Tiago Ruivo Correia

Production and characterization of chitosan/gelatin/β-TCP scaffolds for improved bone tissue regeneration.

Recently, bone tissue engineering emerged as a viable therapeutic alternative, comprising bone implants and new personalized scaffolds to be used in bone replacement and regeneration. In this study, biocompatible scaffolds were produced by freeze-drying, using different formulations (chitosan, chitosan/gelatin, chitosan/β-TCP and chitosan/gelatin/β-TCP) to be used as temporary templates during bone tissue regeneration. Sample characterization was performed through attenuated total reflectance-Fourier transform infrared spectroscopy, X-ray diffraction and energy dispersive spectroscopy analysis. Mechanical characterization and porosity analysis were performed through uniaxial compression test and liquid displacement method, respectively. In vitro studies were also done to evaluate the biomineralization activity and the cytotoxic profile of the scaffolds. Scanning electron and confocal microscopy analysis were used to study cell adhesion and proliferation at the scaffold surface and within their structure. Moreover, the antibacterial activity of the scaffolds was also evaluated through the agar diffusion method. Overall, the results obtained revealed that the produced scaffolds are bioactive and biocompatible, allow cell internalization and show antimicrobial activity against Staphylococcus aureus. Such, make these 3D structures as potential candidates for being used on the bone tissue regeneration, since they promote cell adhesion and proliferation and also prevent biofilm development at their surfaces, which is usually the main cause of implant failure.

Natural melanin: a potential pH-responsive drug release device.

This work proposes melanin as a new nanocarrier for pH-responsive drug release. Melanin is an abundant natural polymer that can be easily extracted from cuttlefish as nanoparticles with a suitable size range for drug delivery. However, despite its high potentiality, the application of this biopolymer in the pharmaceutical and biomedical fields is yet to be explored. Herein, melanin nanoparticles were impregnated with metronidazole, chosen as model antibiotic drug, using supercritical carbon dioxide. The drug release profile was investigated at acidic and physiologic pH, and the dominant mechanism was found to follow a non-Fickian transport. Drug release from melanin shows a strong pH dependency, which allied to its biocompatibility and lack of cytotoxicity envisages its potential application as nanocarrier in formulations for colon and intestine targeted drug delivery.

Functionalization of polydimethylsiloxane membranes to be used in the production of voice prostheses

Abstract The voice is produced by the vibration of vocal cords which are located in the larynx. Therefore, one of the major consequences for patients subjected to laryngectomy is losing their voice. In these cases, a synthetic one-way valve set (voice prosthesis) can be implanted in order to allow restoration of speech. Most voice prostheses are produced with silicone-based materials such as polydimethylsiloxane (PDMS). This material has excellent properties, such as optical transparency, chemical and biological inertness, non-toxicity, permeability to gases and excellent mechanical resistance that are fundamental for its application in the biomedical field. However, PDMS is very hydrophobic and this property causes protein adsorption which is followed by microbial adhesion and biofilm formation. To overcome these problems, surface modification of materials has been proposed in this study. A commercial silicone elastomer, SylgardTM 184 was used to prepare membranes whose surface was modified by grafting 2-hydroxyethylmethacrylate and methacrylic acid by low-pressure plasma treatment. The hydrophilicity, hydrophobic recovery and surface energy of the produced materials were determined. Furthermore, the cytotoxicity and antibacterial activity of the materials were also assessed. The results obtained revealed that the PDMS surface modification performed did not affect the materials biocompatibility, but decreased their hydrophobic character and bacterial adhesion and growth on its surface.

Development of UV cross-linked gelatin coated electrospun poly(caprolactone) fibrous scaffolds for tissue engineering

Cardiovascular disease is the leading cause of morbidity and mortality among industrialized countries. Vascular grafts are often required for the surgical treatments. Considering the limitations associated with the use of autografts and with the currently available synthetic materials, a growing demand in tissue engineered vascular grafts has been registered. During the work here described, electrospinning technique was used to prepared fibrous matrices to be applied as vascular implants. For that purpose, electrospun polycaprolactone (PCL) fibrous mats were produced and afterwards coated with different hydrogel formulations based in photocrosslinkable gelatin (GelMA) and the macromers poly(ethylene glycol) acrylate (PEGA) and poly(ethylene glycol) diacrylate (PEGDA). These were further photocrosslinked under UV irradiation using Irgacure® 2959 (by BASF) as the photoinitiator. The suitability of the coated scaffolds for the intended application, was evaluated by assessing their chemical/physical properties as well as their interaction with blood and endothelial cells.

Photocurable bioadhesive based on lactic acid.

Novel photocurable and low molecular weight oligomers based on l-lactic acid with proven interest to be used as bioadhesive were successfully manufactured. Preparation of lactic acid oligomers with methacrylic end functionalizations was carried out in the absence of catalyst or solvents by self-esterification in two reaction steps: telechelic lactic acid oligomerization with OH end groups and further functionalization with methacrylic anhydride. The final adhesive composition was achieved by the addition of a reported biocompatible photoinitiator (Irgacure® 2959). Preliminary in vitro biodegradability was investigated by hydrolytic degradation in PBS (pH=7.4) at 37 °C. The adhesion performance was evaluated using glued aminated substrates (gelatine pieces) subjected to pull-to-break test. Surface energy measured by contact angles is lower than the reported values of the skin and blood. The absence of cytoxicity was evaluated using human fibroblasts. A notable antimicrobial behaviour was observed using two bacterial models (Staphylococcus aureus and Escherichia coli). The cured material exhibited a strong thrombogenic character when placed in contact with blood, which can be predicted as a haemostatic effect for bleeding control. This novel material was subjected to an extensive characterization showing great potential for bioadhesive or other biomedical applications where biodegradable and biocompatible photocurable materials are required.

Production of new 3D scaffolds for bone tissue regeneration by rapid prototyping

The incidence of bone disorders, whether due to trauma or pathology, has been trending upward with the aging of the worldwide population. The currently available treatments for bone injuries are rather limited, involving mainly bone grafts and implants. A particularly promising approach for bone regeneration uses rapid prototyping (RP) technologies to produce 3D scaffolds with highly controlled structure and orientation, based on computer-aided design models or medical data. Herein, tricalcium phosphate (TCP)/alginate scaffolds were produced using RP and subsequently their physicochemical, mechanical and biological properties were characterized. The results showed that 60/40 of TCP and alginate formulation was able to match the compression and present a similar Young modulus to that of trabecular bone while presenting an adequate biocompatibility. Moreover, the biomineralization ability, roughness and macro and microporosity of scaffolds allowed cell anchoring and proliferation at their surface, as well as cell migration to its interior, processes that are fundamental for osteointegration and bone regeneration.

New drug-eluting lenses to be applied as bandages after keratoprosthesis implantation.

Corneal tissue is the most commonly transplanted tissue worldwide. This work aimed to develop a new drug-eluting contact lens that may be used as a bandage after keratoprosthesis. During this work, films were produced using poly(vinyl alcohol) (PVA) and chitosan (CS) crosslinked with glyoxal (GL). Vancomycin chlorhydrate (VA) was impregnated in these systems by soaking. Attenuated total reflectance - Fourier transform infrared spectroscopy was used to confirm crosslinking. The cytotoxic and drug release profile, hydrophilicity, thermal and biodegradation as well as swelling capacity of the samples were assessed through in vitro studies. PVA and PVA/CS films were obtained by crosslinking with GL. The films were transparent, flexible with smooth surfaces, hydrophilic and able to load and release vancomycin for more than 8h. Biodegradation in artificial lachrymal fluid (ALF) with lysozyme at 37°C showed that mass loss was higher for the samples containing CS. Also, the samples prepared with CS showed the formation of pores which were visualized by SEM. All samples revealed a biocompatible character after 24h in contact with cornea endothelial cells. As a general conclusion it was possible to determine that the 70PVA/30CS film showed to combine the necessary features to prepare vancomycin-eluting contact lenses to prevent inflammation after corneal substitution.

Poly(ester amide)s based on (L)-lactic acid oligomers and α-amino acids: influence of the α-amino acid side chain in the poly(ester amide)s properties.

Novel biodegradable and low cytotoxic poly(ester amide)s (PEAs) based on α-amino acids and (L)-lactic acid (L-LA) oligomers were successfully synthesized by interfacial polymerization. The chemical structure of the new polymers was confirmed by spectroscopic analyses. Further characterization suggests that the α-amino acid plays a critical role on the final properties of the PEA. L-phenylalanine provides PEAs with higher glass transition temperature, whereas glycine enhances the crystallinity. The hydrolytic degradation in PBS (pH = 7.4) at 37 °C also depends on the α-amino acid, being faster for glycine-based PEAs. The cytotoxic profiles using fibroblast human cells indicate that the PEAs did not elicit an acute cytotoxic effect. The strategy presented in this work opens the possibility of synthesizing biodegradable PEAs with low citotoxicity by an easy and fast method. It is worth to mention also that the properties of these materials can be fine-tuned only by changing the α-amino acid.

3D Printed scaffolds with bactericidal activity aimed for bone tissue regeneration

Nowadays, the incidence of bone disorders has steeply ascended and it is expected to double in the next decade, especially due to the ageing of the worldwide population. Bone defects and fractures lead to reduced patients quality of life. Autografts, allografts and xenografts have been used to overcome different types of bone injuries, although limited availability, immune rejection or implant failure demand the development of new bone replacements. Moreover, the bacterial colonization of bone substitutes is the main cause of implant rejection. To vanquish these drawbacks, researchers from tissue engineering area are currently using computer-aided design models or medical data to produce 3D scaffolds by Rapid Prototyping (RP). Herein, Tricalcium phosphate (TCP)/Sodium Alginate (SA) scaffolds were produced using RP and subsequently functionalized with silver nanoparticles (AgNPs) through two different incorporation methods. The obtained results revealed that the composite scaffolds produced by direct incorporation of AgNPs are the most suitable for being used in bone tissue regeneration since they present appropriate mechanical properties, biocompatibility and bactericidal activity.

Surface modification of an intraocular lens material by plasma-assisted grafting with 2-hydroxyethyl methacrylate (HEMA), for controlled release of moxifloxacin

ABSTRACT Endophthalmitis, an inflammation of the eye due to perioperative infection, may occur after cataract surgery. Intraocular lenses (IOLs) loaded with an antibiotic have been proposed as an alternative to the conventional postoperative endophthalmitis prophylaxis, since the antibiotic is delivered directly to the target site. In this work, an IOL‐based antibiotic releasing system was prepared from a copolymer used in the production of IOLs and a fluoroquinolone used in endophthalmitis prophylaxis (moxifloxacin, MFX). Argon plasma‐assisted grafting with 2‐hydroxyethyl methacrylate (HEMA) in the presence of MFX was the approach selected for surface modification, with MFX loaded both by entrapment in the grafted polyHEMA coating and by soaking. Surface and bulk properties were evaluated before and after surface modification and the MFX release profiles were obtained both in batch mode (sink conditions) and under hydrodynamic conditions, employing a purpose‐built microfluidic cell, which simulated the hydrodynamic conditions around the eye lens. The effect of storage on the release profile of the best system was also assessed. The best system released MFX for ca. 15 days above the minimum inhibitory concentration for Staphylococcus aureus and Staphylococcus epidermidis. The released MFX showed antimicrobial activity against these bacteria and was non‐cytotoxic against corneal endothelial cells.