3D-printed scaffold composites for the stimuli-induced local delivery of bioactive adjuncts

Polysaccharide scaffolds have been successfully employed to reconstruct environments that sustain skin tissue regeneration after injuries. Three-dimensional (3D) advanced additive manufacturing technologies allow creating scaffolds with controlled and reproducible macro- and micro-structure that improve the quality of the restored tissue to favor spontaneous repair. However, when persistent inflammation occurs, the physiological tissue healing capacity is reduced, like in the presence of pathologies like diabetes, vascular diseases, chronic infection, and others. In these circumstances, the bioavailability of therapeutic adjuncts like the growth factors in addition to the standard treatments represents undoubtedly a promising strategy to accelerate the healing of skin lesions. Precisely designed polysaccharide scaffolds obtained by 3D printing represent a robust platform that can be further implemented with the controlled delivery of bioactive adjuncts. Human elastin-like polypeptides (HELPs) are stimuli-responsive biopolymers. Their structure allows the integration of domains endowed with biological functionality, making them attractive compounds to prepare composites with smart properties. In the present study, 3D-printed alginate and chitosan scaffolds were combined with the HELP components. The HELP biopolymer was fused to the epidermal growth factor (EGF) as the bioactive domain. Different constructs were prepared and the stimuli-responsive behavior as well as the biological activity were evaluated, suggesting that these smart bioactive composites are suitable to realize multifunctional dressings that sustain the local release of therapeutic adjuncts

Alginate/human elastin-like polypeptide composite films with antioxidant properties for potential wound healing application

In this contribution we describe the preparation and characterization of a series of cross-linked films based on the combination of an elastin-derived biomimetic polypeptide (Human elastin-like polypeptide (HELP)) with alginate (ALG) to obtain a composite with enhanced properties. ALG/HELP composite films loaded with the hydrophobic natural antioxidant curcumin were prepared by solvent casting method followed by the cross-linking with calcium chloride. The compatibility between the two components as well as the final properties was evaluated. The micro-morphological study of films showed a homogeneous structure, but the film tensile strength decrease with HELP content and elongation at break was adversely affected by biopolymer addition. Spectroscopic and thermal analyses confirmed an interaction between ALG and HELP which also causes a modification in swelling kinetics and faster degradation. Moreover, the study of curcumin release showed a controlled delivery up to 10\u202fdays with a faster release rate in the presence of HELP. Human Dermal Fibroblasts (hDF) were used to test the in vitro cytocompatibility. The antioxidant activity correlated to the increase of HELP content suggested the applicability of these composites to develop smart biomaterials. Overall, these features indicated how this composite material has considerable potential as customizable platforms for various biomedical applications

Three-Dimensional (3D) Printed Silver Nanoparticles/Alginate/Nanocrystalline Cellulose Hydrogels: Study of the Antimicrobial and Cytotoxicity Efficacy

Here, a formulation of silver nanoparticles (AgNPs) and two natural polymers such as alginate (ALG) and nanocrystalline cellulose (CNC) was developed for the 3D printing of scaffolds with large surface area, improved mechanical resistance and sustained capabilities to promote antimicrobial and cytotoxic effects. Mechanical resistance, water content, morphological characterization and silver distribution of the scaffolds were provided. As for applications, a comparable antimicrobial potency against S. aureus and P. aeruginosa was demonstrated by in vitro tests as function of the AgNP concentration in the scaffold (Minimal Inhibitory Concentration value: 10 mg/mL). By reusing the 3D system the antimicrobial efficacy was demonstrated over at least three applications. The cytotoxicity effects caused by administration of AgNPs to hepatocellular carcinoma (HepG2) cell culture through ALG and ALG/CNC scaffold were discussed as a function of time and dose. Finally, the liquid chromatography-mass spectrometry (LC-MS) technique was used for targeted analysis of pro-apoptotic initiation and executioner caspases, anti-apoptotic and proliferative proteins and the hepatocyte growth factor, and provided insights about molecular mechanisms involved in cell death induction

3D Printed Chitosan/Alginate Hydrogels for the Controlled Release of Silver Sulfadiazine in Wound Healing Applications: Design, Characterization and Antimicrobial Activity

The growing demand for personalized medicine requires innovation in drug manufacturing to combine versatility with automation. Here, three-dimensional (3D) printing was explored for the production of chitosan (CH)/alginate (ALG)-based hydrogels intended as active dressings for wound healing. ALG hydrogels were loaded with 0.75% w/v silver sulfadiazine (SSD), selected as a drug model commonly used for the therapeutic treatment of infected burn wounds, and four different 3D CH/ALG architectures were designed to modulate the release of this active compound. CH/ALG constructs were characterized by their water content, elasticity and porosity. ALG hydrogels (Young’s modulus 0.582 ± 0.019 Mpa) were statistically different in terms of elasticity compared to CH (Young’s modulus 0.365 ± 0.015 Mpa) but very similar in terms of swelling properties (water content in ALG: 93.18 ± 0.88% and in CH: 92.76 ± 1.17%). In vitro SSD release tests were performed by using vertical diffusion Franz cells, and statistically significant different behaviors in terms of the amount and kinetics of drugs released were observed as a function of the construct. Moreover, strong antimicrobial potency (100% of growth inhibition) against Staphylococcus aureus and Pseudomonas aeruginosa was demonstrated depending on the type of construct, offering a proof of concept that 3D printing techniques could be efficiently applied to the production of hydrogels for controlled drug delivery

Development of three-dimensional printed scaffold prototypes as dressings and implants intended for the treatment of skin tissue regenerative-related pathologies

Tissue engineering is a more and more maturing field of medicine, needing increasingly automated manufacturing processes for the creation of tissue repair solutions. Herein this thesis, design and development of biocompatible materials as well as 3D printing (3DP) technology were focused and deepened in particular for the realization of hydrogel-based advanced medication prototypes, mostly dedicated to the skin regeneration but also suitable for dentistry purposes rather than for further applications. In brief, overall pre and post-3D printing process parameters were investigated, in terms of correct material processing through the developed instrumentation, as well as chemico-physical features characterizing the hydrogels. Biocompatibility, cell proliferation enhancement were also probed by inoculating human fibroblasts and/or keratinocytes and measuring their viability, distribution and morphology. In vivo experimental on diabetic ulcerated rats was assessed to prove their validity as wound dressing prototypes in comparison to a commercial patch, selected as model, and spontaneous healing. Further, alternative antimicrobial compounds to antibiotics were included in hydrogel constructs, studied in this case like controlled drug delivery systems for the treatment of infected wounds. Development and functionalization of polymer hydrogels can boost tissue regeneration, in this work, film materials for dentistry were doped with fibronectin and its engineered binding aptamer for bone reconstruction or epithelial repair. A Further functionalization, by collagen coating, was instead adjusted for 3D printed chitosan scaffolds; several characterizations and assays were performed in order to explore their potentiality for the purpose targeted. Moreover, an analytical approach based on mass spectrometry, defined as hydrogen/deuterium exchange (HDX-MS), was tuned to better evaluate proper material functionalization with fibronectin. Sterilization of developed prototype materials is not trivial for potentially marketable hydrogels, gamma irradiation was assessed and preliminary studied as promising suitable procedure. The 3DP overall process was retained innovative in terms of obtainable hydrogel forms, furthermore, their general technical features seemed to be prone to favor cell growth through the 3D polymer structures. In vivo trials even more efficiently demonstrated their effectiveness as medications for ameliorating tissue regeneration, proved in particular by histological staining and microscopical analysis conducted. In vitro antimicrobial activity of silver sulfadiazine-loaded scaffolds was fully achieved on P. aeruginosa and S. aureus, very common wound bed infecting pathogens. The functionalization of these hydrogel materials for dentistry aims resulted particularly effective in terms of in vitro osteoblast and epithelial cells growth enhancement as well as for those cultured on 3D printed chitosan-collagen scaffolds. Moreover, the HDX-MS analysis confirmed fibronectin could selectively recognize the aptamer, indicating an efficient docking of the complex on the biomaterial. Mass spectrometry was used also for the determination of collagen composition of decellularized rat thyroid matrixes, revealing which matrix preparation protocol was more appropriate for scaffold re-colonization. Gamma rays sterilization procedure seems in a preliminary way to be feasible, but further technical investigations are required. A step forward in the research on biocompatible materials intended for several purposes particularly focusing on wound dressings as well as their processing through additive manufacturing has been made, giving a small but hopefully significant contribute for the prototyping of hydrogels-based three-dimensional scaffolds. As additional application, resourcefulness of 3DP was exploited also for the production of biocompatible wastewaters filtering system models for detoxification from amoxicillin as pollutant pharmaceutical. TiO2 was included in form of powder in the material ad hoc developed for the purpose as drug photocatalytic agent. Characterizations and utilization tests were conducted in order to assay the feasibility of the idea. The formulation and 3D development of models of wastewaters filters surprised for its effectiveness and re-usability, further tests on other drugs are running to finely ameliorate the promising system

¿Fue sin querer queriendo? : El Chavo del 8: un reflejo de la sociedad latinoamericana

El Chavo del Ocho es una famosa serie mexicana de la década del ´70, cuya repercusión a nivel mundial, permitió la llegada a nuestro país y su perpetuación a lo largo de los años. La tesis se concentra en el análisis socio-comunicacional de la serie, a través de los ejes familia, educación y vecindad, con el fin de problematizar la construcción de la sociedad latinoamericana de la década del ´70. Se retomarán algunas perspectivas teórico- metodológicas con las cuales han construido sus objetos de estudio autores de estudios culturales y de América Latina, como así también los conceptos que han aportado al campo de la comunicación. El trabajo está dividido en cinco capítulos: Nacimiento de la serie, comparación y descripción; contexto sociopolítico y económico de México y América Latina en la década del `70; marco teórico; marco metodológico; unidades de análisis: Familia, educación y vecindad.Facultad de Periodismo y Comunicación Socia

Molecular switches of fluorescence operating through metal centred redox couples

Three-component molecular systems (redox active subunit)-spacer-(light-emitting fragment) can operate as fluorescence switches, following the alternate addition of an oxidizing agent and a reducing agent (or the adjustment of the potential of the working electrode in an electrolysis experiment). The redox active subunit typically consists of a metal centred redox couple (M(n+1)+/Mn+), encircled by a macrocyclic receptor, and switching efficiency requires that one of the two oxidation states quenches the proximate fluorophore and the other does not. Four ON/OFF systems, based on either the CuII/CuI or NiIII/NiII couple, will be discussed. The nature of the quenching process responsible for the OFF state, either electron transfer or energy transfer, is related to the length and to the flexibility-rigidity of the spacer

3D Printed Chitosan/Alginate Hydrogels for the Controlled Release of Silver Sulfadiazine in Wound Healing Applications: Design, Characterization and Antimicrobial Activity

The growing demand for personalized medicine requires innovation in drug manufacturing to combine versatility with automation. Here, three-dimensional (3D) printing was explored for the production of chitosan (CH)/alginate (ALG)-based hydrogels intended as active dressings for wound healing. ALG hydrogels were loaded with 0.75% w/v silver sulfadiazine (SSD), selected as a drug model commonly used for the therapeutic treatment of infected burn wounds, and four different 3D CH/ALG architectures were designed to modulate the release of this active compound. CH/ALG constructs were characterized by their water content, elasticity and porosity. ALG hydrogels (Young’s modulus 0.582 ± 0.019 Mpa) were statistically different in terms of elasticity compared to CH (Young’s modulus 0.365 ± 0.015 Mpa) but very similar in terms of swelling properties (water content in ALG: 93.18 ± 0.88% and in CH: 92.76 ± 1.17%). In vitro SSD release tests were performed by using vertical diffusion Franz cells, and statistically significant different behaviors in terms of the amount and kinetics of drugs released were observed as a function of the construct. Moreover, strong antimicrobial potency (100% of growth inhibition) against Staphylococcus aureus and Pseudomonas aeruginosa was demonstrated depending on the type of construct, offering a proof of concept that 3D printing techniques could be efficiently applied to the production of hydrogels for controlled drug delivery

Surface modification of chitosan films with a fibronectin fragment–DNA aptamer complex to enhance osteoblastic cell activity: A mass spectrometry approach probing evidence on protein behavior

RationaleProtein dynamics are fundamental for biological activity. Direct surface functionalization with these biomolecules often creates regions presenting non-homogeneous and altered protein functionality. Aptamers have been shown to be an effective method to enhance the biocompatibility of biomaterials by acting as docking points for biomolecules capable of improving cell responses and the colonization/proliferation processes. Here mass spectrometry (MS) was successfully applied as an analytical approach to study the interactions between a fibronectin fragment (FN) and anti-fibronectin aptamers in solution and on a chitosan film.MethodsA 30kDa N-terminal fibronectin fragment, a ssDNA anti-fibronectin 40 nucleotide long aptamer and chitosan 95/50 (degree of deacetylation 92.6-97.5%; molecular weight 80-220kDa) were used. First, the dynamics of the FN in aqueous solution, in the presence or absence of anti-FN-specific DNA aptamers, were described. Thus, the same study was carried out on 2% (w/w) chitosan-based films, functionalized with FN alone or through aptamers as selective spacers.ResultsAmide hydrogen/deuterium exchange mass spectrometry (HDX-MS) analysis identified the SYRIGDTWSKKDNRGNL and YRVGDTYERPKDSMI, YVVGETWEKPYQGWMM and WERTYLGNAL fragments as presumably involved in the interaction of FN with aptamers. MS findings indicated the improved functionality of FN on chitosan when aptamers were used as selective spacer, and this may explain why cells preferentially attached to aptamer-bound FN rather than on chitosan-FN films.ConclusionsAptamers did not affect the amount of adsorbed FN, but influenced its conformation enhancing its biological activity toward adhesion and growth of osteoblasts. HDX-MS data allowed the identification of the FN/aptamer interaction regions. Differences in FN flexibility on the chitosan film in the presence or absence of the aptamer were established providing insights at the molecular level to better understand the protein biological functionality on cell proliferation