In this review paper, the definition of the tissue engineering (TE) was comprehensively explored towards scaffold fabrication techniques and applications. Scaffold properties and features in TE, biological aspects, scaffold material composition, scaffold structural requirements, and old and current manufacturing technologies were reported and discussed. In almost all the reviewed reports, the TE definition denotes renewal, development, and repairs of damaged tissues caused by various factors such as disease, injury, or congenital disabilities. TE is multidisciplinary that combines biology, biochemistry, clinical medicine, and materials science whose application in cellular systems such as organ transplantation serves as a delivery vehicle for cells and drug. According to the previous literature and this review, the scaffold fabrication techniques can be classified into two main categories: conventional and modern techniques. These TE fabrication techniques are applied in the scaffold building which later on are used in tissue and organ structure. The benefits and drawbacks of each of the fabrication techniques have been described in conjunction with current areas of research devoted to deal with some of the challenges. To figure out, the highlighted aspects aimed to define the advancements and challenges that should be addressed in the scaffold design for tissue engineering. Additionally, this study provides an excellent review of original numerical approaches focused on mechanical characteristics that can be helpful in the scaffold design assessment in the analysis of scaffold parameters in tissue engineering.
The term “tissue engineering” (TE) was initially introduced by Professor Robert Nerem in 1988 at UCLA Symposia on Molecular and Cellular Biology [
The conventional method of tissue regeneration and healing is the auto graft method and is mainly dependent on the availability of donor tissues, coupled with other additional effects such as pain and risks to patients such as donor tissue morbidity and infectious diseases [
In scaffold fabrication, the extracellular matrix (ECM) has always received considerable attention among researchers because of its high biological compatibility, biological degradability, and the possibility of rapid remodeling in vivo [
3D printer assembly and fabrication of the scaffold: (a) servo motor driving: (1) CO2 laser, (2) laser scanner, (3) working platform, (4) scraper, and (5) feeder; (b) paving material; (c) start of 3D printing; (d) end of 3D printing [
Additionally, Figure
Even though there exist several methodologies for scaffold fabrication, however, most methods are characterized by low efficiency because of the challenges involved in the making a scaffold that promotes 3D healing and forming a blood vessel within the scaffold [
TE is an interdisciplinary field based on a broad range of areas, where the life sciences and engineering principles are applied to the development of biosubstitutes to restore, maintain, or improve the function of tissue or organ. Thus, TE is a multidisciplinary study combining biology, biochemistry, clinical medicine, and materials science along with materials science to achieve clinical applications [
Scaffolds can serve as cellular systems or as delivery vehicles for cells and drug in cell and tissue regeneration; thus, the cellular material must be capable of adequately colonizing the host cell to meet the needs of regeneration and repair. The other alternative is to have the scaffolds combined with various types of cells that can improve the formation of tissues in vivo by osteogenic lineage or release specific soluble molecules for lineage. These cells may be selectively expanded ex vivo before implanting into the target site [
In cell regeneration, different types of cells (expanded or nonexpanded) extracted from a donor or a patient are included in the scaffold. Adult stem cells, such as bone marrow, fatty tissue, teeth, blood cells, embryonic stem cells, induced pluripotent stem cells (iPS cells), peripheral blood-derived stem, and genetically engineered cells, are the source of extended cells, while bone marrow aspirate-derived platelet-rich plasma cells are the main source for nonexpended cells [
Even though several studies have reported on numerous discoveries in TE, commercialization of these newly discovered functions have significantly increased due to the medical applicability of these findings. Thus, to improve the acceptance of clinical applications of such technologies, it is essential to incorporate specific biological, clinical, and mechanical aspects, which are not only theoretical but can play a role in practical implementation. An appropriate scaffold must be capable to repair body tissues with minimum requirements, for cell growth, vascularization, proliferation, and host integration, and finally, materials should be degraded naturally during or after the healing process [
The biological aspects of scaffolds include their biocompatibility and nontoxicity properties. Cells grown in scaffolds must be able to reproduce and discriminate freely without interference to produce a new matrix [
Biocompatibility allows simultaneous formation of new tissue along with the degradation of the matrix. The matrix should not be toxic so that the system can dispose of it without disturbing other members [
Biological tissue is an incredibly complex 3D structure with complex mechanical functions associated with mass transport characteristics. Therefore, the critical objective of TE is to abridge this structural complexity and function using biological scaffolds that provide cells, proteins, and genes for tissue reconstruction. It is clear that the biological materials and structures cannot replicate complex tissue environments, including numerous cell types that interact with a variety of cytokines to produce extracellular matrices within cells with hierarchical properties that show mechanical function that exhibits high nonlinearity and two-phase [
Typically, most scaffolds consist of polymers, bioceramics, and hybrid materials, whether natural or human-made [
Polymers are of two types, natural polymers or synthetic polymers. Natural polymers, like hyaluronic acid, fibrin, chitosan, and collagen, have good biological compatibility, low immunogenicity, and osteoconductivity. However, they suffer from free degradation rates and low mechanical stability [
Some in vitro studies have reported that the material itself can destroy the results of ex vivo tissue formation compared to natural tissue matrices. Also, in in vivo situation, impaired regeneration can be strongly affected by material immunogenicity, unexpected degradation time, and side effects stemmed from degradation products. Depending on these considerations, the matrices closest to the natural extracellular matrix are the most promising in TE. Therefore, the recently developed approaches in the process of extracorporeal tissue engineering are to prevent nonbiodegradable scaffolds that are reabsorbed at time rate different from skeletal tissue regeneration. Hence, new methods have been developed to overcome these problems by abandoning scaffolds.
At the in vivo level, a tissue consists of 3D units repeated with a scale ranging from 100 to 1000
In practice, the techniques of the fabrication of 3D scaffolds are subdivided into a conventional or rapid prototyping (RP) method (Table
Classification of the scaffold fabrication technologies.
Fabrication technique | Advantages | Disadvantages | |
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Conventional fabrication techniques | 1. Freeze-drying | 1. Use in a variety of purposes |
1. High energy consumption |
2. Solvent casting and practical leaching | 1. Fits thin membranes of thin wall three-dimensional specimens |
1. Time consuming since thin membranes are only used | |
3. Gas foaming | 1. Porosity up to 85% | 1. If the fabrication process did not change, the product obtained might have a closed pore structure or a solid polymeric skin | |
4. Electrospinning | 1. Essential technique for developing nanofibrous scaffolds for the TE |
1. Used solvents can be toxic | |
5. Thermal-induced phase separation | 1. Construction of the thermoplastic crystalline polymer scaffold |
Only used for thermoplastic | |
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Rapid prototyping (RP) | 1. Stereolithography (SLA) | 1. Enables to overcome the challenges related to wastage in subtractive fabrication methods |
1. Has limitations in the process of photopolymerization |
2. Selective laser sintering (SLS) | 1. Using ultrahigh-molecular-weight polyethylene |
1. Steps after processing the spin of the phase are required to remove injected powder | |
3. Solvent-based extrusion freeforming (SEF) | 1. It can be utilized to make ceramic, metal, and metal/ceramic composite part |
1. Temperature extrusion. Their design includes a change to the factors affecting extrusion pressure, including nozzle length-to-diameter ratio, a paste formulation, and the extrusion velocity | |
4. Bioprinting | 1. Low costs |
Depends on existence of cells | |
5. Fused deposition modeling (FDM) | 1. Useful in the scaffold design under the different aspect of scaffold fabrication. Low-temperature deposition | Has limitations in its application to biodegradable polymers [ |
A significant number of scaffolds have been developed conventionally for drug delivery, but they have subsequently been used in 3D cell culture in the context of TE [
In this technique, a solvent combined with uniformly distributed salt particles of a certain size is used to dissolve the polymer solution. The solvent evaporates leaving a matrix containing salt particles. The matrix is then submerged in water, and the salt leaches away to form a structure with high porosity. The solvent casting with particle leaching only fits thin membranes of thin wall three-dimensional specimens; otherwise, the soluble particles cannot be separated from within the polymer matrix [
Thanh et al. [
One of the drawbacks of this fabrication technique is its time consumption since it only uses thin membranes. Layers of porous sheets allow only a defined number of pore networks between them and may, therefore, limit its suitability to use because of the limited porous size [
The process of freeze-drying is also known as lyophilization; it involves the use of a synthetic polymer that is first dissolved in an appropriate solvent. After dissolution, the polymer solution is cooled under the freezing point, resulting in a solid solvent that is evaporated by sublimation to leave a solid scaffold with numerous interconnected pores [
Freeze-drying technique is a more suitable method in biomedical application because of the use of water and ice crystals instead of an organic solvent during scaffold fabrication; however, this methodology is challenged in the fabrication of hierarchical structured scaffolds such as vascular systems in biomedicine [
TIPS is a low-temperature method designed to force phase separation via the temperature alternate related to setting the homogeneous polymer solution with a high temperature in a decrease temperature environment to induce phase separation so that a polymer-rich phase, as well as a poor polymer phase, is achieved [
Phase separation holds great potential in fabrication of 3D nanofibrous scaffolds with uniform pore structures through dual or multiple phase separation processes compared to electrospinning [
Gas foaming technique is a technique that has been evolved to cope with using high temperature and organic cytotoxic solvents. This technique uses relatively inert gas foaming agents such as carbon dioxide or nitrogen to pressurize modeled biologically degradable polymer with water or fluoroform until they are saturated or full of gas bubbles. This technique usually produces structures like a sponge with a pore size of 30 to 700
Electrospinning is known as a technique for making fibers from a solution by using electricity. This technique is vital for developing nanofibrous scaffolds in TE. Electrospinning is a very complicated technique in which charging liquid under high voltage leads to the interaction between the surface tension and electrostatic repulsion that causes droplets on spinneret to erupt and stretch. A standard electrospinning system consists of four main components: a spinner with a syringe pump, a metallic needle, a high-voltage power supply, and a grounded collector, as shown in Figure
Electrospinning device.
Bofan et al. [
Even though electrospinning is a simple and quick method in fabrication of nanofibrous scaffolds, there still exists a challenge in fabrication of scaffolds with complex structures such as homogeneous distribution of pores, hence limited applications in biomedicine [
Rapid prototyping (RP) technologies, also known as solid free-from fabrication (SFF), are a set of manufacturing processes that can generate direct forms directly from computer-aided design (CAD) models of an object without needing specific tooling or knowledge. The RP systems combine powder, liquid, and sheet materials to make parts compared to machining methods (e.g., milling and drilling). Layer by layer, rapid prototyping machine can produce wood, ceramic, plastic and metal objects using thin horizontal cross sections from a computer-generated model [
Stereolithography method is basically used to creating solid, three-dimensional objects by consecutively printing a thin layer of ultraviolet (UV) curable material layer-by-layer. A stereolithography system (Figure
Stereolithography (SLA).
Studies have reported that stereolithography technique has the potential to fabricate different types of cellular machines that could have applications in a broad spectrum of disciplines, such as biosensing, environmental remediation, drug discovery, and energy harvesting, making it a powerful biofabrication technology. However, the feature size of a scaffold that can be fabricated is limited to the beam width of the laser [
In FDM technique, a solid polymer is cast into a hot extrusion nozzle to be melted and extruded on the surface of 3D object using a computer-controlled extrusion and deposition processes; the scaffold is made from multiple layers of adjacent microfilaments. FDM has been utilized to process thermoplastic biopolymers, Hutmacher et al. [
SLS was developed in 1986 by Texas University of Austin. This technique uses laser as the power source to sinter powdered material defined by a 3D model in thin layers. Due to the use of a laser, this technique has been utilized to make various materials, such as polymers, metals, or ceramics [
3DP is a process of creating tools and functional prototype features directly from the computer models, as described in Figure
Three-dimensional printing (3DP).
Studies have demonstrated the design of poly (dopamine) coatings for 3DP poly (lactic acid) scaffolds for bone tissue engineering [
The method for fabricating bioactive PEEK/HA bone scaffolds.
Step | Process | Diagram | Method |
---|---|---|---|
1 | Preparation of ceramic paste |
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Adhesive binder polyvinyl butyral (PVB) and plasticizer polyethyleneglycol (PEG) are fully dissolved in propan-2-ol solvent with the ratio of 75% (w/v) PVB and 25% (w/v) PEG. HA ceramic powder is then added to the solution (with 60% (v/v) of ceramic based on the dried paste) and stirred for 2 hours to achieve a well-dispersed solution |
2 | Solvent evaporation |
|
Excess solvent is evaporated by fast stirring and blowing hot air (such as using hair dryer) until a viscous ceramic paste is achieved |
3 | 3D printing |
|
Ceramic paste is loaded into a syringe for 3D printing. The extrusion process forms lattice-shaped 3D scaffolds by incrementing regularly arranged 2D layers in the vertical axis |
4 | Drying, debinding, and sintering of the scaffold |
|
The scaffold is left at room temperature for 24 hours to allow evaporation of excess solvent and subsequently to place the scaffold in an oven for debinding and sintering. Different heating procedures can be applied depending on the type of ceramic, such as the maximum sintering temperature for HA is 1300°C with a dwelling time of two hours. The bioceramic scaffold is then obtained |
5 | Compression moulding of PEEK powder into the HA scaffold |
|
Static loading: the mould is heated up to 250°C and then load is applied until the temperature reaches 400°C, maintained for a further 20 minutes (dwelling time) and then heating is stopped, and the mould is left to cool under pressure. Dynamic loading: the mould is heated up to 400°C and maintained for 20 minutes. Load is applied for 5 seconds before heating is stopped and then the mould is left to cool under pressure, whereby the PEEK matrix crystallized and solidified |
6 | Obtaining bioactive PEEK/HA composite |
|
Composites are removed from the mould when the temperature has fallen to just below the glass transition temperature (143°C), followed by cooling to room temperature, thus mitigating thermal stress and cracking |
Bioprinting is a 3DP technique, defined as “using material transfer processes for developing a biological pattern and assembly of relevant materials, cells, molecules, tissues, and biodegradable biomaterials with a prescribed structure to achieve some biological functions” [
Schematic illustration of the bone tissue engineering paradigm.
There are numerous different ways of 3D bioprinting, among which autonomous self-assembly, biomimicry, and minitissue building block are based on [
In contrast, microextrusion bioprinting includes a temperature-managed material handling and dispensing system and stage, with either one or both being able to move along the
Laser-assisted bioprinting is a technique based on laser-induced forward transfer. A typical system includes a pulsed laser beam coupled with a focusing system; a “ribbon” with donor transport support covered with a layer of gold or titanium able to absorb laser energy and a cell- and- hydrogel-containing layer of biological material; and a receiving substrate facing the ribbon. The laser-assisted bioprinter directs laser pulses on the laser-absorbing gold layer of the ribbon leading high-pressure bubble, which in turn drives the cell-containing materials to the collector substrate. One of the benefits of this method is that it has nothing to do with the problem of nozzle clogging with cells or material because it is nozzle free. Moreover, it shows compatibility with some biomaterial’s viscosities (1–300 mPa/s). It has been first applied to print a nano-HA scaffold in mice calvarial defects, using a workstation specified to the high throughput biological laser. Before laser printing experiments, it was shown, by MRI, the nonexistence of inflammation due to laser irradiation on mice dura matter. Preliminary results indicated that in vivo bioprinting is feasible and can be utilized for future medical robots and computer-assisted interventions [
Tissue engineering is an interdisciplinary field constructed on a broad range of areas, so the development of this field has been obtained by numerous biomedical 3D scaffold fabrication techniques comprising conventional and current scaffold manufacturing technologies. The conventional methods are solvent casting and particle leaching, freeze-drying, TIPS, gas foaming, and electrospinning, while the modern methods (rapid prototyping) have included stereolithography, fused deposition modeling (FDM), selective laser sintering, and three-dimensional printing. Based on the previous literature and this review, the TE technique is a modern technique in the construction of scaffolds to be used in tissue and organ structure. Both benefits and drawbacks of each of the fabrication techniques mentioned above have been described in conjunction with current areas of research devoted to deal with some of the challenges.
To sum up, the highlighted aspect is aimed to define the advancements and challenges that should be addressed in the scaffold design for tissue engineering. This study provides an excellent review of original numerical approaches focused on individual characteristics of the fabrication techniques that can be helpful in the choosing the suitable scaffold design method for assessment and analysis of scaffold parameters in tissue engineering.
According to all mentioned information in this text, we found that the prototyping techniques represented the modern aspect of fabrication of scaffolds for different applications. In addition, nonstop growing of technology and synthesizing materials will help in appearance of other new methods. And among prototyping techniques, 3D printing will play an important role in large applications that concern the scaffold-based field.
The authors declare that they have no conflicts of interest.
The authors are grateful to the College of Engineering, Nanjing Agricultural University, for their support. This study was also supported by the State Key Laboratory for Manufacturing Systems Engineering at Xi’an Jiaotong University of China (Grant no. sklms2017008).