Biopolymers and Bioprinting for Dental Tissue Engineering: Comparison
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Please note this is a comparison between Version 2 by Sirius Huang and Version 1 by Hyeongjin Lee.

Dental tissues are composed of multiple tissues with complex organization, such as dentin, gingiva, periodontal ligament, and alveolar bone. These tissues have different mechanical and biological properties that are essential for their functions. Therefore, dental diseases and injuries pose significant challenges for restorative dentistry, as they require innovative strategies to regenerate damaged or missing dental tissues. Biomimetic bioconstructs that can effectively integrate with native tissues and restore their functionalities are desirable for dental tissue regeneration.

  • polymers
  • 3D printing
  • dental tissues
  • tissue engineering

1. Introduction

The field of tissue engineering has witnessed remarkable progress in recent years, offering new avenues for regenerative medicine [1,2][1][2]. In dentistry, tissue engineering holds immense potential for the restoration and regeneration of dental tissues, revolutionizing conventional approaches to dental treatments [3,4,5,6][3][4][5][6]. Dental tissue engineering aims to overcome the limitations associated with traditional restorative techniques by promoting the growth and regeneration of dental tissues, including dentin, cementum, and periodontal ligaments cultured on various scaffolds [7].
The regeneration of dental tissues poses significant challenges because of their complex hierarchical structures, functional requirements, and innate regenerative limitations [8]. Advancements in biomaterials, cell-based therapies, and tissue engineering strategies have provided innovative solutions to these challenges [9,10,11][9][10][11]. Dental tissue engineering endeavors to replicate the natural regenerative processes that occur during tooth development and repair. It involves the use of biocompatible scaffolds, growth factors, and stem cells to create an optimal microenvironment that supports cell adhesion, proliferation, and differentiation [12,13][12][13]. Scaffolds act as three-dimensional (3D) frameworks, providing structural support and guiding the growth and organization of cultured cells. By mimicking the composition of the extracellular matrix (ECM) and architecture of dental tissues, these scaffolds facilitate the formation of functional tissue constructs [14].
A variety of polymer-based biomaterials have recently been explored for scaffold fabrication in dental tissue engineering, including the biodegradable polymers poly(lactic-co-glycolic acid) (PLGA) [15[15][16],16], polycaprolactone (PCL) [17], and collagen-based materials [18]. These biomaterials have demonstrated favorable properties, including biocompatibility, tunable degradation rates, and mechanical integrity. Furthermore, the incorporation of bioactive molecules, including growth factors, peptides, and nanomaterials, into these scaffolds has shown promising results in enhancing cell behavior, tissue regeneration, and mineralization [19,20,21][19][20][21].
In recent years, the emergence of 3D bioprinting has revolutionized the fabrication of complex scaffolds for dental tissue engineering [22]. This additive manufacturing technique enables precise control over scaffold architecture, porosity, and spatial distribution of cells and bioactive factors. By employing computer-aided design software and a layer-by-layer deposition of biomaterials, 3D bioprinting allows for the creation of patient-specific scaffolds with tailored mechanical and biological properties [23]. Furthermore, the incorporation of multiple cell types within these constructs has shown potential for the regeneration of multi-tissue interfaces, including the dentin-pulp or periodontal ligament-bone complex [24].
The future of dental tissue engineering holds great promise. Advancements in biomaterials, scaffold design, stem cell research, and biofabrication techniques have enabled the development of efficient and effective strategies for dental tissue regeneration. The integration of innovative technologies, including gene editing, tissue-on-a-chip systems, and in vitro organogenesis, is likely to further augment the regenerative capabilities of dental tissue engineering approaches.

2. Bioprinting for Dental Tissue Engineering

Advances in tissue engineering have led to the emergence of 3D bioprinting as a promising technique for fabricating complex dental structures [40][25]. Three-dimensional bioprinting enables the precise spatial organization of cells, biomaterials, and growth factors, allowing the creation of patient-specific and functional dental tissues. This section explores various 3D bioprinting technologies used in dental tissue engineering.
Inkjet-based bioprinting is a non-contact printing technique that utilizes thermal, piezoelectric, or microvalve processes to dispense droplets of dilute solutions. It operates similarly to traditional inkjet printing, but uses bioinks containing cells and biomaterials [41][26]. This technology allows for a high spatial resolution, ranging from 50 to 300 μm [42][27], but the presence of cell aggregation within the bioink can affect droplet formation and trajectory, leading to a decrease in print quality [43][28]. Rider et al. utilized a reactive inkjet printing method to obtain high-resolution nanosized hydroxyapatite-incorporated silk fibroin membranes [44][29]. However, the use of low-concentration solutions can limit the construction of 3D structures for dental tissue engineering.
Laser-assisted bioprinting utilizes the power of laser beams to achieve the meticulous deposition of bioink substances onto a substrate. Stereolithography apparatus (SLA) is an example of laser-assisted 3D printing. Son’s team (2021) used SLA 3D printing to fabricate interim crowns for dental implantation [45][30]. Through localized pressure generated by the laser, the bio-ink was propelled in the form of droplets onto a designated target area. This cutting-edge technology offers the ability to achieve high-resolution printing, surpassing the threshold of 20 µm [48][31].
Such precision allows for the accurate placement of cells and biomaterials, thereby promoting intricate biological constructs. To ensure a successful process, the precursor material should be a hydrogel with a viscosity within a moderate range [49][32]. Furthermore, this method has demonstrated the potential to enable precise multicell positioning and facilitate intricate cellular arrangements [50][33]. However, the complexity involved in selecting the optimal printing conditions, such as gelation time and laser fluence, poses challenges, potentially resulting in a compromised cell viability. The requirement for expensive apparatus further complicates the widespread adoption of this technique [51][34].
Operating on a principle such as the SLA, the direct light processing (DLP) fabrication method utilizes localized light to solidify a photocross-linkable liquid to obtain 3D structures. DLP fabrication methods are regularly used to convert 3D models into 3D structures for dental implants. Furthermore, DLP can polymerize each layer of resin much more rapidly compared to SLA, which is a preferable process [46][35].
Extrusion bioprinting is driven by a piston, screw, or pneumatic pressure mechanisms such that highly viscous bioinks can be printed through micronozzles [52][36]. Conventional extrusion-based printing systems are regularly used to fabricate scaffolds for guided dental regeneration. For instance, researchers illustrated the incorporation of salt microparticles in biopolymers (PCL), as recently investigated in 2023 [47][37]. The subsequent leaching of salt particles after fabrication allows for the formation of micro/macroporous scaffolds for dental tissue engineering. This technique is versatile and compatible with a wide range of biomaterials including hydrogels and biopolymers. In particular, for cell-laden structures, this technique enables the printing of very high cell densities with a fast rate of fabrication [53,54][38][39]. However, there are potential cell apoptotic effects induced during and after printing owing to the pressure drop associated with extrusion through a micronozzle [51,55][34][40].
Bioprinting can be used in the regeneration of intricate dental structures with the precise spatial organization of diverse tissues. This facilitates the regeneration of vascularized pulp-like tissue and the formation of mineralized tissue within stem cell constructs by employing DPSCs and stem cells from the apical papilla [56][41]. Bioprinting also plays a role in the regeneration of integrated cementum on the surface of the roots of a human tooth. This was achieved by utilizing growth-factor-releasing scaffolds containing PDLSCs, which were incorporated into 3D-printed PCL scaffolds. The research team (2017) has further enhanced the scaffold with PLGA microspheres encapsulating connective tissue growth factor, BMP-2, or BMP-7 [57][42].
The application of 3D bioprinting in dental tissue engineering offers great potential for personalized dental treatment and the rapid prototyping of dental structures. Although some challenges remain, including the scalability of the technique, optimization of bioink formulations, and long-term functionality of printed tissues, 3D bioprinting is a rapidly evolving field that holds extensive promise.

3. Polymeric Materials and Their Printed Scaffolds for Dental Tissue Engineering

The choice of scaffold material is critical for dental tissue engineering, as it affects the cellular activities and the tissue regeneration. Both biological and mechanical properties are important factors for dental tissue regeneration. The biological properties influence the bioactivities of the cells, while the mechanical properties ensure structural stability under dynamic loadings. Generally, natural polymers can provide suitable biochemical cues, thus enhancing the bioactivities of cells, whereas synthetic polymers can provide adequate mechanical properties. This section discusses various studies that use natural, synthetic, and hybrid polymers for dental tissue engineering.

3.1. Natural Polymers

Natural scaffold materials derived from biological sources offer several advantages, including biocompatibility, biodegradability, and the ability to mimic the composition and structure of native ECM. Commonly used natural scaffold materials in dental tissue engineering include collagen, gelatin, chitosan, hyaluronic acid, fibrin, and decellularized ECM (dECM).
Collagen, derived from several sources, including bovine, porcine, or human tissues, is a versatile scaffold material that supports cell adhesion and promotes the regeneration of dental tissues, including dentin, periodontal ligament, and gingiva. This is due to its anatomically similar structure and chemical properties to the predominant structural proteins existing in the ECM of dental tissues [58,59][43][44].
Although collagen has high tensile strength and can be used in fibrous forms and load-bearing applications, it lacks sufficient mechanical strength for pulp regeneration [60][45]. Crosslinking collagen with glutaraldehyde or genipin enhances its mechanical properties. The combination of a collagen scaffold, DPSCs, and DMP1 improved the formation of ECM in pulp tissue [61][46], while Pandya et al. incorporated erythropoietin (EPO), a glycoprotein hormone that stimulates red blood cell production, into collagen scaffolds in 2021 [62][47]. They compared this with a commercially available BioOss inorganic bovine bone xenograft to investigate its potential for alveolar ridge augmentation. The incorporation of EPO into the collagen scaffold resulted in a two-fold increase in blood vessel formation and improvements in the deposition of ECM. Recently, Chang’s research team (2023) investigated a novel injectable cell-laden hydrogel consisting of collagen and riboflavin for the treatment of periodontal defects. They implemented a dental light-emitting diode crosslinking method to reinforce the injected hydrogel [63][48].
Gelatin, a derivative of collagen, possesses a porous structure that allows for cellular infiltration, making it suitable for regenerating dentin, periodontal tissues, and oral mucosa. Gelatin has been extensively used in both preclinical and clinical settings [64,65][49][50]. Gelatin offers distinct advantages over collagen, including reduced immune responses, more controllable physical properties, and a lower risk of unpredictable pathogen transmission. Importantly, gelatin exhibits favorable engineering properties, including ease of fabrication and the ability to control its mechanical properties during crosslinking [66,67][51][52]. Consequently, gelatin materials are increasingly considered in the application of advanced tissue engineering, particularly for manufacturing highly porous scaffolds that facilitate tissue repair and regeneration [68][53].
Alginate, a polysaccharide produced by a wide variety of brown seaweeds, is an effective cell carrier, scaffold, and delivery system for growth factors and bioactive materials used in regenerative endodontics. For example, Dobie et al. (2002) reported that TGF1 can upregulate the matrix secretion of dentin-pulp ECM, while the alginate polymeric matrix can serve as a delivery platform for various growth factors and bioactive components [69][54]. Zhang et al. (2020) fabricated alginate/laponite hydrogel microspheres that encapsulated DPSCs and vascular endothelial growth factor (VEGF) to develop injectable cell-laden microspheres for endodontic regeneration. The injection of these biomicrospheres induced vascularization after the subcutaneous implantation of tooth slices in nude mice. Additionally, the upregulation of odontogenic-related genes was observed [70][55].
Hyaluronic acid, a glycosaminoglycan found in the ECM presents hydration and lubrication properties. It has been widely used in scaffolds for cell adhesion and has shown promising results for regenerating periodontal tissues and oral mucosa. Park et al. (2003) has demonstrated that hyaluronic acid hydrogel modified with arginine–glycine–aspartic acid (RGD) has exhibited significant potential in promoting cell attachment and proliferation [71][56]. This highlights the favorable effects of RGD-modified hyaluronic acid hydrogels on cellular processes. Furthermore, the injectable nature of hyaluronic acid hydrogel allows it to penetrate narrow canals, making it highly suitable for applications in endodontics and pulp regeneration.
Fibrin, a structural component of blood clots, serves as a 3D framework that supports cell attachment, migration, and tissue formation. Compared to other natural polymers such as collagen, fibrin exhibits superior properties in terms of cell adhesion, biocompatibility, and immune response. However, it also possesses disadvantages, including high shrinkage rates, rapid degradation, and low mechanical strength [72][57]. To address these limitations, composite scaffolds comprising fibrin and biocompatible reinforcements, including hyaluronic acid, calcium phosphate, and polyurethane, significantly enhanced the mechanical properties of fibrin [73][58].
The use of fibrin as a biomaterial has several advantages. First, fibrinogen within its structure undergoes TGF-β transformation, leading to collagen formation [74][59]. Fibrin also provides a suitable environment for angiogenesis, exhibits the potential to control the release of proangiogenic growth factors [75][60], and offers injectability and the capacity to construct 3D structures. Because of these advantages, Zhang et al. (2020) utilized fibrin-based hydrogels as a delivery system for extracellular vesicles extracted from mesenchymal stromal cells to investigate the angiogenic effects in dental pulp regeneration [76][61] and observed favorable ECM deposition and a high angiogenic response.
dECM scaffolds are prepared by removing cellular components from natural tissues while preserving the structure and composition of the ECM. These scaffolds provide a biomimetic environment for tissue regeneration, and dental tissue-specific dECM scaffolds have been used to guide the regeneration of dental tissues. They provide structural support, promote cellular activity, and guide the regeneration of specific dental tissues, contributing to advancements in dental regenerative therapies.

3.2. Synthetic Polymers

Synthetic polymers, including PLGA, PLA, and PCL, are widely used for dental tissue engineering. These polymers exhibit tailorable mechanical properties, degradation rates, and biocompatibility. They can be fabricated in various forms, including films, fibers, and porous scaffolds, using multiple techniques, including electrospinning, solvent casting, and 3D printing. Synthetic polymer scaffolds have been utilized for the regeneration of several dental tissues, including dentin, periodontal ligament, and alveolar bone.
In 2021, alveolar bone regeneration was investigated through the incorporation of β-tricalcium phosphate (β-TCP) and platelet-rich plasma (PRP) into a PCL scaffold. Subsequently, bone marrow stem cells (bMSCs) were seeded onto a scaffold and implanted into mandibular bony defects in miniature pigs [84][62]. The incorporation of bMSCs and PRP into the PCL-TCP scaffold significantly increased the bone–implant contact ratio, the height of newly formed bone, and new bone formation, compared with the conventional PCL-TCP scaffold. Similarly, Li et al. demonstrated that the incorporation of PRP into PCL scaffolds can induce substantial osteogenic activity in DPSCs, in an experiment conducted in 2017 [93][63]. Tatullo et al. (2019) evaluated the osteo/odontogenic properties of a PLA scaffold modified with dicalcium phosphate dihydrate and/or hydraulic calcium silicate [85][64]. Human periapical cyst-derived mesenchymal stem cells (MSCs) cultured on the scaffold exhibited a 2.5-fold upregulation of DMP-1 gene expression compared to those cultured on pristine scaffold. However, the authors noted that the acidic degradation of PLA could hinder regeneration efficacy. Park et al. (2017) utilized an extrusion-based 3D printing system to fabricate PCL/β-TCP scaffolds [94][65]. Owing to the osteoinductive properties of β-TCP, significantly superior osteogenic properties were observed in MSCs lines than in pristine PCL. In 2018, implantation of the scaffold into alveolar defects in rats resulted in the improved formation of new bone [95][66].
Through the utilization of highly porous polymeric scaffolds, researchers have successfully regenerated dental tissues, including dentin and pulp tissues, in preclinical trials [98,99][67][68]. Additionally, the incorporation of drug-loaded polymeric scaffolds holds great promise in controlling the rates of release of gene vectors, proteins, and growth factors, and in creating spatiotemporal microenvironments that support tissue growth and regulate cell activities, including differentiation, proliferation, and migration [100][69]. For instance, in 2018, biodentine (an FDA-approved drug used for dentin repair) was incorporated with PCL to develop bioactive scaffolds for dental/bone regeneration [96][70]. The incorporation of biodentine significantly improves the cellular proliferation and osteogenic activities of human dental pulp cells. However, a key challenge in dental tissue engineering is the development of strategic approaches to optimize tissue formation using complex micron-scale geometries and biodegradable polymeric materials [101,102][71][72]. Thus, there is significant demand for customized or flexible methodologies that employ biopolymers to address complex geometries or non-standardized and unpredictable defects in the field of periodontal tissue engineering.
Synthetic hydrogels, including poly(ethylene glycol) (PEG) and poly(vinyl alcohol), consist of hydrophilic polymers that form 3D networks that can retain significant amounts of water and undergo mild biodegradation. These hydrogels create a hydrated environment similar to the native tissues, enabling cell encapsulation, nutrient diffusion, and growth factor distribution. Through chemical modifications, synthetic hydrogels can incorporate bioactive signals to enhance cell adhesion, proliferation, and differentiation. In dental tissue engineering, these hydrogels have found applications in regeneration of dental pulp and periodontal tissue engineering. Notably, RGD peptide-modified PEG hydrogels have demonstrated elevated cell adhesion and proliferation, while modifications in the molecular weight and photocross-linking of PEG have improved its mechanical properties without detrimental effects on encapsulated cells [103,104,105][73][74][75].
One noteworthy example involves fibrin-loaded PEG hydrogels, which exhibit significant potential as scaffolds for the growth and proliferation of DPSCs and PDLSCs. These hydrogels combined the mechanical support and angiogenic properties of PEG with the benefits of fibrin hydrogels [105][75].

3.3. Polymer-Based Hybrid or Composite Materials

Composite scaffolds combine the advantages of multiple materials to achieve synergistic effects. In dental tissue engineering, composite scaffolds often comprise a combination of natural and synthetic materials. For example, a composite scaffold can be formed by incorporating natural polymers, including collagen or chitosan, in synthetic polymers, including PLGA or PCL. These hybrid scaffolds offer improved mechanical properties, enhanced biocompatibility, and controlled degradation. Composite scaffolds have been utilized for the regeneration of several dental tissues, including enamel, dentin, periodontal ligaments, and bone. Ducret et al. (2019) incorporated chitosan into a fibrin polymeric matrix to prevent the growth of endodontic bacteria and promote the regeneration of dental pulp tissue [86][76]. The authors stated that chitosan-incorporated fibrin polymeric hydrogels could be injected into the endodontic space for antibacterial effects. Furthermore, chitosan–gelatin composite hydrogels have been investigated for alveolar bone regeneration. The implantation of bone MSCs seeded with chitosan–gelatin scaffolds into tooth sockets in rats resulted in significant improvements in new bone formation and neovascularization [91][77]. Yu et al. investigated the effects of 3D-printed alginate/gelatin hybrid scaffolds using human dental pulp cells. Owing to the improved printability of the hybrid bioink, complex 3D geometries were successfully fabricated, while the cellular proliferation and osteo/odontogenic activities of human dental pulp cells were elevated.
Bioactive ceramics, including hydroxyapatite, tricalcium phosphate, and bioglass have been widely used in endodontic applications because of their similarity to the mineral components of natural dental tissues and their high bioactivity [106][78]. Hybrid scaffolds are created by combining bioactive ceramics with synthetic polymers or natural scaffolds. The combination of bioactive ceramics with polymers enhances the mechanical properties and provides a favorable environment for cell attachment, proliferation, and mineralization [107,108][79][80]. Bioactive ceramic-based scaffolds have been employed for the regeneration of dental tissues, including enamel, dentin, and alveolar bone. In 2012, improved characteristics were demonstrated in an alginate scaffold incorporated with a nano-bioglass ceramic [109][81]. The modified scaffold exhibited enhanced attachment, growth, and alkaline phosphatase (ALP) activity in hPDLF. For example, the incorporation of calcium silicate and calcium sulfate into PCL scaffolds greatly improves calcium deposition in human dental pulp cells, as indicated by the more intense Alizarin Red S (ARS) expression, as recently demonstrated in 2022 [110][82]. Similarly, Choi et al. (2022) incorporated calcium silicate cement into photocross-linkable methacrylated gelatin (GelMA) to investigate the cellular activity of human DPSCs [97][83]. As a result, Dentin Sialophosphoprotein (DSPP) and DMP-1 genes were significantly upregulated, indicating the efficient odontogenic activity of the cells. Nejad et al. (2012) fabricated a 3D PCL/calcium sodium phosphoslicate Bioglass (BG) composite and PCL/hydroxyapatite (HA) scaffold to investigate dentin and pulp tissues [111][84]. The incorporation of BG greatly upregulated osteo/odontogenic-related genes, including DSPP, osteocalcin (OCN), and DMP-1, compared to PCL/HA, indicating the significant potential of BG in alveolar bone regeneration.
Nanocomposite scaffolds incorporate nanoscale materials, including nanoparticles or nanofibers, into a scaffold matrix. These nanomaterials can be either natural (e.g., nanocellulose) or synthetic (e.g., carbon nanotubes). The integration of nanomaterials imparts unique properties to scaffolds, including enhanced mechanical strength, improved bioactivity, and controlled drug release. Nanocomposite scaffolds have shown promise in dental tissue engineering for applications in dentin regeneration, enamel remineralization, and drug delivery systems. Jiang et al. (2015) demonstrated the incorporation of electrospun aligned PCL-PEG nanofibers into a porous chitosan scaffold to evaluate the regenerative efficacy of periodontal ligaments. Two months after implantation into the periodontal defect in rats, the significant formation of aligned periodontal ligaments was observed, owing to the appropriate contact guidance for the cells [92][85]. 

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