SUITE an Innovative Bioreactor Platform for in vitro Experiments

In-vitro cell cultures are a fundamental step in preclinical drug testing and are of great interest to the pharmaceutical industry. The most common method for culturing cells is in cell culture incubators. These are large and cumbersome and all mechanical stimuli are absent. They are nevertheless used ubiquitously and their results quoted as "standards" of in-vitro protocols. Several alternative culture methods have been proposed, and many systems are currently available commercially. Indeed, systems and devices for maintaining cells and tissues in controlled physical conditions, or bioreactors, have become an important tool in many areas of research. This is not only due to the growing interest in tissue engineering but also because it is now being increasingly recognised that cells respond not only to their biochemical, but also to their physical environment, and both cues are necessary to create a biomimetic habitat. However most bioreactors for cell culture and tissue engineering are cumbersome and only provide a few cues such as flow or strain, allowing limited control and flexibility. Since drug testing involves a large number of tests on identical cell cultures, a single well culture is inadequate and costly both in time and money. The High Throughput Screening (HTS), is a methodology for scientific experimentation widely used in drug discovery, based on a brute-force approach to collect a large amount of experimental data in less time and using less animals. The parallel nature of HTS makes it possible to collect a large amount of data from a small number of experiments and in a very short time. HTS, however, suffers from a significant problem that may affect the relevance of tests: the environment discrepancy problem. Another problem related with the actual drug testing and tissue engineering experiments is the enormous number of animals that have to be scarified every year. The aim of this study was to develop a generic platform or SUITE (Supervising Unit for In-vitro TEsting) for cell, tissue and organ culture composed of two main components: a universal control unit and an array of bioreactor chambers. The platform provides a biomimetic habitat to cells and tissues since the environment in the chambers is controlled and regulated to provide biomechanical and biophysical stimuli similar to those found in-vivo. In this work I describe how a new concept of cell culture bioreactor was developed by integrating different technologies and research fields. The data extracted using this new cell culture approach is more predictive of the in vivo response with respect to the multi-well approach, particularly for drug related studies. The starting point was a thorough analysis of currently used in-vitro methods; their pros and cons were assessed to exploit their advantages and overcome or circumvent their disadvantages. As far as the culture chamber is concerned, the approach was to use the methods and materials commonly employed in microfluidic fabrication, but at scales compatible with classical culture systems such as petri-dishes and multiwells. This renders the bioreactors more amenable to use by biologists and enables the use of cell densities comparable with classic systems as well as the use of conventional assaying techniques. In most cases, the cell culture chambers are thus made out of PDMS (Polydimethylsiloxane), using soft-moulding with micro- or mini-machined masters, or what we call Soft Milli-molding. A system on a plate Multi Compartmental Modular Bioreactor (MCmB) was developed using this technology. The MCmB is a modular chamber for high throughput multi compartmental bioreactor experiments. It is designed to be used in a wide range of applications and with various cell types. A precise stimulus application is also very important to better understand the correlation between physical variables and pathologies allowing a more accurate study of the tissue physiology and pathologies. For this reason in these thesis three additional stimulation chambers for vascular and articular cartilage stimulation respectively were also designed and tested. The control system was developed to be user-friendly, flexible and expandable to include new stimuli and was based on modular components, including motors and sensors. Importantly a single software interface was designed to allow data acquisition and monitoring of several chambers in series or in parallel. Using SUITE, high throughput experiments can be performed in an in vivo-like simulated environment for a long time to simulate different physiological or pathological scenarios or for toxicity testing of cells, tissues or in-vitro organ models

Ottimizzazione di un bioreattore High Throughput con strategia di controllo autonoma.

Negli ultimi anni si è assistito ad una vera e propria rivoluzione dal punto di vista tecnologico. Questa rivoluzione ha portato a notevoli cambiamenti nelle metodiche di testing biologico e cellulare. Basti pensare all’avvento delle nuove metodologie di analisi High Throughput, grazie alle quali è oggi possibile eseguire esperimenti in tempi molto brevi, estraendo da questi molti dati. Purtroppo non siamo ancora riusciti ad eliminare una delle piaghe della ricerca medico biologica: la sperimentazione animale; si stima che, nel mondo intero, vengano uccisi ogni anno almeno 300 milioni di esemplari. Appurato che, al giorno d’oggi, la sperimentazione animale è indispensabile per la ricerca, è dovere della comunità scientifica lavorare affinché si riesca a ridurre il numero di cavie animali immolate. In questo contesto si inserisce perfettamente il presente lavoro di tesi, che si prefigge lo scopo di ottimizzare un bioreattore High Throughput al fine di trasformarlo da prototipo in fase di testing, a macchinario finito e commercializzabile. Nel 2000, presso il Centro Interdipartimentale di Ricerca E.Piaggio, è stato realizzato il primo prototipo di bioreattore capace di controllare la pressione, il flusso, la temperatura ed il pH di una coltura cellulare in modo automatico e per intervalli di tempo prolungati. Questo bioreattore permette inoltre l’osservazione delle colture cellulari sotto microscopio, in quanto non necessita di essere inserito in un incubatore per il mantenimento delle variabili ambientali necessarie alla sopravvivenza della coltura cellulare. Durante questo lavoro di tesi si è provveduto ad una completa ristrutturazione di questo prodotto, al fine di renderlo un sistema di coltura e testing cellulare commercializzabile e utilizzabile anche da utenti non aventi conoscenze di tipo ingegneristico. Si è provveduto all’ottimizzazione della camera di miscelazione e del sistema di riscaldamento del terreno che, grazie alle nuove modifiche, sono diventati perfettamente compatibili con le nuove metodologie di analisi High Througput. E’ stato realizzato un sistema di riscaldamento capace di mantenere in temperatura contemporaneamente 4 camere di miscelazione con l’utilizzo di un unico bagnetto termostatico, così da risparmiare sugli spazi necessari per l’installazione del sistema. La grande innovazione apportata durante questo lavoro è, però, nel sistema di controllo del bioreattore; in precedenza l’intero apparato era controllato mediante una scheda di acquisizione dati della National Instruments collegata ad un PC. Nel nuovo bioreattore la scheda National Instruments è stata sostituita con una scheda di controllo Wildfire, capace di controllare in modo autonomo il bioreattore che adesso non necessita più di un PC dedicato, ma può essere collegato alla rete Ethernet. L’esperimento verrà osservato poi mediante un’apposita interfaccia anch’essa sviluppat nel corso di questo lavoro di tesi. La suddetta modifica ha portato a dover completamente riscrivere il software di controllo e gestione del bioreattore. È stato scritto quindi un vero e proprio sistema operativo per la scheda di controllo Wildfire. Questo sistema operativo è stato realizzato utilizzando una libreria di controllo per robot, il Robotics4.NET, sviluppata dal Dipartimento di Informatica dell’Università di Pisa. Sarà possibile, mediante il nuovo sistema, condurre contemporaneamente esperimenti su più bioreattori, simulando così varie cavie, permettendo quinid una riduzione dei soggetti animali necessari alla sperimentazione nonchè un naumento delle performance dei test di laboratorio. Attualmente i test cellulari vengono condotti in ambiente statico; questo approccio di coltivazione è in realtà una vaga approssimazione della realtà biologica, che le colture dovrebbero rappresentare. Questa approssimazione rende molto difficile l’esportazione dei dati ricavati dalle colture cellulari ai tessuti in vivo e quindi all’uomo. Grazie al nuovo sistema di bioreattore High Throughput sarà invece possibile effettuare colture cellulari sottoponendole a regimi dinamici tipici dei tessuti in vivo, con un conseguente aumento dell’attendibilità degli esperimenti svolti

Touch sensor for social robots and interactive objects affective interaction

The recognised importance of physical experience in empathic exchanges has led to the development of touch sensors for human–robot affective interaction. Most of these sensors, implemented as matrix of pressure sensors, are rigid, cannot be fabricated in complex shapes, cannot be subjected to large deformations, and usually allow to capture only the contact event, without any information about the interaction context. This paper presents a tactile flux sensor able to capture the entire context of the interaction including gestures and patterns. The sensor is made of alternate layers of sensitive and insulating silicone: the soft nature of the sensor makes it adaptable to complex and deformable bodies. The main features from electrical signals are extracted with the principal component analysis, and a self-organising neural network is in charge for the classification and spatial identification of the events to acknowledge and measure the gesture. The results open to interesting applications, which span from toy manufacturing, to human-robot interaction, and even to sport and biomedical equipment and applications

Improving African healthcare through open source biomedical engineering

The lack of accessible quality healthcare is one of the biggest problems in Africa and other developing countries. This is not only due to the unavailability of resources, but also to the absence of a structured formative process for the design and management of healthcare facilities. Crucial to the effective and efficient exploitation of healthcare facilities and biomedical technology is the support of Biomedical engineers, who form the link between technology and medical practice. Indeed Biomedical engineers, together with nurses and doctors, form the pillars of healthcare systems in the developed world. In this paper, the Open Source for BioMedical Engineering (OS4BME) project and its kick off summer school are presented. The OS4BME project aims at developing a new generation of biomedical engineers, able to exploit emerging technologies generated by the recent "Makers" revolution. During the one week summer school, students from various sub-Saharan countries were introduced to these new design, development and sharing paradigms. Students worked together to identify new simple biomedical devices, which could help in daily clinical practice in their countries. A cheap and easy-to-use neonatal monitoring device was chosen as a Crowd design project. The OS4BME Baby Monitor was designed and assembled by the students during the one week summer school, demonstrating the creative potential of the new generation of biomedical engineers empowered with the paradigms of crowdsourcing and rapid prototyping

SEAI: Social Emotional Artificial Intelligence Based on Damasio's Theory of Mind

A socially intelligent robot must be capable to extract meaningful information in real-time from the social environment and react accordingly with coherent human-like behaviour. Moreover, it should be able to internalise this information, to reason on it at a higher abstract level, build its own opinions independently and then automatically bias the decision-making according to its unique experience. In the last decades, neuroscience research highlighted the link between the evolution of such complex behaviour and the evolution of a certain level of consciousness, which cannot leave out of a body that feels emotions as discriminants and prompters. In order to develop cognitive systems for social robotics with greater human-likeliness, we used an "understanding by building" approach to model and implement a well-known theory of mind in the form of an artificial intelligence, and we tested it on a sophisticated robotic platform. The name of the presented system is SEAI (Social Emotional Artificial Intelligence), a cognitive system specifically conceived for social and emotional robots. It is designed as a bio-inspired, highly modular, hybrid system with emotion modelling and high-level reasoning capabilities. It follows the deliberative/reactive paradigm where a knowledge-based expert system is aimed at dealing with the high-level symbolic reasoning, while a more conventional reactive paradigm is deputed to the low-level processing and control. The SEAI system is also enriched by a model which simulate the Damasio's theory of consciousness and the theory of Somatic Markers. After a review of similar bio-inspired cognitive systems, we present the scientific foundations and their computational formalisation at the basis of the SEAI framework. Then, a deeper technical description of the architecture is disclosed underlining the numerous parallelisms with the human cognitive system. Finally, the influence of artificial emotions and feelings, and their link with the robot's beliefs and decisions have been tested in a physical humanoid involved in Human-Robot Interaction (HRI)