Cryptographic techniques for managing computational effort

Availability has become a primary goal of information security and is as significant as other goals, in particular, confidentiality and integrity. Maintaining availability of essential services on the public Internet is an increasingly difficult task in the presence of sophisticated attackers. Attackers may abuse limited computational resources of a service provider and thus managing computational costs is a key strategy for achieving the goal of availability. In this thesis we focus on cryptographic approaches for managing computational costs, in particular computational effort. We focus on two cryptographic techniques: computational puzzles in cryptographic protocols and secure outsourcing of cryptographic computations. This thesis contributes to the area of cryptographic protocols in the following ways. First we propose the most efficient puzzle scheme based on modular exponentiations which, unlike previous schemes of the same type, involves only a few modular multiplications for solution verification; our scheme is provably secure. We then introduce a new efficient gradual authentication protocol by integrating a puzzle into a specific signature scheme. Our software implementation results for the new authentication protocol show that our approach is more efficient and effective than the traditional RSA signature-based one and improves the DoSresilience of Secure Socket Layer (SSL) protocol, the most widely used security protocol on the Internet. Our next contributions are related to capturing a specific property that enables secure outsourcing of cryptographic tasks in partial-decryption. We formally define the property of (non-trivial) public verifiability for general encryption schemes, key encapsulation mechanisms (KEMs), and hybrid encryption schemes, encompassing public-key, identity-based, and tag-based encryption avors. We show that some generic transformations and concrete constructions enjoy this property and then present a new public-key encryption (PKE) scheme having this property and proof of security under the standard assumptions. Finally, we combine puzzles with PKE schemes for enabling delayed decryption in applications such as e-auctions and e-voting. For this we first introduce the notion of effort-release PKE (ER-PKE), encompassing the well-known timedrelease encryption and encapsulated key escrow techniques. We then present a security model for ER-PKE and a generic construction of ER-PKE complying with our security notion.

Electrospraying of polymers with therapeutic molecules : state of the art

The encapsulation and release of bioactive molecules from polymeric vehicles represents the holy grail of drug and growth factor delivery therapies, whereby sustained and controlled release is crucial in eliciting a positive therapeutic effect. To this end, electrospraying is rapidly emerging as a popular technology for the production of polymeric particles containing bioactive molecules. Compared with traditional emulsion fabrication techniques, electrospraying has the potential to reduce denaturation of protein drugs and affords tighter regulation over particle size distribution and morphology. In this article, we review the importance of the electrospraying parameters that enable reproducible tailoring of the particles' physical and in vitro drug release characteristics, along with discussion of existing in vivo data. Controlled morphology and monodispersity of particles can be achieved with electrospraying, with high encapsulation efficiencies and without unfavorable denaturation of bioactive molecules throughout the process. Finally, the combination of electrospraying with electrospun scaffolds, with an emphasis on tissue regeneration is reviewed, depicting a technique in its relative infancy but holding great promise for the future of regenerative medicine.

The effect of polystyrene sodium sulfonate grafting on polyethylene terephthalate artificial ligaments on in vitro mineralisation and in vivo bone tissue integration

This study investigates the impact of polystyrene sodium sulfonate (PolyNaSS) grafting onto the osseo-integration of a polyethylene terephthalate artificial ligament (Ligament Advanced Reinforcement System, LARS™) used for Anterior Cruciate Ligament (ACL). The performance of grafted and non-grafted ligaments was assessed in vitro by culturing human osteoblasts under osteogenic induction and this demonstrated that the surface modification was capable of up-regulating the secretion of ALP and induced higher level of mineralisation as measured 6 weeks post-seeding by Micro-Computed Tomography. Grafted and non-grafted LARS™ were subsequently implanted in an ovine model for ACL reconstruction and the ligament-to-bone interface was evaluated by histology and biomechanical testings 3 and 12 months post-implantation. The grafted ligaments exhibited more frequent direct ligament-to-bone contact and bone formation in the core of the ligament at the later time point than the non-grafted specimens, the grafting also significantly reduced the fibrous encapsulation of the ligament 12 months post-implantation. However, this improved osseo-integration was not translated into a significant increase in the biomechanical pull-out loads. These results provide evidences that PolyNaSS grafting improved the osseo-integration of the artificial ligament within the bone tunnels. This might positively influence the outcome of the surgical reconstructions, as higher ligament stability is believed to limit micro-movement and therefore permits earlier and enhanced healing.

Zeta-potential and morphology of electrospun nano- and microfibers from biopolymers and their blends used as scaffolds in tissue engineering

Electrostatic spinning or electrospinning is a fiber spinning technique driven by a high-voltage electric field that produces fibers with diameters in a submicrometer to nanometer range.1 Nanofibers are typical one-dimensional colloidal objects with an increased tensile strength, whose length can achieve a few kilometers and the specific surface area can be 100 m2 g–1 or higher.2 Nano- and microfibers from biocompatible polymers and biopolymers have received much attention in medical applications3 including biomedical structural elements (scaffolding used in tissue engineering,2,4–6 wound dressing,7 artificial organs and vascular grafts8), drug and vaccine delivery,9–11 protective shields in speciality fabrics, multifunctional membranes, etc. Other applications concern superhydrophobic coatings,12 encapsulation of solid materials,13 filter media for submicron particles in separation industry, composite reinforcement and structures for nano-electronic machines.

Osteogenic differentiation of bone marrow MSCs by β-tricalcium phosphate stimulating macrophages via BMP2 signalling pathway

Immune reactions play important roles in determining the in vivo fate of bone substitute materials, either in new bone formation or inflammatory fibrous tissue encapsulation. The paradigm for the development of bone substitute materials has been shifted from inert to immunomodulatory materials, emphasizing the importance of immune cells in the material evaluation. Macrophages, the major effector cells in the immune reaction to implants, are indispensable for osteogenesis and their heterogeneity and plasticity render macrophages a primer target for immune system modulation. However, there are very few reports about the effects of macrophages on biomaterial-regulated osteogenesis. In this study, we used b-tricalcium phosphate (b-TCP) as a model biomaterial to investigate the role of macrophages on the material stimulated osteogenesis. The macrophage phenotype switched to M2 extreme in response to b-TCP extracts, which was related to the activation of calcium-sensing receptor (CaSR) pathway. Bone morphogenetic protein 2 (BMP2) was also significantly upregulated by the b-TCP stimulation, indicating that macrophage may participate in the b-TCP stimulated osteogenesis. Interestingly, when macrophageconditioned b-TCP extracts were applied to bone marrow mesenchymal stem cells (BMSCs), the osteogenic differentiation of BMSCs was significantly enhanced, indicating the important role of macrophages in biomaterial-induced osteogenesis. These findings provided valuable insights into the mechanism of material-stimulated osteogenesis, and a strategy to optimize the evaluation system for the in vitro osteogenesis capacity of bone substitute materials.

Publicly verifiable ciphertexts

In many applications, where encrypted traffic flows from an open (public) domain to a protected (private) domain, there exists a gateway that bridges the two domains and faithfully forwards the incoming traffic to the receiver. We observe that indistinguishability against (adaptive) chosen-ciphertext attacks (IND-CCA), which is a mandatory goal in face of active attacks in a public domain, can be essentially relaxed to indistinguishability against chosen-plaintext attacks (IND-CPA) for ciphertexts once they pass the gateway that acts as an IND-CCA/CPA filter by first checking the validity of an incoming IND-CCA ciphertext, then transforming it (if valid) into an IND-CPA ciphertext, and forwarding the latter to the recipient in the private domain. “Non-trivial filtering'' can result in reduced decryption costs on the receivers' side. We identify a class of encryption schemes with publicly verifiable ciphertexts that admit generic constructions of (non-trivial) IND-CCA/CPA filters. These schemes are characterized by existence of public algorithms that can distinguish between valid and invalid ciphertexts. To this end, we formally define (non-trivial) public verifiability of ciphertexts for general encryption schemes, key encapsulation mechanisms, and hybrid encryption schemes, encompassing public-key, identity-based, and tag-based encryption flavours. We further analyze the security impact of public verifiability and discuss generic transformations and concrete constructions that enjoy this property.

Design and flight testing of an integrated solar powered UAV and WSN for remote gas sensing

This paper describes a generic and integrated solar powered remote Unmanned Air Vehicles (UAV) and Wireless Sensor Network (WSN) gas sensing system. The system uses a generic gas sensing system for CH4 and CO2 concentrations using metal oxide (MoX) and non-dispersive infrared sensors, and a new solar cell encapsulation method to power the UASs as well as a data management platform to store, analyse and share the information with operators and external users. The system was successfully field tested at ground and low altitudes, collecting, storing and transmitting data in real time to a central node for analysis and 3D mapping. The system can be used in a wide range of outdoor applications, especially in agriculture, bushfires, mining studies, opening the way to a ubiquitous low cost environmental monitoring. A video of the bench and flight test performed can be seen in the following link https://www.youtube.com/watch?v=Bwas7stYIxQ.

A bioengineered 3D ovarian cancer model for the assessment of peptidase-mediated enhancement of spheroid growth and intraperitoneal spread

Cancer-associated proteases promote peritoneal dissemination and chemoresistance in malignant progression. In this study, kallikrein-related peptidases 4, 5, 6, and 7 (KLK4-7)-cotransfected OV-MZ-6 ovarian cancer cells were embedded in a bioengineered three-dimensional (3D) microenvironment that contains RGD motifs for integrin engagement to analyze their spheroid growth and survival after chemotreatment. KLK4-7-cotransfected cells formed larger spheroids and proliferated more than controls in 3D, particularly within RGD-functionalized matrices, which was reduced upon integrin inhibition. In contrast, KLK4-7-expressing cell monolayers proliferated less than controls, emphasizing the relevance of the 3D microenvironment and integrin engagement. In a spheroid-based animal model, KLK4-7-overexpression induced tumor growth after 4 weeks and intraperitoneal spread after 8 weeks. Upon paclitaxel administration, KLK4-7-expressing tumors declined in size by 91% (controls: 87%) and showed 90% less metastatic outgrowth (controls: 33%, P<0.001). KLK4-7-expressing spheroids showed 53% survival upon paclitaxel treatment (controls: 51%), accompanied by enhanced chemoresistance-related factors, and their survival was further reduced by combination treatment of paclitaxel with KLK4/5/7 (22%, P=0.007) or MAPK (6%, P=0.006) inhibition. The concomitant presence of KLK4-7 in ovarian cancer cells together with integrin activation drives spheroid formation and proliferation. Combinatorial approaches of paclitaxel and KLK/MAPK inhibition may be more efficient for late-stage disease than chemotherapeutics alone as these inhibitory regimens reduced cancer spheroid growth to a greater extent than paclitaxel alone.

Gelatine methacrylamide-based hydrogels : an alternative three-dimensional cancer cell culture system

Modern cancer research requires physiological, three-dimensional (3-D) cell culture platforms, wherein the physical and chemical characteristics of the extracellular matrix (ECM) can be modified. In this study, gelatine methacrylamide (GelMA)-based hydrogels were characterized and established as in vitro and in vivo spheroid-based models for ovarian cancer, reflecting the advanced disease stage of patients, with accumulation of multicellular spheroids in the tumour fluid (ascites). Polymer concentration (2.5-7% w/v) strongly influenced hydrogel stiffness (0.5±0.2kPa to 9.0±1.8kPa) but had little effect on solute diffusion. The diffusion coefficient of 70kDa fluorescein isothiocyanate (FITC)-labelled dextran in 7% GelMA-based hydrogels was only 2.3 times slower compared to water. Hydrogels of medium concentration (5% w/v GelMA) and stiffness (3.4kPa) allowed spheroid formation and high proliferation and metabolic rates. The inhibition of matrix metalloproteinases and consequently ECM degradability reduced spheroid formation and proliferation rates. The incorporation of the ECM components laminin-411 and hyaluronic acid further stimulated spheroid growth within GelMA-based hydrogels. The feasibility of pre-cultured GelMA-based hydrogels as spheroid carriers within an ovarian cancer animal model was proven and led to tumour development and metastasis. These tumours were sensitive to treatment with the anti-cancer drug paclitaxel, but not the integrin antagonist ATN-161. While paclitaxel and its combination with ATN-161 resulted in a treatment response of 33-37.8%, ATN-161 alone had no effect on tumour growth and peritoneal spread. The semi-synthetic biomaterial GelMA combines relevant natural cues with tunable properties, providing an alternative, bioengineered 3-D cancer cell culture in in vitro and in vivo model systems.

Formation of specialized propagules resistant to desiccation and cryopreservation in the threatened moss Ditrichum plumbicola (ditrichales, bryopsida)

Background and Aims Successful cryopreservation of bryophytes is linked to intrinsic desiccation tolerance and survival can be enhanced by pre-treatment with abscisic acid (ABA) and sucrose. The pioneer moss Ditrichum plumbicola is naturally subjected to desiccation in the field but showed unexpectedly low survival of cryopreservation, as well as a poor response to pre-treatment. The effects of the cryopreservation protocol on protonemata of D. plumbicola were investigated in order to explore possible relationships between the production in vitro of cryopreservation-tolerant asexual propagules and the reproductive biology of D. plumbicola in nature. Methods Protonemata were prepared for cryopreservation using a four-step protocol involving encapsulation in sodium alginate, pre-treatment for 2 weeks with ABA and sucrose, desiccation for 6 h and rapid freezing in liquid nitrogen. After each stage, protonemata were prepared for light and electron microscopy and growth on standard medium was monitored. Further samples were prepared for light and electron microscopy at intervals over a 24-h period following removal from liquid nitrogen and re-hydration. Key Results Pre-treatment with ABA and sucrose caused dramatic changes to the protonemata. Growth was arrested and propagules induced with pronounced morphological and cytological changes. Most cells died, but those that survived were characterized by thick, deeply pigmented walls, numerous small vacuoles and lipid droplets in their cytoplasm. Desiccation and cryopreservation elicited no dramatic cytological changes. Cells returned to their pre-dehydration and cryopreservation state within 2 h of re-hydration and/or removal from liquid nitrogen. Regeneration was normal once the ABA/sucrose stimulus was removed. Conclusions The ABA/sucrose pre-treatment induced the formation of highly desiccation- and cryopreservation-tolerant propagules from the protonemata of D. plumbicola. This parallels behaviour in the wild, where highly desiccation-tolerant rhizoids function as perennating organs allowing the moss to endure extreme environmental conditions. An involvement of endogenous ABA in the desiccation tolerance of D. plumbicola is suggested.

Controlling microencapsulation and release of micronized proteins using poly(ethylene glycol) and electrospraying

The fabrication of tailored microparticles for delivery of therapeutics is a challenge relying upon a complex interplay between processing parameters and materials properties. The emerging use of electrospraying allows better tailoring of particle morphologies and sizes than current techniques, critical to reproducible release profiles. While dry encapsulation of proteins is essential for the release of active therapeutics from microparticles, it is currently uncharacterized in electrospraying. To this end, poly(ethylene glycol) (PEG) was assessed as a micronizing and solubilizing agent for dry protein encapsulation and release from electrosprayed particles made from polycaprolactone (PCL). The physical effect of PEG in protein-loaded poly(lactic-co-glycolic acid) (PLGA) particles was also studied, for comparison. The addition of 5–15 wt% PEG 6 kDa or 35 kDa resulted in reduced PCL particle sizes and broadened distributions, which could be improved by tailoring the electrospraying processing parameters, namely by reducing polymer concentration and increasing flow rate. Upon micronization, protein particle size was reduced to the micrometer domain, resulting in homogenous encapsulation in electrosprayed PCL microparticles. Microparticle size distributions were shown to be the most determinant factor for protein release by diffusion and allowed specific control of release patterns.

The influence of extracellular matrix on the generation of vascularized, engineered, transplantable tissue

In a recently described model for tissue engineering, an arteriovenous loop comprising the femoral artery and vein with interposed vein graft is fabricated in the groin of an adult male rat, placed inside a polycarbonate chamber, and incubated subcutaneously. New vascularized granulation tissue will generate on this loop for up to 12 weeks. In the study described in this paper three different extracellular matrices were investigated for their ability to accelerate the amount of tissue generated compared with a no-matrix control. Poly-D,L-lactic-co-glycolic acid (PLGA) produced the maximal weight of new tissue and vascularization and this peaked at two weeks, but regressed by four weeks. Matrigel was next best. It peaked at four weeks but by eight weeks it also had regressed. Fibrin (20 and 80 mg/ml), by contrast, did not integrate with the generating vascularized tissue and produced less weight and volume of tissue than controls without matrix. The limiting factors to growth appear to be the chamber size and the capacity of the neotissue to integrate with the matrix. Once the sides of the chamber are reached or tissue fails to integrate, encapsulation and regression follow. The intrinsic position of the blood supply within the neotissue has many advantages for tissue and organ engineering, such as ability to seed the construct with stem cells and microsurgically transfer new tissue to another site within the individual. In conclusion, this study has found that PLGA and Matrigel are the best matrices for the rapid growth of new vascularized tissue suitable for replantation or transplantation.

Delivery of therapeutic molecules using electrosprayed polymeric particles for applications in tissue engineering

This thesis has developed an innovative technology, electrospraying, that allows biodegradable microparticles to deliver pharmaceuticals that aid bone regeneration. The establishment, characterisation and optimisation of the technique are a step forward in developing an affordable and safe alternative to the products used currently in the clinical setting for the treatment of musculoskeletal disorders. The researcher has also investigated electrospraying as a coating technique on biodegradable structures that are used to replace damaged tissues, in order to provide localised and efficient drug delivery in the site of the defect to help tissue reconstruction.

Preparation and in vitro release of zein microparticles loaded with prednisolone for oral delivery

Zein has been proposed as a polymer for targeted-drug delivery via the oral route. Zein microparticles were loaded with prednisolone and evaluated as an oral delivery system. Microparticles were formulated using phase separation. Starting quantities of zein and prednisolone, along with the agitation method and temperature were found to significantly impact drug loading and loading efficiency. Vortex mixing produced the highest drug loading and loading efficiency. Drug release was measured in simulated conditions of the stomach and small intestine using the microparticles made with the method that best improved drug loading. In simulated stomach and small intestine conditions, prednisolone release reached almost 70 over 3 and 4h, respectively. While a clinically relevant dose may be delivered using c. 100mg of zein microparticles, prednisolone release from the microparticles indicates that they may not be suited as a controlled-or targeted-delivery system.

Microparticle-mediated gene delivery for the enhanced expression of a 19-KDa fragment of merozoite surface protein 1 of Plasmodium falciparum

The 19 kDa carboxyl-terminal fragment of merozoite surface protein 1 (MSP119) is a major component of the invasion-inhibitory response in individual immunity to malaria. A novel ultrasonic atomization approach for the formulation of biodegradable poly(lactic-co-glycolic acid) (PLGA) microparticles of malaria DNA vaccines encoding MSP119 is presented here. After condensing the plasmid DNA (pDNA) molecules with a cationic polymer polyethylenimine (PEI), a 40 kHz ultrasonic atomization frequency was used to formulate PLGA microparticles at a flow rate of 18 mL h1. High levels of gene expression and moderate cytotoxicity in COS-7 cells were achieved with the condensed pDNA at a nitrogen to phosphate (N/P) ratio of 20, thus demonstrating enhanced cellular uptake and expression of the transgene. The ability of the microparticles to convey pDNA was examined by characterizing the formulated microparticles. The microparticles displayed Z-average hydrodynamic diameters of 1.50-2.10 lm and zeta potentials of 17.8-23.2 mV. The encapsulation efficiencies were between 78 and 83%, and 76 and 85% of the embedded malaria pDNA molecules were released under physiological conditions in vitro. These results indicate that PLGA-mediated microparticles can be employed as potential gene delivery systems to antigen-presenting cells in the prevention of malaria.