Longitudinal and Transverse Wakefields Simulations and Studies in Dielectric-Coated Circular Waveguides

In recent years, there has been a growing interest and rapid experimental progress on the use of e.m. fields produced by electron beams passing through dielectric-lined structures and on the effects they might have on the drive and witness bunches. Short ultra-relativistic electron bunches can excite very intense wakefields, which provide an efficient acceleration through the dielectric wakefield accelerators (DWA) scheme with higher gradient than that in the conventional RF LINAC. These beams can also generate high power narrow band THz coherent Cherenkov radiation. These high gradient fields may create strong instabilities on the beam itself causing issues in plasma acceleration experiments (PWFA), plasma lensing experiments and in recent beam diagnostic applications. In this work we report the results of the simulations and studies of the wakefields generated by electron beams at different lengths and charges passing on and off axis in dielectric-coated circular waveguides. We also propose a semi-analytical method to calculate these high gradient fields without resorting to time consuming simulations

Correction of Mutant p63 in EEC Syndrome Using siRNA Mediated Allele-Specific Silencing Restores Defective Stem Cell Function

Ectrodactyly-Ectodermal dysplasia-Clefting (EEC) syndrome is a rare autosomal dominant disease caused by heterozygous mutations in the p63 gene and characterized by limb defects, orofacial clefting, ectodermal dysplasia, and ocular defects. Patients develop progressive total bilateral limbal stem cell deficiency, which eventually results in corneal blindness. Medical and surgical treatments are ineffective and of limited benefit. Oral mucosa epithelial stem cells (OMESCs) represent an alternative source of stem cells capable of regenerating the corneal epithelium and, combined with gene therapy, could provide an attractive therapeutic avenue. OMESCs from EEC patients carrying the most severe p63 mutations (p.R279H and p.R304Q) were characterized and the genetic defect of p.R279H silenced using allele-specific (AS) small interfering RNAs (siRNAs). Systematic screening of locked nucleic acid (LNA)-siRNAs against R279H-p63 allele in (i) stable WT-\u394Np63\u3b1-RFP and R279H-\u394Np63\u3b1-EGFP cell lines, (ii) transient doubly transfected cell lines, and (iii) p.R279H OMESCs, identified a number of potent siRNA inhibitors for the mutant allele, which had no effect on wild-type p63. In addition, siRNA treatment led to longer acquired life span of mutated stem cells compared to controls, less accelerated stem cell differentiation in vitro, reduced proliferation properties, and effective ability in correcting the epithelial hypoplasia, thus giving rise to full thickness stratified and differentiated epithelia. This study demonstrates the phenotypic correction of mutant stem cells (OMESCs) in EEC syndrome by means of siRNA mediated AS silencing with restoration of function. The application of siRNA, alone or in combination with cell-based therapies, offers a therapeutic strategy for corneal blindness in EEC syndrome

Observation of Azimuth-Dependent Suppression of Hadron Pairs in Electron Scattering Off Nuclei

We present the first measurement of dihadron angular correlations in electron-nucleus scattering. The data were taken with the CLAS detector and a 5.0 GeV electron beam incident on deuterium, carbon, iron, and lead targets. Relative to deuterium, the nuclear yields of charged-pion pairs show a strong suppression for azimuthally opposite pairs, no suppression for azimuthally nearby pairs, and an enhancement of pairs with large invariant mass. These effects grow with increased nuclear size. The data are qualitatively described by the gibuu model, which suggests that hadrons form near the nuclear surface and undergo multiple scattering in nuclei. These results show that angular correlation studies can open a new way to elucidate how hadrons form and interact inside nuclei

Conceptual Design of a Soft X‐ray SASE‐FEL Source

FELs based on SASE are believed to be powerful tools to explore the frontiers of basic sciences, from physics to chemistry to biology. Intense R&D programs have started in the USA and Europe in order to understand the SASE physics and to prove the feasibility of these sources. The allocation of considerable resources in the Italian National Research Plan (PNR) brought about the formation of a CNR‐ENEA‐INFN‐University of Roma "Tor Vergata" study group. A conceptual design study has been developed and possible schemes for linac sources have been investigated, bringing to the SPARX proposal. We report in this paper the results of a preliminary start to end simulation concerning one option we are considering based on an S‐band normal conducting linac with high brightness photoinjector integrated in a RF compressor

Advanced cell therapy strategies to correct corneal disorders

Introduction Epithelial homeostasis is guaranted by somatic stem cells wich, through different patways, such as p63 and Notch signaling, self-renew, differentiate and control tissue function and integrity. In vivo behavior of epithelial stem cells (ESCs) from different epithelia reflects differences in physiological role of the tissue. Limbal stem cells deficiency (LSCD) is characterized by conjunctival epithelial ingrowth, neovascularization, recurrent corneal erosion and persistent ulcers, as well as corneal scarring and ultimately leads to visual impairment and blindness. When the cornea is entirely covered with a fibro-vascular tissue, the chances of success of a traditional penetrating keratoplasty are virtually absent. Transplantation of autologous limbal epithelial stem cells (LESCs) or oral mucosa epithelial stem cells (OMESCs) cultured on fibrin glue has shown to be successful for unilateral or bilateral LSCD treatment, respectively. Despite that, fibrin presented several limitations including inability to repair or replace damaged corneal stroma. Aim The purpose of this work is to better understand the potentiality of somatic epithelial stem cells in order to identify possible approaches to exploit this potentiality. In the light of this, my research group has been able to identify and set up a cell therapy approach for a unique homozygous-heterozygous mosaicism of EEC Syndrome, demonstrating that epithelial stem cells have an intrinsic potential for regenerative medicine that can be exploited with a deeper characterization. We, also, aimed to test human keratoplasty lenticules (HKLs): particularly attractive, full-thickness scaffolds for corneal epithelial and stromal reconstruction that provides an interesting organotypic culture system for evaluation of growth, proliferation, and differentiation processes of epithelial stem cells (ESCs). Results During the three years of PhD I had the opportunity to collect and review cellular biology data of four diverse types of primary epithelial cells, derived from four different epithelia (skin, oral mucosa, limbus/cornea and conjunctiva) and, to better investigate the exhaustion of clonogenic potential and the self-renewal of epithelial stem cells we took in consideration also p63-defective oral mucosa primary cell lines obtained from three patients affected by EEC syndrome, already known to cause an acceleration in epithelial aging. Demonstrating that epithelial stem cells have an intrinsic potential for regenerative medicine that can be exploited with a deeper characterization, we have also been able to apply a cell therapy protocol to a patient affected by EEC syndrome in a rare form of mosaicism. We isolated the cell population with a mild phenotype from this patient, enriching these primary cells in vitro and producing well-organized and stratified epithelial sheets. The novelty and the importance of this case was related to the possibility to start a customized cell therapy approach for this unique case of EEC syndrome, based solely on epithelial stem cell manipulation. Limbal stem cells (H-LESCs) expanded onto HKLs gave rise to a keratinized stratified squamous epithelium morphologically similar to that of normal corneas. To set up the cohort of animal patients, we proceded with the characterization of cell lines obtained from biopsies of the cornea (C-CESCs) and oral mucosa (C-OMESCs) of canine origin. Primary lines were serially propagated until exhaustion in order to get life span data to compare their behavior with human limbal stem cells. For each passage, we also performed colony forming efficiency assays (CFE) to estimate the proportion of clonogenic cells present in the culture. Results showed a trend of canine cell lines comparable to human limbal cells trend, with a similar decrease of clonogenic cells number, a similar percentage of aborted colonies during serial cultivation and a similar replicative senescence. Then, primary human and canine epithelial stem cells were seeded onto HKLs. The resulting epithelia was well organized and stratified into four to five cell layers with basal cuboidal cells differentiating upward to winged cells. The layer of basal cuboidal cells was firmly attached to the underlying ECM and to the basement membrane through integrinβ4. Maintenance of stemness potential and differentiation pathways were assessed checking the expression of the stem cells marker p63 and the terminally differentiated cells marker Involucrine. Importantly, expression of the different markers resembled that observed in normal epithelia, thus suggesting that HKLs are able to support the growth and maintain the differentiation pathways of epithelial stem cells. Discussion and conclusions Our findings demonstrate that primary epithelial cells have unique characteristics with an inimitable potential that makes them a malleable tool, able to adapt to different necessities. And so on, although these data are intriguing, further investigation could provide more and more useful data for their clinical application in regenerative medicine. As a proof-of-principle that epithelial stem cells have an intrinsic potential for regenerative medicine that can be exploited with a deeper characterization, my group have been able to apply, in vitro, a cell therapy protocol to a patient affected by a rare mosaic form of EEC syndrome. HKLs appear to be particularly attractive, animal-free (feeder-free) and full-thickness scaffolds for corneal reconstruction. We have already started with the the recruitment of canine patients to assess the transplantability and functionality of these organotypic structures and, through the collaboration with different veterinary departments, we are creating a small cohort on which to start with first transplantation trials.Introduzione L'omeostasi epiteliale è garantita da cellule staminali somatiche che, attraverso diversi pathways, come quello di p63 o di Notch, si auto-rinnovano, differenziano e controllano la funzione e l'integrità del tessuto. Il comportamento in vivo delle cellule staminali epiteliali (ESC) di diversi epiteli riflette le differenze nel ruolo fisiologico che queste cellule hanno nel tessuto di origine. Il deficit di cellule staminali limbali (LSCD) è caratterizzato da crescita epiteliale congiuntivale, neovascolarizzazione, erosione corneale ricorrente e ulcere persistenti, nonché cicatrici corneali, e porta infine a deficit visivo e cecità. Quando la cornea è interamente coperta da un tessuto fibro-vascolare, le probabilità di successo di una cheratoplastica penetrante tradizionale sono praticamente assenti. Il trapianto di cellule staminali epiteliali autologhe del limbus (LESCs) o di cellule staminali epiteliali della mucosa orale (OMESCs) coltivate su scaffold di fibrina ha dimostrato di avere successo per il trattamento unilaterale o bilaterale di LSCD, rispettivamente. Nonostante ciò, la fibrina presentava numerose limitazioni tra cui l'incapacità di riparare o sostituire lo stroma corneale danneggiato. Scopo Lo scopo di questo lavoro è quello di comprendere meglio la potenzialità delle cellule staminali epiteliali somatiche al fine di identificare possibili approcci per sfruttare questa potenzialità. Alla luce di ciò, il mio gruppo di ricerca è stato in grado di identificare e impostare un approccio di terapia cellulare per un caso unico di mosaicismo omozigote-eterozigote della Sindrome EEC, dimostrando che le cellule staminali epiteliali hanno un potenziale intrinseco per la medicina rigenerativa che può essere sfruttata con una caratterizzazione più profonda. Un secondo scopo della tesi è stato quello di testare gli Human Keratoplasty Lenticules (HKL): scaffolds particolarmente attraenti per la ricostruzione a tutto spessore della superficie anteriore dell’occhio che fornisce, anche, un interessante sistema di coltura organotipica per la valutazione dei processi di crescita, proliferazione e differenziazione delle cellule staminali epiteliali (ESCs ). Risultati Durante i tre anni di dottorato ho avuto l'opportunità di raccogliere e rivedere i dati di biologia cellulare di quattro diversi tipi di cellule epiteliali primarie, derivate da quattro diversi epiteli (pelle, mucosa orale, limbus/cornea e congiuntiva) e, per studiare meglio l'esaurimento del potenziale clonogenico e dell'auto-rinnovamento delle cellule staminali epiteliali abbiamo preso in considerazione anche delle linee cellulari primarie di mucosa orale con difetti genetici del gene p63 già noti per causare un'accelerazione nell'invecchiamento epiteliale, ottenute da tre pazienti affetti da sindrome EEC. Dimostrando che le cellule staminali epiteliali hanno un potenziale intrinseco per la medicina rigenerativa che può essere sfruttato con una caratterizzazione più profonda, siamo stati anche in grado di applicare un protocollo di terapia cellulare a un paziente affetto da sindrome EEC in una rara forma di mosaicismo. Abbiamo isolato la popolazione cellulare con un fenotipo lieve da questo paziente, arricchendo queste cellule primarie in vitro e producendo foglietti epiteliali ben organizzati e stratificati. La novità e l'importanza di questo caso è legata alla possibilità di iniziare un approccio di terapia cellulare personalizzata per questo caso unico di sindrome EEC, basata esclusivamente sulla manipolazione delle cellule staminali epiteliali. Le cellule staminali limbari (H-LESC) coltivate su HKL hanno dato origine a un epitelio squamoso stratificato cheratinizzato morfologicamente simile a quello delle cornee normali. Per costituire la coorte di pazienti animali, abbiamo proceduto alla caratterizzazione di linee cellulari ottenute da biopsie della cornea (C-CESCs) e mucosa orale (C-OMESCs) di origine canina. Le linee primarie sono state propagate in serie fino ad esaurimento al fine di ottenere dati di life span per confrontare il loro comportamento con le cellule staminali limbari umane. Per ogni passaggio, abbiamo anche eseguito analisi di Colony Forming Efficiency (CFE) per stimare la proporzione di cellule clonogeniche presenti nella coltura. I risultati hanno mostrato una tendenza delle linee cellulari canine comparabile all'andamento delle cellule limbari umane, con una diminuzione simile del numero di cellule clonogeniche, una percentuale simile di colonie abortive durante i passaggi in coltura e una simile senescenza replicativa. Quindi, cellule staminali epiteliali primarie umane e canine sono state seminate su HKL. L'epitelio risultante era ben organizzato e stratificato in quattro o cinque strati cellulari con cellule cuboidali basali che differenziano le cellule verso l'alto e quelle alate. Lo strato di cellule cuboidali basali era saldamente attaccato alla matrice extracellulare sottostante e alla membrana basale attraverso l'integrina-4. Sono stati valutati il mantenimento del pool di cellule staminali e i corretti processi di differenziamento controllando l'espressione del marcatore di cellule staminali, p63, e del marcatore di cellule terminalmente differenziate, Involucrina. È importante sottolineare che l'espressione dei diversi marcatori assomiglia a quella osservata negli epiteli normali, suggerendo quindi che gli HKL sono in grado di supportare la crescita e il mantenimento di tessuti epiteliali ricostruiti. Discussione e Conclusioni I nostri risultati dimostrano che le cellule epiteliali primarie hanno caratteristiche uniche con un potenziale inimitabile che le rende uno strumento malleabile, in grado di adattarsi alle diverse necessità. Sebbene questi dati siano intriganti, ulteriori indagini potrebbero fornire dati sempre più utili per la loro applicazione clinica nella medicina rigenerativa. Come prova di principio che le cellule staminali epiteliali hanno un potenziale intrinseco per la medicina rigenerativa che può essere sfruttato con una caratterizzazione più profonda, il mio gruppo è stato in grado di applicare, in vitro, un protocollo di terapia cellulare a un paziente affetto da un raro mosaicismo di sindrome EEC. Le HKL sembrano essere scaffold particolarmente attraenti (animal-free) per la ricostruzione a tutto spessore della cornea. Abbiamo già iniziato con il reclutamento di pazienti canini per valutare la trapiantabilità e la funzionalità di queste strutture organotipiche e, attraverso la collaborazione con diversi dipartimenti veterinari, stiamo creando una piccola coorte sulla quale iniziare con le prime prove di trapianto

Simulations of coherent synchrotron radiation effects in electron machines

Coherent synchrotron radiation (CSR) generated by high intensity electron beams can be a source of undesirable effects limiting the performance of storage rings. The complexity of the physical mechanisms underlying the interplay between the electron beam and the CSR demands for reliable simulation codes. In the past, codes based on Lie algebraic techniques have been very efficient to treat transport problems in accelerators. The extension of these methods to the non linear case is ideally suited to treat wakefields - beam interaction. In this paper we report on the development of a numerical code, based on the solution of the Vlasov equation, which includes the non linear contribution due to wakefields. The proposed solution method exploits an algebraic technique that uses the exponential operators. We show that, in the case of CSR wakefields, the integration procedure is capable of reproducing the onset of an instability which leads to microbunching of the beam thus increasing the CSR at short wavelengths. In addition, considerations on the threshold of the instability for Gaussian bunches is also reported

A vlasov solver for collective effects in particle accelerators

Integration techniques based on Lie algebraic methods have been successfully used in beam transport codes for particle accelerators. Generally these methods have been applied to problems of single-particle beam dynamics. Here we present an application of Lie algebraic techniques to the development of a Vlasov solver suitable for problems of beam transport in the presence of non-negligible particle self-fields. The solver we discuss is suitable for modelling a variety of collective effects that may arise at high current. In particular, we consider the case of coherent synchrotron radiation effects in magnetic bunch compressors which can cause instabilities limiting performance of high current accelerators

A four-dimensional Vlasov solver for microbunching instability in the injection system for X-ray FELs

The micro-bunching instability (MBI) is seeded by small charge-density fluctuations in the electron bunches, and is successively amplified by the combined effect of space charge and coherent synchrotron radiation, as the beam travels through magnetic compressors. The quantitative understanding of this effect demands for accurate numerical simulations. Here we report on the progress of an upgrading of a 2D Vlasov solver code toward a 4D grid-based Vlasov solver, including also the transversedynamics. The goal is to provide an accurate characterization of the MBI seeded by random noise present in the bunch distribution. We also comment on the advantages of our procedure with respect to other approaches, e.g. macroparticle simulations