Dissertations / Theses on the topic 'Biomedical and industrial applications'

Modern smartphone technology has created a myriad of opportunities in the field of RF and Microwave. Specifically, Chipless RFID sensor, compact microwave filter, antenna based on a microstrip structure, and many more. In this thesis, innovative ideas for the industrial and biomedical device has been explored. The work presents the reconfigurable filter design, Switch-beam antenna, Microwave interferometer, X-band Rotman Lens antenna, Ultra-wideband antenna based on SIW resonator, L-band Stepped Frequency Continuous Wave antenna, development of a wireless sensor system for environmental monitoring, Indoor Air Quality monitoring, and Wildfire Monitoring based on the modulated scattering technique (MST). The MST sensor probes are based on the scattering properties of small passive antennas and radiate part of the impinging electromagnetic field generated by an interrogating antenna, which also acquires the backscattered signal as information. The MST probes are able to deliver data without a radio frequency front end. They use a simple circuit that alternatively terminates the antenna probe on suitable loads to generate a low modulation signal on the backscattered electromagnetic wave. The antenna presented in this work has been designed in ADS Software by Keysight Technologies. The designed antenna has been assessed numerically and experimentally. The experimental measurement data demonstrate the effectiveness of the individual system. Simultaneously, the MST sensor system has been proposed to obtain the best performance in communication range, load efficiency, and power harvesting. The MST sensor has been fabricated and assessed in practical scenarios. The proposed prototype, able to provide a communication range of about 15 m, serves as a proof-of-concept. The acquired measurements of MST demonstrate the accuracy of the data without radio frequency front end or bulky wired connection with the same efficiency of standard wireless sensors such as radio frequency identifier (RFID) or wireless sensor networks (WSN).

In the modern industry, data processing systems must be able to receive, aggregate, and process information from different sources to achieve complex tasks of production control and coordination. Examples are the real-time monitoring of the quality and quantity of products, biometric data acquisition in the rehabilitation procedures. Energy efficiency in the data communication system is essential in wireless networks. Reduce power consumption in the data exchange can prolong the operating life of battery-powered devices and save energy on a global scale. In this direction, a fundamental step is to accurately model the energy consumption for data communication over a wireless link for the system of interest. The first part concerns the application scenario of the Body Sensor Network for motion reconstruction applications. Wireless systems that use wearable sensors have developed rapidly in recent years, and the requirements in terms of throughput and timing accuracy are challenging. This thesis presents a new general-purpose Inertial Measure Unit that exploits a dual-core architecture. A core offers processing capability, and the other one is a radio interface IEEE 802.15.4. I propose the whole system and a protocol to maximize the throughput, reduce the packet loss, and improve the robustness of wireless sensor nodes communication. In the second part of the thesis, I move the attention to the Low Power Wide Area Network in the IoT scenario. Today, the most promising long-range communication technologies are LoRaWAN and Narrow Band IoT (NB-IoT), which are driving a vast IoT ecosystem. A dedicated chapter evaluates the performance of LoRaWAN and NB-IoT with accurate in-field measurements using the same monitoring application for a comparison in terms of energy efficiency, lifetime, quality of service (QoS), and coverage. Finally, the last part provides configuration guidelines for future industrial applications with harsh requirements of long-range and low power wireless connectivity.

This dissertation will be focused on the characterization of an atmospheric pressure plasma jet source with an application oriented diagnostic approach and the description of processes supported by this plasma source. The plasma source investigated is a single electrode plasma jet. Schlieren images, optical emission spectra, temperature and heat flux profiles are analyzed to deeply investigate the fluid dynamic, the chemical composition and the thermal output of the plasma generated with a nanosecond-pulsed high voltage generator. The maximum temperature measured is about 45 °C and values close to the room temperature are reached 10 mm down the source outlet, ensuring the possibility to use the plasma jet for the treatment of thermosensitive materials, such as, for example, biological substrate or polymers. Electrospinning of polymeric solution allows the production of nanofibrous non-woven mats and the plasma pre-treatment of the solutions leads to the realization of defect free nanofibers. The use of the plasma jet allows the electrospinnability of a non-spinnable poly(L-lactic acid) (PLLA) solution, suitable for the production of biological scaffold for the wound dressing.

This dissertation will be focused on the characterization of an atmospheric pressure plasma jet source with an application oriented diagnostic approach and the description of processes supported by this plasma source. The plasma source investigated is a single electrode plasma jet. Schlieren images, optical emission spectra, temperature and heat flux profiles are analyzed to deeply investigate the fluid dynamic, the chemical composition and the thermal output of the plasma generated with a nanosecond-pulsed high voltage generator. The maximum temperature measured is about 45 °C and values close to the room temperature are reached 10 mm down the source outlet, ensuring the possibility to use the plasma jet for the treatment of thermosensitive materials, such as, for example, biological substrate or polymers. Electrospinning of polymeric solution allows the production of nanofibrous non-woven mats and the plasma pre-treatment of the solutions leads to the realization of defect free nanofibers. The use of the plasma jet allows the electrospinnability of a non-spinnable poly(L-lactic acid) (PLLA) solution, suitable for the production of biological scaffold for the wound dressing.

This research entailed the development of resonant dielectric sensors that utilising alternating electric fields at microwave frequencies. Characterisation and identification of biological samples is currently an expensive and time consuming procedure due to the extreme variation of seemingly similar biological systems. The work describes the development of resonant dielectric sensors for five distinct applications. The aim of the research was to develop non-invasive, affordable and compact dielectric spectroscopy sensors that may be used in-situ to characterise organic and biological systems. The suitability of using dielectric spectroscopy to a number of applications has been investigated. This is to address disadvantages of conventional laboratory analysis such as lengthy, costly and labour intensive assessment methods that require intermittent sampling and/or off site analysis. The research is structured into a series of progressive stages (work packages) that integrate and culminate into feasible inline analytical procedures that can quickly, safely and inexpensively assess a specified variable in organic and biological systems. Overall, a number of sensors using electromagnetic radiation at microwave frequencies were developed that have proven to be sensitive and extremely versatile to a number of detection and monitoring applications. Miniaturisation of microwave generation and analysis circuits will enable the sensors to be incorporated into feasible compact devices for in situ analysis of the analyte in question.

Over the last years, modern technologies such as radiofrequency identification (RFID), wireless sensor networks (WSN), and wireless power transfer (WPT) are increasingly gaining attraction, both for the biomedical and the industrial fields of study, intending to achieve the paradigm of the internet of things (IoT). Within this research, different systems have been designed and realized by exploiting these typical IoT applications. For what concerns the biomedical sphere of interest, it is proving to be more and more urgent to continuously monitor the behaviors and the vital parameters of elderly people to detect as soon as possible any sort of disease or problem. Therefore, a customized 2.45 GHz RFID localization system has been realized in order to simultaneously perform 3-D tracking of multiple tagged people, static or dynamic, in indoor environments, i.e., the retirement homes. Moreover, a 5.8 GHz wearable device for human breath detection has been conceived, making use of the self-injection locked (SIL) radar technique. Finally, focusing on predictive maintenance, which is increasingly playing a crucial role for industrial, and in particular automotive, applications, it has been presented the design and the validation of a WPT system seamlessly integrated with a WSN platform for remote monitoring of important parts of the engine, placed in a typical electromagnetically harsh, metal-rich environment, e.g., the engine compartment of a car. Energy is provided wirelessly by means of an RF power source at 2.45 GHz to the low-power wireless sensor nodes located in difficult-to-be-reached positions, allowing to eliminate their periodic battery replacement.

Present PhD thesis aimed to investigate relatively unknown properties of halloysite nanoparticles, as well as to further examine HNTs as potential drug nanocarriers. NPs loading and release characteristics were studied using model active molecules: magnesium monoperoxyphthalate (MMPP), aspirin and epirubicin. The research was fulfilled with formation of complex multi-functional nanoarchitectures, which apart from ability to deliver incorporated drugs, showed the potential of controlled and sustain release of therapeutics, biocompatible and bioresorbable characteristics as well as potential targeting abilities. Great attention was dedicated to characterization of formed halloysite-based nanoarchitectures in qualitative as well as quantitative manner. Investigations performed in this thesis also faced the problem of exceeding dimensions of halloysite units, nanoparticles aggregation, poor loading capability and dose dumping effect. Subsequently, studies for trying to find a solution to these obstacles were undertaken. Fully characterized halloysite nanoconstructs were further examined in biological field, employing different cancer cell lines. Studies on pristine halloysite nanotubes: Physico-chemical and biological properties of halloysite nanoparticles were evaluated using microscopic techniques, spectroscopic analysis, surface studies regarding charge, porosity and wettability. The thermal and time-based examination of pristine halloysite was performed as well, showing stability of HNTs alumino-silicate skeletons up to ~400 ℃ and over a long period of time (2 years) at room temperature, however with a variable amount of incorporated water molecules. Biological performance of HNTs was determined in vitro in multiple cellular systems by toxicity, cellular uptake, colocalization and accumulation studies using [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] tetrazolium reduction (MTT) assay and set of microscopic techniques. Aiming to deeply characterize halloysite nanoparticles, the study proceeded with employment of non-standard techniques, as multiphoton microscopy that drove to discovery of novel NPs promising capabilities. It was revealed that halloysite is able to convert light to its second harmonic, at twice of the frequency (and therefore half of the wavelength) while using high intensity femtosecond pulsed laser. Halloysite Second Harmonic Generation (SHG) signal was detected over a broad wavelength range, showed stability over a long period of time, polarization properties and quadratic dependence on the intensity of incident light. The analysis also pointed out characteristic structure properties of the nanoparticle that is lack of the center of symmetry and the high crystalline structure organization. Among a wide spectrum of domains where discovered HNTs characteristics can be utilized (e.g. optoelectronics, biosensors), we have explored its application in alternative label-free bioimaging. The proposed multiphoton method of analysis showed advantages over the standard confocal microscopy, since e.g. nanoparticles did not have to be stained prior the analysis, thus no possible alterations of HNTs including size, surface chemistry and consequent cellular uptake were induced. Therefore, for the first time, halloysite nanotubes were exploited as imaging agents, taking advantage of their endogenous properties. Along the research it was revealed that the length of pristine HNTs and the strong aggregation limit their ability to pass intracellular membranes and thus minimize their effectiveness as drug nanocarriers. Therefore, efforts were devoted to the development of facile methodology to efficiently disperse and shorten HNTs units. Set of characterizations techniques, such as Scanning Electron Microscopy (SEM) analysis with size distribution profile and nitrogen adsorption Brunauer–Emmett–Teller (BET) method revealed that the applied ultrasonication procedure resulted with longest tubes breaking and favored obtaining HNTs below 300 nm in length (39.1 % to 76 % of the batch). The number of voids among the pristine nanoparticles when packing together (123–43 nm) greatly increased (total pore volume from 0.23 cm^3/g to 0.30 cm^3/g), meaning that the nanomaterial was efficiently disaggregated as well. In vitro internalization and colocalization studies by Scanning Electron and Multiphoton Microscopy demonstrated that the sonicated halloysite were preferentially internalized via macropinocytosis within 60 min and accumulated in the perinuclear region within 24 h. Halloysite application in nanomedicine: To study halloysite potential as a carrier for drugs, we set up the preparation and characterization of hybrid nanoconstructs with model molecules such as magnesium monoperoxyphthalate hexahydrate (MMPP), a negatively charged oxidizing agent, aspirin, an anti-inflammatory drug and epirubicin, a chemotherapeutic. Chosen molecules were incorporated with 3.5 %wt, 1.1 %wt and 5.1 %wt capacity, respectively for MMPP, aspirin and epirubicin. Loading efficiency (LE) improvement was achieved through the choice of the right solvent (1), enhancement of electrostatic forces between nanoparticle and the drug, via functionalization of HNT surfaces with an active linker (2), as well as NPs structure modification leading to increase of inner lumen volume (3). Specifically, the use of water: EtOH (7:3 v/v) as a solvent instead of water, increased MMPP loading capacity up to 6.1 %wt. Poor incorporation of aspirin was improved by enhancing electrostatic forces between deprotonated aspirin molecules and modified HNTs inner walls with amine-rich organosilane. It was also demonstrated that by enlarging volume of the NPs cavities, more molecules could be loaded. To do that, pristine HNTs were treated with 0.1M aqueous solution of NaOH, which resulted in an exfoliation of bilayers located inside the lumen. At the same time, outer surface of the halloysite tubules was preserved. As a consequence of the base treatment, halloysite cylinders gained more volume in the inner cavity as concluded from Transmission Electron Microscopy (TEM) and nitrogen adsorption BET analysis. The actual test on loading capacity using model MMPP molecule revealed increased MMPP incorporation from 6.1 % wt to 11.7 %wt. To evaluate if the activity of MMPP as an oxidizing agent remained unchanged upon incorporation and release from halloysite, and therefore to demonstrate the inactivity of the inorganic skeleton towards carried molecule, we tested HNT-MMPP nanoconstruct with selective fluorescent 1,3-diphenylisobenzofuran (DPBF) probe. Among available modifications of halloysite nanoparticles via covalent bond, the surface silanization is commonly recognized as one of the most efficient and widespread reaction while HNT manipulation. Up to date, the halloysite nanotubes functionalized with silanes have been used as a support for versatile applications in diverse scientific domains, including enzymes immobilization and biosensing. Willing to explore the halloysite functionalization with those active linkers, we have performed grafting reactions with representative organosilanes carrying the same backbone, while varying in the content of terminal groups, namely (3-aminopropyl)triethoxysilane (APTES), 3-(2-aminoethylamino)propyldimethoxymethylsilane (AEAPS), (3-mercaptopropyl)trimethoxysilane (MPS). Successful HNTs surfaces functionalization with organosilanes was demonstrated by means of quantitative thermogravimetric analysis (TGA) that allowed to estimate the loading capacity of organosilanes to be of 5.7 %wt for APTES, 7.4 %wt for AEAPS and 0.7 %wt for MPS. In addition, particular attention was dedicated to further quantify incorporated organosilane (APTES), since only one method has been so far reported, that is the destructive thermogravimetric analysis (TGA). For this reason, we set up and optimized a Fmoc based method by performing the following three reactions: (i) synthesis of “APTES-Fmoc” molecule; (ii) halloysite functionalization with “APTES-Fmoc”; and (iii) time-dependent Fmoc deprotection reaction in piperidine: EtOH (20 %) solution, resulting in dibenzofulvenepiperidine adduct (DBF-pip) formation. The UV-visible spectroscopic analysis of supernatant solutions demonstrated that the DBF-pip deprotection from halloysite support needs 5 h to be completed. Therefore, it was evidenced that HNT Fmoc-method showed strong coherence with already existing TGA method (± 2 % measurement error) and stood out as a valuable complementary technique for quantification of silane grafting on HNTs surface with additional low-cost and nondestructive advantages. The possibility of using halloysite nanotubes as a non-viral gene delivery nanosystem for therapeutic treatments was studied as well. Aiming to immobilize plasmid DNA (pDNA) based on the Green Fluorescent Protein (GFP) on HNTs support, the layer-by-layer (LbL) adsorption technique was applied. Obtained multi-component assembly was characterized qualitatively by monitoring variation in nanoparticle physico-chemical properties including surface charge, mass weight, presence of functional groups at each step of hybrid formation, which confirmed the successful nanoarchitecture formation. In order to additionally demonstrate the presence of GFP encoding plasmid (pGFP) on HNTs, the nanoarchitecture was treated with the bovine pancreatic deoxyribonuclease (DNase) enzyme, which induced the pGFP degradation through hydrolytic cleavage of phosphodiester linkages in DNA backbone. Thus, as expected, such nanoform with deposed genetic material varied in physico-chemical properties, expressing similar ones of the nanoconstruct without pGFP plasmid attached. The biological efficiency of HNTs-pGFP nanosystem was checked by means of Multiphoton microscopy. Successful pGFP plasmid transportation into cells was verified by detection of GFP expression, which yielded fluorescence emission. The interesting and innovative aspect of this case study was the simultaneous observation of GFP expression via fluorescence detection, and colocalization of halloysite nanoparticles by their SHG signal. This study proved that halloysite can act as an efficient carrier of genetic material, since free pGFP cannot be internalized by same cells, due to its large size and significant charge. Drug-loaded halloysite nanoconstructs (HNT-MMPP, HNT-APTES-aspirin) were also examined on the drug release kinetics, demonstrating long-term MMPP leakage taking 18 days and aspirin over 60 min. However, great drug liberation into the solvent of release was observed in the first minutes, followed by desired sustained drug release. The initial molecule liberation (dose dumping effect) is known to entail local toxicity. Herein, trying to find a solution to this problem, the coating of HNTs with the natural collagen polymer was investigated. Two strategies for the loading with this biopolymer were studied: (i) formation of a covalent bond between collagen and APTES-modified HNTs using glutaraldehyde cross-linker or (ii) noncovalent adsorption of collagen into pores of NPs. Immobilization of collagen on the surface of HNTs was estimated to be 3.7 %wt (i) and 1.8 %wt (ii). Other supplementary characterization techniques, such as water contact angle, ζ–potential analysis, Kaiser test, ultraviolet and visible (UV-vis) spectroscopy and Fourier Transform Infrared Spectroscopy (FTIR) were in accordance and proved nanoarchitectures formation. For the visualization purpose of HNTs encapsulated in collagen shell, the innovative characterization technique was implemented, namely 3D Multiphoton microscopy. It revealed that the biopolymer coating blocked the entrances of the hollow tubes thus, entrapping the drug in NPs. Mimicking tumor microenvironment (TME), the pH and/or enzyme triggered release was performed. LC-Mass analysis revealed that the collagen coating slowed down the release of aspirin from HNTs. Studies on cells showed that the collagen coating on HNTs is biocompatible and cell viability assay performed on 5637 urinary bladder and HeLa cervical cancer cell lines demonstrated the sustained release of the entrapped epirubicin chemotherapeutic agent in the biological context. Industrial application of halloysite: During a stage in BASF SE (USA), validation and properties enhancement of halloysite-based products potentially manufacturable in the company on an industrial scale were studied. In particular, the research was dedicated to aspects such as the pH-dependent dispersion behavior of halloysite nanotubes and iron coarse impurities removal from bulk samples. Applied methodologies and set of physico-chemical characterization techniques generated and revealed decreased percentage of present aggregates, maintained low shear viscosity under the threshold value and increased solids loading capacity in final halloysite-based products. Conclusions: In conclusion, PhD studies here reported contributed to the exploration of halloysite nanotubes for their application in the nanomedical and industrial fields. The investigations suggest a facile manipulation and functionalization of HNTs, useful for properties modification and improved NPs performance. Specifically, the study was directed toward formation of multi-functional nanocarriers with controlled drug delivery and release properties, together with targeting and imaging abilities. Moreover, the research was completed with halloysite-related technology transfer to the BASF SE, for the purpose of knowledge increase in the halloysite-field and bringing forward placement of halloysite-based products on the market. The systematic study on HNTs characterization and application performed in this PhD thesis will contribute to the development of HNTs as a high performance structural and functional material.

In this dissertation are reported the most relevant results obtained during my three years Ph.D. project. An open-air plasma source has been developed to treat plastic and metallic films typically used in food packaging manufacturing. Among others, the DBD configuration was chosen due to its many advantages such as high intensity and uniformity of the treatment, possibility of operating in ambient air as well as ease of scale up. Biological experiments were performed to assess the microbial reduction induced by the plasma treatment. Different operative conditions have been tested in order to identify the most efficient configuration and two distinct behaviours have been observed: low-power density treatment allowed to achieve microbial inactivation values below log 2 independently on treatment time; high-power density treatment where the microbial reduction grew with increasing treatment time. Subsequently, the plasma discharge has been characterized by means of three investigation methods: thermal, electrical and optical absorption spectroscopy (OAS) analysis. The thermal and electrical analyses were employed to identify the best dielectric materials for food packaging manufacturing purposes. Once defined the optimal DBD configuration, OAS was used to measure the absolute concentration of ozone and nitrogen dioxide. Results showed that at low-power density the chemistry is governed by ozone; while at high-power density ozone is consumed by the poisoning effect and only nitrogen dioxide is detectable. Lastly, a numerical simulation has been used to deeper investigate the chemistry governing the plasma discharge; by means of PLASIMO a global model and a fluid model were implemented.

We introduce and study a novel graph optimization problem to search for multiple cliques with the maximum overall weight, to which we denote as the Maximum Weighted Multiple Clique Problem (MWMCP). This problem arises in research involving network-based data mining, specifically, in bioinformatics where complex diseases, such as various types of cancer and diabetes, are conjectured to be triggered and influenced by a combination of genetic and environmental factors. To integrate potential effects from interplays among underlying candidate factors, we propose a new network-based framework to identify effective biomarkers by searching for "groups" of synergistic risk factors with high predictive power to disease outcome. An interaction network is constructed with vertex weight representing individual predictive power of candidate factors and edge weight representing pairwise synergistic interaction among factors. This network-based biomarker identification problem is then formulated as a MWMCP. To achieve near optimal solutions for large-scale networks, an analytical algorithm based on column generation method as well as a fast greedy heuristic have been derived. Also, to obtain its exact solutions, an advanced branch-price-and-cut algorithm is designed and solved after studying the properties of the problem. Our algorithms for MWMCP have been implemented and tested on random graphs and promising results have been obtained. They also are used to analyze two biomedical datasets: a Type 1 Diabetes (T1D) dataset from the Diabetes Prevention Trial-Type 1 (DPT-1) Study, and a breast cancer genomics dataset for metastasis prognosis. The results demonstrate that our network-based methods can identify important biomarkers with better prediction accuracy compared to the conventional feature selection that only considers individual effects.

In recent years, the increasing public concern about the environmental pollution caused by persistent plastic wastes has stimulated the interest in replacing synthetic polymers by biopolymers. The term biopolymer refers to polymers that are renewable or biodegradable or both. Biopolymers are capable of bio-assimilation at accelerated rates so they are biocompatible with the environment. Furthermore, their sustainability is of exceptional interest; the renewable origin of biopolymers makes them inexhaustible in contrast with the uncertain accessibility at medium-term of synthetic polymers produced from fossil sources. Nevertheless, biopolymers often have inferior properties compared to commodity polymers. Modification is a way to improve properties and achieve property combinations required for specific applications. Therefore the synthesis, characterization and property evaluation of new biopolymers derivatives are essential tasks that have to be done for the development of new materials able to replace the traditional plastics in areas such as industrial, medical, food, consumer products, and pharmaceutical applications. In the present Thesis, the chemical modification of two kinds of carboxylic biopolymers has been studied to respond to the necessity of creating new biopolymer derivatives with advanced properties at reasonable cost. Poly (gamma-glutamic acid) (PGGA) and hyaluronic acid (HyA) were the biopolymers selectedin this Thesis for their capacity to form stable ionic complexes with cationic surfactants to generate stable materials with new properties. These complexes are currently object of intensive research in our group due to their outstanding features. They are easily prepared and they tend to be selfassembled in well-ordered amphiphilic structures able to respond reversibly to thermal effects. This behaviour is of high scientific interest and also of practical relevance in the design of medical devices for thermally and chemically controlled store and delivery of drugs. The main goal of this Thesis is the preparation of ionic complexes of the two mentioned polyacids using different cationic surfactants depending on the desired final properties. The first part of the work is devoted to provide physicochemical knowledge of the structure and properties of alkyltrimethylphosphonium surfactants which have potential interest for novel applications. Then these surfactants were coupled to both PGGA and HyA to obtain the respective ionic complexes with biocide activity and thermal stability higher than those made by their ammonium analogs. PGGA complexes, abbreviated as nATMP·PGGA, have high interest as food preserving and packaging applications displaying as main advantage the edibility of the polymer and the possibility of improving their basic properties through blending with nanoclays. On the other hand, HyA complexes, nATMP·HyA, are useful to obtain HyA derivatives with antimicrobial activity. In addition, the preparation of nanoparticles of nATMP·HyA with antimicrobial properties was feasible using the ionotropic gelation method. The s econd part of the Thes is was devoted to the preparation of “greener” complexes of hyaluronic acid. For this regard, alkanoylcholine surfactants were used to prepare the ionic complexes nACh·HyA. These complexes constitute a highly promising biocompatible/biodegradable platform for the design of systems suitable for drug transport and targeting delivery in anticancer chemotherapy because it was demonstrated that non-cytotoxic nanoparticles can be prepared from these systems. The third part of the Thesis is dedicated to the preparation of biocompatible antimicrobial complexes using as cationic surfactant one of the most potent food preservative agents that is known today, that is the ethyl alpha-N-lauroyl L-arginate chloride surfactant (also known as LAE). These complexes (LAE·PGGA and LAE·HyA) are shown to be potential candidates to develop antimicrobial materials.
En els últims anys s’ha produït un augment de la preocupació sobre la contaminació produïda pels plàstics persistents que ha estimulat l’interès per reemplaçar els polímers sintètics pels biopolímers. El terme biopolímer es refereix als polímers que son o bé renovables o biodegradables, o ambdós. A més, és de excepcional interès la seva sostenibilitat; l’origen renovable dels biopolímers els fa inesgotables al contrari dels polímers sintètics, que al ser produïts a partir de fonts fòssils presenten una accessibilitat incerta a mig termini. No obstant, els biopolímers presenten de vegades unes propietats inferiors si se'ls comparem amb els polímers tradicionals. La modificació és una via per millorar les propietats i arribar a les combinacions de propietats requerides per a aplicacions específiques. Per tant, la síntesi, caracterització i avaluació de propietats de nous derivats de biopolímers són tasques essencials que s’han de realitzar pel des envolupament de nous materials capaços de reemplaçar els plàstics tradicionals en sectors com ara la indústria química, medicina, alimentació, productes de consum i farmàcia. En la present Tes i, s’ha estudiat la modificació química de dos tipus de biopolímers carboxílics per respondre a la necessitat de crear derivats de biopolímers amb propietats avançades i a un preu raonable. L’àcid poli(gamma-glutámic) (PGGA) i l’àcid hialurònic (HyA) van ser seleccionats per la seva capacitat per formar complexes iònics estables amb surfactants catiònics per generar materials estables amb noves propietats. Aquests complexes es preparen fàcilment i tendeixen a autoassemblar-se en estructures amfifíliques ordenades, capaços de respondre reversiblement a efectes tèrmics. Aquest comportament és d’elevat interès científic i de rellevància pràctica en el disseny de dispositius mèdics per l’emmagatzematge i alliberament de fàrmacs de manera tèrmica i químicament controlada. El principal objectiu d’aquesta Tes i es la preparació de complexes iònics dels dos poliàcids mencionats utilitzant diferents surfactants catiònics depenent de les propietats finals desitjades. La primera part del treball està dedicada a proporcionar el coneixement físic-químic de l’estructura i propietats dels surfactants d’alquiltrimetilfos foni, els quals tenen un interès potencial per a noves aplicacions. Posteriorment, aquests surfactants foren acoblats als dos biopolímers, PGGA i HyA, per obtenir els respectius complexes iònics amb propietats biocides i de major estabilitat tèrmica que aquells complexes preparats pels seus anàlegs d’amoni. Els complexes de PGGA, tenen un alt interès com a preservadors d’aliments i aplicacions d’envasat tenint com a principal avantatge la comestibilitat d’aquest polímer i la possibilitat de millorar les seves propietats bàsiques mesclant aquests compostos amb nanoargiles. Per altra banda, els complexes de HyA, són útils per obtenir derivats de l’àcid hialurònic amb activitat antimicrobiana. A més, s’ha dut a terme la preparació de nanopartícules de nATMP·HyA amb propietats antimicrobianes utilitzant el mètode “ionotropic gelation”. La segona part de la Tesi es va focalitzar en la preparació de complexes sostenibles d’àcid hialurònic. Per a realitzar-ho es van utilitzar surfactants d’alcanoilcolines per preparar els complexes iònics nACh·HyA. Aquests complexes constitueixen una prometedora plataforma biocompatible/biodegradable pel disseny de sistemes capaços de transportar i alliberar de forma específica els fàrmacs anticancerígens de quimioteràpia. La tercera part de la Tesi està dedicada a explorar la preparació de complexes biocompatibles amb propietats antimicrobianes utilitzant com a surfactant catiònic un dels preservants alimentaris més potents que es coneixen actualment, el LAE. Aquests complexes (LAE·PGGA i LAE·HyA) són candidats potencials per desenvolupar films comestibles amb propietats microbianes i materials d’àcid hialurònic amb propietats microbianes respectivament

Our research mainly consisted by two parts. First, apply p-norm error measure instead of 1-norm measure in a linear programming discrimination, which generates a linear hyperplane to classify two data sets. With this p-norm error measure, the errors generated by the classifier are not treated equally but rather biased. For 1, the bigger one error is, the more weight it obtains in the objective function. Second, investigation is conducted on a psychophysiological data set. Various methods are tested on this multi-dimensional time-series data set, from the linear programming method to the neural network method. With the help of DFT, The data is able to be transferred from the time domain to the frequency domain, in which the data set has interesting patterns

The aim of this thesis is to push forward the synthesis of well-defined materials containing polar monomers. The ATRP of polar monomers was investigated with the aim to obtain living and well-defined materials. Block copolymers with pre-determinable composition and unimodal distribution of molecular weight were synthesized. Furthermore, the Atom Transfer Radical Co-Polymerization of NVCL and NVP with non-polar monomers was investigated with the aim to obtain amphiphilic material with tunable polarity. The ATRP of vinyl acetate (VAc), which was poorly optimized, was studied trying to obtain poly(VAc) with low polydispersity (<1.25), pre-determinable molecular weight and living character. The optimization of the ATRP of VAc and the synthesis of several block copolymers, synthesized in presence of different experimental conditions, can significantly expand the field of materials and applications of poly(VAc) and poly(vinyl alcohol)-based products. Moreover, the synthesis of pH and temperature polymers was investigated with the aim to obtain products suitable for the development of drug-delivery systems which can be applied in anti-cancer applications. For this purpose Pluronic F127, which is thermosensitive, and poly(ethylene glycol)s were modified with pH poly[2-(N,N-dimethylamino)ethyl methacrylate] (PDMAEMA), poly[2-(N,N-diethylamino)ethyl methacrylate] (PDEAEMA) and poly[2-(N,N-diisopropylamino)ethyl methacrylate] (PDIAEMA). The methacrylic moieties have different pKa, and they give to the synthesized materials the desired pH responsiveness. The gelation behavior of the obtained products was investigated by rheological measurements; the dimension of the polymeric aggregates in water solutions at different pH was studied by DLS and the drug-incorporation as a function of pH was determined in systems with stable pH and in systems in which the pH was decreased progressively. All the cited investigation allowed to well-characterized the behavior and the structure of polymeric aggregates in water solution and they also allowed to determine their pH and temperature responsiveness.

The aim of this thesis is to push forward the synthesis of well-defined materials containing polar monomers. The ATRP of polar monomers was investigated with the aim to obtain living and well-defined materials. Block copolymers with pre-determinable composition and unimodal distribution of molecular weight were synthesized. Furthermore, the Atom Transfer Radical Co-Polymerization of NVCL and NVP with non-polar monomers was investigated with the aim to obtain amphiphilic material with tunable polarity. The ATRP of vinyl acetate (VAc), which was poorly optimized, was studied trying to obtain poly(VAc) with low polydispersity (<1.25), pre-determinable molecular weight and living character. The optimization of the ATRP of VAc and the synthesis of several block copolymers, synthesized in presence of different experimental conditions, can significantly expand the field of materials and applications of poly(VAc) and poly(vinyl alcohol)-based products. Moreover, the synthesis of pH and temperature polymers was investigated with the aim to obtain products suitable for the development of drug-delivery systems which can be applied in anti-cancer applications. For this purpose Pluronic F127, which is thermosensitive, and poly(ethylene glycol)s were modified with pH poly[2-(N,N-dimethylamino)ethyl methacrylate] (PDMAEMA), poly[2-(N,N-diethylamino)ethyl methacrylate] (PDEAEMA) and poly[2-(N,N-diisopropylamino)ethyl methacrylate] (PDIAEMA). The methacrylic moieties have different pKa, and they give to the synthesized materials the desired pH responsiveness. The gelation behavior of the obtained products was investigated by rheological measurements; the dimension of the polymeric aggregates in water solutions at different pH was studied by DLS and the drug-incorporation as a function of pH was determined in systems with stable pH and in systems in which the pH was decreased progressively. All the cited investigation allowed to well-characterized the behavior and the structure of polymeric aggregates in water solution and they also allowed to determine their pH and temperature responsiveness.

The possibility to tune material properties up to nanoscale represents a great opportunity for the scientific community to obtain devices capable to fulfill the always new medical demands. During this Thesis project micro and nano-structured polymeric materials have been used in the field of drug delivery and tissue engineering. Three different research lines have been explored: (i) the use of polymeric fibrous systems as drug and nanoparticles carriers, (ii) design and evaluation of novel shape memory polymers to produce shape memory scaffolds and (iii) development of smart affinity membranes. Electrospinning were exploited to obtain polymeric fibrous carriers made of different biodegradable and bioresorbable polymers, such us Poly(lactic-acid) and Poly(lactic-co-glycolic). The obtained biodegradable carriers have been exploited to achieve controllable particles release as well as, to obtain composites capable to deliver two drugs simultaneously with controllable and predictable kinetics. The possibility to obtain electrospun scaffolds capable of interconverting between a temporary and a permanent shape with the application of a thermal stimulus was explored. In this context, two polymers have been designed to behave as shape memory materials in the range of human body temperature. Finally smart affinity membranes have been studied. This kind of materials are capable to detect specific molecules or biomacromolecules from complex mixtures, finding useful applications in the biomedical field as diagnostic and therapeutic devices. Smart affinity membranes might be used for example to detect specific kind of cells by exploiting the binding interaction between an antibody and cell receptors. During this thesis project poly(L-lactic acid) electrospun scaffolds conjugated with antibodies have been produced and the efficacy of different functionalization methods to generate the –COOH group necessary to perform the antibodies conjugation was investigated.

Nella ricerca scientifica e medica, l'uso degli animali ha apportato notevoli benefici all’uomo. Sebbene la loro fisiologia non imiti perfettamente il corpo umano, essi agiscono come perfetti "modelli" per lo studio delle malattie umane e lo sviluppo di nuovi farmaci e trattamenti. Tuttavia, secondo la Food and Drug Administration (FDA), solo l'8% dei farmaci testati sugli animali è considerato sicuro ed efficace per l'uso sull’essere umano, il 92% non lo è. Questi tassi di fallimento estremamente elevati nello sviluppo di farmaci, insieme ai costi in forte aumento, hanno portato molti ricercatori a rivalutare il valore degli studi in vivo sugli animali. Questo è stato il motivo per cui, negli ultimi tre decenni, gli scienziati hanno lavorato duramente per trovare valide alternative alla sperimentazione animale in vivo, che potessero essere più efficaci, più affidabili e più umane. Lo sviluppo e l'uso di modelli in vitro sono diventati rapidamente un'alternativa molto popolare agli esperimenti sugli animali nella ricerca farmaceutica e biomedica. Essi coinvolgono la crescita di cellule al di fuori del corpo (umano o animale) in un ambiente chiuso e controllato. Tra i diversi tipi di approcci in vitro studiati nel corso degli anni, la medicina rigenerativa e l'ingegneria dei tessuti hanno proposto la coltura cellulare 3D come un passaggio cruciale per colmare il divario tra i risultati in vitro e in vivo nei test preclinici. In particolare, la coltura cellulare in ambiente dinamico, attraverso l’utilizzo di bioreattori, è stata considerata un valido passaggio intermedio tra l'approccio statico convenzionale in vitro e gli studi sugli animali in vivo. La progettazione di un bioreattore dovrebbe ispirarsi alle osservazioni dell'ambiente nativo, cercando di riprodurre i numerosi e complessi stimoli che agiscono a livello cellulare, in modo controllato e modulabile. Tutti questi aspetti rendono ancora oggi lo sviluppo e l’utilizzo dei bioreattori una grande sfida per i bioingegneri. Nel mondo della ricerca, nel frattempo, hanno fatto la loro comparsa le nanotecnologie e i nanomateriali, aiutando e migliorando le scienze attraverso la manipolazione della materia su scala nanometrica. L’implementazione della nanotecnologia nei campi della biologia e della medicina ha avuto un impatto positivo sulla salute umana. La nanobiotecnologia possiede un enorme potenziale per le scienze mediche, grazie all’utilizzo di nanoparticelle come nuove sonde per il “bioimaging”, come potenziali biosensori e mezzi per la somministrazione di farmaci in modo più efficace a livello cellulare, e infine per diagnosticare le malattie in vitro in modo più rapido e sensibile. Il seguente studio è centrato sulla progettazione e lo sviluppo di un bioreattore a perfusione dotato di doppio flusso, in cui un’efficiente e regolabile perfusione a flusso radiale viene applicata per consentire la diffusione di nano-sistemi attraverso matrici polimeriche microporose. Il lavoro proposto mostra anche un'ampia sperimentazione riguardante la sintesi di strutture polimeriche al fine di realizzare un modello 3D in vitro altamente performante per le coltura cellulari. Tutto questo, insieme alla produzione di potenziali nanoparticelle utili per molteplici applicazioni, come sonde per bioimaging o vettori per la somministrazione di farmaci. Con la combinazione di micro- e nano-strutture in un ambiente controllato, il bioreattore è proposto come un potenziale apparato in vitro per nuovi test pre-screening di farmaci e per studiare nuove terapie, con più benefici costi/tempo e meno problemi etici rispetto a modelli animali in vivo.
The use of animals in science and medicine research has significantly benefited human beings. Although their physiology does not identically mimic the human body, they act as perfect “models” for studying human diseases and developing potential new drugs and medical treatments. However, according to the U.S. Food and Drug Administration (FDA), just 8% of drugs tested on animals are considered safe and effective for human use, 92% are not. These extremely high failure rates together with dramatically rising costs in drug development have led many researchers to re-evaluate the value of animal studies. Because of that, over the past three decades, scientists worked hard for finding valid alternatives to in vivo animal testing that could be more effective, more reliable, and more humane. The development and use of in vitro models, which involve the growth of cells outside the body in a closed and controlled environment, quickly became a very popular alternative to animal experiments in pharmaceutical and biomedical research. Among the variety of in vitro approaches studied during the years, 3D in vitro cell culture was proposed by Regenerative medicine and Tissue engineering as a crucial and essential step for bridging the gap between the in vitro and in vivo outcomes in pre-clinical testing. Particularly, dynamic cell culture using bioreactors was considered a good intermediate step between the conventional in vitro static approach and in vivo animal studies. The design of a bioreactor should be inspired by observations of the native environment trying to reproduce the numerous and complex stimuli on cell behaviour in a controlled and scalable way. All these aspects still make the design and operation of bioreactors a big challenge for bioengineers. In the meantime, Nanotechnology and nanomaterials came into play helping and improving the life sciences with the manipulation of matter over the scale of a nanometer. The subsequent implementation of nanotechnology in biology and medicine fields showed an enormous positive impact on human health. Nanobiotechnology held enormous potential for healthcare, by using nanoparticles as new probes for bioimaging, potential biosensors and vehicles for delivering drugs more effectively in cellular systems, and in vitro diagnosing diseases more rapidly and sensitively. This study is focused on the design and development of an advanced dual-flow perfusion bioreactor, in which an efficient and tunable radial flow perfusion is applied to enable the diffusion of nano-systems through micro-porous polymeric matrices. At the same time, the proposed work presents a wide investigation concerning the synthesis of “ad hoc” polymeric structures for achieving the most performant 3D in vitro model for cell culture, together with the production of powerful and versatile nanoparticles useful for multiple applications as probes for bioimaging or vectors for drug delivery. With the combination of micro- and nano-structures in a controlled environment, the bioreactor is suggested here as a potential in vitro apparatus for new drug pre-screening testing and for studying new human therapies with less cost/time consuming and less ethical concerns compared to in vivo animal models.

This thesis describes two main sections of work, an examination of a commercial product, Intrasite Gel, and the development of an algorithm for variable selection using projected latent structures.Following on from the successful development of a variable selection procedure for multivariate linear regression this work looks at transferring this idea for use with projected latent structures. The first part of this thesis will show how the variable selection algorithm was developed and used with three different data sets. The algorithm will be shown to be superior to standard projected latent structures, for linear multi-component data. Although the final algorithm developed requires considerable computing resources to carry out this is compensated for by significantly improved model predictions and robustness. The final algorithm developed is written to run using MATLAB on any computer platform that supports this application, though the principles of operation could be transferred to another method of execution, for example custom code written in C or Pascal. The approach used in the development of this method is that the ability of the model to predict unknownsamples is of far greater importance than the internal performance of the model. All the assessments of the procedures developed are based on the ability of the model to predict accurately and precisely samples that were not presented to the model during the training stage.The second section of this thesis is concerned with the study of Intrasite Gel, produced by Smith & Nephew Ltd. Hull. The material in question is a medical device intended to assist in the treatment and healing of wounds that are necrotic, sloughy or granulating. The product is characterised by its ability to maintain moisture equilibrium in a wound environment and to provide a suitable medium to encourage the growth of new cell tissue. Medical devices require registration, and as part of that registration a number of tests are made on samples to ensure that the material meets the required specifications. There was some concern at Smith & Nephew that the tests they were required to carry out as part of the device registration were not providing appropriate information about the product. Of particular interest was the fluid absorption property as it was suspected that the test has a large amount of random error associated with it and an investigation was required to examine this test and to provide an alternative procedure should the fluid absorption test prove inadequate. Also of interest to Smith & Nephew was the issue of sampling frequency, as it was felt that this should also be examined to determine whether the correct rate of sampling to ensure product quality was being carried out. The work reported here shows that the fluid absorption test as it stands is insufficient to the task of monitoring this property of Intrasite gel and that an alternative test should be considered. This work also showed that current sampling rate was too high and that the high sampling rate may in fact cause misleading assumptions as to the stability and quality of the product.

Modern day medicine is on the brink of a new age of therapy, which aims to harness the natural power of molecular biology for disease treatment. This therapy could include replacement of dysfunctional genes that cause disorders such as cystic fibrosis (Lommatzsch and Aris, 2009), or silencing the overexpression of genes that cause disorders such as cancer (Pelengaris and Khan, 2003). In both examples, the treatment of these genetic diseases lies in the delivery of synthetic nucleic acids into diseased cells, the former being called gene replacement therapy (Dobson, 2006a), and the latter being called RNA interference (RNAi) therapy (Whitehead et al., 2009). While these techniques have long been in use as genetic research tools for gene transfection or silencing in vitro, their translation for use in clinical disease treatment has yet to be achieved. The main problem facing the development of these novel therapies is the specific delivery of nucleic acids into diseased cells within the body. It is hoped that nanoparticles (NPs) can be used to overcome this problem, by acting as vehicles to transport nucleic acids through the body for specific delivery into diseased cells. This feat can be aided by the attachment of additional functional molecules such as cell penetrating peptides (CPPs), targeting peptides, additional drug types and molecules for imaging during treatment. Many different NP design strategies are currently under development. It is essential for new designs to be extensively tested for toxicity and efficiency in human cells before they can be successfully released into the clinic. As part of this effort, this PhD project has investigated two different NP design strategies for drug delivery: 1) the use of a magnetic field (MF) and a CPP to increase the delivery of iron oxide magnetic NPs (mNPs) to cells grown in tissueequivalent 3D collagen gels, and 2) gold NPs (AuNPs) for the delivery of siRNA to silence the c-myc oncogene for cancer treatment. In the first investigation, a MF and the CPP penetratin were found to increase mNP delivery to cells grown in 3D. In the second investigation, AuNPs were assessed in a range of different cell types (grown in 2D) for their performance in 4 main areas; cellular toxicity, cellular uptake, c-myc knockdown and effect on the cell cycle.

Surface topography is known to be important in biomedical applications such as scaffolds for tissue regeneration and has been shown to affect wettability and cell behaviour. Traditionally, topographical effects such as surface texturing have been generated using methods such as photolithography, soft lithography, thermal embossing, and laser/electron beam techniques. This thesis introduces a relatively new technique known as photoembossing to create surface texturing for biomedical applications. Photoembossing is used to produce surface texturing on polymer surfaces by patterned ultraviolet (UV) exposure of a photopolymer blend without an etching step or an expensive mould. After a short general introduction and a literature review, the first experimental chapters describe surface patterning of poly(methyl methacrylate) (PMMA) photopolymer substrates by photoembossing. PMMA is blended with an acrylate monomer and photoinitiator by dissolution in a volatile solvent and processed into films by wire bar coating, and fibres are produced by electrospinning. Surface texture is achieved on both films and fibres by photoembossing. Endothelial cell culture shows that the substrates are biocompatible and cells readily adhere to the surface. In tissue regeneration applications, scaffold degradation is often important to allow tissue in-growth. Thus, in subsequent studies polylactide-co-glycolide (PLGA) is used as a polymer binder. PLGA blended with a triacrylate monomer showed partial degradation after 10 weeks, with a cross-linked acrylate network remaining. Endothelial cell adhesion was even better on the PLGA photopolymer substrates compared to PMMA. Furthermore, surface texture improved cell adhesion and proliferation on the PLGA photopolymer. To obtain completely degradable substrates, thiol monomer was used in addition to the acrylate to produce ester bonds after the thiol-ene reaction, which is cleavable by hydrolysis. Accelerated degradation in sodium hydroxide (NaOH) showed complete degradation of this photopolymer system. The degradation rate of the photopolymer could be tuned by the molecular weight of the acrylate monomer, with low molecular weight monomers degrading more slowly than high molecular weight species. Furthermore, the height of the surface relief structures could be enhanced by using low-molecular-weight acrylate monomers. Endothelial cell culture revealed biocompatibility of the blend and cells were able to adhere after 24 hours of seeding. This thesis demonstrates that photoembossing is a viable technique in producing surface texture for tissue engineering applications. This surface texture can be achieved on both biocompatible and biodegradable photopolymer films and fibres.

Chitosan, a copolymer of glucosamine and N-acetyl glucosamine, is a polycationic, biocompatible and biodegradable polymer. In addition, chitosan has different functional groups that can be modified with a wide array of ligands. Because of its unique physicochemical properties, chitosan has great potential in a range of biomedical applications, including tissue engineering, non-viral gene delivery and enzyme immobilization. In our work, the primary amine groups of chitosan were utilized for chitosan modification through biotinylation using N-hydroxysuccinimide chemistry. This was followed by the addition of avidin which strongly binds to biotin. Biotinylated ligands such as polyethylene glycol (PEG) and RGD peptide sequence, or biotinylated enzymes such as trypsin, were then added to modify the surface properties of the chitosan for a variety of purposes. Modified chitosans were formulated into nano-sized particles or cast into films. Different factors affecting fabrication of chitosan particles, such as the pH of the preparation, the inclusion of polyanions, the charge ratios and the degree of deacetylation and the molecular weight of chitosan were studied. Similarly, parameters affecting the fabrication of chitosan films, such as cross-linking, were investigated for potential applications in tissue engineering and enzyme immobilization. It was found that the inclusion of dextran sulfate resulted in optimum interaction between chitosan and DNA, as shown by the high stability of these nanoparticles and their high in vitro transfection efficiencies in HEK293 cells. When applying these formulations as DNA vaccines in vivo, chitosan nanoparticles loaded with the ovalbumin antigen and the plasmid DNA encoding the same antigen resulted in the highest antibody response in C57BL/6 mice. Furthermore, engineering of the surface of chitosan nanoparticles was done by utilizing the avidin-biotin interaction for attaching PEG and RGD. The modified formulations were tested for their in vitro gene delivery properties and it was found that these ligands improved gene transfection efficiencies significantly. Chitosan nanoparticles were optimized further for enzyme immobilization purposes using sodium sulfate and glutaraldehyde as physical and chemical cross-linking agents, respectively. These particles and chitosan films were used for immobilizing trypsin utilizing several techniques. Enzyme immobilization via avidin-biotin interaction resulted in high immobilization efficiency and high enzymatic activity in different reaction conditions. Additionally, the immobilized trypsin systems were stable and amenable to be regenerated for multiple uses. Finally, glutaraldehyde cross-linked chitosan films were modified with PEG and RGD for their cell repellant and cell adhesion properties, respectively, using avidin-biotin interaction. This method was again effective in engineering chitosan surfaces for modulating cell adhesion and proliferation. In conclusion, using avidin-biotin technique to modify biotinylated chitosan surfaces is a facile method to attach a wide variety of ligands in mild reaction conditions, while preserving the functionality of these ligands.

[Truncated abstract] In the past decade, the synthesis of superparamagnetic iron oxide nanoparticles (SPIONs) has received considerable attention due to their potential applications in biomedical fields. However, success in size and shape control of the SPIONs has been mostly achieved through organic routes using large quantities of toxic or/and expensive precursors in organic reaction medium at high reaction temperature. This has limited the biomedical applications of SPIONs and therefore, development of a synthetic method under aqueous condition that is reproducible, scalable, environmentally benign, and economically feasible for industrial production is of paramount importance in order to fully realise their practical applications. Spinning Disc Processing (SDP) has been used to synthesise superparamagnetic magnetite (Fe3O4) nanoparticles at room temperature via a modified chemical precipitation method under continuous flow condition and offer a potential alternative to be applied to SPIONs production. SDP has extremely rapid mixing under plug flow conditions, effective heat and mass transfer, allowing high throughput with low wastage solvent efficiency. The synthesis process involves passing ammonia gas over a thin aqueous film of Fe2+/3+ which is introduced through a jet feed close to the centre of a rapidly rotating disc (500-2500 rpm). Synthetic parameters such as precursor concentrations, temperature, flow rate, disc speed, and surface texture influence the particle sizes. ... Magnetic silica microspheres are receiving great attention for possible applications in magnetic targeting drug delivery, bioseparation and enzyme isolation. However, the current available methods for preparation suffer from the setback of low loading of Fe3O4 nanoparticles in the silica microsphere, which result in low magnetic moment, thereby limiting their practical applications. Therefore it is of considerable importance to develop new alternative synthetic methods for fabricating magnetic silica microspheres with high magnetic nanoparticles loading. Superparamagentic Fe3O4 nanoparticles (8-10 nm diameter) and curcumin have been encapsulated in mesoporous silica in a simple multiplestep self assembly approach process with high Fe3O4 nanoparticles loading (37%). The synthesis involves loading of curcumin in the Cetyltrimethylammonium bromide (CTAB) micellar rod in the presence of superparamagnetic Fe3O4 nanoparticles via a parallel synergistic approach. The synthesised magnetic mesoporous silica composite material is stable, superparamagnetic with high saturation magnetisation before and after curcumin leaching experiment. Under physiological pH in phosphate buffer, the curcumin is slowly released over several days. These magnetic mesoporous silica are expected to have great potential as targeted drug delivery systems.

Switchable oligopeptides, able to expose of conceal biomolecules on a surface, upon the application of an electrical potential, represent a versatile tool for the development of novel devices, presenting potential biomedical applications. Recently, several studies have demonstrated the applicability of smart devices for the control of protein binding and cellular response. In this work; a detailed analysis of the steric requirements necessary to develop a mixed oligopeptide Self-Assembled Monolayer (SAM) presenting an optimum switching ability will be described. The influence of both the SAM components surface ratio and the switching unit length on the mixed SAMs switching performance will be investigated. The findings of this investigation will be used to develop, for the first time, a platform, based on electrically switchable oligopeptides, able to control the interaction between an antigen and its relative antibody. The influence of the biological medium on the oligopeptide switching ability will also be investigated. Finally, an orthogonal functionalisation strategy, will be investigated in detail, together with a new platform able to promote human sperm cells adhesion. The results of this research thesis will also represent the first building blocks towards the development of glass-gold rnicropattemed surfaces able to control the calcium signalling in human sperm cells, presenting potential applications in the improvement of in-vitro fertilisation (NF) treatments success rates.

Thesis: Ph. D., Massachusetts Institute of Technology, Department of Materials Science and Engineering, 2017.
This electronic version was submitted by the student author. The certified thesis is available in the Institute Archives and Special Collections.
Cataloged from student-submitted PDF version of thesis.
Includes bibliographical references (pages 170-176).
Research on biomedical applications of magnetic nanoparticles (MNPs) has increasingly sought to demonstrate noninvasive actuation of cellular processes and material responses using heat dissipated in the presence of an alternating magnetic field (AMF). By modeling the dependence of hysteresis losses on AMF amplitude and constraining AMF conditions to be physiologically suitable, it can be shown that MNPs exhibit uniquely optimal driving conditions that depend on controllable material properties such as magnetic anisotropy, magnetization, and particle volume. "Magnetothermal multiplexing," which relies on selecting materials with substantially distinct optimal AMF conditions, enables the selective heating of different kinds of collocated MNPs by applying different AMF parameters. This effect has the potential to extend the functionality of a variety of emerging techniques with mechanisms that rely on bulk or nanoscale heating of MNPs. Experimental investigations on methods for actuating deep brain stimulation, drug release, and shape memory polymer response are summarized, with discussion of the feasibility and utility of applying magnetothermal multiplexing to similar systems. The possibility of selective heating is motivated by a discussion of various models for heat dissipation by MNPs in AMFs, and then corroborated with experimental calorimetry measurements. A heuristic method for identifying materials and AMF conditions suitable for multiplexing is demonstrated on a set of iron oxide nanoparticles doped with various concentrations of cobalt. Design principles for producing AMFs with high amplitude and ranging in frequency from 15kHz to 2.5MHz are explained in detail, accompanied by a discussion of the outlook for scalability to clinically relevant dimensions. The thesis concludes with a discussion of the state of the field and the broader lessons that can be drawn from the work it describes.
by Michael G. Christiansen.
Ph. D.

Thesis: S.M. in Engineering and Management, Massachusetts Institute of Technology, System Design and Management Program, 2018.
This electronic version was submitted by the student author. The certified thesis is available in the Institute Archives and Special Collections.
Cataloged from student-submitted PDF version of thesis.
Includes bibliographical references (pages 77-79).
Identifying patients with aggressive cancers is a major healthcare challenge in resource-limited settings such as sub-Saharan Africa. Holographic imaging techniques have been shown to perform diagnostic screening at low cost in order to meet this clinical need, however the computational and logistical challenges involved in deploying such systems are manifold. This thesis aims to make two specific contributions to the field of point-of-care diagnostics. First, it documents the design and construction of low-cost holographic imaging hardware which can serve as a template for future research and development. Second, it presents a novel deep-learning architecture that can potentially lower the computational burden of digital holography by replacing existing image reconstruction methods. We demonstrate the effectiveness of the algorithm by reconstructing biological samples and quantifying their structural similarity relative to spatial deconvolution methods. The approaches explored in this work could enable a standalone holographic platform that is capable of efficiently performing diagnostic screening at the point of care.
by Ismail Degani.
S.M. in Engineering and Management

Measuring chemical concentration is vital for understanding normal and disease physiology involving metabolism and signalling, but monitoring chemical concentrations in living systems poses a unique challenge because of biological heterogeneity. The purpose of this work is to develop a system able to monitor chemical concentrations in primary cultured cells, and apply it to the detection of oxygen, a nutritional status marker, and nitric oxide which is a signalling molecule. Both of these electroactive species are involved in angiogenesis which is the growth of new blood vessels, and a hallmark of cancer. The approach used in this study is to grow porcine endothelial cells onto bronectin-coated gold microelectrode arrays with diameter 25 μm and perform electrochemical measurement on them. An experimental protocol is developed for measurements of dissolved oxygen and nitric oxide around cells in their normal cell-culture environment. It includes developing instrumentation like a heating platform and silver reference microelectrode; data processing for automation and normalisation; and optimising voltammetry techniques. Culture medium is found to a ect electrochemical measurements by changing double layer capacitance, reaction rate constant and di usion. The measurement system is used to detect oxygen reduction around cells, and this is used to estimate their oxygen consumption rate. Nitric oxide produced by cells is also measured, and this is used to identify an angiogenic pathway leading to nitric oxide production by endothelial cells. Variability in cell measurements is shown to originate from the biological system rather than from sensor design. A novel electroanalytical technique for determining parameters of reversible redox systems is developed by experimentally testing an analytical solution for the current response to a large-amplitude sinusoidal voltage input. The technique is used to nd estimates for double layer capacitance, half wave potential and di usion coe cients for both potassium ferrocyanide and ruthenium hexaamine.

Dans cette thèse, nous nous sommes intéressés aux systèmes intelligents pour des applicationsmédicales et cosmétiques. Ainsi, nous avons conçu et réalisé trois instruments. Le premier estdédié à la mesure de la mouillabilité de la peau. L’originalité de ce dispositif réside en sa capacité àdonner une image 3D de la goûte de la surface de la peau explorée et de donner le comportementdynamique de la goûte. Cette stratégie nous donnera la possibilité de créer de nouvelles basesde données relatives à la mouillabilité de tout le corps humain. En effet, nous disposons que desdonnées sur la mouillabilité de l’avant-bras. Le deuxième instrument intelligent concerne la mesurede la réflectance d’une surface. Ce dispositif assure une mesure de très haute résolution angulairede la BRDF et une très bonne répétabilité de la mesure. Il a été validé sur la peau pour la mesurede l’ éclat. Et enfin le troisième instrument, basé sur une méthode originale de mesure de vibrationà l’aide d’un système de stéréo-vision associée à un motif périodique. Il a été appliqué pour lamesure du mouvement thoracique et abdominal lors de la respiration. Notre principale motivationpour développer ce système fut la réduction des artefacts, dus aux mouvements d’un patient lorsd’un examen radiologique
Smart Devices have been widely used by health care and cosmetics professionals. Indeed, they helpin many aspects of clinical practice by providing an efficient way for medical diagnosis, supportingbetter clinical decision-making and improving patient outcomes. In this thesis, we have beeninterested in three applications. The first one is related to the wettability measurement, especially forthe human skin. So we propose, a held-hand device that is based on the contact angle measurementto determine skin wettability. Besides, the device allows the visualization of the liquid dropletspreading in both dynamic and static modes. Moreover, it can measure the top and the left views ofthe droplet and provides the 3D droplet and the skin explored area profiles. The second applicationpermits the skin radiance measurement. For this purpose, we propose a miniaturized device havingan original method for the BRDF measurement associated with 3D profile measurement of the areastudied. As regards the third application, it is a non-invasive method for breath measurement that usesa stereovision system and a pseudo-periodic pattern. This system allows a high-resolution threedimensionaldisplacement measurement for the recording of the thoracoabdominal wall respiratorymovement. The devices developed during this research gives us a high accuracy, a good resolutionand repeatability of measurements

Body-Centric Wireless Communication (BCWC) is a central topic in the development of healthcare and biomedical technologies. Increasing healthcare quality, in addition to the continuous miniaturisation of sensors and the advancement in wearable electronics, embedded software, digital signal processing and biomedical technologies, has led to a new era of biomedical devices and increases possibility of continuous monitoring, diagnostic and/or treatment of many diseases. However, the major difference between BCWC, particularly implantable devices, and conventional wireless systems is the radio channel over which the communication takes place. The human body is a hostile environment from a radio propagation perspective. This environment is a highly lossy and has a high effect on the antenna elements, the radio channel parameters and, hence a dramatic drop in the implanted antenna performance. This thesis focuses on how to improve the gain of implanted antennas. In order to improve the gain and performance of implanted antennas, this thesis uses a combination of experimental and electromagnetic numerical investigations. Extensive simulation and experimental investigations are carried out to study the effects of various external elements on the performance improvement of implanted antennas. The thesis also shows the design, characterisation, simulation and measurements of four different antennas to work at ISM band and seventeen different scenarios for body wireless communication. A 3- layer (skin, fat and muscle) and a liquid homogenise phantom were used for human body modelling in both simulation and measurements. The results shows that a length of printed line and a grid can be used on top of the human skin in order enhance the performance of the implanted antennas. Moreover, a ring and a hemispherical lens can be used externally in order to enhance the performance of the implanted antenna. This approach yields a significant improvement in the antenna gain and reduces the specific absorption rate (SAR) in most cases and the obtained gain varies between 2 dB and 8 dB.

Biocompatible polymers are used exhaustively within the biomedical arena, demonstrating a mechanical and chemical diversity that few other materials possess. As polymer technologies evolves to cater for new medical demands, even the most niche biomedical application becomes an achievable reality. However, the discovery of new polymers is hindered by the complexity and intricacy in which the biological milieu interacts with a new substrate, reducing the ability to predict the appropriateness of a certain polymer for a specific application. This drawback can be countered by the high-throughput evaluation of large numbers of chemically diverse polymer candidates. In this thesis, the use of polymer microarrays is invoked to address two separate medically-relevant issues: the control of inflammation, and the improvement of cancer screening. In addition, I provide details of how polymer microarray techniques and technology can be employed to expand the repertoire of biomaterials research. Mitochondrial DNA (mtDNA) is an alarm molecule that contributes to the cytokine storm observed during severe tissue injury. An application where control of this systemic inflammation is achieved through scavenging of mtDNA by a polymer was proposed. Primary screening highlighted that 166 out of the 380 polymers evaluated bound to blood cells, making them unsuitable for a blood-based application. The remaining 214 blood-compatible polymers were cross-examined for mtDNA binding. Through polymer microarray and subsequent scale-up of promising candidates, a poly(methoxyethyl methacrylate-co-di(ethylamino)ethyl acrylate-co-methoxyethyl acrylate) was found to have a remarkable ability to scavenge mtDNA. Removal of cell-free mtDNA using this polymer is proposed to remove a key trigger of systemic inflammation. Cervical cancer screening includes the cytological evaluation of patient material for developed or developing abnormalities. An application was sought that would enrich for cancerous/pre-cancerous cells and improve upon current standards for detection. Four cancerous cervical cell lines (HeLa, CaSki, SiHa, and C33a) and four precancerous cell lines (W12E, W12G, W12GPX, and W12GPXY) were interrogated to identify polymers with consistent binding that may improve routine cytological evaluation. A short-list of 24 polymers was assembled, and cells from liquid based cytology samples from healthy patient were spiked with DiI-labelled cancerous/precancerous cells and the short-listed polymers were re-evaluated for preferential binding. An enrichment of abnormal cervical cells was observed with three polymers, which could form the foundation for improved screening resources. Inkjet printing can be a useful tool in developing patterned substrates, such as polymer microarrays. A piezoelectric drop-on-demand printer was used to explore the methods in which these can be fabricated. A wettability assay using picolitre volumes was developed and used to characterise O2 plasma treatment of glass slides. Additionally, the printing of a cell-binding polymer using this approach enabled the decoration of cells with precise spatial resolution.

In my PhD polymer microarrays have been central in discovery of new materials for cardiovascular repair, cartilage tissue engineering and bacteria resistant medical devices. This has led to the work described in the following four chapters of my thesis. In the first part of my thesis polymers for the development of novel heart valve leaflets were identified. Diseased heart valves are currently replaced with the either synthetic or bioprosthetic (acellular xenografts) valve prostheses. While synthetic prosthesis have excellent durability, thromboembolic complications are frequent, requiring patients to undergo lifelong anti-coagulation therapy. On the other hand, the leaflets of bioprosthetic valves undergo structural deterioration, resulting in the patients having to undergo follow-up replacement surgeries. In order to overcome these shortcomings, the aim of this part of my PhD was to discover polymers that will enable the development of a ‘bio-synthetic’ heart valve, with the durability of synthetic valves and the biocompatibility of bioprosthetic vales. Polymers that bind valve interstitial cells (cells with a plastic fibroblast / myofibroblast phenotype that renew the extracellular matrix components of the valve leaflets) and also enable stable expression of key markers were identified. Immunohistochemistry and RNA expression analysis identified polymers for coating 3-D scaffolds, with the coated scaffolds showing excellent cell invasion, viability and maintenance of valve interstitial cell markers. To mimic the regions of the valve leaflets with differing stiffness, the response of valve interstitial cells to substrate stiffness was studied with various crosslinked gels. Thus, polymeric gels, prepared with the same chemical composition but with different Young’s modulus (covering 3 orders of magnitude) showed valve interstitial cell attachment with the cells showing differing behaviour based on the stiffness of the gels. In the second part of this thesis, polymers were identified for cartilage repair. Hyaline articular cartilage has very low potential for self-renewal, therefore cell-based therapies with autologous chondrocyte implantation are desired. Due to limited availability from biopsies, chondrocytes have to be expanded by in vitro culture; and fully defined synthetic culture substrates are essential for regulatory approvals. Using the high throughput approach I identified ‘hit’ polymers that allowed adhesion, proliferation and long-term culture of primary human chondrocytes and also chondrocytes derived from Mesenchymal stem cells. 2-D scale-up identified 2 lead polymers that supported long-term attachment and maintenance of chondrocyte markers. Since prolonged monolayer culture is known to induce loss of chondrocyte phenotype (dedifferentiation), 3D versions of the polymers were prepared and their potential for their long-term maintenance of chondrocytes via immunohistochemistry and RNA expression was demonstrated. The 3D gels were also used to encapsulate chondrocytes and their long-term maintenance of phenotype within these matrices, offers the exciting possibility of using these matrices for cartilage regeneration. The third part and fourth parts of the thesis focussed on reducing medical device associated infections. Thus polymers identified that prevented binding of a variety of bacteria including clinical isolates from infected medical devices, were used to coat two commercially available central venous catheters resulting in up to 96% reduction in bacterial binding. This non-binding was enhanced by the generation of polymeric nanocapsules containing the anti-bacterial eugenol (or its natural source clove oil). A coating consisting of eugenol nanocapsules entrapped within an interpenetrating network of the best bacteria repellent polymer, allowed slow-release of eugenol and further improved its performance.

La resposta del nostre cos als biomaterials suposa una gran obstacle per la efectivitat de múltiples teràpies basades en biomaterials. Accionats per la absorció inespecífica de biomolècules a la superfície del material, barreres com el sistema immune o les superfícies mucoses eliminen els materials del cos, evitant que arribin al seu destí i realitzin la seva funció. Els materials zwitteriònics han emergit en els últims anys com a materials antiadherents prometedors per a superar les mencionades barreres. Tot i que molts sistemes han utilitzat els materials zwitteriònics com a recobriments, les seves propietats úniques de superhidrofilicitat i versatilitat química suggereixen múltiples beneficis en utilitzar-los com a material principal. Aquí, dos sistemes diferents basats en materials zwitteriònics són presentats. En primer lloc una plataforma de alliberament de fàrmac antiadherent basat en copolímers de bloc amfifílics (CBA) és desenvolupada. Els CBAs zwitteriònics són sintetitzats i optimitzats perquè s’auto-organitzin en nanopartícules zwitteriòniques. Les propietats antiadherents d’aquestes nanopartícules es demostren, al igual que el seu potencial per a esdevenir un sistema d’alliberament de fàrmac oral. Seguidament, el sistema s’utilitza com a portador per a fàrmacs contra la malària i el càncer. Les nanopartícules mostren internalització en eritròcits infectats per Plasmòdium, i nanopartícules carregades amb curcumina demostren la seva eficàcia contra la malària in vitro. S’observa absorció oral de polímer i curcumina in vivo utilitzant un model de ratolí, indicant el potencial del sistema per a esdevenir una teràpia oral contra la malària. Quan s’optimitza el sistema per la teràpia contra el càncer, nanopartícules carregades de Paclitaxel exhibeixen activitat anti-cancerígena en models in vitro de cèl·lules canceroses. En segon lloc, microrobots zwitteriònics no-immunogènics que poden evitar el reconeixement per el sistema immune són introduïts. Es desenvolupa una fotoresistència zwitteriònica per a la microimpressió de microrobots zwitteriònics a través de la polimerització de dos fotons amb una ample funcionalització: propietats mecàniques variables, anti-bioadhesió i propietats no-immunogèniques, funcionalització per a la actuació magnètica, encapsulació de biomolècules i modificació superficial per a l’alliberament de fàrmac. Els robots invisibles eviten que els macròfags del sistema immune els detectin després d’una inspecció exhaustiva (de més de 90 hores), fet que no s’ha aconseguit fins el moment en cap sistema microrobòtic. Aquests materials zwitteriònics versàtils eliminen un dels grans obstacles en el desenvolupament de microrobots biocompatibles, i serviran com una caixa d’eines de materials no-immunogènics per a crear robots biomèdics i altres dispositius per a la bioenginyeria i per a aplicacions biomèdiques.
La respuesta de nuestro cuerpo a los biomateriales supone un gran obstáculo para la efectividad de múltiples terapias basadas en los biomateriales. Accionados por la absorción de biomoléculas en la superficie del material, barreras como el sistema inmune o las superficies mucosas eliminan los materiales del cuerpo, evitando que lleguen a su destino y realicen su función. Los materiales zwitteriónicos han emergido en los últimos años como materiales antiadherentes prometedores para superar las barreras mencionadas. Aunque muchos sistemas utilizan materiales zwitteriónicos como recubrimientos, sus propiedades únicas de superhidrofilicidad i versatilidad química sugieren múltiples beneficios en utilizarlos como material principal. Aquí, dos sistemas basados en materiales zwitteriónicos son presentados. En primer lugar, una plataforma para la liberación de fármaco antiadherente basada en copolímeros de bloque amfifílicos (CBA) es desarrollada. Los CBA zwitteriónicos son sintetizados y optimizados para que se auto-organicen en nanopartículas zwitteriónicas. Las propiedades antiadherentes de estas nanopartículas son probadas, al igual que su potencial para convertirse en un sistema oral de liberación de fármaco. Seguidamente, el sistema se utiliza como portador para fármacos animalarios y anticancerígenos. Las nanopartículas muestran internalización en eritrocitos infectados por Plasmodio, y nanopartículas cargadas con curcumina demuestran su eficacia contra la malaria in vitro. Se observa la absorción oral de polímero y curcumina in vivo utilizando un modelo de ratón, indicando el potencial del sistema para convertirse en una terapia oral contra malaria. Cuando se optimiza el sistema para la terapia contra el cáncer, las nanopartículas cargadas con Paclitaxel exhiben actividad anticancerígena en modelos in vitro de células cancerosas. En segundo lugar, microrobots zwitteriónicos no-inmunológicos que pueden evitar el reconocimiento por parte del sistema inmune son introducidos. Se desarrolla una fotoresisténcia zwitteriónica para la microimpresión de microrobots zwitteriónicos a través de la polimerización de dos fotones con una amplia funcionalización: propiedades mecánicas variables, anti-bioadhesión i propiedades no-inmunogénicas, funcionalización para la actuación magnética, encapsulación de biomoléculas i modificación superficial para la liberación de fármaco. Los robots invisibles evitan que los macrófagos del sistema inmune innato los detecten después de una inspección exhaustiva (de más de 90 horas), hecho que no se ha conseguido hasta la fecha por ningún sistema microrobótico. Estos materiales zwitteriónicos versátiles eliminan uno de los grandes obstáculos en el desarrollo de microrobots biocompatibles, y servirán como una caja de herramientas de materiales no-inmunogénicos para crear robots biomédicos y otros dispositivos para la bioingeniería y para las aplicaciones biomédicas.
Body response to biomaterials suppose a major roadblock for the effectiveness of multiple biomaterial-based therapies. Triggered by unspecific absorption of biomolecules in the material surface, barriers such as immune system or mucosal surfaces clear foreign materials from the body, preventing them to reach their target and perform their function. Zwitterionic materials have emerged in the last years as promising antifouling materials to overcome the mentioned barriers. Although many systems have used zwitterionic materials as coatings, the unique properties of superhydrophilicity and chemical versatility suggest multiple benefits of using zwitterionic polymers as bulk materials. Here, two different systems based on zwitterionic materials are presented. In first place, an antifouling drug delivery platform based on zwitterionic amphiphilic polymers (ABC) is developed. Zwitterionic ABCs are synthetized and optimized to self-assemble in zwitterionic nanoparticles. The antifouling properties of zwitterionic nanoparticles are proved, together with their potential to become an oral drug delivery system. Next, the system is used as a drug carrier for antimalarial and anticancer drugs. Nanoparticles show internalization in Plasmodium infected erythrocytes, and curcumin-loaded nanoparticles prove their antimalarial efficacy in vitro. Oral absorption of polymer and curcumin is also observed in vivo using mice model, indicating the potential of this system to become oral therapy against malaria. When optimizing the system for anticancer therapy, Paclitaxel-loaded nanoparticles exhibit anticancer activity in in vitro cancer cell models. Second, non‐immunogenic stealth zwitterionic microrobots that avoid recognition from immune cells are introduced. Zwitterionic photoresist are developed for the 3D microprinting of zwitterionic hydrogel microrobots through 2-photon polymerization with ample functionalization: tunable mechanical properties, anti-biofouling and non-immunogenic properties, functionalization for magnetic actuation, encapsulation of biomolecules, and surface functionalization for drug delivery. Stealth microrobots avoid detection by macrophage cells of the innate immune system after exhaustive inspection (> 90 h), which has not been achieved in any microrobotic platform to date. These versatile zwitterionic materials eliminate a major roadblock in the development of biocompatible microrobots, and will serve as a toolbox of non-immunogenic materials for medical microrobot and other device technologies for bioengineering and biomedical applications.

Biomedicine is an area in which computers have long been expected to play a significant role. Although many of the early claims have proved unrealistic, computers are gradually becoming accepted in the biomedical, clinical and research environment. Within these application areas, expert systems appear to have met with the most resistance, especially when applied to image interpretations. In order to improve the acceptance of computerised decision support systems it is necessary to provide the information needed to make rational judgements concerning the inferences the system has made. This entails an explanation of what inferences were made, how the inferences were made and how the results of the inferences are to be interpreted. Furthermore there must be a consistent approach to the combining of information from low level computational processes through to high level expert analyses. Until recently ad hoc formalisms were seen as the only tractable approach to reasoning under uncertainty. A review of some of these formalisms suggests that they are less than ideal for the purposes of decision making. Belief networks provide a tractable way of utilising probability theory as an inference formalism by combining the theoretical consistency of probability for inference and decision making, with the ability to use the knowledge of domain experts. The potential of belief networks in biomedical applications has already been recognised and there has been substantial research into the use of belief networks for medical diagnosis and methods for handling large, interconnected networks. In this thesis the use of belief networks is extended to include detailed image model matching to show how, in principle, feature measurement can be undertaken in a fully probabilistic way. The belief networks employed are usually cyclic and have strong influences between adjacent nodes, so new techniques for probabilistic updating based on a model of the matching process have been developed. An inference shell called FLAPNet has been implemented and used to apply the belief network formalism to the tasks of model based image matching and the incremental aggregation of disparate information for diagnosis. The domains of application are fetal ultrasound imaging and cervical screening respectively. It is argued that belief networks combine the necessary quantitative features required of a decision support system with desirable qualitative features that will lead to improved acceptability of expert systems in the biomedical domain.

La télémétrie biomédicale et l’interfaçage neuronal à base de dispositifs miniatures et autonomes sans fil constituent de nouvelles applications en émergence. Elles visent à répondre à de nombreux enjeux y compris dans les domaines de la santé, du sport et bien être, ou encore de la sécurité au travail et de la défense. Parmi les applications typiques de biotélémétrie, nous pouvons citer le monitoring de certains paramètres physiologiques : température corporelle, pression artérielle, rythme cardiaque, taux de glucose et d’anticorps, détection d’agents chimiques, etc. En ce qui concerne l’interfaçage neuronal, il permet de restaurer les informations sensorielles, d’aider à la réadaptation des amputés, des personnes atteintes de paralysie ou des patients atteints de maladies neurodégénératives. L’objectif principal de cette thèse est de contribuer au développement de dispositifs miniaturisés et communicants pour le monitoring, en continu, de variables physiologiques d’humains ainsi que d’animaux. Ces dispositifs innovants nécessitent un système de communication fiable. Plus particulièrement, il s’agit d’analyser le milieu de propagation à l’intérieur des tissus biologiques et de développer des antennes miniatures innovantes ainsi que des méthodes pour leur analyse et leur caractérisation. Le verrou majeur concerne le rendement des antennes miniatures. Les effets de forte hétérogénéité, dispersion, pertes très élevées des milieux biologiques et les contraintes de miniaturisation et d’intégration dans des dispositifs in-body limitent la portée des systèmes existants à quelques dizaines de centimètres. Tout d’abord, des outils spécifiques de modélisation et d’optimisation ont été développés en collaboration avec l’Université de Bohème de l’Ouest. Ces outils sont indispensables pour l’analyse des composants de systèmes antennaires complexes : le code Agros2D (CAO interne) utilise des méthodes entièrement adaptatives. Cette approche permet de réduire la complexité d’optimisation des antennes in-body jusqu’un seul dégrée de liberté. Puis, la limite fondamentale de rendement des antennes pour les applications in-body a été définie ; les liens entre cette limite et la taille de l’antenne, sa fréquence de fonctionnement, la polarisation et les matériaux utilisés (dont hypothétiques) ont été quantifiés pour la première fois. Ce travail fondamental a d’abord pour objectif l’optimisation des performances de l’antenne actuelle de la capsule e-Celsius de l’entreprise BodyCAP pour accroître la portée de la gélule, en prenant en compte les caractéristiques des matériaux et le milieu de propagation que constituent les tissus biologiques. Dans cette étape on inclut également la fabrication des prototypes de gélules télémétriques ainsi que leurs mesures d’impédance. L’antenne optimisée a une portée trois fois plus importante que celle actuelle tout en occupant le même volume. En utilisant ces principes de conception, nous avons développé et caractérisé une antenne à 434 MHz adaptée à une large gamme d'applications in-body. Des dimensions ultra-miniatures, une robustesse et un rendement accrus permettent de l'utiliser à la fois pour des applications des capsules à implanter et à avaler. Enfin, en développant davantage les méthodes de conception et d’optimisation, nous avons conçu une antenne double-bande. Ayant la même robustesse que son équivalent actuel mono-bande, elle présente également un rendement encore plus élevé, permettant ainsi de fonctionner au-delà de 10 m. La caractéristique double-bande permet de concevoir les dispositifs in-body rechargeables sans fil dans le corps. Les antennes proposées contribuent au développement ultérieur d'une nouvelle génération de dispositifs miniatures in-body qui impliquent une intégration complexe et dense des capteurs, de la logique et de la source d'alimentation
Emerging wireless biotelemetry using miniature implantable, ingestible or injectable (in-body) devices allows continuously monitor and yield human or animal physiological parameters while maintaining mobility and quality of life. Recent advances in microelectromechanical systems and microfluidics—along with ongoing miniaturization of electronics—have empowered numerous innovations in biotelemetry devices, creating new applications in medicine, clinical research, wellness, and defense. Among the typical applications, I can mention, for example, the monitoring of physiological variables: body temperature, blood pressure, heart rate, detection of antibodies, chemical, or biological agents. Biotelemetry devices require a reliable communication system: robust, efficient, and versatile. Improving the transmission range of miniature in-body devices remains a major challenge: for the time being, they are able to operate only up to a few meters. Among the main issues to face are low radiation efficiencies (< 0.1%), antenna impedance detuning, and strong coupling to lossy and dispersive biological tissues. Thus, the main goal of the thesis is to conduct a multi-disciplinary study on development, optimization and characterization of antennas for in-body biotelemetry devices. After state-of-the-art and the context, I start with the development on both physical and numerical approaches to account for the effect of human tissues on the antenna. I propose the methodology to achieve given electromagnetic properties at a given frequency based on the full factorial experiment and surface response optimization. In addition, I describe the spherical physical phantom for the far-field characterization along with a combination of feed decoupling techniques. I proceed by reviewing the trough-body propagation mechanisms and deriving the optimal frequency for the in-body devices. I formulate the problem using four phantoms (homogeneous and heterogeneous) and perform full-wave analysis using an in-house hp-FEM code Agros 2D. Next, I study the existing antenna used by the BodyCap Company for its e-Celsius capsule and the ways on how to improve its operating range and robustness under strict integration and material constraints. The mechanisms of antenna–body coupling are analyzed and the found solution improves the antenna IEEE gain by 11 dBi (the operating range is at least tripled). The existing matching circuit and balun are optimized too for the given application reducing its size from eleven to seven discrete elements. In the following chapters, I continue studying the decoupling of antennas from a body using specific microstrip designs and dielectric loading via capsule shell. By applying the developed approaches, a high robustness and radiation efficiency can be achieved. At first, I develop a proof-of-concept antenna that demonstrates that the perfect matching (detuning immunity) is achievable for the operation within all human tissues. Based on these results, I develop a miniature and versatile biotelemetry platform: a 17 mm x 7 mm alumina capsule containing a conformal 434 MHz antenna. The antenna is well matched to 50 Ohm within the majority of human tissues and operates with an arbitrary device circuitry. Like this, one can use it ''as is,'' applying it for a wide range of in-body applications. Then, I develop a low profile conformal dual-band antenna operating in 434 MHz and 2.45 GHz bands. Such antenna can integrate both data transmission and wireless powering functionality increasing the available space inside an in-body device and increasing its scope of applications. Finally, I present the perspective developments including in-body sensing methodology. The obtained results contributes to further development of a new generation of miniature in-body devices that involve complex and dense integration of sensors, logic, and power sources

Doutoramento em Engenharia Informática
A exigente inovação na área das aplicações biomédicas tem guiado a evolução das tecnologias de informação nas últimas décadas. Os desafios associados a uma gestão, integração, análise e interpretação eficientes dos dados provenientes das mais modernas tecnologias de hardware e software requerem um esforço concertado. Desde hardware para sequenciação de genes a registos electrónicos de paciente, passando por pesquisa de fármacos, a possibilidade de explorar com precisão os dados destes ambientes é vital para a compreensão da saúde humana. Esta tese engloba a discussão e o desenvolvimento de melhores estratégias informáticas para ultrapassar estes desafios, principalmente no contexto da composição de serviços, incluindo técnicas flexíveis de integração de dados, como warehousing ou federação, e técnicas avançadas de interoperabilidade, como serviços web ou LinkedData. A composição de serviços é apresentada como um ideal genérico, direcionado para a integração de dados e para a interoperabilidade de software. Relativamente a esta última, esta investigação debruçou-se sobre o campo da farmacovigilância, no contexto do projeto Europeu EU-ADR. As contribuições para este projeto, um novo standard de interoperabilidade e um motor de execução de workflows, sustentam a sucesso da EU-ADR Web Platform, uma plataforma para realizar estudos avançados de farmacovigilância. No contexto do projeto Europeu GEN2PHEN, esta investigação visou ultrapassar os desafios associados à integração de dados distribuídos e heterogéneos no campo do varíoma humano. Foi criada uma nova solução, WAVe - Web Analyses of the Variome, que fornece uma coleção rica de dados de variação genética através de uma interface Web inovadora e de uma API avançada. O desenvolvimento destas estratégias evidenciou duas oportunidades claras na área de software biomédico: melhorar o processo de implementação de software através do recurso a técnicas de desenvolvimento rápidas e aperfeiçoar a qualidade e disponibilidade dos dados através da adopção do paradigma de web semântica. A plataforma COEUS atravessa as fronteiras de integração e interoperabilidade, fornecendo metodologias para a aquisição e tradução flexíveis de dados, bem como uma camada de serviços interoperáveis para explorar semanticamente os dados agregados. Combinando as técnicas de desenvolvimento rápidas com a riqueza da perspectiva "Semantic Web in a box", a plataforma COEUS é uma aproximação pioneira, permitindo o desenvolvimento da próxima geração de aplicações biomédicas.
The demand for innovation in the biomedical software domain has been an information technologies evolution driver over the last decades. The challenges associated with the effective management, integration, analyses and interpretation of the wealth of life sciences information stemming from modern hardware and software technologies require concerted efforts. From gene sequencing hardware to pharmacology research up to patient electronic health records, the ability to accurately explore data from these environments is vital to further improve our understanding of human health. This thesis encloses the discussion on building better informatics strategies to address these challenges, primarily in the context of service composition, including warehousing and federation strategies for resource integration, as well as web services or LinkedData for software interoperability. Service composition is introduced as a general principle, geared towards data integration and software interoperability. Concerning the latter, this research covers the service composition requirements within the pharmacovigilance field, namely on the European EU-ADR project. The contributions to this area, the definition of a new interoperability standard and the creation of a new workflow-wrapping engine, are behind the successful construction of the EUADR Web Platform, a workspace for delivering advanced pharmacovigilance studies. In the context of the European GEN2PHEN project, this research tackles the challenges associated with the integration of heterogeneous and distributed data in the human variome field. For this matter, a new lightweight solution was created: WAVe, Web Analysis of the Variome, provides a rich collection of genetic variation data through an innovative portal and an advanced API. The development of the strategies underlying these products highlighted clear opportunities in the biomedical software field: enhancing the software implementation process with rapid application development approaches and improving the quality and availability of data with the adoption of the Semantic Web paradigm. COEUS crosses the boundaries of integration and interoperability as it provides a framework for the flexible acquisition and translation of data into a semantic knowledge base, as well as a comprehensive set of interoperability services, from REST to LinkedData, to fully exploit gathered data semantically. By combining the lightness of rapid application development strategies with the richness of its "Semantic Web in a box" approach, COEUS is a pioneering framework to enhance the development of the next generation of biomedical applications.

Doutoramento em Ciência e Engenharia de Materiais
The increased longevity of humans and the demand for a better quality of life have led to a continuous search for new implant materials. Scientific development coupled with a growing multidisciplinarity between materials science and life sciences has given rise to new approaches such as regenerative medicine and tissue engineering. The search for a material with mechanical properties close to those of human bone produced a new family of hybrid materials that take advantage of the synergy between inorganic silica (SiO4) domains, based on sol-gel bioactive glass compositions, and organic polydimethylsiloxane, PDMS ((CH3)2.SiO2)n, domains. Several studies have shown that hybrid materials based on the system PDMS-SiO2 constitute a promising group of biomaterials with several potential applications from bone tissue regeneration to brain tissue recovery, passing by bioactive coatings and drug delivery systems. The objective of the present work was to prepare hybrid materials for biomedical applications based on the PDMS-SiO2 system and to achieve a better understanding of the relationship among the sol-gel processing conditions, the chemical structures, the microstructure and the macroscopic properties. For that, different characterization techniques were used: Fourier transform infrared spectrometry, liquid and solid state nuclear magnetic resonance techniques, X-ray diffraction, small-angle X-ray scattering, smallangle neutron scattering, surface area analysis by Brunauer–Emmett–Teller method, scanning electron microscopy and transmission electron microscopy. Surface roughness and wettability were analyzed by 3D optical profilometry and by contact angle measurements respectively. Bioactivity was evaluated in vitro by immersion of the materials in Kokubos’s simulated body fluid and posterior surface analysis by different techniques as well as supernatant liquid analysis by inductively coupled plasma spectroscopy. Biocompatibility was assessed using MG63 osteoblastic cells. PDMS-SiO2-CaO materials were first prepared using nitrate as a calcium source. To avoid the presence of nitrate residues in the final product due to its potential toxicity, a heat-treatment step (above 400 °C) is required. In order to enhance the thermal stability of the materials subjected to high temperatures titanium was added to the hybrid system, and a material containing calcium, with no traces of nitrate and the preservation of a significant amount of methyl groups was successfully obtained. The difficulty in eliminating all nitrates from bulk PDMS-SiO2-CaO samples obtained by sol-gel synthesis and subsequent heat-treatment created a new goal which was the search for alternative sources of calcium. New calcium sources were evaluated in order to substitute the nitrate and calcium acetate was chosen due to its good solubility in water. Preparation solgel protocols were tested and homogeneous monolithic samples were obtained. Besides their ability to improve the bioactivity, titanium and zirconium influence the structural and microstructural features of the SiO2-TiO2 and SiO2-ZrO2 binary systems, and also of the PDMS-TiO2 and PDMS-ZrO2 systems. Detailed studies with different sol-gel conditions allowed the understanding of the roles of titanium and zirconium as additives in the PDMS-SiO2 system. It was concluded that titanium and zirconium influence the kinetics of the sol-gel process due to their different alkoxide reactivity leading to hybrid xerogels with dissimilar characteristics and morphologies. Titanium isopropoxide, less reactive than zirconium propoxide, was chosen as source of titanium, used as an additive to the system PDMS-SiO2-CaO. Two different sol-gel preparation routes were followed, using the same base composition and calcium acetate as calcium source. Different microstructures with high hydrophobicit were obtained and both proved to be biocompatible after tested with MG63 osteoblastic cells. Finally, the role of strontium (typically known in bioglasses to promote bone formation and reduce bone resorption) was studied in the PDMS-SiO2-CaOTiO2 hybrid system. A biocompatible material, tested with MG63 osteoblastic cells, was obtained with the ability to release strontium within the values reported as suitable for bone tissue regeneration.
O aumento da longevidade dos seres humanos e a procura de uma melhor qualidade de vida têm conduzido a uma pesquisa contínua de novos materiais para implantes. O desenvolvimento científico, juntamente com uma crescente multidisciplinaridade entre as ciências dos materiais e as ciências da vida deram origem a novas abordagens, como a medicina regenerativa e a engenharia de tecidos. A busca de um material com propriedades mecânicas próximas das do osso humano produziu uma nova família de materiais híbridos que tiram partido da sinergia entre os domínios inorgânicos de sílica (SiO4), com base em composições de vidros bioativos obtidos por sol-gel, e os domínios orgânicos de polidimetilsiloxano, PDMS ((CH3)2.SiO2)n. Vários estudos têm demonstrado que os materiais híbridos baseados no sistema PDMS-SiO2 constituem um grupo de biomateriais promissores com várias aplicações potenciais tais como a regeneração de tecido ósseo e a recuperação do tecido cerebral, passando por revestimentos bioativos e sistemas de libertação controlada de fármacos. O objetivo do presente trabalho foi preparar materiais híbridos para aplicações biomédicas com base no sistema PDMS-SiO2 e contribuir para uma melhor compreensão das relações entre as condições de processamento sol-gel, as estruturas químicas, a microestrutura e as propriedades macroscópicas. Para alcançar tal objetivo, foram usadas diferentes técnicas de caracterização: espectroscopia de infravermelho por transformada de Fourier, ressonância magnética nuclear no estado sólido e no estado líquido, difração de raios-X, dispersão de raios-X de baixo ângulo, dispersão de neutrões de baixo ângulo, análise da área de superfície pelo método de Brunauer–Emmett–Teller, microscopia eletrónica de varrimento e microscopia eletrónica de transmissão. A rugosidade e a molhabilidade das superfícies foram analisadas por perfilometria óptica 3D e por medidas de ângulo de contacto, respectivamente. A bioatividade in vitro foi avaliada através de testes de imersão em plasma sintético e posterior observação da superfície dos materiais e análise do líquido sobrenadante por espectrometria de emissão atômica por plasma acoplado Indutivamente. A biocompatibilidade in vitro foi avaliada usando células osteoblásticas MG63. Materiais do sistema PDMS-SiO2-CaO foram inicialmente preparados usando o nitrato como fonte de cálcio. Para eliminar os resíduos de nitrato no produto final, devido à sua potencial toxicidade, é necessária uma etapa de tratamento térmico (acima dos 400° C). A fim de aumentar a estabilidade térmica dos materiais submetidos a altas temperaturas, foi adicionado titânio ao sistema híbrido. Obteve-se assim um material híbrido contendo cálcio, sem vestígios de nitrato, mantendo-se uma quantidade significativa de grupos metilo. A dificuldade de obter amostras monolíticas de híbridos PDMS-SiO2-CaO por síntese sol-gel e posterior tratamento térmico para eliminação de nitratos, criou um novo objetivo: a procura de fontes alternativas de cálcio. Novas fontes de cálcio foram avaliadas para substituir o nitrato tendo-se escolhido o acetato de cálcio devido à sua boa solubilidade em água. Estabeleceram-se protocolos de preparação por sol-gel a partir dos quais se obtiveram amostras monolíticas homogéneas. Além de melhorar a bioatividade, o titânio e o zircónio influenciam as características estruturais e microestruturais dos sistemas binários SiO2-TiO2 e SiO2-ZrO2, bem como dos sistemas PDMS-TiO2 e PDMS-ZrO2. Neste contexto, foram estudadas diferentes condições experimentais no processo sol-gel, de modo a compreender o papel destes aditivos no sistema SiO2-PDMS. Concluiu-se que o titânio e o zircónio influenciam a cinética do processo sol-gel devido à diferente reatividade dos despectivos alcóxidos, conduzindo à obtenção de xerogéis híbridos com diferentes características e morfologias. O isopropóxido de titânio, menos reativo do que o propóxido de zircónio, foi escolhido como fonte de titânio, usado como aditivo no sistema PDMS-SiO2CaO. Dois procedimentos diferentes de preparação por sol-gel foram seguidos, utilizando a mesma composição de base e o acetato de cálcio como fonte de cálcio. Foram obtidas diferentes microestruturas muito hidrofóbicas e ambas mostraram ser biocompatíveis após serem testadas com células osteoblásticas MG63. Finalmente, foi avaliado o papel do estrôncio (conhecido nos biovidros por favorecer a formação de tecido ósseo e reduzir a sua reabsorção) no sistema híbrido PDMS-CaO-SiO2-TiO2. O material produzido revelou-se biocompatível, através de testes com células osteoblásticas MG63, e com a capacidade de libertar estrôncio dentro dos limites considerados adequados para a reparação do tecido ósseo.

Recently, the interest in implantable devices for biomedical telemetry has significantly increased. Amongst the different components of the implantable device, the antenna plays the most significant role in the wireless data transmission. However, the human body around the antenna alters its overall characteristics and absorbs most of its radiation. Therefore, this thesis is mainly focused on improving the antenna characteristics (bandwidth and radiation efficiency) to overcome the human body effect and investigating new structures that reduce the power absorption by the human body tissues. A novel antenna design methodology is developed and used to design new flexible implantable antennas of much lighter weight, larger radiation efficiency, and wider bandwidth than existing embedded antennas. These antennas work for multiple ((401-406 MHz) MedRadio, 433 MHz and 2.45 GHz ISM) bands which satisfy the requirements of low power consumption and wireless power transfer. This has been combined with thorough investigations of the antenna performance in the anatomical human body. New effective evaluation parameters such as the antenna orientation are investigated for the first time. New structures inspired by complementary and multiple split ring resonators (CSRRs and MSRRs) are designed. The structures are found to reduce the electric near field and hence the absorbed power which increases the radiated power accordingly. This new promising function of metamaterial based structures for implantable applications is investigated for the first time. The path loss (between pacemaker and glucose monitoring implantable antennas inside the anatomical body model) and (between an implantable and external antennas for a wireless power channel at 433 MHz) are estimated. Moreover, the optimum antenna type for on-in body communication is investigated. Loop antennas are found to outperform patch antennas in close proximity to the human body.

The main aim of this project was to assess auxetic (negative Poisson's ratio) materials for potential in biomedical devices. Specifically, a detailed comparative indentation study has been undertaken on auxetic and conventional foams for hip protector devices; radially-gradient one-piece foams having auxetic character have been produced for the first time and shown to have potential in artificial intervertebral disc (IVD) implant devices; and auxetic honeycomb geometries have been assessed for the stem component in hip implant devices. For the hip protector application, combined compression and heat treatment of conventional polyurethane open-cell foam was used to produce monolithic auxetic foams. The foams were characterised structurally using optical microscopy, and mechanically using mechanical testing combined with videoextensometry. Static indentation using six different indenter shapes on each of the six faces of the foam specimens has been undertaken. The key conclusion here is that the enhanced indentation resistance for the converted foam is not a consequence of increased density accompanied by the usual significant increase in foam stiffness. The enhanced indentation resistance is consistent with the auxetic effect associated with the increased density, providing a localised densification mechanism under indentation (i.e. material flows under the indenter). At higher indentation displacement the Poisson’s ratios for both the unconverted and converted foams tend towards zero. In this case, the increase in foam stiffness for the converted foams at higher strain may also contribute to the indentation enhancement at high indentation displacement. New radially-gradient foams mimicking the core-sheath structure of the natural IVD have been produced through the development of a novel thermo-mechanical manufacturing route. Foam microstructural characterisation has been undertaken using optical and scanning electron microscopy, and also micro-CT scans performed by collaborators at the University of Manchester. Detailed x-y strain mapping using combined mechanical testing and videoextensometry enabled the local and global Young's modulus and Poisson's ratio responses of these new materials to be determined. In one example, global auxetic response is achieved in a foam having a positive Poisson's ratio core and auxetic sheath. It is suggested this may be a more realistic representation of the properties of natural IVD tissue. Analytical and Finite Element (FE) models have been developed to design honeycomb geometries for the stems in new total hip replacement implants. FE models of the devices implanted within bone have been developed and the auxetic stems shown to lead to reduced stress shielding effect.

Cette thèse décrit le développement de nouvelles méthodes pour obtenir des nanoparticules fonctionnelles polyvalentes qui peuvent potentiellement être utilisées pour des applications biomédicales telles que la vectorisation de médicaments, des essais biologiques et la bio-imagerie. Les nanomatériaux sont des outils polyvalents qui ont trouvé des applications comme vecteurs de médicaments, la bio-imagerie ou les biocapteurs. En particulier, les nanoparticules de type core-shell ont attiré beaucoup d'attention en raison de leur petite taille, une relation surface/volume élevée, et une biocompatibilité. Dans ce contexte, nous proposons dans la première partie de la thèse (Chapitre 2), une nouvelle méthode pour obtenir des nanoparticules core-shell via la polymérisation radicalaire en émulsion et vivante combinées. Des particules cœurs de polystyrène de 30 à 40 nm, avec une distribution de taille étroite et portant à la surface des groupements iniferter ont été utilisés pour amorcer la polymérisation supplémentaire d'une couche de polymère. Des nanoparticules core-shell ont été préparées de cette façon. Différents types d’enveloppes : anionique, zwitterioniques, à empreintes moléculaires, thermosensibles, ont ainsi été greffées. Notre méthode est une plate-forme polyvalente permettant d'ajouter des fonctionnalités multiples soit dans le noyau et/ou l'enveloppe pour les études d'interaction cellulaire et de toxicité, ainsi que des matériaux récepteurs pour l'imagerie cellulaire. Dans la deuxième partie de la thèse (Chapitre 3), nous décrivons un procédé nouveau et polyvalent pour la modification de surface des nanoparticules de conversion ascendante (UCP). Ce sont des nanocristaux fluorescents dopés de lanthanides qui ont récemment attiré beaucoup d'attention. Leur fluorescence est excitée dans le proche infrarouge, ce qui les rend idéales comme marqueurs dans des applications biomédicales telles que les tests biologiques et la bio-imagerie, l'auto-fluorescence étant réduite par rapport à des colorants organiques et les quantum dots. Cependant, les UCP sont hydrophobes et non-compatible avec les milieux aqueux, donc une modification de leur surface est essentielle. La stratégie que nous proposons utilise l'émission UV ou visible après excitation en proche infrarouge des UCP, comme source de lumière secondaire pour la photopolymérisation localisée de couches minces hydrophiles autour les UCP. Notre méthode offre de grands avantages comme la facilité d'application et la fonctionnalisation de surface rapide pour fixer divers ligands, et fournit une plateforme pour préparer des UCP encapsulée de polymères pour des différentes applications. Des hydrogels stimuli-sensibles sont des matériaux qui changent leurs propriétés physicochimiques en réponse à des stimuli externes tels que la température, le pH ou la lumière. Ces matériaux intelligents jouent un rôle critique dans des applications biomédicales telles que la vectorisation de médicaments ou l'ingénierie tissulaire. La troisième partie de cette thèse (Chapitre 4) propose un nouveau procédé de préparation d'hydrogels photo et pH sensible. Deux composantes, l'un photosensible à base dl'acide 4-[(4-méthacryloyloxy) phénylazo] benzoïque et l'autre cationic contenant des unités 2-(diéthylamino)éthyl méthacrylate, ont été synthétisés. Leur association donne des particules monodispersées de 100 nm photo et pH sensibles. Ces nanoparticules peuvent être potentiellement utilisées pour la vectorisation de médicaments, en particulier de biomolécules telles que protéines ou siARN. En conclusion, nous avons conçu plusieurs nouvelles méthodes efficaces, polyvalentes, génériques et facilement applicables pour obtenir des nanoparticules et nanocomposites de polymères fonctionnels qui peuvent être appliqués dans de différents domaines biomédicaux comme la vectorisation de médicaments, les biocapteurs, les tests biologiques et la bio-imagerie
This thesis describes the development of novel methods to obtain versatile, functional nanoparticles that can potentially be used for biomedical applications such as drug delivery, bioassays and bioimaging. Nanomaterials are versatile tools that have found applications as drug carriers, bioimaging or biosensing. In particular, core-shell type nanoparticles have attracted much attention due to their small size, high surface to volume ratio and biocompatibility. In this regard, we propose in the first part of the thesis (Chapter 2), a novel method to obtain core-shell nanoparticles via combined radical emulsion and living polymerizations. Polystyrene core seeds of 30-40 nm, with a narrow size distribution and surface-bound iniferter moieties were used to further initiate polymerization of a polymer shell. Core-shell nanoparticles were prepared in this way. Different types of shells : anionic, zwitterionic, thermoresponsive or molecularly imprinted shells, were thus grafted. Our method is a versatile platform with the ability to add multi-functionalities in either the core for optical sensing or/and the shell for cell interaction and toxicity studies, as well as receptor materials for cell imaging. In the second part of the thesis (Chapter 3), we describe a novel and versatile method for surface modification of upconverting nanoparticles (UCPs). UCPs are lanthanide-doped fluorescent nanocrystals that have recently attracted much attention. Their fluorescence is excitated in the near infrared, which makes them ideal as labels in biomedical applications such as bioimaging and bioassays, since the autofluorescence background is minimized compared to organic dyes and quantum dots. However, UCPs are hydrophobic and non-compatible with aqueous media, therefore prior surface modification is essential. The strategy that we propose makes use oft he UV or Vis emission light of near-infrared photoexcited upconverting nanoparticles, as secondary light source for the localized photopolymerization of thin hydrophilic shells around the UCPs. Our method offers great advantages like ease of application and rapid surface functionalization for attaching various ligands and therefore can provide a platform to prepare polymeric-encapsulated UCPs for applications in bioassays, optical imaging and drug delivery. Stimuli responsive hydrogels are materials that can change their physico-chemical properties in response to external stimuli such as temperature, pH or light. These smart materials play critical roles in biomedical applications such as drug delivery or tissue engineering. The third part of the thesis (Chapter 4) proposes a novel method for obtaining photo and pH-responsive supramolecularly crosslinked hydrogels. Two building blocks, one containing photoresponsive 4-[(4-methacryloyloxy)phenylazo] benzoic acid and the other, consisting of cationic 2-(diethylamino)ethyl methacrylate units, were first synthesized. Combining the two building blocks yielded photo and pH responsive monodisperse 100-nm particles. These nanoparticles can be eventually utilized for drug delivery, especially delivery of biomolecules such as siRNAs or proteins. In conclusion, we have designed several new efficient, versatile, generic and easily applicable methods to obtain functionalized polymer nanoparticles and nanocomposites that can be applied in various biomedical domains like drug delivery, biosensing, bioassays and bioimaging

The application of mass spectrometry to verification of the structure of 3-methyluridine (m³U) isolated by HPLC from normal human urine is described. m³U has been used as an internal standard for studies of urinary nucleosides, a practice that is discouraged with the confirmation of m³U as a naturally occurring compound. Mass spectrometry has been used for the identification of 5'-deoxyxanthosine (5'-dX) a novel nucleoside in normal human urine. Initial concern over availability of a reference sample of 5'-dX prompted investigations of the structure/fragmentation relationships of the TMS deratives of 2'-, 3'-, and 5'-deoxynucleosides toward differentiation between the three deoxynucleosides. Results are presented which allow discrimination between the model compounds, deoxyanalogs of adenosine. Subsequent to the deoxynucleoside fragmentation studies, a biosynthetically produced reference sample of 5'-dX became available for direct comparison of mass spectra and chromatographic retention times which, when combined with observations from the deoxynucleoside studies established the structure of 5'-dX. In response to the large number of mass spectra produced from the GC-MS analysis of a TMS derivatized urine sample, computer software has been written to aid in spectral analysis. Examples are shown in which the software uses established fragmentation rules to assign structure to ions in the mass spectrum and suggest modifications in the sugar portion of two urinary nucleosides. The structure/fragmentation relationships of the unique antitumor drug taxol has been studied by EI, CI and FAB mass spectrometry. Information is presented showing characteristic fragmentation of the side-chain and verification of functional groups attached to the taxane ring. Studies have been conducted to determine the relationship between target temperature and matrix and sample lifetime in the source of the mass spectrometer. Results are presented showing that cooling the target permits the use of matrix materials that are too volatile at ambient temperatures thus extending the range of compounds that can be studied by mass spectrometry. A recently constructed four-sector mass spectrometer is described with a detailed discussion of instrumental capabilities. Results of experiments designed to apply these capabilities to the structural analysis of TMS nucleosides using FAB ionization are discussed with an emphasis on the fragmentation unique to 4-sector daughter ion experiments compared with conventional studies and 2-sector daughter ion results.

Acousto-electric (AE) effect comes from an interaction between electrical current and acoustic pressure generated when acoustic waves travel through a conducting material. It currently has two main application areas, ultrasound current source density imaging (UCSDI) and AE hydrophone. UCSDI can detect the current direction by modulating the dipole field with ultrasound pulse, and it is now used to form 3D imaging of dipole changing in one period of treatment, such as arrhythmia in the heart and epilepsy in the brain. As ultrasound pulse passes through electrical field, it convolutes or correlates with the inner product of the electric fields formed by the dipole and detector. The polarity of UCSDI is not determined by Doppler effect that exists in pulse echo (PE) signal, but the gradient of lead field potentials created by dipole and recording electrode, making the base-banded AE voltage positive at the anode and negative at cathode. As convolution shifts spectrum lower, the base band frequency for polarity is different from the center frequency of AE signal. The simulation uses the principles of UCSDI, and helps to understand the phenomena in the experiment. 3-D Fast Fourier Transform accelerates the computing velocity to resolve the correlation in the simulation of AE signal. Most single element hydrophones depend on a piezoelectric material that converts pressure changes to electricity. These devices, however, can be expensive, susceptible to damage at high pressure, and/or have limited bandwidth and sensitivity. An AE hydrophone requires only a conductive material and can be constructed out of common laboratory supplies to generate images of an ultrasound beam pattern consistent with more expensive hydrophones. Its sensitivity is controlled by the injected bias current, hydrophone shape, thickness and width of sensitivity zone. The design of this device needs to be the tradeoff of these parameters. Simulations were made to optimize the design with experimental validation using specifically fabricated devices composed of a resistive element of indium tin oxide (ITO).

Les propriétés des nanozéolithes, à savoir leur grande surface, leur stabilité hydrothermale et leur nature non toxique, permettent leur utilisation dans des applications prospectives, notamment la biomédecine (capteurs, administration de médicaments et de gaz) et la microbiologie (agents antibactériens). De nombreuses recherches ont été consacrées à l’étude de nouvelles applications biomédicales utilisant des matériaux zéolithiques, toutefois leur plein potentiel n’a toujours pas été pleinement dévoilé.Il est bien connu que la résistance croissante aux traitements établis de tumeurs et d’infections bactériennes par radiothérapie et antibiotiques est un problème de première importance. Par conséquent, le développement de nouvelles stratégies thérapeutiques pour résoudre ces problèmes est très démandé.L'objectif de cette recherche de doctorat est de synthétiser et de modifier post-synthétiquement des zéolithes nanométriques pour des applications biomédicales. Cela implique l'échange d'ions de zéolithe avec divers cations pour trouver celui qui convient le mieux aux applications souhaitées : le traitement antimicrobien, la réoxygénation des tissus tumoraux et l’administration de gaz.Dans cette étude, nous rapportons: (i) l'effet de la zéolithe FAU de type nanométrique modifiée au cuivre sur les bactéries de type ESKAPE (chapitre 3), (ii) l’utilisation de nanozéolithes contenant du métal comme outil d'oxygénation et de visualisation tissulaire (chapitre 4) et enfin (iii) l'utilisation de nanozéolithes FAU comme vecteur de l'oxyde nitrique et du dioxyde de carbone pour prévenir des maladies potentiellement létales (chapitre 5)
The properties of nanozeolites, namely, large surface area, hydrothermal stability and non-toxic nature, enable their utilization in forward-looking applications, including biomedicine (sensors, drug and gas delivery) and microbiology (antibacterial agents). Hence, a lot of research has been devoted to study the new biomedical applications using zeolitic materials, their full potential has still not been fully unveiled.It is well recognised that growing resistance to already established treatments of tumors and bacterial infections using radiotherapy and antibiotics is a distressing matter. Therefore, the development of new therapeutic strategies towards above issues is of great demand.The goal of this PhD research is to synthesise and post-synthetically modify nanosized zeolites for biomedical applications. This involves the ion-exchange of zeolite with various cations to find the most suitable one for desired applications in regards to antimicrobial treatment, tumour tissue reoxygenation and gas delivery.In this study, we report: (i) the effect of copper modified nanosized FAU type zeolite on ESKAPE type bacteria (Chapter 3), (ii) the metal containing nanozeolite as a tool for tissue oxygenation and visualisation using MRI (Chapter 4), and lastly (iii) the use of FAU nanozeolite as nitric oxide and carbon dioxide gas vector to prevent life threatening conditions (Chapter 5)

Objects with complex shape and functions have always attracted attention and interest. The morphological diversity and complexity of naturally occurring forms and patterns have been a motivation for humans to copy and adopt ideas from Nature to achieve functional, aesthetic and social value. Biomimetics is addressed to the design and development of new synthetic materials using strategies adopted by living organisms to produce biological materials. In particular, biomineralized tissues are often sophisticate composite materials, in which the components and the interfaces between them have been defined and optimized, and that present unusual and optimal chemical-physical, morphological and mechanical properties. Moreover, biominerals are generally produced by easily traceable raw materials, in aqueous media and at room pressure and temperature, that is through cheap process and materials. Thus, it is not surprising that the idea to mimic those strategies proper of Nature has been employed in several areas of applied sciences, such as for the preparation of liquid crystals, ceramic thin films computer switches and many other advanced materials. On this basis, this PhD thesis is focused on the investigation of the interaction of biologically active ions and molecules with calcium phosphates with the aim to develop new materials for the substitution and repair of skeletal tissue, according to the following lines: I. Modified calcium phosphates. A relevant part of this PhD thesis has been addressed to study the interaction of Strontium with calcium phosphates. It was demonstrated that strontium ion can substitute for calcium into hydroxyapatite, causing appreciable structural and morphological modifications. The detailed structural analysis carried out on the nanocrystals at different strontium content provided new insight into its interaction with the structure of hydroxyapatite. At variance with the behaviour of Sr towards HA, it was found that this ion inhibits the synthesis of octacalcium phosphate. However, it can substitute for calcium in this structure up to 15 atom %, in agreement with the increase of the cell parameters observed on increasing ion concentration. A similar behaviour was found for Magnesium ion, whereas Manganese inhibits the synthesis of octacalcium phosphate and it promotes the precipitation of dicalcium phosphate dehydrate. It was also found that Strontium affects the kinetics of the reaction of hydrolysis of α-TCP. It inhibits the conversion from α-TCP to hydroxyapatite. However, the resulting apatitic phase contains significant amounts of Sr2+ suggesting that the addition of Sr2+ to the composition of α-TCP bone cements could be successfully exploited for its local delivery in bone defects. The hydrolysis of α-TCP has been investigated also in the presence of increasing amounts of gelatin: the results indicated that this biopolymer accelerates the hydrolysis reaction and promotes the conversion of α-TCP into OCP, suggesting that its addition in the composition of calcium phosphate cements can be employed to modulate the OCP/HA ratio, and as a consequence the solubility, of the set cement. II. Deposition of modified calcium phosphates on metallic substrates. Coating with a thin film of calcium phosphates is frequently applied on the surface of metallic implants in order to combine the high mechanical strength of the metal with the excellent bioactivity of the calcium phosphates surface layers. During this PhD thesis, thank to the collaboration with prof. I.N. Mihailescu, head of the Laser-Surface-Plasma Interactions Laboratory (National Institute for Lasers, Plasma and Radiation Physics – Laser Department, Bucharest) Pulsed Laser Deposition has been successfully applied to deposit thin films of Sr substituted HA on Titanium substrates. The synthesized coatings displayed a uniform Sr distribution, a granular surface and a good degree of crystallinity which slightly decreased on increasing Sr content. The results of in vitro tests carried out on osteoblast-like and osteoclast cells suggested that the presence of Sr in HA thin films can enhance the positive effect of HA coatings on osteointegration and bone regeneration, and prevent undesirable bone resorption. The possibility to introduce an active molecule in the implant site was explored using Matrix Assisted Pulsed Laser Evaporation to deposit hydroxyapatite nanocrystals at different content of alendronate, a bisphosphonate widely employed in the treatments of pathological diseases associated to bone loss. The coatings displayed a good degree of crystallinity, and the results of in vitro tests indicated that alendronate promotes proliferation and differentiation of osteoblasts even when incorporated into hydroxyapatite. III. Synthesis of drug carriers with a delayed release modulated by a calcium phosphate coating. A core-shell system for modulated drug delivery and release has been developed through optimization of the experimental conditions to cover gelatin microspheres with a uniform layer of calcium phosphate. The kinetics of the release from uncoated and coated microspheres was investigated using aspirin as a model drug. It was shown that the presence of the calcium phosphate shell delays the release of aspirin and allows to modulate its action.

Objects with complex shape and functions have always attracted attention and interest. The morphological diversity and complexity of naturally occurring forms and patterns have been a motivation for humans to copy and adopt ideas from Nature to achieve functional, aesthetic and social value. Biomimetics is addressed to the design and development of new synthetic materials using strategies adopted by living organisms to produce biological materials. In particular, biomineralized tissues are often sophisticate composite materials, in which the components and the interfaces between them have been defined and optimized, and that present unusual and optimal chemical-physical, morphological and mechanical properties. Moreover, biominerals are generally produced by easily traceable raw materials, in aqueous media and at room pressure and temperature, that is through cheap process and materials. Thus, it is not surprising that the idea to mimic those strategies proper of Nature has been employed in several areas of applied sciences, such as for the preparation of liquid crystals, ceramic thin films computer switches and many other advanced materials. On this basis, this PhD thesis is focused on the investigation of the interaction of biologically active ions and molecules with calcium phosphates with the aim to develop new materials for the substitution and repair of skeletal tissue, according to the following lines: I. Modified calcium phosphates. A relevant part of this PhD thesis has been addressed to study the interaction of Strontium with calcium phosphates. It was demonstrated that strontium ion can substitute for calcium into hydroxyapatite, causing appreciable structural and morphological modifications. The detailed structural analysis carried out on the nanocrystals at different strontium content provided new insight into its interaction with the structure of hydroxyapatite. At variance with the behaviour of Sr towards HA, it was found that this ion inhibits the synthesis of octacalcium phosphate. However, it can substitute for calcium in this structure up to 15 atom %, in agreement with the increase of the cell parameters observed on increasing ion concentration. A similar behaviour was found for Magnesium ion, whereas Manganese inhibits the synthesis of octacalcium phosphate and it promotes the precipitation of dicalcium phosphate dehydrate. It was also found that Strontium affects the kinetics of the reaction of hydrolysis of α-TCP. It inhibits the conversion from α-TCP to hydroxyapatite. However, the resulting apatitic phase contains significant amounts of Sr2+ suggesting that the addition of Sr2+ to the composition of α-TCP bone cements could be successfully exploited for its local delivery in bone defects. The hydrolysis of α-TCP has been investigated also in the presence of increasing amounts of gelatin: the results indicated that this biopolymer accelerates the hydrolysis reaction and promotes the conversion of α-TCP into OCP, suggesting that its addition in the composition of calcium phosphate cements can be employed to modulate the OCP/HA ratio, and as a consequence the solubility, of the set cement. II. Deposition of modified calcium phosphates on metallic substrates. Coating with a thin film of calcium phosphates is frequently applied on the surface of metallic implants in order to combine the high mechanical strength of the metal with the excellent bioactivity of the calcium phosphates surface layers. During this PhD thesis, thank to the collaboration with prof. I.N. Mihailescu, head of the Laser-Surface-Plasma Interactions Laboratory (National Institute for Lasers, Plasma and Radiation Physics – Laser Department, Bucharest) Pulsed Laser Deposition has been successfully applied to deposit thin films of Sr substituted HA on Titanium substrates. The synthesized coatings displayed a uniform Sr distribution, a granular surface and a good degree of crystallinity which slightly decreased on increasing Sr content. The results of in vitro tests carried out on osteoblast-like and osteoclast cells suggested that the presence of Sr in HA thin films can enhance the positive effect of HA coatings on osteointegration and bone regeneration, and prevent undesirable bone resorption. The possibility to introduce an active molecule in the implant site was explored using Matrix Assisted Pulsed Laser Evaporation to deposit hydroxyapatite nanocrystals at different content of alendronate, a bisphosphonate widely employed in the treatments of pathological diseases associated to bone loss. The coatings displayed a good degree of crystallinity, and the results of in vitro tests indicated that alendronate promotes proliferation and differentiation of osteoblasts even when incorporated into hydroxyapatite. III. Synthesis of drug carriers with a delayed release modulated by a calcium phosphate coating. A core-shell system for modulated drug delivery and release has been developed through optimization of the experimental conditions to cover gelatin microspheres with a uniform layer of calcium phosphate. The kinetics of the release from uncoated and coated microspheres was investigated using aspirin as a model drug. It was shown that the presence of the calcium phosphate shell delays the release of aspirin and allows to modulate its action.

In this thesis, pH and potassium all-solid-state ISE based on potentiometry and bioimpedance sensors were designed, fabricated and integrated in a miniaturized array for its application in endoscopic surgery for in vivo ischemia detection inside the stomach. To achieve this goal, the developed array withstood the low pH and corrosive condition in the gastric juice of the stomach, by modifying the surface with a conductive Ag/AgCl ink containing hydrophilic and hydrophobic groups. That creates a stable and robust candidate for low pH applications. However, these sensors have to demonstrate besides stability, high sensitivity, and selectivity. For this purpose, different ionophores specific to a single ion were tested. Octadecyl isonicotinate was the one that shown better results as pH ionophore and valinomycin, bis [(benzo-15-crown-4)-4-ylmethyl] pimelate for potassium detection. All these ionophores were embedded in PVC polymer membrane containing also plasticizers such as 2-nitrophenyl octyl ether, bis (1-butylpentyl) adipate (BBPA) and liphophilic anionic additives such as potassium tetrakis (4-chlorophenyl) borate (KTpClPB). The specific compositions of membranes to detect potassium or pH were optimized for the better performance of the sensors. pH ISE sensor shows a nernstian behavior (-54,38 mV/pH) at low pH and a nearly nernstian behavior at physiological pH (-34,899 mV/pH). Bioimpedance sensor was tested and optimized in vitro with different solutions of ions concentration to mimic ischemia detection and with different kinds of tissues from different nature. For this purpose, chicken fat and breast tissues were taken as a model for mimicking non-ischemic and ischemic states respectively. The effect of electrodes insulation as well as the pressure applied on the tissue was studied. The dependence of the impedance response with different pressure applied to the sensor was overcome by applying magnetic field attachment. The sensor array was modified with ring magnets which were attracted by an external magnet, giving stable and reliable signal discarding mechanical motion. The shape and size of the sensor array were designed for being adapted to the commercially available gastroendoscopes. Round shaped cylinder of 7 mm diameter was fabricated with 12 electrodes pin of 600 µm diameter, containing 3 RE, 3 pH and 2 potassium all-solid-state sensors and 4 electrodes in a row for impedance measurements. The sensor array was successfully integrated in commercial endoscope and inserted inside the pig stomach. The blood flow of certain area of the stomach was interrupted by ligating or crossclamping vessels and organ wall. Ischemia and reperfusion steps were sensed successfully with potassium and pH sensors. These results also indicate that information about hypoxic tissue damage can be collected with this array. Ischemia was also sensed on small intestine tissue by opening the abdominal part of the body and getting the sensor array in contact with the intestine. By crossclamping of mesenteric artery by tourniquets and scissors, ischemic and reperfusion states were controlled. Results proved that ischemia and reperfusion can be monitored by our integrated sensor array. As a conclusion, a novel all-solid-state potentiometric, miniaturized, low cost and mass producible pH, potassium all-solid-state ISE and impedance sensors integrated in an array was successfully fabricated for detecting ischemia inside the stomach by means of endoscopic techniques and also on small intestine. This array was tested in vitro and vivo giving reproducible and reliable results. The developed all-solid-state pH sensors permit low pH sensing from 0.7-2.5, which is the only example in the literature that allows so low pH detection, and so make this sensor a unique device for stomach detection.
El diagnóstico médico es uno de los campos que han obtenido más ventajas de la capacidad de los electrodos selectivos de iones (ESI) para la detección de iones, ya que los cambios en la concentración de estos elementos están directamente relacionados con diferentes enfermedades. La detección de isquemia es una de las favorecidas por estos sensores. La isquemia es una disminución del suministro de sangre a un órgano y se requiere una detección rápida y precisa. Los métodos de detección in situ en el tejido de los órganos conllevan una detección temprana de la isquemia y el estómago es uno de los órganos más importantes en la detección de Ischemia. Sin embargo, el bajo pH del jugo gástrico del estómago hace difícil la fabricación de sensores de estado sólido con ESI estables y funcionales, principalmente debido a la interferencia de aniones y al problema de la adhesión entre la membrana ESI y la superficie del electrodo. En esta tesis, se han diseñado y fabricado electrodos selección de iones de pH y potasio ESI de estado sólido basados en la potenciometría y sensores de bioimpedancia y se han integrado en una matriz en miniatura para su aplicación en la cirugía endoscópica para la detección de isquemia in vivo en el interior del estómago. El conjunto de sensores se integró con éxito en endoscopio comercial y se inserto en el interior del estómago de un cerdo. El flujo de sangre de cierta área del estómago se interrumpió mediante la ligación o pinzamiento de los vasos sanguineos y la pared del órgano. Los pasos de isquemia y reperfusión fueron detectados con éxito con los sensores de potasio y de pH. Estos resultados también indican que se puede obtener información sobre el daño en el tejido hipóxico recogido con esta matriz. Los sensores de pH de sólido desarrollados permiten la detección pH bajos de 0,7 a 2,5, que es el único ejemplo en la literatura de detección de pH tan bajos con este tipo de sensores y por lo tanto hacen que sea este sensor de un dispositivo único para la detección de isquemia en el estómago.

Els cossos d’inclusió bacterians (CIs) són dipòsits proteics que apareixen normalment durant el procés de producció de proteïna recombinant. Històricament, aquests agregats han estat classificats com a sub-productes de rebuig i, per tant, descartats. No obstant, aquesta percepció ha canviat al llarg de les dues darreres dècades. Nombrosos estudis han proporcionat evidències que demostren que aquestes partícules són, de fet, estructures supramoleculars formades per contactes estereoespecífics proteïna-proteïna, morfològicament i fisicoquímicament estables, fàcils de purificar i que mostren un alt nivell de plasticitat pel que fa a activitat biològica, mida i propietats fisicoquímiques. En el present treball s’ha explorat la possibilitat d’aprofitar les propietats úniques dels CIs per desenvolupar noves aplicacions biomèdiques. En aquest sentit, ens hem centrat en l’estudi de la idoneïtat dels CIs com a nanomaterial particulat per tal de generar nanotopografies capaces de controlar la resposta de cèl·lules animals. La modificació a nivell nanomètric de superfícies per tal d’estimular una resposta cel·lular específica resulta molt prometedora pels camps de l’enginyeria tissular i la medicina regenerativa en general. En aquest treball es presenten evidències de que els CIs es poden dipositar sobre superfícies de cultiu, obtenint nanotopografies modificades amb efecte biològic. Al cultivar cèl·lules de mamífer sobre aquestes, s’aprecia un elevada capacitat d’adhesió cel·lular. A més a més, s’ha observat que el CIs són capaços d’estimular la proliferació cel·lular mitjançant una cascada de senyalització basada en la mecanotransducció. Tant l’adhesió com la proliferació cel·lulars són factors crucials per la colonització de la superfície d’implants. S’ha demostrat també que les topografies formades per CIs són capaces de dirigir cèl·lules mare mesenquimals (MSCs) indiferenciades cap a osteoblasts. És important esmentar que l’adhesió, la proliferació i la diferenciació cel·lular mediades per CIs poden ser modulades per la correcta elecció del fons genètic de la soca utilitzada per a produir els CIs, obtenint d’aquesta manera agregats fisicoquímica i morfològicament diferents. Finalment els CIs han demostrat ser capaços d’alliberar quantitats significatives de la proteïna que els forma, en una versió activa. Aquest fet ha propiciat el seu estudi com plataforma d’entrega de fàrmacs proteics, des de nanoestructures de la superfície del substrat, cap als cultius diana. Aquesta darrera aplicació seria particularment interessant en tractaments a llarg termini, ja que el CIs proporcionarien un alliberament de la proteïna sostingut. A més, aquesta capacitat d’entrega afegiria un nivell extra de complexitat i control a les aplicacions prèviament esmentades ja que combinaria l’efecte mecànic de les topografies basades en CIs amb l’activitat biològica de la proteïna formadora de l’agregat.
Bacterial Inclusion Bodies (IBs) are protein deposits usually observed in recombinant bacteria during protein production processes. These aggregates have been historically regarded as waste by-products and therefore discarded. However, in the last two decades this perception has changed. Numerous studies have provided evidence that these particles are in fact supramolecular structures formed by stereospecific cross-molecular protein-protein interactions, morphologically and mechanically stable, easy to purify and highly tunable in terms of biological activity, size and physicochemical properties,. In the present work it has been explored to exploit appealing IB properties for biomedical applications. In this regard, we have focused on the suitability of IBs as particulate protein nanomaterial to produce engineered nanotopographies able to modulate mammalian cell responses. Nanomodification of scaffolds to stimulate a specific cell response is a promising field for tissue engineering and regenerative medicine. In this regard, we have provided evidence that bacterial IBs can be effectively deposited onto cell culture surfaces generating altered nanotopographies. These IB-based topographies, when used as cell culture substrate, exhibited higher capability for cell adhesion in four distinct cell lines. Moreover, IBs were shown to be able of actively stimulating cell proliferation through mechanotransductive events, being these two activities crucial for cell colonization of implant materials. On the other hand, it has been also proved how IB-based topographies are able to direct mesenchymal stem cells (MSCs) to osteogenesis. Noteworthy, cell adhesion, cell proliferation and cell differentiation can be controlled by the proper choice of Escherichia coli producing strain genetic background, rendering physicochemically and morphologically distinct IBs. Finally, IBs have been shown capable of releasing significant amounts of their forming protein, prompting their use as a delivery platform of therapeutic protein drugs, from nanostructured surfaces to cell cultures. This last application would be particularly appealing for long term treatments, since IBs provide a sustained release of their active component. Moreover, this delivery capacity adds an extra level of complexity and control to the previous mentioned IB applications, through the combination of the mechanical effect of the IB-based topographies with the biological activity of the IB-forming protein