Bioprinting

Bioprinting is an emerging field in the areas of tissue engineering and regenerative medicine. It is defined as the printing of structures consisting of living cells, biomaterials and active biomolecules. The ultimate aim is to produce implantable organs and tissues to replace the use of autografts, which cause donor site morbidity and require two invasive surgeries. Not only is bioprinting aimed at the restoration of tissue, it has significant potential for drug delivery and cancer studies. Bioprinting provides control over cell placement and therefore creates a homogenous distribution of cells correlating to a uniform tissue ingrowth. Another attribute of bioprinting is the production of patient-specific spatial geometry, controllable microstructures and a high degree of reproducibility and scalability between designs. This book chapter will discuss the many parameters of bioprinting; manufacturing techniques, precursor materials, types of printed cells and the current research

Estruturas tridimensionais eletro-estimuláveis à base de nanoestruturas de carbono/hidrogel para engenharia de tecido neuronal

The main objective of the present work consists of the optimization of the production of three-dimensional electro-responsive carbon-reinforced hydrogels, to study their cytocompatibility with neural stem cells (NSCs) for neural tissue engineering. For that matter, initially vertically aligned carbon nanotubes (VA-CNTs) with two different patterns were prepared by thermal chemical vapor deposition (T-CVD): (1) VA-CNTs dense forest and (1) VA-CNTs micropillars. Furthermore, the substrates previously described were studied after acetone vapor treatment, resulting in a cellular and “flower-like” pattern morphology, respectively. Structural characterization of the respective samples was made using scanning electron microscopy (SEM), transmission electron microscopy (TEM) and the measurement of the water contact angle (WCA). The integration with gelatinmethacryloyl (GelMA) -based hydrogels were explored in the different studied samples. The influence of the different VA-CNTs prepared patterns was studied by the evaluation of the cell behavior with resort to NSCs. By immunocytochemical staining, cell viability assays and SEM, it was observed the cells affinity for the diverse carbon structures, in comparison to the silicon (Si) substrate. Besides, it was also verified the suitability of the VA-CNTs platforms for cell viability and proliferation. The collapsed VA-CNTs substrate made evident the tendency for cell differentiation into neurons, possibly due to their superficial roughness at the nanoscale, which favors this biological mechanism. The results obtained demonstrated that VA-CNTs based structures favors the proliferation and differentiation of NSCs, making them promising as future threedimensional electroresponsive structures with excellent performances for neural tissue engineering.O principal objetivo do presente trabalho constituiu na otimização da produção de estruturas tridimensionais eletro-estimuláveis à base de nanoestruturas de carbono/hidrogel, estudando a sua citocompatibilidade com células estaminais para engenharia de tecido neuronal. Nesse sentido foram primeiramente preparados dois padrões de nanotubos de carbono verticalmente alinhados (VA-CNTs) por deposição química em fase vapor (T-CVD): (1) floresta densa de VA-CNTs e (2) micropilares de VA-CNTs. Além disso, foram também estudados os substratos anteriormente descritos após tratamento por vapor de acetona, resultando na formação de VA-CNTs e micropadrões colapsados, apresentando uma morfologia com um padrão celular e uma semelhante a uma "flor", respetivamente. As respetivas amostras foram caracterizadas por microscopia eletrónica de varrimento (SEM), de transmissão (TEM) e foi medido o ângulo de contacto com a água (WCA). As diferentes amostras estudadas foram exploradas na integração com hidrogéis à base de gelatina metacrilada (GelMA). A influência dos diferentes padrões de VA-CNTs preparados foi estudada através da avaliação do comportamento celular com o recurso a células estaminais neurais (NSCs). Por ensaios de imunocitoquímica, viabilidade celular e SEM, foi observada a afinidade das células para com as diversas estruturas de carbono, em comparação com o substrato de silício (Si). Para além disso foi também verificada a aptidão das diversas estruturas baseadas em VA-CNTs como plataformas para proliferação e diferenciação de NSCs. Os substratos de VA-CNTs colapsados evidenciaram uma propensão para induzir a diferenciação celular em neurónios, possivelmente devido à sua rugosidade superficial à nanoescala favorecer este mecanismo biológico. Os resultados obtidos demonstraram que as estruturas baseadas em VA-CNTs favorecem a proliferação e diferenciação das células estaminais neurais, podendo futuramente ser aplicados como estruturas tridimensionais eletroestimuláveis com elevado desempenho para engenharia de tecido neural.Mestrado em Materiais e Dispositivos Biomédico

4D Printing: The Development of Responsive Materials Using 3D-Printing Technology

Additive manufacturing, widely known as 3D printing, has revolutionized the production of biomaterials. While conventional 3D-printed structures are perceived as static, 4D printing introduces the ability to fabricate materials capable of self-transforming their configuration or function over time in response to external stimuli such as temperature, light, or electric field. This transformative technology has garnered significant attention in the field of biomedical engineering due to its potential to address limitations associated with traditional therapies. Here, we delve into an in-depth review of 4D-printing systems, exploring their diverse biomedical applications and meticulously evaluating their advantages and disadvantages. We emphasize the novelty of this review paper by highlighting the latest advancements and emerging trends in 4D-printing technology, particularly in the context of biomedical applications.The authors would like to acknowledge grants from the Universidad de Buenos Aires, UBACYT 20020150100056BA and PIDAE 2022 (Martín F. Desimone), and from CONICET PIP 0826 (Martín F. Desimone), and PIBAA 28720210100962CO (Sofia Municoy), which supported this work

Nanoengineered Biomaterials for Cell and Therapeutic Delivery

Direct-write extrusion bioprinting, a form of additive manufacturing, is a useful technique to recapitulate anatomical complexity for tissue engineering applications. However, bioprinting has hit a bottleneck in progress due to the lack of available bioinks with high printability, mechanical strength, and biocompatibility. Here, we report a family of hydrogel-based bioinks for extrusion bioprinting from poly (ethylene glycol) (PEG) and two-dimensional (2D) nanoparticles. PEG, a non-fouling easily modifiable polymer, combined with biocompatible Laponite XLG nanoparticles (2D nanosilicates) to obtain shear-thinning hydrogel bioinks. Electrostatic interactions between nanoparticles and hydrogen-bonding between polymer and nanoparticles govern the flow behavior and printability of bioink. The evaluation of hydrogel bioink using flow sweeps, peak holds, and dynamic oscillatory rheology, suggest that minimum shear-thinning index of ~0.3, solution viscosities >1000 Pa·s, and 80% recovery within 30s are necessary for printing high fidelity constructs. Mechanically stiff 3D printed structures are obtained by covalently crosslinking polymeric chains using ultraviolet (UV) light. Modifications to the PEG system through inclusion of dithiothreitol linkage or combining with gelatin methacrylate are used to control matrix degradation, cell adhesion properties, and therapeutic release. We envision that PEG bioinks can be used to print complex, large-scale, cell-laden tissue constructs with high structural fidelity and mechanical stiffness for applications in custom bioprinted scaffolds and tissue engineered implants

Nanoengineered Ionic-Covalent Entanglement (NICE) Reinforced Bioinks for 3D Bioprinting

Three-dimensional (3D) bioprinting is emerging as a promising method for rapid fabrication of biomimetic cell-laden constructs for tissue engineering using cell-containing hydrogels, called bioinks, that can be cross-linked to form a hydrated matrix for encapsulated cells. Bioprinting currently enables precise deposition of viable cells in 2 dimensions, however, their printability in the Z-axis is severely limited because the inks are too weak to support additional layers or do not have the flow properties necessary to fabricate stable many-layered structures. Thus, extrusion-based 3D bioprinting has hit a bottleneck in progress over the lack of suitable bioinks. My research has focused on overcoming this limitation by developing a bioink able to bioprint in all 3 dimensions. Nanoengineered Ionic-Covalent Entanglement (NICE) bioink formulations combine nanocomposite and ionic-covalent entanglement (ICE) strengthening mechanisms to print customizable cell-laden constructs for tissue engineering with high structural fidelity and mechanical stiffness. Nanocomposite and ICE strengthening mechanisms complement each other through synergistic interactions, improving mechanical strength, elasticity, toughness, and flow properties beyond the sum of the effects of either reinforcement technique alone. NICE bioinks can be used to bioprint complex, large-scale, cell-laden constructs for tissue engineering with high structural fidelity and mechanical stiffness for applications in custom bioprinted scaffolds and tissue engineered implants. Next, we transform this platform technology into a specialized bioink for recreating missing bone tissue by testing bioink components to create a highly printable bioink with appropriate mechanical and degradation properties for osteogenic tissue formation. Then, bone marrow derived stem cells are encapsulated and bioprinted into custom structures using patient scans, and are closely followed for stem cell differentiation, proliferation, histological changes, and blood vessel ingrowth. The overall effect of this research is the development of a new range of bioinks capable of replicating large 3D tissue structures, and the demonstration of their use for rapidly fabricating cell-containing custom scaffolds for bone tissue regeneration. I envision my research’s continued development towards a realistic clinical process for bioprinting patient-specific bone tissue

4D Printing : The Development of Responsive Materials Using 3D-Printing Technology

Additive manufacturing, widely known as 3D printing, has revolutionized the production of biomaterials. While conventional 3D-printed structures are perceived as static, 4D printing introduces the ability to fabricate materials capable of self-transforming their configuration or function over time in response to external stimuli such as temperature, light, or electric field. This transformative technology has garnered significant attention in the field of biomedical engineering due to its potential to address limitations associated with traditional therapies. Here, we delve into an in-depth review of 4D-printing systems, exploring their diverse biomedical applications and meticulously evaluating their advantages and disadvantages. We emphasize the novelty of this review paper by highlighting the latest advancements and emerging trends in 4D-printing technology, particularly in the context of biomedical applications

Recent progress in extrusion 3D bioprinting of hydrogel biomaterials for tissue regeneration: a comprehensive review with a focus on advanced fabrication techniques

Over the last decade, 3D bioprinting has received immense attention from research communities for developing functional tissues. Thanks to the complexity of tissues, various bioprinting methods are exploited to figure out the challenges of tissue fabrication, in which hydrogels are widely adopted as a bioink in cell printing technologies based on the extrusion principle. Thus far, there is a wealth of the literature proposing the crucial parameters of extrusion-based bioprinting of hydrogel biomaterials (e.g., hydrogel properties, printing conditions, and tissue scaffold design) toward enhancing performance. Despite the growing research in this field, numerous challenges that hinder advanced applications still exist. Herein, the most recently reported hydrogel-based bioprinted scaffolds, i.e., skin, bone, cartilage, vascular, neural, and muscular (including skeletal, cardiac, and smooth), are systematically discussed with an emphasis on the advanced fabrication techniques from tissue engineering perspective. Methods covered include the multiple-dispenser, coaxial, and hybrid 3D bioprinting. The present work is a unique study to figure out the opportunities of the novel techniques to fabricate complicated constructs with structural and functional heterogeneity. Finally, the principal challenges of current studies and a vision of future research are presented

Nanoengineered Biomaterials for Cell and Therapeutic Delivery

Direct-write extrusion bioprinting, a form of additive manufacturing, is a useful technique to recapitulate anatomical complexity for tissue engineering applications. However, bioprinting has hit a bottleneck in progress due to the lack of available bioinks with high printability, mechanical strength, and biocompatibility. Here, we report a family of hydrogel-based bioinks for extrusion bioprinting from poly (ethylene glycol) (PEG) and two-dimensional (2D) nanoparticles. PEG, a non-fouling easily modifiable polymer, combined with biocompatible Laponite XLG nanoparticles (2D nanosilicates) to obtain shear-thinning hydrogel bioinks. Electrostatic interactions between nanoparticles and hydrogen-bonding between polymer and nanoparticles govern the flow behavior and printability of bioink. The evaluation of hydrogel bioink using flow sweeps, peak holds, and dynamic oscillatory rheology, suggest that minimum shear-thinning index of ~0.3, solution viscosities >1000 Pa·s, and 80% recovery within 30s are necessary for printing high fidelity constructs. Mechanically stiff 3D printed structures are obtained by covalently crosslinking polymeric chains using ultraviolet (UV) light. Modifications to the PEG system through inclusion of dithiothreitol linkage or combining with gelatin methacrylate are used to control matrix degradation, cell adhesion properties, and therapeutic release. We envision that PEG bioinks can be used to print complex, large-scale, cell-laden tissue constructs with high structural fidelity and mechanical stiffness for applications in custom bioprinted scaffolds and tissue engineered implants

3D Bioprinting In Bone And Cartilage Regeneration Review

 Bone and articular cartilage degeneration and damage are the most common causes of musculoskeletal disability. 3D bioprinting can help regenerate these structures. Autologous/allogeneic bone and cartilage transplantation, vascularized bone transplantation, autologous chondrocyte implantation, mosaicplasty, and joint replacement are all common clinical and surgical procedures. In vitro layer-by-layer printing of biological materials, living cells, and other biologically active substances using 3D bio printing technology is anticipated to replace the aforementioned repair methods. With the ability to prepare various organs and tissue structures, 3D bio printing has largely solved the issue of insufficient organ donors. Researchers use biomedical materials and cells as discrete materials. Bioprinting cell selection and its use in bone and cartilage repair are the primary topics of discussion in this paper

Efeitos de nanopartículas contendo galantamina após lesão da medula espinal em ratos

Distúrbios do sistema nervoso central apresentam dificuldades de tratamento devido à baixa capacidade regenerativa das células deste sistema. Dentre eles, a lesão da medula espinal (LME) é uma condição que leva a grandes impactos na vida do paciente acometido, como situações de paraplegia, e não há tratamentos eficazes até o presente momento. A galantamina é um fármaco utilizado na doença de Alzheimer. Estudo prévio realizado pelo grupo demonstrou a eficácia em promover melhora funcional em um modelo de LME em ratos. Entretanto, administrações sistêmicas de medicamentos produzem mais efeitos colaterais e levam a menor biodisponibilidade do fármaco no tecido-alvo. Assim, o objetivo deste trabalho foi produzir e avaliar o potencial terapêutico de nanopartículas contendo galantamina para LME. As partículas de poli(ácido lático-co-glicólico) (PLGA) e galantamina foram produzidas por electrospraying e caracterizadas por microscopia eletrônica de varredura e por espalhamento de luz dinâmico para avaliação do tamanho, potencial zeta e índice de polidispersão (PdI). A liberação de galantamina das partículas foi avaliada por 35 dias através de cromatografia líquida de alta eficiência (HPLC). O potencial neuroprotetor das partículas foi avaliado in vitro em uma cultura de células PC12. In vivo, utilizou-se um modelo de contusão de LME em ratos e as formulações farmacêuticas foram aplicadas no local da lesão. Os animais foram separados em 5 grupos: Sham, lesão, galantamina, partículas de PLGA e partículas de PLGA contendo galantamina (PG). Os efeitos desse tratamento foram avaliados após 3 e 42 dias, quando os parâmetros oxidativos e inflamatórios das medulas espinais foram avaliados, assim como os parâmetros histológicos, após 42 dias. Ainda, a locomoção dos animais foi avaliada semanalmente, por 6 semanas. As partículas poliméricas apresentaram morfologia bicôncava e tamanho adequado, assim como potencial zeta e PdI também adequados para administração local. A galantamina apresentou uma liberação no perfil de burst, mas manteve uma liberação controlada ao longo de 35 dias. In vitro, apenas o tratamento com galantamina livre reduziu a perda de viabilidade das células expostas a peróxido de hidrogênio e nenhuma das formulações farmacêuticas promoveu a redução da produção de espécies reativas de oxigênio (EROs) após a exposição. Por outro lado, em estudo in vivo, o único grupo que apresentou melhoras significativas na função motora após 42 dias foi o grupo tratado com PG. Três dias após a lesão, a administração de galantamina diminuiu os níveis de peroxidação lipídica, ao passo que PG diminuiu os níveis de EROs e IL-1β, além dos níveis de peroxidação lipídica. Além disso, ao analisar os efeitos dos tratamentos em 42 dias após a lesão, o grupo no qual administrou-se apenas a galantamina apresentou diminuição dos níveis de EROs, enquanto o grupo PG diminuiu tanto EROs quanto IL-1β. Assim, o tratamento com PG demonstrou melhoras inflamatórias e oxidativas, bem como melhora funcional após LME.Central nervous system disorders are particularly difficult to treat due to the limited regenerative capacity of the cells from this system. Among them, spinal cord injury (SCI) is a condition that provokes great impact on the patient’s life, such as paraplegy, and currently there are no effective treatments for this disorder. Galantamine is a drug used for Alzheimer’s disease. A previous study from the group demonstrated its efficacy in promoting functional improvements in a rat model of SCI. However, systemic drug administration can cause side effects and lead to decreased drug bioavailability at the target tissue. Hence, the aim of this study has been to produce and evaluate the therapeutic potential of galantamine nanoparticles for SCI. Poly(lactic acid-co-glycolic acid) (PLGA) and galantamine particles were produced using electrospraying and characterized by scanning electron microscopy and dynamic light scattering for assessment of size, zeta potential and polydispersity index (PdI). Galantamine release from the particles was evaluated for 35 days using High Performance Liquid Chromatography (HPLC). The particles’ neuroprotective potential was assessed in vitro in PC12 cell culture. A contusion model of SCI in rats was used in vivo, and the pharmaceutical formulations were applied at the lesion site. The animals were divided into 5 groups: sham, injury, galantamine, PLGA particles and PLGA particles containing galantamine (PG). The treatment effects were evaluated after 3 and 42 days, when oxidative and inflammatory parameters were assessed, as well as histological parameters after 42 days. In addition, the animals' locomotion was evaluated weekly for 6 weeks. The particles presented a biconcave shape and adequate size, as well as adequate zeta potential and PdI for local administration. Galantamine was released in a burst profile, but maintained a controlled release for 35 days. In vitro, only galantamine reduced the viability loss in the cells exposed to hydrogen peroxide, and no pharmaceutical formulation reduced reactive oxygen species (ROS) production after the exposure. In vivo, on the other hand, the only group that presented significant improvements in motor function was in the PG treatment. Three days after the injury, the administration of galantamine resulted in a decrease in lipid peroxidation levels, whereas the use of PG improved levels of reactive oxygen species (ROS) and IL-1β, in addition to lipid peroxidation levels. Furthermore, when analyzing the treatment effects 42 days after the injury, galantamine treatment was able to reduce ROS, while PG reduced both ROS and IL-1β. Therefore, PG treatment showed not only inflammatory and oxidant improvements, but also significant functional recovery after SCI