Henrique A. Almeida

Biomanufacturing for tissue engineering: Present and future trends

Tissue engineering, often referred to as regenerative medicine and reparative medicine, is an interdisciplinary field that necessitates the combined effort of cell biologists, engineers, material scientists, mathematicians, geneticists, and clinicians toward the development of biological substitutes that restore, maintain, or improve tissue function. It has emerged as a rapidly expanding approach to address the organ shortage problem and comprises tissue regeneration and organ substitution. Cells placed on/or within constructs is the most common strategy in tissue engineering. Successful cell seeding depends on fast attachment of cell to scaffolds, high cell survival and uniform cell distribution. The seeding time is strongly dependent on the scaffold material and architecture. Scaffolds provide an initial biochemical substrate for the novel tissue until cells can produce their own extra-cellular matrix (ECM). Thus scaffolds not only define the 3D space for the formation of new tissues, but also serve to provide tissues with appropriate functions. These scaffolds are often critical, both in vivo (within the body) or in vitro (outside the body) mimicking in vivo conditions. Additive fabrication processes represent a new group of non-conventional fabrication techniques recently introduced in the biomedical engineering field. In tissue engineering, additive fabrication processes have been used to produce scaffolds with customised external shape and predefined internal morphology, allowing good control of pore size and pore distribution. This article provides a comprehensive state-of-the-art review of the application of biomanufacturing additive processes in the field of tissue engineering. New and moving trends in biomanufacturing technologies and the concept of direct cell-printing technologies are also discussed.

Rapid prototyping and manufacturing for tissue engineering scaffolds

The controlled fabrication of the scaffold structures for tissue engineering is becoming increasingly important as a viable vehicle in future for regenerative medicine. This paper provides a brief description of the conventional techniques used to manufacture scaffolds and the associated limitations, particularly the lack of full control of the pore morphology and architecture as well as reproducibility. Rapid Prototyping and Manufacturing (RPM laser sintering; extrusion and Three-Dimensional (3D) printing, are described in detail along the main research efforts deployed towards the fabrication of simple and complex 3D scaffolds.

Advanced Processes to Fabricate Scaffolds for Tissue Engineering

Tissue engineering is an interdisciplinary field that necessitates the combined effort of cell biologists, engineers, material scientists, mathematicians, geneticists, and clinicians toward the development of biological substitutes that restore, maintain, or improve tissue function (Fig. 8.1). It comprises tissue regeneration and organ substitution (Table 8.1). The first definition of tissue engineering was provided by Skalak and Fox (1988) who stated it to be “the application of principles and methods of engineering and life sciences toward the fundamental understanding of structurefunction relationships in normal and phatological mammalian tissues and the development of biological substitutes to restore, maintain, or improve tissue function”. An historical overview of this field can be found in a recent report published by National Science Foundation, USA (2003). Three strategies have been explored for the creation of a new tissue (Fuchs et al. 2001; Langer, 1997; Langer and Vacanti, 1993):

Design of tissue engineering scaffolds based on hyperbolic surfaces: structural numerical evaluation.

Tissue engineering represents a new field aiming at developing biological substitutes to restore, maintain, or improve tissue functions. In this approach, scaffolds provide a temporary mechanical and vascular support for tissue regeneration while tissue in-growth is being formed. These scaffolds must be biocompatible, biodegradable, with appropriate porosity, pore structure and distribution, and optimal vascularization with both surface and structural compatibility. The challenge is to establish a proper balance between porosity and mechanical performance of scaffolds. This work investigates the use of two different types of triple periodic minimal surfaces, Schwarz and Schoen, in order to design better biomimetic scaffolds with high surface-to-volume ratio, high porosity and good mechanical properties. The mechanical behaviour of these structures is assessed through the finite element method software Abaqus. The effect of two parametric parameters (thickness and surface radius) is also evaluated regarding its porosity and mechanical behaviour.

Additive manufacturing techniques for scaffold-based cartilage tissue engineering

Articular cartilage damage is of great concern as it creates chronic pain and reduction of joint movement, leading to osteoarthritis. In current treatments, the resulting healing tissues lack structural organisation of cartilage and consequently have inferior mechanical properties when compared to native cartilage, therefore being prone to failure. Tissue engineering has long worked on cartilage regeneration and several requirements have been identified for the engineered structures to meet the desired function, by combining biodegradable and biocompatible materials, cells and growth factors, aiming at the production of biological structures closely resembling the native tissue. Within the scaffold based techniques for cartilage tissue production, conventional methods have shown limitations, especially regarding the control over the micro-structure and repeatability of the produced constructs. Therefore, additive manufacturing techniques grew popular, allowing for a high level of control over the internal scaffold architecture and external shape of the construct, as well as guaranteeing its reproducibility.

Innovative Developments in Design and Manufacturing: Advanced Research in Virtual and Rapid Prototyping -- Proceedings of VRP4, Oct. 2009, Leiria, Portugal

The use of rapid prototyping has increasingly begun to reveal itself as a tool of great value in supporting medical activity. From two-dimensional medical images from computed tomography (CT) or magnetic resonance imaging (MRI) it is possible to obtain three-dimensional models. The models produced by rapid prototyping technologies are useful both in educational and medical-surgical environments. It can simplify the diagnosis of certain diseases, the development of complex surgical procedures, the prostheses and medical devices manufacture and the visualization of anatomical structures in educational environment produce models and prototype parts (Alves & Braga 2001). One of the main applications of Rapid Prototyping is the fast way that is allowed in verifying new concept projects in the earlier stages or even in advanced phases of conception. In all Rapid Prototyping processes, a 3D CAD model is used that is translated into an STL (Stereolithography) format file, (Souza et al. 2003) where all the model surfaces are converted in a triangle mesh. In Biomedical Engineering field, using Rapid Prototyping techniques it is possible to produce several types of anatomical models and implant replica with educational purposes or to better understand a specific patient pathology. The models, depending of available techniques, can be made of paper, wax, ceramic, plastic or metal (Antas & Lino 2008). These models can be produced without finishing or color or have these finishing operations done later to improve visualization. For educational purpose it is possible to manufacture implant replica with much lower cost than the implant value. A great interest can be found in anatomical models manufacture from patient tomographic images. These models allow students from biomedical field to have an easier view of a specific pathology and compare it with normal anatomical models. To better understand image techniques and anatomy, it is also possible to simultaneously compare the original image (TC or MRI) and 3D solid model. Medical professionals cooperate with other field professionals to optimize pre-surgical pathology analysis, shorten surgical times, create personalized tools, facilitate the communication with patients and, simultaneously, to explore the capabilities this technology offers in personalized prosthesis design (Antas & Lino 2008). Vertebral Spine replica are particularly useful to diagnose, plan and simulate surgical procedures as it also allows the patients to understand the nature of their pathologies as well the need for surgical procedures (Madrazo et al. 2008) Several manufacturing processes are available today, as Fused Deposition Modeling (FDM), Stereolithography (SLA), Selective Laser Sintering (SLS), Tridimensional Printing (TDP or 3DP) and Laminated Object Manufacturing (LOM) among other specific processes. A brief description of the most used Rapid Prototyping processes is presented as follows: • Fused Deposition Modeling (FDM): This prototyping process build the prototypes by depositing an extruded thermoplastic material. The injection head draw transversal section perimeters and fills them building, this way, each layer. The most used material is ABS once it has good mechanical properties. More recently have been developed equipments that allow the used of materials such as polycarbonate and polyphenilsulfone (PPSU) that have better mechanical and thermic properties than ABS. • Stereolitography (SLA): This system builds the prototype by polymerizing a photosensitive liquid resin by applying an ultraviolet light formed by a laser. The solidifying process is made layer by layer, allowing obtaining a good surface finished prototype. • Selective Laser Sintering (SLS): This process allows physical models building by using dust materials like ceramics or metal. These materials are processed in an inert and thermally controlled environment inside a chamber. In here, the melting point (sintering) is achieved by action of a CO2 laser. After one layer being sinterized, another layer is deposited until the prototype is finished. This method demands a post-processing work to obtain a better surface. • Three Dimensional Printing (TDP or 3DP): In this process, models are built from a dust material (which can be a blend using materials like composite, cellulose among others) infiltrated with a liquid binder. This binder is applied through a printing head as used in traditional printing. The prototype is removed having the dust blended with the binder and needing operations of cleaning and medium consolidation. • Laminated Object Manufacturing (LOM): In LOM, most of the times, the models are obtained by gluing successive layers of paper which are cut by a laser beam. All the paper not used in the model is cut in square or rectangle forms to make easier prototype remove. To ensure the needed rigidity a frame is also built. Model definition will result from paper thickness and quality. Sometimes, instead paper there can be, also, used glass fibres, ceramics or metal (Alves & Braga 2001).

Structural and Vascular Analysis of Tissue Engineering Scaffolds, Part 2: Topology Optimisation

Rapid prototyping technologies were recently introduced in the medical field, being particularly viable to produce porous scaffolds for tissue engineering. These scaffolds should be biocompatible, biodegradable, with appropriate porosity, pore structure, and pore distribution, on top of presenting both surface and structural compatibility. Surface compatibility means a chemical, biological, and physical suitability with the host tissue. Structural compatibility corresponds to an optimal adaptation to the mechanical behaviour of the host tissue. This chapter presents a computer tool to support the design of scaffolds to be produced by rapid prototyping technologies. The software enables to evaluate scaffold mechanical properties as a function of porosity and pore topology and distribution, for a wide rage of materials, suitable for both hard and soft tissue engineering.

Biofabrication of Hydrogel Constructs

Hydrogels are three dimensional (3D) hydrophilic networks with the ability to absorb and retain large amounts of water without dissolution, as a result of the establishment of physical or chemical bonds between the polymeric chains. Hydrogels obtained from either natural or synthetic polymers are attractive materials for tissue engineering applications due to their excellent biocompatibility, biodegradability, elasticity and compositional similarities to the extracellular matrix. Several techniques have been explored to produce hydrogel meshes, films or 3D constructs for cell attachment, differentiation and proliferation or to release drugs and growth factors according to specific release profiles. This chapter describes the current state-of-the-art of biomanufacturing additive processes to produce hydrogel constructs for tissue engineering. Biomanufacturing processes are described in detail and the major advantages and limitations outlined.

Structural and Vascular Analysis of Tissue Engineering Scaffolds, Part 1: Numerical Fluid Analysis

Rapid prototyping technologies were recently introduced in the medical field, being particularly viable to produce porous scaffolds for tissue engineering. These scaffolds should be biocompatible, biodegradable, with appropriate porosity, pore structure, and pore distribution on top of presenting both surface and structural compatibility. This chapter presents the state-of-the-art in tissue engineering and scaffold design using numerical fluid analysis for optimal vascular design. The vascularization of scaffolds is an important aspect due to its influence regarding the normal flow of biofluids within the human body. This computational tool also allows to design either a scaffold offering less resistance to the normal flow of biofluids or reducing the possibility for blood coagulation through forcing the flow toward a specific direction.

Computer modelling and simulation of a bioreactor for tissue engineering

A conventional approach to tissue engineering involves the implantation of porous, biodegradable and biocompatible scaffolds seeded with cells into the defect site. In some strategies, tissue engineering requires the in vitro culture of tissue-engineering constructs for implantation later. In this case, bioreactors are used to grow 3D tissues under controlled and monitored conditions. However, the quality of the resulting 3D tissue is highly dependent on the design and dimensions of the bioreactor, as well on the operating conditions. In this work, a computational fluid dynamic software package was used to investigate the influence of cylindrical bioreactor dimensions (length and diameter) on the fluid flow and scaffold shear stress. Computer simulations were performed using three different rotational movements (horizontal, vertical and biaxial rotation) and appropriate boundary conditions. Results show that the effect of the bioreactor length on the scaffold shear stress is more important than the diameter, while high length is associated to low scaffold shear stress. On the other hand, the fluid flows within the bioreactor and scaffold shear stresses are dependent on the rotational movement, being more uniform in the biaxial rotation due to the combination of rotational movements.