Biofunctionalization of zirconia based materials by immobilization of alp in tissue engineering applications

The aim of this project is to modify Zirconia surface to perform as a bioactive material in contact with biological tissue with high stability and activity. Alumina was also modified in order to compare the properties with Zirconia in the same laboratory condition. The protein used in this test is alkaline phosphatase (ALP) which improves mineralization and creation of hydroxyapatite in bone formation process. Adsorption and covalent bonding immobilization were investigated on both ceramics. The modified ceramic surfaces also tested in contact with and without human body cells, in biological condition in vitro. This project contains laboratory tests for powders and planar surfaces. Characterizations of zirconia powder, such as IEP (isoelectric point) were investigated after ALP immobilization. In addition, ALP functionalized planar surface and its behaviour was studied in vitro in simulated body conditions with and without presence of osteoblast-like cells. Main results of these studies supported that high mineralization accomplished by ALP functionalized zirconia in compare with non-functionalized zirconia in in vitro. In summary, the results of this research indicate the successful immobilization and surface modification of zirconia. In addition, zirconia powder silanized by APTES, as the initial step of surface modification, showed acceptable stability in room temperature during 28 days. In case of comparison between physical adsorption and covalent bonding methods, it can be observed that physical adsorption represents higher enzyme attachment. However, these attachments do not show suitable stability. In all the experiments regarding modification of the particle surface, Zirconia powder illustrates higher potential for protein (ALP) immobilization compare to Alumina

Electrospun Fibrous Architectures for Drug Delivery, Tissue Engineering and Cancer Therapy

The versatile electrospinning technique is recognized as an efficient strategy to deliver active pharmaceutical ingredients and has gained tremendous progress in drug delivery, tissue engineering, cancer therapy, and disease diagnosis. Numerous drug delivery systems fabricated through electrospinning regarding the carrier compositions, drug incorporation techniques, release kinetics, and the subsequent therapeutic efficacy are presented herein. Targeting for distinct applications, the composition of drug carriers vary from natural/synthetic polymers/blends, inorganic materials, and even hybrids. Various drug incorporation approaches through electrospinning are thoroughly discussed with respect to the principles, benefits, and limitations. To meet the various requirements in actual sophisticated in vivo environments and to overcome the limitations of a single carrier system, feasible combinations of multiple drug-inclusion processes via electrospinning could be employed to achieve programmed, multi-staged, or stimuli-triggered release of multiple drugs. The therapeutic efficacy of the designed electrospun drug-eluting systems is further verified in multiple biomedical applications and is comprehensively overviewed, demonstrating promising potential to address a variety of clinical challenges.Peer reviewe

Gelatin-Lysozyme Nanofibrils Electrospun Patches with Improved Mechanical, Antioxidant and Bioresorbability Properties for Myocardial Regeneration Applications

Biopolymeric patches show enormous potential for the regeneration of infarcted myocardium tissues. However, most of them usually lack appropriate mechanical performance, stability in water, and important functionalities; for instance, antioxidant activity. Protein nanofibrils, such as lysozyme nanofibrils (LNFs), are biocompatible nanostructures with excellent mechanical performance, water insolubility, and antioxidant activity exploited to fabricate materials for different biomedical applications. In this study, LNFs are used to produce gelatin electrospun nanocomposite cardiac patches with improved properties. The addition of the LNFs to the gelatin electrospun patches enhance their mechanical properties, increasing the patches Young's modulus from 3 to 6 MPa, in their wet state, which agrees with the requirements of myocardial contractility. Additionally, it is observed an increment of the antioxidant activity to 80%, by adding only 5% (w/w) of LNFs, and the bioresorbability rate is shortened to 30-35 d, compared to 45 d for the gelatin-only patches, while maintaining their morphology, and biocompatibility toward cardiomyoblasts and fibroblasts. Furthermore, 15% of a model drug is burst released from the patches and preserved for 21 d. Overall, these results demonstrate that LNFs have a great potential as functional reinforcements to fabricate biopolymeric electrospun patches for myocardial infarcted tissue regeneration.Peer reviewe

Acetylated nanocellulose for single-component bioinks and cell proliferation on 3D-printed scaffolds

Nanocellulose has been demonstrated as a suitable material for cell culturing, given its similarity to extracellular matrices. Taking advantage of the shear thinning behavior, nanocellulose suits three-dimensional (3D) printing into scaffolds that support cell attachment and proliferation. Here, we propose aqueous suspensions of acetylated nanocellulose of a low degree of substitution for direct ink writing (DM). This benefits from the heterogeneous acetylation of precursor cellulosic fibers, which eases their deconstruction and confers the characteristics required for extrusion in DIW. Accordingly, the morphology of related 3D printed architectures and their performance during drying and rewetting as well as interactions with living cells are compared with those produced from typical unmodified and TEMPO-oxidized nanocelluloses. We find that a significantly lower concentration of acetylated nanofibrils is needed to obtain bioinks of similar performance, affording more porous structures. Together with their high surface charge and axial aspect, acetylated nanocellulose produces dimensionally stable monolithic scaffolds that support drying and rewetting, required for packaging and sterilization. Considering their potential uses in cardiac devices, we discuss the interactions of the scaffolds with cardiac myoblast cells. Attachment, proliferation, and viability for 21 days are demonstrated. Overall, the performance of acetylated nanocellulose bioinks opens the possibility for reliable and scaleup fabrication of scaffolds appropriate for studies on cellular processes and for tissue engineering.Peer reviewe

Microfibers synthesized by wet-spinning of chitin nanomaterials : mechanical, structural and cell proliferation properties

Partially deacetylated chitin nanofibers (ChNF) were isolated from shell residues derived from crab biomass and used to prepare hydrogels, which were easily transformed into continuous microfibers by wet-spinning. We investigated the effect of ChNF solid content, extrusion rate and coagulant type, which included organic (acetone) and alkaline (NaOH and ammonia) solutions, on wet spinning. The properties of the microfibers and associated phenomena were assessed by tensile strength, quartz crystal microgravimetry, dynamic vapor sorption (DVS), thermogravimetric analysis and wide-angle X-ray scattering (WAXS). The as-spun microfibers (14 GPa stiffness) comprised hierarchical structures with fibrils aligned in the lateral direction. The microfibers exhibited a remarkable water sorption capacity (up to 22 g g−1), while being stable in the wet state (50% of dry strength), which warrants consideration as biobased absorbent systems. In addition, according to cell proliferation and viability of rat cardiac myoblast H9c2 and mouse bone osteoblast K7M2, the wet-spun ChNF microfibers showed excellent results and can be considered as fully safe for biomedical uses, such as in sutures, wound healing patches and cell culturing.Peer reviewe

Multifunctional 3D-printed patches for long-term drug release therapies after myocardial infarction

A biomaterial system incorporating nanocellulose, poly(glycerol sebacate), and polypyrrole is introduced for the treatment of myocardial infarction. Direct ink writing of the multicomponent aqueous suspensions allows multifunctional lattice structures that not only feature elasticity and electrical conductivity but enable cell growth. They are proposed as cardiac patches given their biocompatibility with H9c2 cardiomyoblasts, which attach extensively at the microstructural level, and induce their proliferation for 28 days. Two model drugs (3i‐1000 and curcumin) are investigated for their integration in the patches, either by loading in the precursor suspension used for extrusion or by direct impregnation of the as‐obtained, dry lattice. In studies of drug release conducted for five months, a slow in vitro degradation of the cardiac patches is observed, which prevents drug burst release and indicates their suitability for long‐term therapy. The combination of biocompatibility, biodegradability, mechanical strength, flexibility, and electrical conductivity fulfills the requirement of the highly dynamic and functional electroresponsive cardiac tissue. Overall, the proposed cardiac patches are viable alternatives for the regeneration of myocardium after infarction through the effective integration of cardiac cells with the biomaterial.Peer reviewe

Fabrication and Characterization of Drug-Loaded Conductive Poly(glycerol sebacate)/Nanoparticle-Based Composite Patch for Myocardial Infarction Applications

Heart tissue engineering is critical in the treatment of myocardial infarction, which may benefit from drug-releasing smart materials. In this study, we load a small molecule (3i-1000) in new biodegradable and conductive patches for application in infarcted myocardium. The composite patches consist of a biocompatible elastomer, poly(glycerol sebacate) (PGS), coupled with collagen type I, used to promote cell attachment. In addition, polypyrrole is incorporated because of its electrical conductivity and to induce cell signaling. Results from the in vitro experiments indicate a high density of cardiac myoblast cells attached on the patches, which stay viable for at least 1 month. The degradation of the patches does not show any cytotoxic effect, while 3i-1000 delivery induces cell proliferation. Conductive patches show high blood wettability and drug release, correlating with the rate of degradation of the PGS matrix. Together with the electrical conductivity and elongation characteristics, the developed biomaterial fits the mechanical, conductive, and biological demands required for cardiac treatment.Peer reviewe

Conductive vancomycin-loaded mesoporous silica polypyrrole-based scaffolds for bone regeneration

Bone tissue engineering is considered an alternative approach for conventional strategies available to treat bone defects. In this study, we have developed bone scaffolds composed of hydroxyapaptite (HAp), gelatin and mesoporous silica, all recognized as promising materials in bone tissue engineering due to favorable biocompatibility, osteoconductivity and drug delivery potential, respectively. These materials were coupled with conductive polypyrrole (PPy) polymer to create a novel bone scaffold for regenerative medicine. Conductive and non-conductive scaffolds were made by slurry casting method and loaded with a model antibiotic, vancomycin (VCM). Their properties were compared in different experiments in which scaffolds containing PPy showed good mechanical properties, higher protein adsorption and higher percentage of VCM release over a long duration of time compared to non-conductive scaffolds. Osteoblast cells were perfectly immersed into the gelatin matrix and remained viable for 14 days. Overall, new conductive composite bone scaffolds were created and the obtained results strongly verified the applicability of this conductive scaffold in drug delivery, encouraging its further development in tissue engineering applications

Development and Characterization of Conductive Drug-loaded Scaffolds for Tissue Engineering Applications

Biomaterials science denotes a multidisciplinary science combining materials engineering, biomedical engineering, and biology. Tissue engineering is defined as an engineering system field using biomaterials to regenerate, restore tissue, or improve its function. This thesis focuses on the fabrication of scaffolds and patches for bone and heart regeneration using a conductive polypyrrole (PPy) polymer. PPy is widely used in different tissue engineering applications due to its biocompatibility, high electrical properties and conductivity, ease to synthesize, and environmental stability, such as in water and air. First, the role of this polymer in attracting protein and osteoblast cells was investigated. The bone scaffold’s mechanical properties using PPy were analyzed. The results showed no difference between conductive and non-conductive bone scaffolds, presenting almost the same young modulus between the two systems. In addition, more proteins were adsorbed on the surface of the conductive polymers. The same results were obtained in conductive cardiac patches, which showed higher blood coverage on the conductive 2D patch using PPy compared to the non-conductive one. In addition, cellulose was used to 3D print and fabricate a porous conductive cardiac patch. The in vitro investigations showed that cell attachment and proliferation on the conductive bone scaffold and cardiac patches were higher. Biomineralization was induced in simulated body fluid on the bone scaffold’s conductive surface. PPy was also combined with bacterial cellulose (BC) to make conductive hydrogel for the treatment of myocardial infarction. The positive chains of PPy improved the loading of negative charged drug-loaded nanoparticles (NPs). All four systems were successfully developed, fabricated, and loaded with different drugs with different methods to make multifunctional bioengineered systems for drug delivery applications. The results showed that PPy controlled the release of the vancomycin, an antibiotic, and the GATA4-targeted compound 3i-1000, inside the pores of 3D bone scaffold and 3D printed cardiac patch, respectively, which suitable for long-term therapy. Furthermore, bone scaffold, 2D cardiac patch, and conductive BC were combined with microparticles of silica, NPs of porous silicon, and acetylated dextran, respectively, presenting the high potential of in situ drug delivery system enabling and dual delivery of drugs. Overall, this thesis demonstrated that PPy-based biocomposites are successfully developed showing high biocompatibility and high functionality within hard scaffolds and soft patches, and thus, they are very promising platforms for future bone and heart tissue regeneration applications in vivo studies.NOT AVAILABL

3D scaffolding of fast photocurable polyurethane for soft tissue engineering by stereolithography: Influence of materials and geometry on growth of fibroblast cells

Tissue engineering can benefit from the availability of three-dimensional (3D) printing technologies that make it possible to produce scaffolds with complex geometry. Chemical, mechanical, and structural properties should be considered in scaffold design and development since these properties affect cell adhesion, proliferation, and differentiation. To this end, in this study, we developed a series of fast photocuring polyurethanes (PUs), using poly(ε-caprolactone) (PCL) and/or polyethylene glycol (PEG) as microdiols, using a solvent-free method and stereolithography strategy for the fabrication of elastic 3D-printed scaffold. The effects of different diols on the hydrolytic degradation, thermal and mechanical properties, and hydrophilicity of PUs were evaluated. The results showed that PEG-containing PUs had higher degradation rates, and the tensile strength of PU/PCL/PEG was 1.4 and 2 times higher than that of PU/PEG and PU/PCL, respectively. Moreover, the effect of different diols and scaffold geometry on toxicity and cell attachment were studied in vitro. The results of MTT and AlamarBlue assays on dermal fibroblast cells showed high proliferation of printed PU/PCL/PEG scaffold with no sign of cytotoxicity. In addition, compared to cast film PUs, relatively high cell attachment was seen on the surface of printed PU/PCL/PEG even after 4 days. Therefore, 3D printed PU/PCL/PEG showed high applicability in soft tissue engineering, especially for scaffold development.Peer reviewe