Characterization of L-type Calcium Channel Binding-Site of a new class of Calcium modulators by a Multidisciplinary approach

Many potential diltiazem related L-VDCC blockers were developed using a multidisciplinary approach. This current study was to investigate and compare diltiazem with to the newly developed compounds by mouse Langendorff-perfused heart, Ca2+-transient and on recombinant L-VDCC. Twenty particular compounds were selected by the ligand-based virtual screening procedure (LBVS). From these compounds, five of them (5b, M2, M7, M8 and P1) showed a potent and selective inotropic activity on guinea-pig left atria driven 1 Hz. Further assays displayed an interesting negative inotropic effect of M2, M8, P1 and M7 on guinea pig isolated left papillary muscle driven at 1 Hz, a relevant vasorelaxant activity of 5b, M2, M7, M8 and P1 on K+-depolarized guinea-pig ileum longitudinal smooth muscle and a significant inhibition of contraction of 5b, M2, M8 and P1 on carbachol stimulated ileum longitudinal smooth muscle. Wild-type human heart and rabbit lung α1 subunits were expressed (combined with the regulatory α2δ and β3 subunits) in Xenopus Leavis oocytes using a two-electrode voltage clamp technique. Diltiazem is a benzothiazepine Ca2+ channel blocker used clinically for its antihypertensive and antiarrhythmic effects. Previous radioligand binding assays revealed a complex interaction with the benzothiazepine binding site for M2, M7 and M8. (Carosati E. et al. J. Med Chem. 2006, 49; 5206). In agreement with this findings, the relative order of increased rates of contraction and relaxation at lower concentrations s(≤10-6M) in unpaced hearts was M7>M2>M8>P1. Similar increases in Ca2+ transient were observed in cardiomyocytes. Diltiazem showed negative inotropic effects whereas 5b had no significant effect. Diltiazem blocks Ca2+current in a use-dependent manner and facilitates the channel by accelerating the inactivation and decelerating the recovery from inactivation. In contrast to diltiazem, the new analogs had no pronounced use-dependence. Application of 100 μM M8, M2 showed ~ 10% tonic block; in addition, M8, M2 and P1 shifted the steady state inactivation in hyperpolarized direction and the current inactivation time was significantly decreased compared with control (219.6 ± 11.5 ms, 226 ± 14.5 vs. 269 ± 12.9 vs. 199.28 ± 8.19 ms). Contrary to diltiazem, the recovery from the block by M8 and M2 was comparable to control. Only P1 showed a significantly decrease of the time for the recovery from inactivation. All of the compounds displayed the same sensitivity on the Ca2+ channel rabbit lung α1 except P1. Taken together, these findings suggest that M8, M2 and P1 might directly decrease the binding affinity or allow rapid dissociation from the benzothiazepine binding site.

A new class of bragg stacks and its principle application

This work is focused on the development of high quality nanoporous 1D photonic crystals –so called Bragg stacks – made by spin-coating of approximately 25 nm large SiO2 and TiO2 nanoparticles bearing interparticle voids large enough to infiltrate reactive species. Therefore, the first part of this work describes the synthesis of well-dispersed TiO2 nanoparticles in this size range (the corresponding SiO2 nanoparticles are commercially available). In the second part, a protocol was developed to prepare nanoporous Bragg stacks of up to 12 bilayers with high quality and precision. Tailor-made Bragg stacks were prepared for different applications such as (i) a surface emitting feedback laser with a FWHM of only 6 nm and (ii) an electrochromic device with absorption reversibly switchable by an external electrical bias independently of the Bragg reflection. In the last chapter, the approach to 1D photonic crystals is transferred to 1D phononic crystals. Contrast in the modulus is achieved by spin-coating SiO2 and PMMA as high and low moduli material. This system showed a band gap of fg = 12.6 GHz with a width of Dfg/fg = 4.5 GHz.

Middleware per Internet of Things: Java Embedded come caso di studio

Grazie al progresso dell'elettronica, ai giorni nostri, è possibile costruire dispositivi elettronici molto piccoli, che col passare del tempo lo sono sempre più. Questo ci permette di poter imboccare nuove strade nel mondo dell'informatica, sfruttando proprio questo fatto. Le dimensioni ridotte dei dispositivi in commercio, come sensori, attuatori, tag e tanto altro, sono particolarmente adatte a nuovi scenari applicativi. Internet of Things è una visione in cui Internet viene esteso alle cose. Facendo largo uso di dispositivi come sensori e tag è possibile realizzare sistemi intelligenti che possono avere riscontri positivi nella vita di tutti i giorni. Tracciare la posizione degli oggetti, monitorare pazienti da remoto, rilevare dati sull'ambiente per realizzare sistemi automatici (ad esempio regolare automaticamente la luce o la temperatura di una stanza) sono solo alcuni esempi. Internet of Things è la naturale evoluzione di Internet, ed è destinato a cambiare radicalmente la nostra vita futura, poichè la tecnologia sarà sempre più parte integrante della nostra vita, aumentando sempre più il nostro benessere e riducendo sempre più il numero delle azioni quotidiane da compiere. Sempre più sono middleware, le piattaforme e i sistemi operativi che nascono per cercare di eliminare o ridurre le problematiche relative allo sviluppo di sistemi di questo genere, e lo scopo di questa tesi è proprio sottolinearne l'importanza e di analizzare gli aspetti che questi middleware devono affrontare. La tesi è strutturata in questo modo: nel capitolo uno verrà fatta una introduzione a Internet of Things, analizzando alcuni degli innumerevoli scenari applicativi che ne derivano, insieme però alle inevitabili problematiche di tipo tecnologico e sociale. Nel secondo capitolo verranno illustrate le tecnologie abilitanti di Internet of Things, grazie alle quali è possibile realizzare sistemi intelligenti. Nel terzo capitolo verranno analizzati gli aspetti relativi ai middleware, sottolineandone l'importanza e prestando attenzione alle funzioni che devono svolgere, il tutto riportando anche degli esempi di middleware esistenti. Nel quarto capitolo verrà approfondito il middleware Java Embedded di Oracle.

Estensione del framework kura per internet of things tramite supporto distribuito coap

Realizzazione di un supporto CoAP per il framework Kura con le seguenti caratteristiche: 1. Ottima scalabilità, ad organizzazione gerarchica, con aggiunta e rimozione dinamica di nodi e gestione automatica delle disconnessioni. 2. Integrazione efficiente di tecnologie CoAP ed MQTT progettate appositamente per l’IoT tramite lo sviluppo di un pattern di comunicazione per la gestione degli scambi delle informazioni. 3. Un limitato uso di risorse con modifiche su entrambe le implementazioni standard dei protocolli usati in modo tale da adattarle agli obiettivi prefissati. Il tutto a un costo bassissimo, dato che si basa su tecnologie open e grazie alla possibilità di utilizzo su Raspberry Pi.

Piattaforme per Internet of Things: Windows IoT Core come caso di studio

Questa tesi si pone l'obiettivo di esplorare alcuni aspetti di uno dei settori più in crescita in questi anni (e nei prossimi) in ambito informatico: \textbf{Internet of Things}, con un occhio rivolto in particolar modo a quelle che sono le piattaforme di sviluppo disponibili in questo ambito. Con queste premesse, si coglie l'occasione per addentrarsi nella scoperta della piattaforma realizzata e rilasciata da pochi mesi da uno dei colossi del mercato IT: Microsoft. Nel primo capitolo verrà trattato Internet of Things in ambito generale, attraverso una panoramica iniziale seguita da un'analisi approfondita dei principali protocolli sviluppati per questa tecnologia. Nel secondo capitolo verranno elencate una serie di piattaforme open source disponibili ad oggi per lo sviluppo di sistemi IoT. Dal terzo capitolo verrà incentrata l'attenzione sulle tecnologie Microsoft, in particolare prima si tratterà Windows 10 in generale, comprendendo \emph{UWP Applications}. Di seguito, nel medesimo capitolo, sarà focalizzata l'attenzione su Windows IoT Core, esplorandolo dettagliatamente (Windows Remote Arduino, Modalità Headed/Headless, etc.). Il capitolo a seguire concernerà la parte progettuale della tesi, comprendendo lo sviluppo del progetto \textbf{Smart Parking} in tutte le sue fasi (dei Requisiti fino ad Implementazione e Testing). Nel quinto (ed ultimo) capitolo, saranno esposte le conclusioni relative a Windows IoT Core e i suoi vantaggi/svantaggi.

Radiolabeled bicyclic somatostatin-based analogs: a novel class of potential radiotracers for SPECT/PET of neuroendocrine tumors

A variety of radiolabeled somatostatin analogs have been developed for targeting of somatostatin receptor (sst)-positive tumors. Bicyclic somatostatin-based radiopeptides have not been studied yet. Hypothesizing that the introduction of conformational constraints may lead to receptor subtype selectivity or may help to delineate structural features determining pansomatostatin potency, we developed and evaluated first examples of this new class of potential radiotracers for imaging or therapy of neuroendocrine tumors.

Prognostic relevance of Bcl-2 overexpression in surgically treated prostate cancer is not caused by increased copy number or translocation of the gene

Overexpression of anti-apoptotic Bcl-2 plays a role in prostate cancer progression, particularly in transformation to androgen-independent disease. Androgen-independent prostate cancers have been shown to harbor Bcl-2 gene copy number gains frequently suggesting that this genetic alteration might play a role in Bcl-2 overexpression. The relation of Bcl-2 overexpression and copy number gains or translocation of the BCL-2 gene in prostate cancer under hormone-naïve conditions is unknown.

Mechanisms of osteoclastogenesis inhibition by a novel class of biphenyl-type cannabinoid CB(2) receptor inverse agonists

The cannabinoid CB(2) receptor is known to modulate osteoclast function by poorly understood mechanisms. Here, we report that the natural biphenyl neolignan 4'-O-methylhonokiol (MH) is a CB(2) receptor-selective antiosteoclastogenic lead structure (K(i) < 50 nM). Intriguingly, MH triggers a simultaneous G(i) inverse agonist response and a strong CB(2) receptor-dependent increase in intracellular calcium. The most active inverse agonists from a library of MH derivatives inhibited osteoclastogenesis in RANK ligand-stimulated RAW264.7 cells and primary human macrophages. Moreover, these ligands potently inhibited the osteoclastogenic action of endocannabinoids. Our data show that CB(2) receptor-mediated cAMP formation, but not intracellular calcium, is crucially involved in the regulation of osteoclastogenesis, primarily by inhibiting macrophage chemotaxis and TNF-α expression. MH is an easily accessible CB(2) receptor-selective scaffold that exhibits a novel type of functional heterogeneity.

Structural basis for the regulation of cysteine-protease activity by a new class of protease inhibitors in Plasmodium

Plasmodium cysteine proteases are essential for host-cell invasion and egress, hemoglobin degradation, and intracellular development of the parasite. The temporal, site-specific regulation of cysteine-protease activity is a prerequisite for survival and propagation of Plasmodium. Recently, a new family of inhibitors of cysteine proteases (ICPs) with homologs in at least eight Plasmodium species has been identified. Here, we report the 2.6 A X-ray crystal structure of the C-terminal, inhibitory domain of ICP from P. berghei (PbICP-C) in a 1:1 complex with falcipain-2, an important hemoglobinase of Plasmodium. The structure establishes Plasmodium ICP as a member of the I42 class of chagasin-like protease inhibitors but with large insertions and differences in the binding mode relative to other family members. Furthermore, the PbICP-C structure explains why host-cell cathepsin B-like proteases and, most likely, also the protease-like domain of Plasmodium SERA5 (serine-repeat antigen 5) are no targets for ICP.

Identification of a host cell target for the thiazolide class of broad-spectrum anti-parasitic drugs

The thiazolide nitazoxanide (NTZ) and some derivatives exhibit considerable in vitro activities against a broad range of parasites, including the apicomplexans Neospora caninum and Toxoplasma gondii tachyzoites. In order to identify potential molecular targets for this compound in both parasites, RM4847 was coupled to epoxy-agarose and affinity chromatography was performed. A protein of approximately 35 kDa was eluted upon RM4847-affinity-chromatography from extracts of N. caninum-infected human foreskin fibroblasts (HFF) and non-infected HFF, but no protein was eluted when affinity chromatography was performed with T. gondii or N. caninum tachyzoite extracts. Mass spectrometry analysis identified the 35 kDa protein as human quinone reductase NQO1 (P15559; QR). Within 8h after infection of HFF with N. caninum tachyzoites, QR transcript expression levels were notably increased, but no such increase was observed upon infection with T. gondii tachyzoites. Treatment of non-infected HFF with RM4847 did also lead to an increase of QR transcript levels. The enzymatic activity of 6-histidine-tagged recombinant QR (recQR) was assayed using menadione as a substrate. The thiazolides NTZ, tizoxanide and RM4847 inhibited recQR activity on menadione in a concentration-dependent manner. Moreover, a small residual reducing activity was observed when these thiazolides were offered as substrates.