Processing and structural properties of random oriented lead lanthanum zirconate titanate thin films

Polycrystalline lead lanthanum zirconate titanate (PLZT) thin films have been prepared by a polymeric chemical route to understand the mechanisms of phase transformations and map the microstructure and elastic properties at the nanoscale in these films. X-ray diffraction, atomic force microscopy (AFM) and ultrasonic force microscopy (UFM) have been used as investigative tools. On one side, PLZT films with mixed-phase show that the pyrochlore phase crystallizes predominantly in the bottom film-electrode interface while a pure perovskite phase crystallizes in top film surface. On the contrary, pyrochlore-free PLZT films show a non-uniform microstrain and crystallite size along the film thickness with a heterogeneous complex grainy structure leading to different elastic properties at nanoscale

Clathrin-coated structures support 3D directed migration through local force transmission

Migrating cells navigate in complex environments through sensing and interpreting biochemical and/or mechanical cues. Here, we report that recently identified tubular clathrin/AP-2 lattices (TCALs), a subset of clathrin-coated structures (CCSs) that pinch collagen fibers, mechanically control directed migration along fibers decorated with ligands of CCS cargoes in three-dimensional (3D) environments. We observed that epidermal growth factor or low-density lipoprotein bound to collagen fibers leads to increased local nucleation and accumulation of TCALs. By using engineered, mixed collagen networks, we demonstrate that this mechanism selectively increases local forces applied on ligand-decorated fibers. We show that these effects depend on the ligand’s receptors but do not rely on their ability to trigger signaling events. We propose that the preferential accumulation of TCALs along ligand-decorated fibers steers migration in 3D environments. We conclude that ligand-regulated, local TCAL accumulation results in asymmetric force distribution that orients cell migration in 3D environments

Interlaboratory performance of a Real-Time PCR method for detection of Ceratocystis platani, the agent of canker stain of Platanus spp

Ceratocystis platani (CP), an ascomycetous fungus, is the agent of canker stain, a lethal vascular disease of Platanus species. Ceratocystis platani has been listed as a quarantine pest (EPPO A2 list) due to extensive damage caused in Southern Europe and the Mediterranean region. As traditional diagnostic assays are ineffective, a Real-Time PCR detection method based on EvaGreen, SYBR Green, and Taqman assays was previously developed, validated in-house, and included in the official EPPO standard PM7/14 (2). Here, we describe the results of a test performance study performed by nine European laboratories for the purpose of an interlaboratory validation. Verification of the DNA extracted from biological samples guaranteed the high quality of preparations, and the stability and the homogeneity of the aliquots intended for the laboratories. All of the laboratories reproduced nearly identical standard curves with efficiencies close to 100%. Testing of blind-coded DNA extracted from wood samples revealed that all performance parameters-diagnostic sensitivity, diagnostic specificity, accuracy and reproducibility-were best fit in most cases both at the laboratory and at the assay level. The previously established limit of detection, 3 fg per PCR reaction, was also validated with similar excellent results. The high interlaboratory performance of this Real-Time PCR method confirms its value as a primary tool to safeguard C. platani-free countries by way of an accurate monitoring, and to investigate the resistance level of potentially canker stain-resistant Platanus genotypes

Can Clinical and Surgical Parameters Be Combined to Predict How Long It Will Take a Tibia Fracture to Heal? A Prospective Multicentre Observational Study: The FRACTING Study

Background. Healing of tibia fractures occurs over a wide time range of months, with a number of risk factors contributing to prolonged healing. In this prospective, multicentre, observational study, we investigated the capability of FRACTING (tibia FRACTure prediction healING days) score, calculated soon after tibia fracture treatment, to predict healing time. Methods. The study included 363 patients. Information on patient health, fracture morphology, and surgical treatment adopted were combined to calculate the FRACTING score. Fractures were considered healed when the patient was able to fully weight-bear without pain. Results. 319 fractures (88%) healed within 12 months from treatment. Forty-four fractures healed after 12 months or underwent a second surgery. FRACTING score positively correlated with days to healing: r = 0.63 (p < 0.0001). Average score value was 7.3 \ub1 2.5; ROC analysis showed strong reliability of the score in separating patients healing before versus after 6 months: AUC = 0.823. Conclusions. This study shows that the FRACTING score can be employed both to predict months needed for fracture healing and to identify immediately after treatment patients at risk of prolonged healing. In patients with high score values, new pharmacological and nonpharmacological treatments to enhance osteogenesis could be tested selectively, which may finally result in reduced disability time and health cost savings

The Role of the Golgi Protein GM130 in Cell Polarity and Tumorigenesis

The Golgi apparatus is linked to the establishment of cell polarity, but the mechanism that allows the organelle to control cell polarity still remains unknown. Research in the area focused primarily on signaling from the plasma membrane to understand how polarity is established, and the small GTPase Cdc42 was identified as a main regulator of this process. Interestingly Cdc42 is mainly localized at the Golgi, thus we investigated the possibility that the Golgi regulates Cdc42 activity and by this mechanism it regulates polarity. We identified a GM130–RasGRF complex as a regulator of Cdc42 at the Golgi. Silencing GM130 results in RasGRF-dependent inhibition of the Golgi pool of Cdc42, but does not affect Cdc42 at the cell surface. Therefore, this is a specific mechanism to control a spatially restricted pool of Cdc42. Furthermore, active Cdc42 at the Golgi is important to sustain asymmetric front–rear Cdc42-GTP distribution in directionally migrating cells. We propose that the Golgi delivers active Cdc42 to the leading edge of migrating cells, thereby establishing asymmetry in Cdc42 activation at the plasma membrane and promoting directional migration. Two further observations supported a possible role for GM130 in cancer. First, concurrent to Cdc42 inhibition, silencing GM130 also results in RasGRF-dependent Ras-ERK pathway activation. Second, depletion of GM130 is sufficient to induce E-cadherin downregulation, indicative of a loss in cell polarity. We found that GM130 expression is frequently lost in colorectal and breast cancer patients. Whether the loss of GM130 solely affects polarity, or whether it affects other processes relevant for tumorigenesis remains unclear. To further investigate the role of GM130 in cancer, we analyzed the effect of GM130 depletion in a panel of breast cancer cells lines looking at processes linked to tumor progression such as survival, proliferation, adhesion, migration and invasion. We show that depletion of GM130 does not drastically affect survival, proliferation and adhesion. However, GM130 depleted cells show increased cellular velocity and increased invasiveness though matrigel, therefore supporting the view that alterations of polarity contribute to tumor progression.These findings establish a previously unrecognized role for a GM130–RasGRF–Cdc42 connection in regulating polarity and tumorigenesis

Studio e valutazione del rischio biologico legato alla diffusione di virus enterici e respiratori in ambito ospedalero

I virus sono gli agenti responsabili del 60% delle infezioni umane a livello mondiale. Le più comuni malattie virali sono causate da virus enterici e respiratori. Questi tipi di virus possono essere trasmessi attraverso molteplici vie, dal contatto diretto con superfici, oggetti o persone contaminate, all’ingestione di cibi, all’esposizione ad aerosol contenente particelle virali. Attività che possono portare alla formazione di bioaerosol sono starnuti, vomito, scarico del bagno, ecc. ecc. Il monitoraggio ambientale è un mezzo essenziale per conoscere le vie di diffusione e valutarne l’importanza ai fini dell’analisi del rischio. La ricerca virologica ambientale presenta difficoltà nell’individuazione di virus, in quanto questi sono presenti solo saltuariamente e, nella maggior parte dei casi, a concentrazioni estremamente ridotte. È perciò necessario utilizzare tecniche di biologia molecolare ad alta sensibilità quali PCR qualitative e quantitative (TaqMan PCR). Lo scopo della tesi è stato quindi lo studio della diffusione di virus a trasmissione enterica (in particolare), prendendo come ambienti di riferimento i bagni di strutture ospedaliere. È stato eseguito un monitoraggio ambientale prima all’interno di ambienti ad uso ufficio per valutare l’efficacia delle tecniche utilizzate e successivamente in ambito ospedaliero con lo scopo di effettuare un risk assessment. Il bagno è stato classificato come punto critico di controllo per la sua caratteristica di essere un luogo condiviso da più persone che lo rende un possibile punto di contatto con agenti di rischio. Sono stati eseguiti campionamenti di superfici per mezzo di tamponi e di aerosol (SAS: Surface Air System) alla ricerca di virus a DNA (TTV e adenovirus) e ad RNA (norovirus, rotavirus, enterovirus, rhinovirus). Associata alla ricerca virologica è stata condotta la ricerca di indicatori normalmente utilizzati nell’analisi del rischio: carica batterica totale e enterobatteri, presenza di colifagi somatici. Per quanto riguarda i batteri, è stata effettuata la semina su piastre utilizzando PCA (plate count agar) per la CBT e MacConkey per gli enterobatteri. In caso di crescita di colonie atipiche sono stati usati terreni selettivi per l’identificazione particolare. Per i colifagi è stata utilizzata la tecnica UNI EN ISO 10705-2:2001. Dai tamponi superficiali sono stati estratti gli acidi nucleici per mezzo dei kit commerciali Qiagen QIAamp viral RNA e Promega IQ System per DNA. Per mezzo del SAS è stata aspirata aria su una piastra di TSA (Trypticase Soy Agar) che è stata poi eluita utilizzando BE (Beef Extract) pH 9. Sull’eluato sono state applicate le tecniche di estrazione degli acidi nucleici già citate. I campioni risultati positivi sono stati sottoposti a sequenziamento per confermare la presenza del virus indagato. Nei locali ad uso ufficio, su un totale di 64 campioni superficiali, il 67% è risultato positivo per almeno un virus. Di questi, l’86% è risultato contaminato da adenovirus ed il 9% da TTV; nel 5% dei casi è stata ritrovata una doppia contaminazione di adenovirus+TTV. Su un totale di 16 campioni di aerosol, il 62% è risultato positivo per almeno un virus. Di questi, il 70% è risultato positivo per adenovirus, il 20% per TTV e nel 10% dei casi è stata riscontrata una doppia contaminazione. In ambito ospedaliero il 67% dei campioni di superfici è risultato positivo per almeno un virus. Di questi il 41% è risultato positivo per adenovirus ed il 17% per TTV; nel 41% dei campioni è stata registrata una doppia contaminazione di adenovirus e TTV. In un solo caso è stato ritrovato norovirus. Per quanto riguarda i campioni di aerosol, il 68% è risultato contaminato. L’83% dei campioni è risultato contaminato da adenovirus, il 13% da TTV e si è registrata una doppia infezione (adenovirus+TTV) nel 4% dei campioni. Non è stata riscontrata correlazione statisticamente significativa tra presenza/assenza di virus e carica batterica o presenza/assenza di colifagi. Sono stati ritrovati alti livelli di adenovirus anche nelle acque degli scarichi dei bagni ed il sequenziamento ha confermato che nella maggior parte dei casi si ritrova lo stesso tipo di adenovirus sia nelle acque che sulle superfici. La contaminazione dell’aerosol è generalmente maggiore della contaminazione delle superfici, dato riscontrato anche in letteratura ed imputabile all’effetto dei trattamenti di disinfezione che vengono utilizzati per le superfici. È stato effettuato un tentativo di calcolare il rischio puntiforme di contaminazione derivante dall’uso dei bagni, attraverso l’uso di equazioni che descrivono il processo di contaminazione. In questo modo, è stato calcolato un rischio derivante dal contatto con le superfici del bagno del 48,8% ed un rischio dovuto alla respirazione di aerosol dell’1,3%. Questo studio è da considerare un punto di partenza per effettuare una QMRA volta a rispondere alle domande che restano aperte riguardo al vero e proprio rischio di infezione

Endomembranes: Unsung Heroes of Mechanobiology?

Mechanical stimuli have profound effects on the cellular architecture and functions. Over the past two decades, considerable progress has been made in unraveling the molecular machineries that confer cells the ability to sense and transduce mechanical input into biochemical signals. This has resulted in the identification of several force-sensing proteins or mechanically activated ion channels distributed throughout most cell types, whereby the plasma membrane, cytoskeleton, and the nucleus have garnered much attention. Although organelles from the endomembrane system make up significant portion of cell volume and play pivotal roles in the spatiotemporal distribution of signaling molecules, they have received surprisingly little attention in mechanobiology. In this mini-review, we summarize results that document participation of the endomembrane system in sensing and responding to mechanical cues

Crosstalk of small GTPases at the Golgi apparatus

Small GTPases regulate a wide range of homeostatic processes such as cytoskeletal dynamics, organelle homeostasis, cell migration and vesicle trafficking, as well as in pathologic conditions such as carcinogenesis and metastatic spreading. Therefore, it is important to understand the regulation of small GTPase signaling, but this is complicated by the fact that crosstalk exists between different GTPase families and that we have to understand how they signal in time and space. The Golgi apparatus represents a hub for several signaling molecules and its importance in this field is constantly increasing. In this review we will discuss small GTPases signaling at the Golgi apparatus. Then, we will highlight recent work that contributed to a better understanding of crosstalk between different small GTPase families, with a special emphasis on their crosstalk at the Golgi apparatus. Finally, we will give a brief overview of available methods and tools to investigate spatio-temporal small GTPase crosstalk

Mobile Agents for QoS Tailoring, Control and Adaptation over the Internet: the ubiQoS Video on Demand Service

Service provision over the Internet has to address the issues of differentiated Quality-ofService (QoS) and ubiquitous accessibility. Internet services should take into consideration the user QoS desiderata together with the various properties of servers providing replicated/partitioned services and of different access devices/points, from workstations over high-capacity ATM links to personal digital assistants over packetswitched 3G mobile phone. The major paper claim is that the provision over best-effort networks of services with negotiated and controlled QoS requires a distributed support infrastructure consisting of intermediate active nodes along the path between clients and servers. The paper deals with the Mobile Agent (MA) technology as suitable for implementing this active infrastructure and, in particular, presents the MA-based design and implementation of the ubiQoS Video on Demand (VoD) middleware. At negotiation time, ubiQoS establishes an active path between the requesting client and the proper VoD server to tailor the quality of VoD flow depending on user profile and device characteristics. At provision time, ubiQoS dynamically controls the offered QoS level to adapt locally when and where network resource availability changes