 and X: 1)High-density SNP association and CNV analysis of two Autism Susceptibility Loci. The International Molecular Genetic Study of Autism Consortium (IMGSAC) previously identified linkage loci on chromosomes 7 and 2, termed AUTS1 and AUTS5, respectively. In this study, we evaluated the patterns of linkage disequilibrium (LD) and the distribution of haplotype blocks, utilising data from the HapMap project, across the two strongest peaks of linkage on chromosome 2 and 7. More than 3000 SNPs have been selected in each locus in all known genes, as well as SNPs in non-genic highly conserved sequences. All markers have been genotyped to perform a high-density association analysis and to explore copy number variation within these regions. The study sample consisted of 127 and 126 multiplex families, showing linkage to the AUTS1 and AUTS5 regions, respectively, and 188 gender-matched controls. Association and CNV analysis implicated several new genes, including IMMP2L and DOCK4 on chromosome 7 and ZNF533 and NOSTRIN on the chromosome 2. Particularly, my contribution to this project focused on the characterization of the best candidate gene in each locus: On the AUTS5 locus I carried out a transcript study of ZNF533 in different human tissues to verify which isoforms and start exons were expressed. High transcript variability and a new exon, never described before, has been identified in this analysis. Furthermore, I selected 31 probands for the risk haplotype and performed a mutation screen of all known exons in order to identify novel coding variants associated to autism. On the AUTS1 locus a duplication was detected in one multiplex family that was transmitted from father to an affected son. This duplication interrupts two genes: IMMP2L and DOCK4 and warranted further analysis. Thus, I performed a screening of the cohort of IMGSAC collection (285 multiplex families), using a QMPSF assay (Quantitative Multiplex PCR of Short fluorescent Fragments) to analyse if CNVs in this genic region segregate with autism phenotype and compare their frequency with a sample of 475 UK controls. Evidence for a role of DOCK4 in autism susceptibility was supported by independent replication of association at rs2217262 and the finding of a deletion segregating in a sib-pair family. 2)Analysis of X chromosome inactivation. Skewed X chromosome inactivation (XCI) is observed in females carrying gene mutations involved in several X-linked syndromes. We aimed to estimate the role of X-linked genes in ASD susceptibility by ascertaining the XCI pattern in a sample of 543 informative mothers of children with ASD and in a sample of 164 affected girls. The study sample included families from different european consortia. I analysed the XCI inactivation pattern in a sample of italian mothers from singletons families with ASD and also a control groups (144 adult females and 40 young females). We observed no significant excess of skewed XCI in families with ASD. Interestingly, two mothers and one girl carrying known mutations in X-linked genes (NLGN3, ATRX, MECP2) showed highly skewed XCI, suggesting that ascertainment of XCI could reveal families with X-linked mutations. Linkage analysis was carried out in the subgroup of multiplex families with skewed XCI (≥80:20) and a modest increased allele sharing was obtained in the Xq27-Xq28 region, with a peak Z score of 1.75 close to rs719489. In this region FMR1 and MECP2 have been associated in some cases with austim and therefore represent candidates for the disorder. I performed a mutation screen of MECP2 in 33 unrelated probands from IMGSAC and italian families, showing XCI skewness. Recently, Xq28 duplications including MECP2, have been identified in families with MR, with asymptomatic carrier females showing extreme (>85%) skewing of XCI. For these reason I used the sample of probands from X-skewed families to perform CNV analysis by Real-time quantitative PCR. No duplications have been found in our sample. I have also confirmed all data using as alternative method the MLPA assay (Multiplex Ligation dependent Probe Amplification). 3)ASMT as functional candidate gene for autism. Recently, a possible involvement of the acetylserotonin O-methyltransferase (ASMT) gene in susceptibility to ASDs has been reported: mutation screening of the ASMT gene in 250 individuals from the PARIS collection revealed several rare variants with a likely functional role; Moreover, significant association was reported for two SNPs (rs4446909 and rs5989681) located in one of the two alternative promoters of the gene. To further investigate these findings, I carried out a replication study using a sample of 263 affected individuals from the IMGSAC collection and 390 control individuals. Several rare mutations were identified, including the splice site mutation IVS5+2T>C and the L326F substitution previously reported by Melke et al (2007), but the same rare variants have been found also in control individuals in our study. Interestingly, a new R319X stop mutation was found in a single autism proband of Italian origin and is absent from the entire control sample. Furthermore, no replication has been found in our case-control study typing the SNPs on the ASMT promoter B.

Fra openness ed embeddedness: network analysis di un distretto industriale glocale.

Il distretto è un luogo relazionale dinamico dove le imprese danno luogo a differenti comportamenti economici di vario genere e natura, cooperando in un certo senso per lo sviluppo e la crescita del distretto stesso. In un primo momento di formazione del distretto si sono delineati comportamenti di tipo path dependent per vantaggi economici dovuti alla distribuzione delle imprese nel territorio, ma con il tempo si sono cominciati ad avere comportamenti espansionistici differenti sia dall'interno che dall'esterno del distretto influendo direttamente sulla struttura del stesso. É ragionevole dunque pensare che gli attori guardino al rapporto “locale/globale” con una sorta di "strabismo", da un lato leggendo il distretto (dall’interno come dall’esterno) come un luogo privilegiato per la formazione di economie di prossimità, dall’altro puntando a disporre le catene produttive nello spazio globale, alla ricerca dei vantaggi derivanti da un minor costo del lavoro o dalla immediata prossimità dei mercati di sbocco. il distretto viene dunque attraversato da dinamiche che lo globalizzano ma, al contempo, ne preservano (almeno per ora) la specificità. Non è più possibile leggere la sua forma economica solo nella logica della embeddedness, e non sarebbe certo corretto farlo solo in chiave di openness. Si tratta dunque di interrogarsi sul rapporto più di integrazione/complementarità che di contrapposizione fra openness ed embeddedness. In questa tesi verrà descritto un metodo d'approccio per dare un valore al fenomeno di Openness e Embeddedness presente nel distretto partendo da un dataset di dati relazionali ricavati da due database economici Amadeus e Aida. Non essendo possibile trovare pubblicamente dati sulle reti di fornitura delle singole aziende, siamo partiti dai dati relazionali di cinque aziende “seme”, ed attraverso una ricerca ricorsiva nelle relazioni di azionariato/partecipazione, siamo riusciti ad ottenere un campione di analisi che ci permette di mettere in luce tramite la custer analysis le principali tipologie di reti di imprese presenti nel distretto ed estese nello spazio globale.

Energia da fonti rinnovabili negli edifici esistenti. Integrazione architettonica di captatori solari ed eolici.

Questa tesi tratta dell'applicazione agli edifici esistenti di dispositivi per la produzione energetica da fonte rinnovabile: solare fotovoltaica, solare termica ed eolica. Dopo aver analizzato i motivi economici ed ambientali che oggi inducono ad affrontare il tema complesso dell'integrazione di queste tecnologie anche negli edifici esistenti, vengono documentate alcune decine di recenti istallazioni, reperite in ambito internazionale, costituendo così un piccolo repertorio di casi applicativi. Il ricorso alle fonti di energia rinnovabili in architettura ed il concetto di sostenibilità applicato alla progettazione sono temi di interesse mondiale che coinvolgono la riqualificazione energetica del parco edilizio esistente. Questo fenomeno sta generando una vera rivoluzione che, da una dimensione culturale promossa dalle politiche energetiche dei vari paesi, travolge svariati ambiti disciplinari, primo tra tutti l'architettura. L’elaborato è idealmente diviso in quattro parti, come di seguito specificato. Nel primo capitolo vengono trattate le problematiche che stanno portando alla diffusione delle energie rinnovabili nel mondo e descritte le varie forme e tecnologie con cui si utilizzano. Nel secondo e terzo capitolo sono trattati i temi dell’energia negli edifici e la sostenibilità nel progetto per la riqualificazione energetica in architettura. Negli ultimi tre capitoli si analizzano le caratteristiche, i componenti ed il funzionamento dei sistemi fotovoltaici, solari termici e micro/mini-eolici applicati ai manufatti edilizi. Si descrivono, inoltre, le tecnologie e le soluzioni per l’integrazione di detti impianti sugli edifici esistenti. L’ultima parte, costituita dall’allegato che contiene le schede di analisi di alcuni casi studio, descrive la realizzazione di progetti di riqualificazione energetica in cui sistemi fotovoltaici, solari termici ed eolici vengono applicati ad edifici di vario genere.

Diabete ed ischemia critica degli arti inferiori. Studio del danno della parete arteriosa e del ruolo dell'endotelio circolante nella riparazione tessutale e nella neoangiogenesi

Objectives In diabetic and non diabetic patients with peripheral artery obstructive disease (PAOD), we sought to establish whether the vascular wall damage, the mature circulating endothelium and the "in situ" neoangiogenesis are related with each other. Design In the peripheral blood of diabetic patients suffering critical ischaemia associated with peripheral artery disease, low levels and poor function of circulating endothelial progenitor cells (EPCs) were observed. Moreover, circulating endothelial cells (CECs) have been described in different conditions of vascular injury. In this type of disorders, which are all characterized by endothelial damage, neoangiogenesis plays a key role. Materials In the study we recruited 22 diabetic and 16 non diabetic patients, all of them suffering PAOD and critical ischaemia; healthy subjects and multiorgan donors have also been considered like controls. Methods Histopathologic characterization was performed on arterial tissue samples under a light microscope. Flow cytofluorimetric analysis was used to quantify CECs in peripheral blood samples. "In situ" expression of the Vascular Endothelial Growth Factor (VEGF) and Metalloproteinase 9 (MMP-9) transcripts was quantified in a Real Time-PCR analysis. Circulating VEGF concentration was determined by an ELISA assay. Results Arterial wall from diabetic patients, compared with non diabetic subjects, revealed a higher incidence of serious lesions (60% vs 47%) and a lower number of capillaries (65% vs 87%). Mean number of CECs/ml was significantly increased in all patients, compared to healthy controls (p=0.001). Compared to healthy subjects, VEGF transcripts expression resulted significantly higher in diabetic patients and in all patients (p<0.05) and a similar result was obtained in the MMP-9 transcripts expression. Serum VEGF concentration was significantly increased in PAOD patients correlated with controls (p=0.0431). Conclusions Our study demonstrates that in all patients considered, probably, regressive phenomenons prevail on reparative ones, causing an inesorable and progressive degeneration of the vascular wall, worse by diabetes. The vascular damage can be monitored by determining CECs number and its severity and development are emphasized by the MMP-9 transcripts expression. The "in situ" VEGF increased expression seems to be the evidence of a parietal cells bid to induce local angiogenesis. This reparing mechanism could induce the EPCs mobilitation by means the release of VEGF from the arterial wall. The mechanism, however, is ineffective like demonstrated by the EPCs reduced number and activities observed in patients suffering PAOD and critical ischaemia.