Scaffolds for carrying growth factors

Introduction

Scaffolds designed for carrying growth factors are fundamental in tissue engineering and regenerative medicine. These scaffolds provide a three-dimensional (3D) structure that mimics the extracellular matrix (ECM), facilitating cellular activities such as adhesion, proliferation, and differentiation. The integration of growth factors into these scaffolds further enhances their regenerative potential by directing cellular behavior and promoting tissue regeneration. Scaffolds in tissue engineering can be classified based on various criteria, including their material composition, structure, and fabrication methods.

Classification of scaffolds based on their composition

Natural Scaffolds:

Collagen: Mimics the natural extracellular matrix.

Chitosan: Derived from chitin, used for its biocompatibility and biodegradability.

Alginate: Extracted from seaweed, often used in hydrogel form.

Synthetic Scaffolds:

Polylactic Acid (PLA): Biodegradable and used for its mechanical properties.

Polyglycolic Acid (PGA): Often used in combination with PLA.

Polycaprolactone (PCL): Known for its slow degradation rate.

Classification of scaffolds based on structure

Porous Scaffolds: Have interconnected pores to facilitate cell migration and nutrient flow.

Fibrous Scaffolds: Mimic the fibrous nature of the extracellular matrix, often created using electrospinning.

Hydrogels: Water-swollen networks that can encapsulate cells and growth factors.

Microsphere Scaffolds: Small spherical particles that can be loaded with growth factors.

Classification of scaffolds based on their fabrication methods

Electrospinning: Produces fibrous scaffolds with high surface area.

3D Printing: Allows precise control over scaffold architecture.

Freeze-Drying: Creates porous structures by sublimating frozen solvents.

Solvent Casting/Particulate Leaching: Uses a solvent to dissolve the polymer and create a porous structure by leaching out embedded particles.

Classification of scaffolds based on their degradation rate

Biodegradable Scaffolds: Designed to degrade over time, matching the rate of tissue regeneration.

Non-Biodegradable Scaffolds: Provide long-term support and are not intended to degrade.

Natural scaffolds

Collagen Scaffolds

Collagen is a highly versatile natural polymer, widely utilized in tissue engineering due to its excellent biocompatibility, biodegradability, and ability to promote cell adhesion and growth. As the primary structural protein in the extracellular matrix (ECM) of various tissues, collagen provides a favorable environment for cellular activities, making it an ideal material for scaffold fabrication. When combined with growth factors, collagen scaffolds can significantly enhance tissue regeneration and repair. Following is the description of the properties, fabrication methods, applications, and future directions of collagen scaffolds loaded with growth factors.

Biological properties of collagen

Collagen is naturally present in the human body, minimizing the risk of immune rejection and adverse reactions. It degrades into non-toxic byproducts that can be resorbed or excreted by the body. It contains bioactive motifs that interact with cell surface receptors, promoting cell adhesion, migration, and proliferation. Collagen scaffolds can be engineered to have varying mechanical properties, making them suitable for different tissue engineering applications. Collagen scaffolds loaded with BMP-2 have shown to enhance bone formation by promoting osteoblast differentiation and mineralization. These scaffolds can be used to treat critical-sized bone defects and fractures. Furthermore, collagen scaffolds loaded with TGF-β have been used to promote chondrocyte proliferation and extracellular matrix production. These scaffolds are suitable for treating articular cartilage injuries and osteoarthritis. Collagen hydrogels loaded with FGF have been shown to accelerate wound healing by enhancing fibroblast proliferation and angiogenesis. These scaffolds can be used for treating chronic wounds and burns.

Fabrication Methods

Electrospinning is the most widely used technique to make scaffolds. This technique produces collagen nanofibers with a high surface area-to-volume ratio, mimicking the natural ECM structure. Growth factors can be incorporated during the electrospinning process or immobilized on the fiber surface. Another method is Freeze-Drying which involves freezing a collagen solution followed by sublimation of the solvent, creating a porous scaffold structure. Growth factors can be mixed into the collagen solution before freeze-drying. Advanced 3D printing techniques which have been recently introduced, allow for the precise fabrication of collagen scaffolds with complex geometries. Growth factors can be incorporated into the collagen bioink or printed in specific regions. Another technique is hydrogel technique. Collagen hydrogels are formed by polymerizing collagen solutions under physiological conditions. Growth factors can be encapsulated within the hydrogel matrix or crosslinked to the collagen fibers.

Applications of collagen scaffolds

Collagen scaffolds have been used in bone regeneration. Collagen scaffolds loaded with BMP-2 have shown to enhance bone formation by promoting osteoblast differentiation and mineralization. These scaffolds can be used to treat critical-sized bone defects and fractures. Also, collagen scaffolds loaded with TGF-β have been used to promote chondrocyte proliferation and extracellular matrix production. These scaffolds are suitable for treating articular cartilage injuries and osteoarthritis. Researchers investigated the use of collagen scaffolds loaded with TGF-β for repairing articular cartilage defects in rabbits. The TGF-β-loaded scaffolds promoted chondrocyte proliferation and extracellular matrix production, leading to improved cartilage repair. These have also been used to promote wound healing. Collagen hydrogels loaded with FGF have been shown to accelerate wound healing by enhancing fibroblast proliferation and angiogenesis. A clinical trial evaluated the efficacy of collagen hydrogels loaded with FGF in treating chronic diabetic foot ulcers. The FGF-loaded hydrogels accelerated wound healing, reduced ulcer size, and improved tissue granulation compared to standard care. These scaffolds can be used for treating chronic wounds and burns. In areas where neo-angiogenesis is required these scaffolds have been used successfully. Scaffolds loaded with VEGF have been used to enhance vascularization in ischemic tissues. These scaffolds can be applied in cardiovascular grafts and tissue-engineered constructs requiring vascular networks.

Chitosan

Chitosan is a versatile biomaterial widely used as a scaffold in tissue engineering, particularly for bone regeneration. It is a natural polysaccharide derived from chitin which is found in the exoskeletons of crustaceans like shrimp and crabs. The process of deacetylation converts chitin into chitosan, which is soluble in acidic solutions. This solubility allows for easy manipulation and fabrication into various forms such as films, fibers, and hydrogels. Its unique properties, such as biocompatibility, biodegradability, and non-toxicity, make it an ideal candidate for use as a scaffold in the delivery of growth factors. Key properties of chitosan include:

Biocompatibility: Chitosan is well-tolerated by the human body, reducing the risk of adverse immune reactions.

Biodegradability: It can be broken down by lysozymes in the body, ensuring that it does not accumulate and cause long-term issues.

Antimicrobial Activity: Chitosan has inherent antimicrobial properties, which can help in preventing infections at the site of implantation.

Fabrication techniques of chitosan scaffolds

Chitosan scaffolds can be fabricated using a variety of techniques, each offering unique advantages and tailored properties for specific applications in tissue engineering. Solvent casting and particulate leaching is an old process of making scaffolds. This method involves dissolving chitosan in an acidic solution and then casting it into a mold. Particulate leaching, often using salt particles, creates a porous structure. After the solvent evaporates, the scaffold is immersed in water to dissolve the salt, leaving behind a porous chitosan scaffold. Second method of making scaffolds is electrospinning. Electrospinning produces nanofibrous scaffolds by applying a high voltage to a chitosan solution, creating fine fibers that are collected on a grounded surface. This technique is particularly useful for creating scaffolds that mimic the extracellular matrix, promoting cell attachment and proliferation. Lyophilization (Freeze-Drying) is another method of making scaffolds. In this process, a chitosan solution is frozen and then subjected to sublimation under vacuum. This removes the solvent, leaving behind a highly porous scaffold. Lyophilization is advantageous for creating scaffolds with controlled pore sizes and high porosity. 3D Printing is a recent advancement in scaffold fabrication. It allows for precise control over the scaffold’s architecture. Chitosan can be printed layer by layer to create complex structures tailored to specific tissue engineering needs. This technique is highly versatile and can produce scaffolds with intricate designs and controlled porosity.

Chitosan can form hydrogels through physical or chemical cross-linking through a process called gelation. Physical gelation involves changes in temperature or pH, while chemical gelation uses cross-linking agents. Hydrogels are particularly useful for soft tissue engineering and drug delivery applications. Another method is emulsification which involves creating an emulsion of chitosan in an oil phase, followed by solvent evaporation or extraction. The result is a scaffold with a highly porous structure, suitable for applications requiring high surface area. One more method is cross-linking. Cross-linking can enhance the mechanical properties and stability of chitosan scaffolds. This can be achieved using agents like glutaraldehyde or genipin, which form covalent bonds between chitosan molecules, improving the scaffold’s durability.

Similar to particulate leaching, the porogen leaching method uses porogens (e.g., paraffin spheres) that are mixed with chitosan. After the scaffold is formed, the porogens are removed, leaving behind a porous structure. This technique allows for precise control over pore size and distribution. Micro-templating method involves using a template to create a scaffold with a specific microstructure. The chitosan solution is cast around the template, which is then removed, leaving behind a scaffold with the desired architecture. This method is useful for creating scaffolds with complex internal geometries.

Application of chitosan scaffolds

Chitosan scaffolds have been extensively studied for their applications in various fields of tissue engineering, including bone, cartilage, skin, and nerve regeneration. Chitosan scaffolds have shown great promise in bone tissue engineering. They can be combined with osteogenic growth factors such as bone morphogenetic proteins (BMPs) to enhance bone formation. Studies have demonstrated that chitosan scaffolds loaded with BMP-2 significantly improve bone regeneration in critical-sized defects (References are available in book). Chitosan-based scaffolds have also been explored for cartilage repair. The incorporation of growth factors like transforming growth factor-beta (TGF-β) into chitosan scaffolds has been shown to promote chondrogenesis and improve the quality of regenerated cartilage. In the field of wound healing, chitosan scaffolds loaded with growth factors such as epidermal growth factor (EGF) have been used to accelerate the healing process. These scaffolds provide a moist environment and promote cell migration and proliferation, leading to faster wound closure. Chitosan scaffolds have been investigated for their potential in nerve regeneration. The delivery of neurotrophic factors like nerve growth factor (NGF) using chitosan scaffolds has shown promising results in promoting nerve repair and functional recovery.

Alginate

Alginate, a naturally occurring polysaccharide derived from brown seaweed, has emerged as a highly versatile biomaterial in the field of tissue engineering and regenerative medicine. Its unique properties, such as biocompatibility, biodegradability, and gel-forming ability, make it an ideal candidate for use as a scaffold for the delivery of growth factors. Alginate is composed of two types of uronic acids: mannuronic acid (M) and guluronic acid (G). The ratio and sequence of these acids determine the physical properties of alginate, such as its gel strength and viscosity. Alginate can form hydrogels in the presence of divalent cations like calcium, which cross-link the polymer chains, creating a three-dimensional network. It is well-tolerated by the human body, reducing the risk of adverse immune reactions. It can be broken down by enzymes in the body, ensuring that it does not accumulate and cause long-term issues. Alginate is non-toxic, making it safe for use in various biomedical applications.

Fabrication techniques of alginate scaffolds

Several techniques can be employed to fabricate alginate scaffolds, each offering unique advantages and tailored properties for specific applications. Sol-Gel method is commonly used to make alginate scaffolds. This involves dissolving alginate in water and then cross-linking it with divalent cations like calcium to form a gel. This method is simple and allows for the incorporation of growth factors during the gelation process. Another method is electrospinning. This technique produces nanofibrous scaffolds by applying a high voltage to an alginate solution, creating fine fibers that are collected on a grounded surface. Electrospinning is particularly useful for creating scaffolds that mimic the extracellular matrix. One recent advancement in making scaffolds is 3D printing. 3D printing allows for precise control over the scaffold’s architecture. Alginate can be printed layer by layer to create complex structures tailored to specific tissue engineering needs. This technique is highly versatile and can produce scaffolds with intricate designs and controlled porosity. Lyophilization (Freeze-Drying) is one more method of making alginate scaffolds. In this process, an alginate solution is frozen and then subjected to sublimation under vacuum. This removes the solvent, leaving behind a highly porous scaffold. Lyophilization is advantageous for creating scaffolds with controlled pore sizes and high porosity.

Application of alginate scaffolds

Alginate scaffolds have been extensively studied for their applications in various fields of tissue engineering. Alginate scaffolds have been used to regenerate bone defects and promote osteogenesis. Alginate scaffolds loaded with BMP-2 have shown to enhance bone formation by promoting osteoblast differentiation and mineralization. These scaffolds can be used to treat critical-sized bone defects and fractures. In a study (references available in book) alginate hydrogels encapsulating BMP-2 and bone marrow stromal cells (BMSCs) were used to repair critical-sized bone defects in a rat model. The combination significantly enhanced new bone formation compared to control groups. Alginate scaffolds loaded with TGF-β have also been used to promote chondrocyte proliferation and extracellular matrix production. These scaffolds are suitable for treating articular cartilage injuries and osteoarthritis. Alginate hydrogels loaded with FGF have been shown to accelerate wound healing by enhancing fibroblast proliferation and angiogenesis. These scaffolds can be used for treating chronic wounds and burns. In a clinical trial, alginate dressings loaded with FGF were applied to chronic diabetic foot ulcers. The FGF-loaded dressings accelerated wound healing, reduced ulcer size, and improved tissue granulation compared to standard care.

Synthetic Scaffolds

Polylactic Acid (PLA)

Polylactic Acid (PLA) is a biodegradable and biocompatible polymer widely used in the field of tissue engineering and regenerative medicine. Its favorable properties make it an excellent candidate for scaffolds designed to deliver growth factors, which are crucial for tissue regeneration and repair. It is derived from renewable resources such as corn starch or sugarcane, making it an environmentally friendly material. It is a linear aliphatic polyester that can be processed into various forms, including fibers, films, and porous scaffolds. PLA is well-tolerated by the human body, reducing the risk of adverse immune reactions. It degrades into lactic acid, a naturally occurring metabolite, which is then absorbed and eliminated by the body. It has good mechanical properties, making it suitable for load-bearing applications.

Fabrication techniques of polylactic acid (PLA) scaffolds

Several techniques can be employed to fabricate PLA scaffolds. The solvent casting and particulate leaching involves dissolving PLA in a solvent and then casting it into a mold. Particulate leaching, often using salt particles, creates a porous structure. After the solvent evaporates, the scaffold is immersed in water to dissolve the salt, leaving behind a porous PLA scaffold. Electrospinning is a popular method used for fabrication of scaffolds. It produces nanofibrous scaffolds by applying a high voltage to a PLA solution, creating fine fibers that are collected on a grounded surface. This technique is particularly useful for creating scaffolds that mimic the extracellular matrix, promoting cell attachment and proliferation. 3D printing is extensively used method for scaffold preparation at present. It allows for precise control over the scaffold’s architecture. PLA can be printed layer by layer to create complex structures tailored to specific tissue engineering needs. This technique is highly versatile and can produce scaffolds with intricate designs and controlled porosity. One more method is lyophilization (freeze-drying). In this process, a PLA solution is frozen and then subjected to sublimation under vacuum. This removes the solvent, leaving behind a highly porous scaffold. Lyophilization is advantageous for creating scaffolds with controlled pore sizes and high porosity.

Application of polylactic acid (PLA) scaffolds

PLA scaffolds have shown great promise in bone tissue engineering. They can be combined with osteogenic growth factors such as bone morphogenetic proteins (BMPs) to enhance bone formation. Studies have demonstrated that PLA scaffolds loaded with BMP-2 significantly improve bone regeneration in critical-sized defects. The incorporation of growth factors like transforming growth factor-beta (TGF-β) into PLA scaffolds has been shown to promote the regeneration of the periodontal ligament, improving the quality of regenerated tissue. PLA scaffolds loaded with growth factors such as platelet-derived growth factor (PDGF) have been used to promote cementum regeneration, which is essential for the attachment of the periodontal ligament to the tooth root.

One of the main limitations of PLA scaffolds is their relatively weak mechanical strength. This can be a drawback in load-bearing applications such as bone regeneration. Researchers are exploring various strategies to enhance the mechanical properties of PLA scaffolds, including the incorporation of reinforcing materials like hydroxyapatite and carbon nanotubes. The production of PLA scaffolds with consistent quality and properties is crucial for their clinical translation. Standardization of the fabrication process and scalability of production are important factors that need to be addressed.

Polyglycolic Acid (PGA)

Polyglycolic Acid (PGA) is a synthetic, biodegradable polymer that has gained significant attention in the field of tissue engineering and regenerative medicine. Its unique properties make it an excellent candidate for scaffolds designed to deliver growth factors, which are crucial for tissue regeneration and repair. is a member of the aliphatic polyester family, constituted by glycolic acid (hydroxyacetic acid) units linked through ester bonds. It is known for its high tensile strength and remarkable hydrolytic stability. PGA is synthesized through ring-opening polymerization of glycolide or direct polycondensation of glycolic acid. PGA is well-tolerated by the human body, reducing the risk of adverse immune reactions. It degrades into glycolic acid, which is then metabolized and eliminated by the body. PGA has excellent mechanical properties, making it suitable for load-bearing applications.

Fabrication techniques of polyglycolic acid (PGA) scaffolds

As discussed in PLA fabrication techniques, essentially same techniques are used to fabricate PGA scaffolds.

Application of polyglycolic acid (PGA) scaffolds

The PGA scaffolds have been used for bone regeneration successfully.  Studies have demonstrated that PGA scaffolds loaded with BMP-2 significantly improve bone regeneration in critical-sized defects. Along with this, incorporation of growth factors like transforming growth factor-beta (TGF-β) into PGA scaffolds has been shown to promote chondrogenesis and improve the quality of regenerated cartilage. PGA scaffolds loaded with PDGF have been shown to enhance the proliferation and differentiation of periodontal ligament cells, promoting the formation of new periodontal ligament fibers. In a study, PGA scaffolds encapsulating PDGF and periodontal ligament stem cells (PDLSCs) were used to repair periodontal defects in a dog model. The combination significantly enhanced new periodontal ligament formation compared to control groups. PGA scaffolds loaded with BMP-2 have been used to promote osteoblast differentiation and new bone formation in periodontal defects. In a study, PGA scaffolds encapsulating BMP-2 were used to treat alveolar bone defects in a rabbit model. The BMP-2-loaded scaffolds significantly enhanced new bone formation and mineralization compared to control scaffolds.

Polycaprolactone (PCL)

Polycaprolactone (PCL) is a synthetic, biodegradable polymer. It is a semi-crystalline polymer with a low melting point (around 60°C) and a glass transition temperature of about -60°C. It is synthesized through the ring-opening polymerization of ε-caprolactone. PCL is known for its excellent biocompatibility, biodegradability, and mechanical properties. It is well-tolerated by the human body, reducing the risk of adverse immune reactions. It degrades into non-toxic byproducts, primarily through hydrolysis of its ester linkages. PCL has good mechanical properties, making it suitable for load-bearing applications. Advantages of using PCL scaffolds for growth factor delivery:

Controlled Release: PCL can be engineered to provide a sustained and controlled release of growth factors, ensuring a prolonged therapeutic effect.

Enhanced Stability: Encapsulation of growth factors within PCL scaffolds can protect them from degradation, enhancing their stability and bioactivity.

Targeted Delivery: PCL scaffolds can be designed to deliver growth factors specifically to the site of injury or defect, improving the efficiency of tissue regeneration.

Fabrication techniques of polycaprolactone (PCL) scaffolds

The polycaprolactone (PCL) scaffolds are prepared by various techniques including electrospinning, solvent casting and particulate leaching, 3D printing and lyophilization (freeze-drying). These techniques have been discussed in the previous sections.

Application of Polycaprolactone (PCL) scaffolds

PCL scaffolds have been extensively studied for their applications in various fields of tissue engineering. PCL scaffolds have shown significant potential in periodontal regeneration, which involves restoring the structure and function of the periodontium, including the gingiva, periodontal ligament, cementum, and alveolar bone. In a study, PCL scaffolds encapsulating PDGF and periodontal ligament stem cells (PDLSCs) were used to repair periodontal defects in a dog model. The combination significantly enhanced new periodontal ligament formation compared to control groups. In another study, PCL membranes encapsulating TGF-β were used in a GTR approach to treat periodontal defects in a dog model. The TGF-β-loaded membranes promoted new bone, periodontal ligament, and cementum formation compared to control membranes.

Challenges in using scaffolds

Growth Factor Stability: Ensuring the stability and bioactivity of growth factors during scaffold fabrication and storage is challenging.

Controlled Release: Achieving a controlled and sustained release of growth factors to match the tissue regeneration timeline is critical.

Scaffold Degradation: Balancing the degradation rate of PCL scaffolds with tissue regeneration is essential to maintain mechanical support and integration.

Immune Response: Minimizing immune responses to growth factors and scaffold materials is necessary to avoid inflammation and tissue rejection.

Future Directions

Advanced Fabrication Techniques: Developing advanced fabrication techniques like 3D printing and electrospinning can create PCL scaffolds with precise architecture and enhanced functionality.

Smart Scaffolds: Designing smart scaffolds that respond to environmental cues (e.g., pH, temperature) to control growth factor release and scaffold degradation.

Multi-Growth Factor Delivery: Incorporating multiple growth factors in a controlled manner to mimic the natural healing process and enhance tissue regeneration.

Clinical Translation: Conducting more preclinical and clinical studies to validate the safety and efficacy of PCL scaffolds loaded with growth factors for periodontal regeneration.

Conclusion

At present, there is a lot of research going on in the field of scaffold designing. Each fabrication technique offers unique advantages and can be selected based on the specific requirements of the tissue engineering application. The choice of method depends on factors such as the desired scaffold architecture, mechanical properties, and the type of tissue being targeted. In near future we shall see a lot of advancements in this field for sure

References

References are available in the hardcopy of the website “Periobasics:  A Textbook of Periodontics and Implantology”.