The determination of skeletal loading conditions in vivo and their relationship to the health of bone tissues, remain an open question. Computational modeling of the musculoskeletal system is the only practicable method providing a valuable approach to muscle and joint loading analyses, although crucial shortcomings limit the translation process of computational methods into the orthopedic and neurological practice. A growing attention focused on subject-specific modeling, particularly when pathological musculoskeletal conditions need to be studied. Nevertheless, subject-specific data cannot be always collected in the research and clinical practice, and there is a lack of efficient methods and frameworks for building models and incorporating them in simulations of motion. The overall aim of the present PhD thesis was to introduce improvements to the state-of-the-art musculoskeletal modeling for the prediction of physiological muscle and joint loads during motion. A threefold goal was articulated as follows: (i) develop state-of-the art subject-specific models and analyze skeletal load predictions; (ii) analyze the sensitivity of model predictions to relevant musculotendon model parameters and kinematic uncertainties; (iii) design an efficient software framework simplifying the effort-intensive phases of subject-specific modeling pre-processing. The first goal underlined the relevance of subject-specific musculoskeletal modeling to determine physiological skeletal loads during gait, corroborating the choice of full subject-specific modeling for the analyses of pathological conditions. The second goal characterized the sensitivity of skeletal load predictions to major musculotendon parameters and kinematic uncertainties, and robust probabilistic methods were applied for methodological and clinical purposes. The last goal created an efficient software framework for subject-specific modeling and simulation, which is practical, user friendly and effort effective. Future research development aims at the implementation of more accurate models describing lower-limb joint mechanics and musculotendon paths, and the assessment of an overall scenario of the crucial model parameters affecting the skeletal load predictions through probabilistic modeling.

Wearable inertial and magnetic measurements units (IMMU) are an important tool for underwater motion analysis because they are swimmer-centric, they require only simple measurement set-up and they provide the performance results very quickly. In order to estimate 3D joint kinematics during motion, protocols were developed to transpose the IMMU orientation estimation to a biomechanical model. The aim of the thesis was to validate a protocol originally propositioned to estimate the joint angles of the upper limbs during one-degree-of-freedom movements in dry settings and herein modified to perform 3D kinematics analysis of shoulders, elbows and wrists during swimming. Eight high-level swimmers were assessed in the laboratory by means of an IMMU while simulating the front crawl and breaststroke movements. A stereo-photogrammetric system (SPS) was used as reference. The joint angles (in degrees) of the shoulders (flexion-extension, abduction-adduction and internal-external rotation), the elbows (flexion-extension and pronation-supination), and the wrists (flexion-extension and radial-ulnar deviation) were estimated with the two systems and compared by means of root mean square errors (RMSE), relative RMSE, Pearson’s product-moment coefficient correlation (R) and coefficient of multiple correlation (CMC). Subsequently, the athletes were assessed during pool swimming trials through the IMMU. Considering both swim styles and all joint degrees of freedom modeled, the comparison between the IMMU and the SPS showed median values of RMSE lower than 8°, representing 10% of overall joint range of motion, high median values of CMC (0.97) and R (0.96). These findings suggest that the protocol accurately estimated the 3D orientation of the shoulders, elbows and wrists joint during swimming with accuracy adequate for the purposes of research. In conclusion, the proposed method to evaluate the 3D joint kinematics through IMMU was revealed to be a useful tool for both sport and clinical contexts.

The high long-term failure rate of synthetic vascular grafts in the replacement of small vessels is known to be associated with the lack of physiological signals to vascular cells causing adverse hemodynamic, inflammatory or coagulatory events. Current studies focus on developing engineered vascular devices with ability of directing cell activity in vitro and in vivo for tissue regeneration. It is also known that controlled molecule release from scaffolds can dramatically increase the scaffold ability of directing cell activities in vitro and in vivo for tissue regeneration. To address the mechanical and biological problems associated with graft materials, we demonstrated a degradable polyester-fibroin composite tubular scaffolds which shows well-integrated nanofibrous structure, endothelial-conducive surface and anisotropic mechanical property, suitable as engineered vascular constructs. Tissue regeneration needs not only functional biomolecules providing signaling cues to cells and guide tissue remodeling, but also an adequate modality of molecule delivery. In fact, healthy tissue formation requires specific signals at well-defined place and time. To develop scaffolds with multi-modal presentation of biomolecules, we patterned electrospun nanofibers over the thickness of the 3-dimensional scaffolds by programming the deposition of interpenetrating networks of degradable polymers poly(ε-caprolactone) and poly(lactide-co-glycolide) acid in tailored proportion. Fluorescent model molecules, drug and growth factors were embedded in the polymeric fibers with different techniques and release profiles were obtained and discussed. Fabrication process resulted in precise gradient patterns of materials and functional biomolecules throughout the thickness of the scaffold. These graded materials showed programmable spatio-temporal control over the release. Molecule release profiles on each side of the scaffolds were used to determine the separation efficiency of molecule delivery, which achieved >90% for proteins in 200µm scaffolds. Gradient-patterned scaffolds were also used to program simultaneous release of two proteins to the opposite sides of the scaffold and sequential release of proteins to a defined space, which further demonstrate the ability of patterned nanofibers to spatially and temporally confine sustained release. Moreover, results showed that temporal release kinetics could be altered by the structural patterns. Thus, the hierarchically-structured scaffolds presented here may enable development of novel multifunctional scaffolds with defined 3D dynamic microenvironments for tissue regeneration.

The term “theranostics” defines the effort to develop individualized therapies by the combination of diagnostic and therapeutic functions in the same agent. Gold-magnetite hybrid nanoparticles (H-NPs) are proposed as innovative theranostic nanotools for imaging-guided radiosensitization of cancers. H-NPs are designed to exert a dual function: (i) to provide contrast enhancement in magnetic resonance imaging (MRI).and (ii) to enhance radiation effects in the cancer. The imaging and the radiosensitization potentialities of H-NPs arise from the superparamagnetic behaviour of magnetite and the large x-ray extinction coefficient of gold, respectively. Hybrid nanoparticles allows cancer theranostics as the biodistibution of nanoparticles can be tracked by MR imaging, providing a real-time picture of the cancer radiosensitivity profile and allowing precise modulation of radiotherapy. The purposes of this work are to synthetize properly designed gold-magnetite hybrid nanoparticles and to provide preliminary in vitro evaluations about the potentialities of nanoparticles as MRI-contrast agents and radiosensitizers. A novel method for the synthesis of hydrophilic and superparamagnetic Tween20-stabilized dumbbell-like gold-magnetite hybrid nanoparticles was set up. Morphology and chemical composition of nanoparticles were assessed by transmission electron microscopy, x-ray diffraction analysis and ion-coupled plasma optical emission spectroscopy. Colloidal stability and magnetic properties of nanoparticles were determined by dynamic light scattering and alternating field magnetometer. The potentialities of H-NPs for MR imaging were studied using a human 4T-MRI scanner. Nanoparticles were proven to induce concentration-dependent contrast enhancement in T2*-weighted MR-images. The biosafety, the cellular uptake and the radiosensitization activity of H-NPs were investigated in human osteosarcoma MG63 cell cultures and murine 3T3 fibroblasts, using specific bioassays and laser scanning confocal microscopy. The results evidenced that nanoparticles were taken up by cells without inducing any cytotoxic effects, even at high nanoparticle concentration. In addition, nanoparticles were proven to induce osteosarcoma-specific reduction of cell viability in clonogenic cell cultures treated with radiotherapy. The experimental results confirmed the potentialities of H-NPs as theranostic tools for MRI-guided radiosensitization. Further studies are needed to confirm our findings and to identify other potential biological targets for MRI-guided radiosensitization.

Cardiovascular disease (CVD) is the most common cause of death in the United States of America, accounting for 24% of all deaths each year,(Anderson et al., 2003) and is projected to rise above 20% globally by 2030.(Mathers and Loncar, 2006) Options for CVD treatment do exist, but are limited by availability of healthy replant tissue from the patient or long term effectiveness and failure rates of both autologous tissue grafts and artificial implants. Grafting failure may often be attributed to the poor mimicry of the site-specific, healthy arterial tissue. While much TEVG research focuses on endothelialization of the graft lumen through chemical signaling, mechanotransduction plays a large role in forming and maintaining a healthy endothelial cell (EC) monolayer. Arteries and grafts interact with hemodynamics to determine flow pulsatility and create healthy, or pathological, mechanical signaling environments. Though arterial tissue is known to be viscoelastic,(Armentano et al., 2006; Bergel, 1961; Bia et al., 2006) the importance of this in developing healthy blood flow is undetermined. Therefore, a gap in the knowledge occurs in the importance of arterial mechanics affecting graft outcomes. To address this we attempt to examine specific shortcomings: 1) Determine whether pathological flow is capable of maintaining EC monolayer in a low arterial compliance model, 2) Establish methods of catering protein hydrogel frequency-dependent properties towards establishing biodegradable materials intended for TEVG, 3) Determine benefits of viscous wall damping in improving hemodynamics towards improved cell response. This proposal centers on improving cell response to pathological hemodynamics through catered viscoelastic material response at the arterial wall. To address this, we hypothesize that maintaining healthy EC monolayer is predicated on hemodynamic mechanotransduction, which results from both graft compliance and viscous damping of the material. To validate this hypothesis, we examine healthy and pathological hemodynamic effects on EC monoculture, and systematically determine the role of viscoelastic material response in maintaining healthy hemodynamics.

The increasing number of lipophilic drug candidates in development in the pharmaceutical industry calls for advanced drug delivery systems with increased bioavailability less day-to-day and food-intake-dependent. Many of these drug candidates possess poor water solubility, so that their dissolution rate in the gastrointestinal tract (GIT) limits their absorption following oral administration. In the past few decades, various lipid-based formulations have been investigated to enhance the bioavailability of such challenging drug candidates and to increase their clinical efficacy when administered orally. Recently, self-emulsifying drug delivery systems (SEDDS) have attracted increasing interests and, in particular, self-nanoemulsifying drug delivery systems (SNEDDS). SEDDS and SNEDDS consist in micro- or nano-emulsions of oil containing the drug that spontaneously form in aqueous media on mild agitation. Usually, they use high amounts of surfactant that may cause degradation and instability of the drugs, being moreover toxic for the gastrointestinal tract. The aim of the present thesis was the preparation of novel self-nanoemulsifying drug delivery systems to overcome the shortages of conventional SEDDS or SNEDDS. To reduce the amount of surfactant, we formulated first a self-nanoemulsifying drug delivery system containing high proportion of essential lemon oil, that was characterized in terms of drug solubility, formulation stability, viscosity, emulsion droplet size, ζ-potential and in vitro drug release. Then, a pH-sensitive SNEDDS was developed that emulsify only at basic pHs. The goal was to protect the lipophilic drugs from the harsh acidic environment in stomach and render it available in the enteric tract where the bioactive compound should be absorbed.

Tissue engineering is an interdisciplinary field aimed to design and engineer an efficient system for tissue and organ regeneration, for instance, for bone healing, based on the combined use of scaffolds, cells, bioactive or signalling molecules. An optimal tissue engineering procedure requires materials and scaffolds fulfilling several requirements, one of those being the ability to trigger and control the crosstalk with the biological environment both in vitro and in vivo, and to induce and control the extracellular matrix production and assembling. Diatomite is one of the most abundant natural sources of hydrated amorphous silica resulting from the accumulation of diatom skeletons. Diatoms possess particular features in structure, morphology as well as composition. Interestingly, it has been recognized that the formation process of diatom skeleton is possibly related to that of human bone. In this study, we wanted to utilize diatoms as silicon donor additives in scaffolds for bone tissue engineering, having been demonstrated the important role of silicon in bone formation. In this first part of the project, we used several methods to eliminate impurities in the raw diatomite. Diatom microparticles (DMPs) and nanoparticles (DNPs) were successfully produced by fragmentation of purified diatoms under alkaline condition. Our result showed that both DMPs and DNPs were able to release silicon, as detected in-vitro by inductively coupled plasma optical emission spectrometry (ICP/OES). In addition, diatom microparticles and nanoparticles - derived from diatom skeletons - showed minimal or non-cytotoxic effects in-vitro as determined by lactate dehydrogenase assays on cell cultures. These findings suggest that diatom particles derived from diatom skeleton as a silicon donor might have potential use for bone tissue engineering. In the second part of this thesis, we studied the effect of diatom particles on some properties of silk fibroin/diatom particles scaffolds. To handle this task, a series of fibroin scaffolds loaded with different amounts and size of diatom particles (microparticles, nanoparticles and their combination) were fabricated by using the salt leaching method. Diatom particles addition influenced scaffold morphology and mechanical properties, and its biological behaviour as assessed on human osteosarcoma cell line MG63 cultures. Scaffolds loaded with diatom particles strongly enhanced cell adhesion, metabolic activity and proliferation. Moreover, the possible beneficial effect of the addition of diatoms particles to silk fibroin on early bone formation was determined through collagen type I synthesis evaluation, osterix expression and alkaline phosphatase induction. Cultures with human mesenchymal stem cells (hMSCs) demonstrated the silk/diatom particles scaffolds were able to induce the differentiation of progenitor cells. In conclusion, our findings provided strong evidence for a potential use of diatom particles- derived from natural diatom skeleton in biological applications, in particular for bone tissue regeneration.

In this dissertation, a report of my PhD research activity is provided. The activity was carried out in Biotech Research Center, part of the Industrial Engineering Department, of the University of Trento (Italy), under the supervision of Prof. Claudio Migliaresi and Dr. Devid Maniglio. Biofabrication, an approach to the bottom-up paradigm of tissue engineering, represents the research topic. This technology is defined as the production of complex biological constructs using cells, components of the ECM, biomolecules, and biomaterials that are assembled with different techniques in an engineered tissue fragment. The general aim of the work was to address some of the problems that currently limited the development and applicability of biofabrication. In particular, two issues were considered in the experimental part: the cryopreservation of cell-laden hydrogel constructs and the development of novel building blocks containing cells using alginate-based hydrogels. Alginate was the material of choice for investigation, as an accepted support for different tissue engineering applications that can sustain several modification and fabrication methods. In the first chapter, the concepts of bottom-up tissue engineering and biofabrication are introduced. The role and state of the art of hydrogels to manufacture cell-laden building blocks, the techniques for cell encapsulation and the commonly used fabrication strategies for biofabrication and bioprinting are reviewed together with their applications. Moreover, the limitations that currently restrict the applicability of hydrogel-based tissue engineering are discussed. In chapter two, the role of alginate hydrogels in tissue engineering and biofabrication is described. In particular, its chemical content, crosslinking behavior, manufacturing capacity, and applications are reviewed with emphasis on the possible modification of alginate hydrogels in order to enhance biocompatibility and functionality of encapsulated cells. The experimental part is described in the following chapters. Chapter three introduces the concept of cryopreservation and in particular the issues concerning the preservation of cell-laden building blocks. Subsequently, the impact of cryopreservation on the viability and functionality of cells encapsulated in alginate matrices is evaluated comparing different cryoprotective agents. The experimental methods for manufacturing and preserving cell-laden alginate fibers and for performing the biological and structural tests are reported. The results are presented, discussed and compared with the state of the art. In chapter four, a novel method for encapsulating cells within alginate-based hydrogel films with micrometer thickness is described. The procedure for immobilizing cells within hydrogel films with different composition is described, together with the performed biological assays aimed at selecting the best matrix composition. The results are reported and discussed, emphasizing the potential applications and future developments of the proposed method.

L'attività di ricerca descritta in questa tesi fornisce linee guida per la progettazione di arti protesici inferiori, con particolare riferimento alla progettazione di protesi a basso costo. La necessità di efficienti protesi a basso costo risulta infatti sentita nei Paesi in via di sviluppo ma anche dalle fasce meno abbienti dei paesi occidentali. Al fine di comprendere le strategie adottate dall'apparato locomotorio per muoversi con le protesi sono analizzati il cammino fisiologico, le protesi presenti sul mercato ed infine le modalità con cui le loro prestazioni sono valutate. Con il presente lavoro, dopo aver osservato la presenza di una scarsa strutturazione della metodologia di progettazione che riguarda specialmente il settore del basso costo, si propone una metodologia il più possibile oggettiva e ripetibile tesa ad individuare quali sono gli aspetti essenziali di una protesi per garantire al paziente una buona qualità di vita. Solo questi aspetti dovranno essere selezionati al fine di ottenere la massima semplificazione della protesi e ridurre il più possibile i costi. Per la simulazione delle attività di locomozione, in particolare del cammino, è stato elaborato un apposito modello spaziale del cammino. Il modello proposto ha 7 membri rigidi (corrispondenti a piedi, tibie, femori e bacino) e 24 gradi di libertà. Le articolazioni e l'appoggio dei piedi al suolo sono modellati con giunti sferici. La pianta del piede consente tre possibili punti di appoggio. I criteri di realizzazione delle simulazioni possono comprendere aspetti energetici, cinematici e dinamici considerati come obiettivo dall'apparato locomotorio. In questa tesi vengono trattati in particolare gli aspetti cinematici ed è mostrata un'applicazione della procedura nella quale vengono dapprima identificati i riferimenti fisiologici del cammino e quindi simulato il cammino in presenza di una menomazione al ginocchio (eliminazione della flessione in fase di appoggio). Viene quindi lasciato a sviluppi futuri il completamento della procedura e la sua implementazione in un codice di calcolo. / Inexpensive and efficient prostheses are needed both for developing and Western countries. The research activity described in this thesis aims at providing guidelines for the development of lower limb prostheses, referring in particular to the design of low cost prostheses. Physiological gait, commercial prostheses and prostheses evaluation methods are analyzed in order to understand the strategies adopted by the human locomotion system to walk with such artificial devices. This work outlines a lack of systematic approaches for the design prostheses, in particular for the low cost ones. This lack is overcome by suggesting a metodology, which is as much objective and repeatable as possible, oriented to the definition of the essential aspects that provide the patient with a good quality of life. Only these aspects should be selected to design the low cost prostheses, i.e. in order to obtain the maximum simplification and thus the maximum cost reduction. A model for the simulation of gait has been implemented. The spatial model presented is made up of 7 rigid members (correspondent to feet, shanks, thighs and pelvis) and has 24 degrees of freedom. The articular joints and the contact of the foot with ground are modeled as spherical joints. The foot sole allows three different supporting points. The simulations are executed utilizing criteria that consider the energetic, kinematic and dynamic issues as addressed by the locomotion system. An application of the procedure is shown. The identification of the kinematic physiological parameters and the simulation of a maimed gait (without stance knee flexion) are presented.

The treatment of the Cerebral Palsy (CP) is considered as the “core problem” for the whole field of the pediatric rehabilitation. The reason why this pathology has such a primary role, can be ascribed to two main aspects. First of all CP is the form of disability most frequent in childhood (one new case per 500 birth alive, (1)), secondarily the functional recovery of the “spastic” child is, historically, the clinical field in which the majority of the therapeutic methods and techniques (physiotherapy, orthotic, pharmacologic, orthopedic-surgical, neurosurgical) were first applied and tested. The currently accepted definition of CP – Group of disorders of the development of movement and posture causing activity limitation (2) – is the result of a recent update by the World Health Organization to the language of the International Classification of Functioning Disability and Health, from the original proposal of Ingram – A persistent but not unchangeable disorder of posture and movement – dated 1955 (3). This definition considers CP as a permanent ailment, i.e. a “fixed” condition, that however can be modified both functionally and structurally by means of child spontaneous evolution and treatments carried out during childhood. The lesion that causes the palsy, happens in a structurally immature brain in the pre-, peri- or post-birth period (but only during the firsts months of life). The most frequent causes of CP are: prematurity, insufficient cerebral perfusion, arterial haemorrhage, venous infarction, hypoxia caused by various origin (for example from the ingestion of amniotic liquid), malnutrition, infection and maternal or fetal poisoning. In addition to these causes, traumas and malformations have to be included. The lesion, whether focused or spread over the nervous system, impairs the whole functioning of the Central Nervous System (CNS). As a consequence, they affect the construction of the adaptive functions (4), first of all posture control, locomotion and manipulation. The palsy itself does not vary over time, however it assumes an unavoidable “evolutionary” feature when during growth the child is requested to meet new and different needs through the construction of new and different functions. It is essential to consider that clinically CP is not only a direct expression of structural impairment, that is of etiology, pathogenesis and lesion timing, but it is mainly the manifestation of the path followed by the CNS to “re”-construct the adaptive functions “despite” the presence of the damage. “Palsy” is “the form of the function that is implemented by an individual whose CNS has been damaged in order to satisfy the demands coming from the environment” (4). Therefore it is only possible to establish general relations between lesion site, nature and size, and palsy and recovery processes. It is quite common to observe that children with very similar neuroimaging can have very different clinical manifestations of CP and, on the other hand, children with very similar motor behaviors can have completely different lesion histories. A very clear example of this is represented by hemiplegic forms, which show bilateral hemispheric lesions in a high percentage of cases. The first section of this thesis is aimed at guiding the interpretation of CP. First of all the issue of the detection of the palsy is treated from historical viewpoint. Consequently, an extended analysis of the current definition of CP, as internationally accepted, is provided. The definition is then outlined in terms of a space dimension and then of a time dimension, hence it is highlighted where this definition is unacceptably lacking. The last part of the first section further stresses the importance of shifting from the traditional concept of CP as a palsy of development (defect analysis) towards the notion of development of palsy, i.e., as the product of the relationship that the individual however tries to dynamically build with the surrounding environment (resource semeiotics) starting and growing from a different availability of resources, needs, dreams, rights and duties (4). In the scientific and clinic community no common classification system of CP has so far been universally accepted. Besides, no standard operative method or technique have been acknowledged to effectively assess the different disabilities and impairments exhibited by children with CP. CP is still “an artificial concept, comprising several causes and clinical syndromes that have been grouped together for a convenience of management” (5). The lack of standard and common protocols able to effectively diagnose the palsy, and as a consequence to establish specific treatments and prognosis, is mainly because of the difficulty to elevate this field to a level based on scientific evidence. A solution aimed at overcoming the current incomplete treatment of CP children is represented by the clinical systematic adoption of objective tools able to measure motor defects and movement impairments. A widespread application of reliable instruments and techniques able to objectively evaluate both the form of the palsy (diagnosis) and the efficacy of the treatments provided (prognosis), constitutes a valuable method able to validate care protocols, establish the efficacy of classification systems and assess the validity of definitions. Since the ‘80s, instruments specifically oriented to the analysis of the human movement have been advantageously designed and applied in the context of CP with the aim of measuring motor deficits and, especially, gait deviations. The gait analysis (GA) technique has been increasingly used over the years to assess, analyze, classify, and support the process of clinical decisions making, allowing for a complete investigation of gait with an increased temporal and spatial resolution. GA has provided a basis for improving the outcome of surgical and nonsurgical treatments and for introducing a new modus operandi in the identification of defects and functional adaptations to the musculoskeletal disorders. Historically, the first laboratories set up for gait analysis developed their own protocol (set of procedures for data collection and for data reduction) independently, according to performances of the technologies available at that time. In particular, the stereophotogrammetric systems mainly based on optoelectronic technology, soon became a gold-standard for motion analysis. They have been successfully applied especially for scientific purposes. Nowadays the optoelectronic systems have significantly improved their performances in term of spatial and temporal resolution, however many laboratories continue to use the protocols designed on the technology available in the ‘70s and now out-of-date. Furthermore, these protocols are not coherent both for the biomechanical models and for the adopted collection procedures. In spite of these differences, GA data are shared, exchanged and interpreted irrespectively to the adopted protocol without a full awareness to what extent these protocols are compatible and comparable with each other. Following the extraordinary advances in computer science and electronics, new systems for GA no longer based on optoelectronic technology, are now becoming available. They are the Inertial and Magnetic Measurement Systems (IMMSs), based on miniature MEMS (Microelectromechanical systems) inertial sensor technology. These systems are cost effective, wearable and fully portable motion analysis systems, these features gives IMMSs the potential to be used both outside specialized laboratories and to consecutive collect series of tens of gait cycles. The recognition and selection of the most representative gait cycle is then easier and more reliable especially in CP children, considering their relevant gait cycle variability. The second section of this thesis is focused on GA. In particular, it is firstly aimed at examining the differences among five most representative GA protocols in order to assess the state of the art with respect to the inter-protocol variability. The design of a new protocol is then proposed and presented with the aim of achieving gait analysis on CP children by means of IMMS. The protocol, named ‘Outwalk’, contains original and innovative solutions oriented at obtaining joint kinematic with calibration procedures extremely comfortable for the patients. The results of a first in-vivo validation of Outwalk on healthy subjects are then provided. In particular, this study was carried out by comparing Outwalk used in combination with an IMMS with respect to a reference protocol and an optoelectronic system. In order to set a more accurate and precise comparison of the systems and the protocols, ad hoc methods were designed and an original formulation of the statistical parameter coefficient of multiple correlation was developed and effectively applied. On the basis of the experimental design proposed for the validation on healthy subjects, a first assessment of Outwalk, together with an IMMS, was also carried out on CP children. The third section of this thesis is dedicated to the treatment of walking in CP children. Commonly prescribed treatments in addressing gait abnormalities in CP children include physical therapy, surgery (orthopedic and rhizotomy), and orthoses. The orthotic approach is conservative, being reversible, and widespread in many therapeutic regimes. Orthoses are used to improve the gait of children with CP, by preventing deformities, controlling joint position, and offering an effective lever for the ankle joint. Orthoses are prescribed for the additional aims of increasing walking speed, improving stability, preventing stumbling, and decreasing muscular fatigue. The ankle-foot orthosis (AFO), with a rigid ankle, are primarily designed to prevent equinus and other foot deformities with a positive effect also on more proximal joints. However, AFOs prevent the natural excursion of the tibio-tarsic joint during the second rocker, hence hampering the natural leaning progression of the whole body under the effect of the inertia (6). A new modular (submalleolar) astragalus-calcanear orthosis, named OMAC, has recently been proposed with the intention of substituting the prescription of AFOs in those CP children exhibiting a flat and valgus-pronated foot. The aim of this section is thus to present the mechanical and technical features of the OMAC by means of an accurate description of the device. In particular, the integral document of the deposited Italian patent, is provided. A preliminary validation of OMAC with respect to AFO is also reported as resulted from an experimental campaign on diplegic CP children, during a three month period, aimed at quantitatively assessing the benefit provided by the two orthoses on walking and at qualitatively evaluating the changes in the quality of life and motor abilities. As already stated, CP is universally considered as a persistent but not unchangeable disorder of posture and movement. Conversely to this definition, some clinicians (4) have recently pointed out that movement disorders may be primarily caused by the presence of perceptive disorders, where perception is not merely the acquisition of sensory information, but an active process aimed at guiding the execution of movements through the integration of sensory information properly representing the state of one’s body and of the environment. Children with perceptive impairments show an overall fear of moving and the onset of strongly unnatural walking schemes directly caused by the presence of perceptive system disorders. The fourth section of the thesis thus deals with accurately defining the perceptive impairment exhibited by diplegic CP children. A detailed description of the clinical signs revealing the presence of the perceptive impairment, and a classification scheme of the clinical aspects of perceptual disorders is provided. In the end, a functional reaching test is proposed as an instrumental test able to disclosure the perceptive impairment. References 1. Prevalence and characteristics of children with cerebral palsy in Europe. Dev Med Child Neurol. 2002 Set;44(9):633-640. 2. Bax M, Goldstein M, Rosenbaum P, Leviton A, Paneth N, Dan B, et al. Proposed definition and classification of cerebral palsy, April 2005. Dev Med Child Neurol. 2005 Ago;47(8):571-576. 3. Ingram TT. A study of cerebral palsy in the childhood population of Edinburgh. Arch. Dis. Child. 1955 Apr;30(150):85-98. 4. Ferrari A, Cioni G. The spastic forms of cerebral palsy : a guide to the assessment of adaptive functions. Milan: Springer; 2009. 5. Olney SJ, Wright MJ. Cerebral Palsy. Campbell S et al. Physical Therapy for Children. 2nd Ed. Philadelphia: Saunders. 2000;:533-570. 6. Desloovere K, Molenaers G, Van Gestel L, Huenaerts C, Van Campenhout A, Callewaert B, et al. How can push-off be preserved during use of an ankle foot orthosis in children with hemiplegia? A prospective controlled study. Gait Posture. 2006 Ott;24(2):142-151.