The Osteogenic Role of Biomaterials Combined with Human-Derived Dental Stem Cells in Bone Tissue Regeneration

Abstract

The use of stem cells in regenerative medicine had great potential for clinical applications. However, cell delivery strategies have critical importance in stimulating the differentiation of stem cells and enhancing their potential to regenerate damaged tissues. Different strategies have been used to investigate the osteogenic potential of dental stem cells in conjunction with biomaterials through in vitro and in vivo studies. Osteogenesis has a broad implication in regenerative medicine, particularly for maxillofacial defects. This review summarizes some of the most recent developments in the field of tissue engineering using dental stem cells.

Introduction

The role of stem cells in regenerative medicine has been widely investigated in the field of tissue engineering, which made a significant difference in the last decade [1]. Tissue engineering techniques are based on the reproduction of tissues by using different combinations of cells, biomaterials, scaffolds, supplements, and growth factors [2].

Bone is a dynamic, highly vascularized connective tissue that constitute the human skeleton. It is classified into two different types based on its architectural arrangement: compact cortical bone, which makes up around 80% of the total body skeleton; and spongy trabecular bone, which constitutes the remaining 20%. Both types of bone tissues are composed of a matrix containing inorganic calcium salts [3]. Bone tissue primarily functions to provide the human body with structural support. Additionally, bone is essential for mineral regulation and storage, in reflection of body demands [2]. Bone tissue is comprised of three main cell types: osteoblasts, osteocytes, and osteoclasts. Osteoblasts are responsible for matrix mineralization [4], while osteocytes are mainly involved in matrix calcification and blood calcium regulation [5] . Bone resorption is the function of osteoclasts [6].

Bone diseases are highly prevalent worldwide and are a major cause of disabilities for many patients. These diseases have become very challenging and demanding for orthopedic surgeons [7]. Joint pain and knee replacement in elderly people are major cause of physical disability. Moreover, 1.6 million people suffer from hip fractures by accidents [8]. These issues would result in high economic and social burdens on communities and health care systems [2, 8].

Bone tissue regeneration can be used to replace the destructed bone tissues, specifically in the field of dentistry. The regeneration procedure focused on the grafting of the bone defect with different combinations of biomaterials, either as scaffolds or membranes, according to the classification of the bone defect [3].

Current therapeutic strategies include bone replacement therapy, which mainly focuses on bone substitution by autografts, allografts, or demineralized bone matrix. Unfortunately, aforementioned procedures are invasive and might carry the risk of post-operative infections. In addition, they have no standardized amounts of materials to be used in the grafting site, and can cause tissue death and immune stimulation [2, 9].

Periodontal diseases are one of the most critical diseases in the field of dentistry as it has a negative impact on patients’ quality of life. Periodontitis, a common and highly distributed disease, is characterized by the damage of tooth-supporting tissues; root cement, periodontal ligaments and the alveolar bone in the oral maxillofacial region [10]. Periodontitis is traditionally treated by restoring periodontal supporting tissues either by surgical or non-surgical interventions [11, 12]. However, the loss of bone tissue is one of the most important consequences of progressive disease. Various technical procedures have been used to repair and regenerate damaged tissues, such as bone replacement, tissue regeneration, growth factors, and biomaterials. However, the process is very challenging as it requires the regeneration of specific functional tissues and restoring their interaction with the surrounding tissues in the environment [13, 14].

In order to tackle previously mentioned issues, cell therapeutic technologies have been developed thereby applying stem cells in bone regeneration. Stem cells have been derived from different sources such as bone marrow, adipose tissue, dental tissues, umbilical cord, embryonic stem cells, and Induced Pluripotent Stem Cells (iPSCs) [15–17]. Stem cells are known to have high proliferative and differentiation potentials; hence, they are considered a new promising and attractive source in cell therapeutic technologies [15–17]. Different studies have investigated the role of stem cells in bone regeneration, as they have a high osteogenic potential [15].

Dental stem cells

Dental-derived stem cells are distinguished from other types of stem cells by their distinctive qualities, which make them a promising source in the field of regenerative medicine. Dental stem cells (DSCs) are simple to isolate and have a high rate of proliferation. There are no ethical requirements because DSCs are retrieved via a non-invasive method and are therefore regarded as medical waste.

Dental-derived stem cells have shown a great potential to differentiate into bone marrow mesenchymal stem cells (BMMSCs), and embryonic stem cells. Dental tissues such as; pulp, ligament, follicle and dental papilla have been identified as feasible sources of ectomesenchymal undifferentiated stem cells for bone regeneration [18].

Dental pulp stem cells

Dental pulp stem cells (DPSCs) were the first DSCs to be isolated in 2000 by Gronthos et al. and can be harvested from adult tooth pulp tissues through enzyme treatment. DPSCs possess the same MSCs features as they express same cell surface markers and possess a similar differentiation potential to BMMSCs. Dental pulp develops from the dental papilla and originates from ectomesenchyme. In comparison to MSCs derived from BMMSCs, DPSCs have higher ability to form colonies (CFU) and higher growth rates, in addition to their superior osteogenic differentiation potential and mineralization ability [19].

Stem cells from human exfoliated deciduous teeth

Derived from primary deciduous teeth from the remnant pulp tissue, stem cells from human exfoliated deciduous teeth (SHED) possess high proliferation potential. Moreover, SHED are positive for all MSC surface markers, and negative for hematopoietic stem cell markers. They have capability to directly differentiate into bone to be transplanted in animal models without differentiation into osteoblasts in vivo [20, 21].

Stem cells from apical papilla

Stem cells from apical papilla (SCAP) are derived from the soft tissue (apical papilla) that is found at the apex of a developing permanent impacted teeth, in which it plays a role in the formation and development of the dental pulp tissue. However, this tissue is separated from the pulp, less vascularized, and processes less cellular components. SCAP have higher mineralization potential when compared to DPSCs [22].

Dental follicle stem cells DFSCs

Dental follicle stem cells (DFSCs) are extracted at early stage of crown formation from loose connective tissue that surrounds the enamel and dental papilla of the teeth germ, prior to its eruption. Stem cells are harvested from follicles of impacted human third molars. During teeth development, dental follicle has essential role in the bone development and resorption of the periodontium [23].

Periodontal ligament stem cells

Periodontal ligament stem cells (PDLSCs) are isolated from periodontal ligament tissue from human third molars. They possess high differentiation potential into adipocytes, cementoblast, and collagen forming cells, which indicated a great contribution for periodontal tissue repair by generating a cementum/PDL structure in vivo [24]. Subsequent investigations demonstrated that PDLSCs promote osteoblastic differentiation and tissue mineralization in vivo under osteogenic inductive conditions, and could regenerate bone in vivo [25, 26]. Immunomodulatory properties of PDLSCs were also reported revealing PDLSCs as candidates for allogeneic stem cell-based therapies [27].

Stem cell delivery as a therapeutic approach

Certain clinical conditions lead to a complete loss of bone structure and periodontal tissues, which cannot be reversed and treated by normal mechanisms. Hence, the direct delivery and implantation of stem cells will be the right choice in order to regenerate and repair the damaged tissues [28].

Various studies have demonstrated the effect of using different types of stem cells for bone regeneration capabilities. However, not only the cell type has a significant impact on the regeneration process, but also cell-mediated delivery method plays a crucial role in the healing process of damaged bone tissues [12, 29].

Scaffolds and biomaterials

Cells can be delivered by using different strategies; either by injecting the cells alone or combined to a biomaterial. The efficiency and the success rate of these strategies largely depends on the cell type and the delivery strategy that has been used [30, 31]. While direct implantation of cells into wound site would be very easy and efficient to regenerate the damaged tissue, the use of biomaterials will generate 3D bio-engineered culture, that is biocompatible with damaged tissue [32].

The interaction between cells and their microenvironment is very crucial to maintain the normal behavior of cells. Thus, the 2D cultures limited these interactions. Since, cells are grown in a plate, leading to a great loss in these interactions such as; cell–cell and cell–matrix interactions resulting in a decline of the cells’ growth rate [33]. Therefore, finding a new model that can allow cells to interact as in their natural environment, enhancing their normal behaviors [34]. The appropriate selection of scaffolding materials, stem cell types, and bioactive substances is of utmost importance for the success of tissue engineering, to repair, regenerate, and enhance the function of damaged tissues. As the selection of the suitable scaffold will be reflected on the cell behaviors and processes such as; growth, migration, adhesion and differentiation, accelerating the regeneration and repair potencies of damages organs and tissues [35].

Scaffolds can be used in two different modes: absorbable, or injectable [36, 37]. Calcium phosphate cement (CPC) is an example of injectable scaffold which is used as a paste on defected bone sites [38]. Moreover, scaffolds can be loaded with different types of bone cells to increase the efficiency of scaffold transplantation procedures. Though, this CPC (scaffold-cells) combination strategy faced different issues regarding the efficacy of loading CPC scaffolds with cells; as the growth rate, penetration, and differentiation potential were very low [39].

One of the key ingredients in creating scaffolds is biomaterials. Tissue bioengineering offers potential outcomes for repairing damaged tissues by mixing scaffolds with various stem cells [40]. An ideal biomaterial for bonding with stem cells has the following properties: biocompatibility in the initial stage, the capacity to exchange gases and nutrients, the capacity to protect cells from immune system invasion and external stress, porosity, pore size, connectivity, stability, electrical conductivity and the capacity to properly create crosstalk between stem cells and neighboring cells [41].

Different procedures and methods have been used to fabricate fibers that have been known as the primary ingredient of 3D scaffolds, these methods include electrospinning, microfluidic-based method., micropatterning and 3D bioprinting [42]. Electrospinning is one of the most popular scaffold fabrication methods for creating nano/micro-fibrous scaffolds. The idea of creating fibers using electrostatic forces is not new. Electrospinning is a relatively simple method for tissue engineering due to its simple process and requirements. Although the procedure appears to be straightforward, several parameters must be carefully specified to achieve good mechanical, morphological, and biological properties. These properties are influenced by three main parameters: process parameters, solution parameters, and environmental parameters. Despite the promise qualities of nanofibers, some restrictions include inadequate mechanical properties, hydrophobicity, inefficient pore-structure controllability, shrinking, and distortion. Therefore,, recent studies have been oriented toward combining more than one strategy together, to get over these drawbacks. [43, 44].

Microfluidics has been promoted as a trustworthy high-tech method for defining micro and nanoscales. The small size of the devices may provide ideal conditions for cell and tissue growth. To be more specific, the examination of the cell's activities in a controlled microenvironment is their primary advantage over other methodologies. Microfluidic devices and microfabrication processes can be used to create a wide range of artificial tissue architectures, such as the synthesis of fibers particles, and hydrogel, in which all these materials can be applied and used as scaffolds in tissue engineering. Moreover, it is possible to create a multi-component surface by selecting various solutions for various channels. Additionally, cells can be deposited simultaneously with the solution in this patterning method rather than being introduced after a protein pattern has been formed. Notably, obtaining surfaces with more than two adhesive components (and consequently more than two cell types to form the pattern) is also feasible using microcontact printing; however, this necessitates the use of multi-leveled stamps, which are very difficult to produce, or sequential patterning of adhesive components onto the same surface, which requires sophisticated manual or automated alignment for accurate superimposition of features reaching a desired level of accuracy [44, 45].

Micropatterning approaches involve controlling cellular attachment, shape and spreading as a function of the engineered spatial properties of the culture surface. While there are a variety of ways to accomplish this, extracellular matrix (ECM) proteins and polyaminoacids are required to mediate the attachment and spreading of anchorage-dependent cell types on an underlying substrate. The most employed strategies for micropatterning involve seeding cells on a surface that exhibit areas (patterns) of differential adhesiveness. However, this approach is very expensive, because of the optimization steps that must be performed before the preparation of the real samples [46, 47].

3D bioprinting, is a recently used technique based on using biological element such as cells to repair damaged tissues and organs [48], by using bio-ink that contains the biomaterial that is encapsulated with the biological component such as cells, to produce 3D biological structure that can be used as an implant in the tissue regeneration [49]. This technology has overcome the limitations of the previously used techniques for scaffold preparation, by stabilizing the potential of the derived cells to activate the osteogenesis pathways in stem cells, in which reflecting on their potential to repair damaged bone tissue [50]. Hence, this approach would bevery useful for screening cell-biomaterial interactions in 3D cultures, and stimulating functional tissue regeneration [51].

Different types of biomaterials have been used in the regeneration of bone. The most commonly used types are natural, synthetics, ceramics and composites. Natural polymers are classified according to their base into; polysaccharide-based polymers such as collagen or protein-based polymers such as chitosan. Collagen is the most commonly used polymer in the regeneration of bone. As different types of collagen with different physical and chemical properties have been used in bone regeneration. Collagen is one of the most frequently used polymer in bone regeneration; due to its biodegradability, flexibility, tensile strength and low immunogenicity. Additionally, Gelatin is a polymer that is resulted from the hydrolysis of collagen with a lower immunogenicity compared to collagen, and is also highly used in bone regeneration. [52, 53]. On the other hand, polysaccharide based natural polymers such as chitosan, that is highly used in bone regeneration as it’s known with its high osteo-conductivity, but it has some drawbacks related to stability and mechanical strength [54]. Synthetic polymers are polymers that are synthesized with specific chemical and physical properties, such as poly(l-lactic acid) (PLLA) and poly(D,L-lactic acid) (PDLA), in which these have a high tensile strength [55]. And Polycaprolactone (PCL), that’s characterized by its biocompatibility and low degradability [55]. Polylactic-co-glycolic acid (PLGA) is a copolymer of PLA and polyglycolic acid (PGA). PLGA is biodegradable, allows for adhesion, and shows good mechanical properties. Its degradation rate can be regulated by varying the percentage of the two polymers [56].

Ceramic scaffolds, are classified according to their reactivity; non-inert, semi-inert and relativity-inert scaffolds. Ceramics have a great application in dental field, as they possess unique features that make them a good candidate in dentistry. As these scaffolds are biocompatible, tough, vulnerable to stress. An example of these ceramics; calcium phosphate which is a non-inert ceramic, it’s one of the mostly used scaffolds due to its capability to enhance the formation of inorganic bone matrix, though it has low tensile strength [57].

Hydroxyapatite (HA), which is a semi inert ceramic, that’s highly used in bone regeneration very similar to the inorganic matrix of the bone, but it has low mechanical resistance [54]. B-tricalcium-phosphate (β-TCP) can be more easily produced than HA and also shows a faster resorption time, but it is more fragile [54].

Moreover, scaffolds with a fibrous nature that is similar the fibrous nature of Collagen. These Fibrous scaffolds have different structure, organization and size. Hydrogel scaffolds results from the crosslinking of monomer or polymer chains through covalent and/or noncovalent interactions, which are hydrophilic polymer networks. They have physicochemical characteristics that are comparable to some human tissues [58]. Additionally, it is feasible to achieve the desired shape, degradation rate, porosity, or release profile by altering the crosslinking method and degree [58–61].

Thus, to effectively respond to host tissue and prevent the induction of an inflammatory response, it is crucial to identify the appropriate biomaterials for scaffold creation. Additionally, altering the scaffolds' surfaces may change their properties, affecting how they interact with MSCs. Numerous mechanical and chemical procedures have been suggested in order to alter the amount of functional groups, surface charge, hydrophilicity, and molecular weight of compounds, as well as changes in surface topography, functional groups, and wettability [62, 63]

Dental stem cells and biomaterials (Fig. 1)

Fig. 1.

Combination of dental stem cells and biomaterials that are used in vitro and in vivo studies

DPSCs and biomaterials

The efficacy of DPSCs in osteogenesis was investigated in combination with various scaffolds and in different defects and animal models, resulting in broad outcomes. Calvarial defects repair and subcutaneous implementation were the most frequent in vivo trials. A comparison between DPSCs and BMSCs seeded in Bio-Oss scaffold showed that (DPSCs-Bio-Oss) group has similar bone regeneration capability, bone mineral density and osteogenesis-related proteins to those for the (BMSCs-Bio-Oss) group in rabbit calvarial defects [64]. On the other hand, another study demonstrated that the use of DPSCs in combination with Bio-Oss scaffold or micro–macro biphasic calcium phosphate (MBCP) resulted in less bone formation than autogenous bone group-the gold standard- or (BMSCs + Bio-Oss) and (BMSCs + MBCP) groups in rabbit calvarial bone defect model [65].

DPSCs can be long-term cryopreserved and still capable of differentiation to produce woven bone in vitro, which converted into a 3D lamellar bone type giving similar results to non-cryopreserved DPSCs when transplanted subcutaneously into the dorsal surface of immunosuppressed rats [66, 67]. Woven bone resulted from culturing of DPSCs for 30 days showed significantly increased expression of osteogenic factors like RUNX2 and vascularization factors like VEGF. Woven bone samples, transplanted subcutaneously into the dorsal hypodermal layer of rats, were consistently remodeled into highly vascularized bone tissue [68].

The combination of DPSCs and hyaluronic-based hydrogel scaffold showed superior bone formation compared to untreated or scaffold-treated groups in calvarial defect of immunocompetent rats [69]. Additionally, a mixture of DPSCs and (B-TCP) or (HA/PLGA) scaffold supported the formation of mineralized jawbone resembling natural rabbit mandibular bone in critical-sized jawbone defects. In vivo implantation of HA/PLGA seeded with DPSCs in a rabbit mandible model was successful in significantly decreasing the lesion size while increasing osteogenic marker expression [70, 71]. Higher osteogenesis and vascularization can be obtained by treating scaffolds to create subtractive pits or microcavities [72]. Porous biphasic hydroxyapatite/ tricalcium phosphate (HA/TCP) is the predominant scaffold material used in most of human dental pulp cells studies [73], while evidence about its effectiveness in bone regeneration in vivo is inconsistent when seeded with DPSCs. It showed limited hard tissue formation [74] and did not significantly improve bone formation when compared with the effect of scaffolds alone [75].

A potential alternative to calcium-based scaffolds is non-calcium based biomaterial–self-assembling peptide nano-fiber hydrogel, which presented very promising results in vivo and dynamic ability in comparison with previous studies [73]. In addition, 3D 45S5 Bioglass scaffolds combined with DPSCs have the potential to promote in vivo bone tissue formation [76].

Additionally, the chitosan/collagen hybrid scaffold was used to stimulate the osteo/odontogenic differentiation of DPSCs in vitro, resulting in an extensive formation of a hydroxyapatite-rich nanocrystalline calcium phosphate. The in vivo results were very promising, where formation of bone-like tissues in the in vivo samples was observed [77]. Moreover, different studies have shown that chitosan/alginate (Ch/Alg) favored fibro/chondrogenic differentiation of DPSC, by upregulating the expression of chondrogenic markers [78].

In a study by Tatsuhiro et al., 3D scaffold free constructs of DPSCs were shown to express bone related genes, and formation of calcified matrix was observed. Cell constructs that were cultured with osteogenic induction had higher expression of bone related genes and had a higher degree of calcified matrix formation when compared with constructs grown without induction [79].

Formation of dental pulp-like tissue has been observed from 3D constructs of DPSCs, implanted subcutaneously in severe combined immunodeficient (SCID) mice. Both previous studies revealed the regenerative and osteogenic potential of 3D cell constructs of DPSCs. Additionally, a major advantage of cell constructs is that they are scaffold free and the cells don’t require growth factors to exhibit an osteogenic behavior [79, 80].

Gelatin has also been used in combination with many other scaffolds to improve bone regeneration [81]. First, combination of gelatin with HNTs/GelMA Hydrogels showed significant increase in bone regeneration and expression of osteogenic genes in vivo [82]. Second, gelatin/hA-TCP scaffolds increased bone regeneration significantly by upregulating the expression of osteogenic genes [83]. Third, encapsulation of DPSCs with HUVEC cells with GelMA hydrogels showed to improve dental pulp regeneration [84].

Collagen has been widely used as a scaffold for bone regeneration; as collagen is one of the main components of bone matrix [85]. Therefore, collagen has been combined with other types of polymers and/or inorganic material to create new scaffolds. Akkouch et al. combined collagen with HA and PLCL and compared this scaffold to a PLCL scaffold. The combined scaffold seeded with DPSCs showed higher cell adhesion, cell viability, bone regeneration, ALP activity, osteoblast growth and differentiation, and mineralization rate. It should be noted that this study used human osteoblast-like cells (differentiated or partly differentiated DPSCs) [86].

Hyaluronic acid is a glycosaminoglycan distributed throughout the body primarily in synovial fluid. Hyaluronic acid was used as a scaffold by Ferroni et al., the scaffold was seeded with DPSCs. In vitro, the cells showed increased expression to specific differentiation markers and were able to have osteogenic phenotypes, there was formation of osteodentin-like tissue in hyaluronan-DPSCs grafts [87]. In a similar study by Schmidt et al., low molecular weight hyaluronic acid was seeded with DPSCs, the DPSCs had fibroblast and angiogenesis-related surface markers [88].

Recently, polypeptides have been shown to be unique biomaterials for 3D cell culture and excellent candidate scaffolds for both soft and mineralized dental tissues in tissue engineering. As previous studies have demonstrated, Self-Assembling Peptide seeded with DPSCs and human umbilical vein endothelial cells (HUVEC) showed an upregulation in the expression of odontogenic genes and mineral nodules. In addition, the cells underwent osteogenic, chondrogenic and adipogenic differentiation, concluding that DPSCs cultured in SAP-based biomimetic scaffold considered as a promising candidate for regeneration of dentin-pulp complex (DPC) [89].

Hydroxyapatite (HA) is considered the main inorganic component of animal bones and teeth, which is one of the first and the most common material used in synthesizing scaffolds that induce osteogenic and odontogenic differentiation of stem cells [90]. Hagar et al. seeded SHED and DPSCs on granular hydroxyapatite (gHA), even though both dental stem cell types showed osteogenic differentiation, SHED exhibited more osteogenic differentiation than DPSCs [91].

Moreover, platelet-rich plasma (PRP) is a scaffold that has been used in tissue engineering as a natural cell delivery method, it is known with its great potential in bone regeneration. Previous studies have shown that PRP scaffold seeded with DPSCs has a great potential to regenerate bone in critical defects, compared to the scaffold solely when implanted in a canine mandible in animal model, in addition to the use of dental implants [92, 93].

Interestingly, DPSCs-cell sheets treated with 4-(4-methoxyphenyl)pyrido[40,30:4,5]thieno[2,3-b]pyridine-2-carboxamide (TH), a helioxanthin derivative, showed significant bone regeneration without the use of scaffolds or growth factors [94]. As the sheet showed the greatest bone volume and the healing of most fractures when implanted in a mouse tibia model. Additionally, TH-treated sheets had induced the osteogenesis of DPSCs and promoted the maturation of differentiated DPSCs, compared the BM derived MSCs [95, 96].

Different studies have shown the potential of E1001(1 k)/B-TCP scaffold, which possesses osteoconductive and osteoinductive properties, seeded with DSCs to stimulate bone regeneration. Moreover, the generation potential was increased when it’s combined to other materials such as B-TCP and dicalcium phosphate dihydrate (DCPD). Moreover, the combination of this scaffold with bone morphogenetic protein-2 (BMP-2) showed a higher differentiation potential into bone and dentine matrix, in addition to its potential to stimulate angiogenesis when DPSCs co-cultured with HUVEC cells, results in improving blood vessel formation and bone regeneration [97, 98].

The most extensively used synthetic polymers in medical application is poly(lactic-co-glycolic acid; PLGA), it has been used recently due to its biodegradability and biocompatibility. The degree of hydrophobicity/hydrophilicity can be modified according to the way of delivery into cells in clinical application [99]. Bone regeneration was stimulated when PLGA scaffold was seeded with these SHED, BMMSCs, DPSCs [100].

Poly(lactic-co-glycolic) acid scaffold was modified by combining it with different groups such as HA to create a HA/PLGA scaffold [70]. The use of 3D printing technology has induced the differentiation potential of PLGA, as 3D printed PLGA/TCP scaffold, seeded with DPSCs promoted the bone regeneration [101].

SHED and biomaterials

Orofacial bone defects have been commonly treated with autologous hBMSCs extracted from axial and appendicular bones. While hBMSCs often resulted in unfavorable outcomes, SHED exhibited equivalent ability to BMSCs in restoring the parietal continuity with significant amounts of bone formation in all transplanted mice in combination with (HA/TCP) scaffold [102]. Superior immunomodulatory effects were achieved by (SHED-HATCP) compared to BMSCs in terms of recovering Tregs/Th17 ratio and reducing Th17 cell levels in peripheral blood when transplanted subcutaneously in mice [103].

Stem Cells from Human Exfoliated Deciduous Teeth (SHED) proved to be autologous and easily accessible stem cell source to repair mandibular defects in swine and rats models in combination with (B-TCP) and PLGA-10% bioactive glass composite scaffolds, respectively [104, 105]. In comparison to DPSCs, subcutaneous transplantation of (SHED-ceramic bovine bone (CBB)) suggested higher capability of mineralization than DPSCs [106].

Hydroxyapatite (HA) is the main inorganic component of animal bones and teeth, this made it one of the first and the most common material used in synthesizing scaffolds that induce osteogenic and odontogenic differentiation of stem cells. Hagar et al. seeding of granular hydroxyapatite (gHA) with SHED and DPSCs showed that SHED exhibited more osteogenic differentiation than DPSCs [91]. Similarly, HA seeded with (SHED- treated with FGF-2) were grafted in SCID mice, the results had shown an increase in the expression level of osteoprotegerin (OPG) and decreased levels of receptor activator of NF-kappaB ligand (RANK-L) expression, this indicates osteoblast-like behavior as OPG inhibits bone resorption [107].

DFSCs and biomaterials

Dental follicle stem cells (DFSCs) seeded in demineralized bone matrix (DBM) showed remarkably enhanced bone formation in mandibular defects of miniature pigs and subcutaneous tissues of mice with no significant difference between fresh and cryopreserved DFSCs [108]. Equivalently, bone tissue formation was observed in vivo with no statistically significant difference among the groups (DFSCs-Collagen 1 scaffold), (DFSCs-polystyrene dishes (Pol-d)) and (BMSCs-Pol-d) in rats calvarial defects model, which makes DFSCs a promising alternative to BMMSCs [109]. Positive results were obtained with (DFSCs-PCL) in repairing critical sized calvarial defects in immunocompetent rats [110]. Moreover, metal nanoparticles have been incorporated into HA based scaffolds; Lucaciu et.al., incorporated titanium nanoparticles in HA scaffolds to form TiHA scaffolds, they seeded the scaffolds with DFSCs, BMP-2 was added to some of the samples. Both samples showed markers of osteogenic differentiation, however, samples without BMP-2 exhibited better osteogenic differentiation [111].

PDLSCs and biomaterials

Periodontal ligament stem cells seeded in HA/TCP scaffold generated relatively loose fiber arrangement with high directionality of fibers which was similar to DPSCs but with no obvious vascular regeneration in vivo [112]. It is suggested that nano-hydroxyapatite/collagen/ poly(Llactide) (nHAC/PLA) can be used as potent alternative for HA/TCP [113]. There was no statistically significant difference in bone formation observed between PDLSCs, BMSCs and DFSCs [109]. Nanohydroxyapatite-coated genipin-chitosan conjunction scaffolds (HGCCS), OsteoBiol® Dual-Block scaffold and Gelfoam absorbable collagen sponge possessed promising outcomes in repairing craniofacial and mandibular defects [114–116].

Additionally, for periodontal regeneration, PLGA was synthesized as biodegradable sponge polymer. It was used to deliver cementoblasts in a rodent periodontal fenestration model, stimulating the mineralization potential of cementoblasts to repair periodontal wound defects [61]. HydroMatrix seeded with PDLSCs showed a strong mineralization potential [117].

Hydroxyapatite (HA) can also be used in combination with other materials to synthesize hybrid scaffolds. E, L.L et al. synthesized a nano-HA (nHA)/Collagen/PLA hybrid scaffold, seeded with rat PDLSCs scaffold, and then implanted into mice. When compared to cells not treated with Estrogen (E2), the cells in mice treated with E2 showed significantly reduced proliferative ability however they had a very intense alkaline phosphatase (Sasportas et al.) staining, and showed calcium deposition and mineralization nodules [119].

Moreover, PDLSC loaded with gelatin methacrylate (GelMA) and poly(ethylene glycol) dimethacrylate (PEGDA), GelMA/PEGDA composite hydrogel by using 3D bioprinting technique, was found to regenerate bone efficiently when implanted in animal model of periodontitis [120].

SCAP and biomaterials

Stem cells from apical papilla (SCAP) seeded in a decellularized human dental pulp scaffold in the presence of rat tail collagen I, induced the differentiation of the cells into differentiated cells with similar structure and function such as dental pulp stem cells [22, 121].

Polycaprolactone (PCL) is a biodegradable polymer available for use as a scaffold. The degradation of PCL is slow and this makes it suitable for long term implantation in the human body. PCL seeded with stem cells from the apical papilla (SCAP) have been shown to form calcification nodules, indicating the capability of this scaffold to induce the osteogenic differentiation of SCAP [58].

Table 1 summarizes all studies related to in vitro and in vivo in respect of using biomaterials combined to the dental stem cells.

Table 1.

Summary of in vitro and animal model in vivo studies

Cell type	Scaffold	In vivo/in vitro	In vivo model/defect	Results	References
DPSCs and BMSCs	Bio-Oss	In vivo	Rabbit calvarial defects	Bone was regenerated in the defect size	[64]
DPSCs and BMSCs	Micro–macro biphasic calcium phosphate (MBCP) and Bio-Oss	In vivo	Rabbit calvarial bone defects	The study results demonstrated that autogenous bone is the gold standard. Both the DPSCs and BMSCs enhanced the osteoconductive capacities of MBCP and Bio-Oss. In addition, the efficiency of the BMSCs combined with MBCP and Bio-Oss was comparable to that of the autogenous bone after 8 weeks of healing	[133]
DPSCs	Woven bone obtained in vitro, no scaffold needed	In vivo	Planted subcutaneously into the dorsal surface of 10–12 week immunosuppressed rats	Dental pulp stem cells and their osteoblast-derived cells can be long-term cryopreserved and may prove to be attractive for clinical applications	[66]
DPSCs	Woven bone obtained in vitro, no scaffold needed	In vivo	Transplanted into the dorsal surface (i.e., subcutaneously) of 10–12 week immunosuppressed rats	These bone samples, after in vivo transplantation into immunosuppressed rats, were remodeled in a lamellar bone containing entrapped osteocytes	[67]
DPSCs	Woven bone obtained in vitro, no scaffold needed	In vivo	Subcutaneously transplanted into the dorsal surface of 10-week-old athymic nude rats	WB samples, fabricated by DPSCs, constitute a noteworthy tool and do not need the use of scaffolds, and therefore they are ready for customized regeneration	[68]
DPSCs	Hyaluronic-based hydrogel scaffold	In vivo	Calvarial defects of immunocompetent rats	Statistically significant superior new bone formation in the biocomplex-treated group, compared to scaffold and nothing, and cell-scaffold and nothing	[69]
DPSCs	Hydroxyapatite matrix and polylactic polyglycolic acid (HA/PLGA)	In vivo	Rabbits with bilateral mandibular critical-sized defects	(HA/PLGA-DPSC) scaffold was an effective in vivo method for mandibular bone regeneration in critical-sized defects induced on rabbit models	[70]
DPSCs	Tyrosine-derived polycarbonate, E1001(1 K)/β-TCP scaffolds	In vivo	Critical sized alveolar bone defects in an in vivo rabbit mandible defect model	Although unseeded scaffolds supported limited alveolar bone regeneration, more robust and homogeneous bone formation was observed in hDPSC/HUVEC-seeded constructs, suggesting that hDPSCs/HUVECs contributed to enhanced bone formation	[71]
DPSCs	(PLGA) scaffolds	In vivo	Transplanted into the dorsal surface (i.e. subcutaneously) of immunocompromised 10–12 week Wistar rats	Surface microcavities appear to support a more vigorous osteogenic response of stem cells and should be used in the design of therapeutic substrates to improve bone repair and bioengineering applications in the future	[130]
DPSCs	Self-assembling peptide nano-fibre hydrogel	In vivo	Subcutaneous pockets were created on both sides of the animals in the flank region, cells transplanted subcutaneously into five nude mice	The cell-gel constructs were transformed into tissue pieces that were mineralised and contained blood capillaries	[73]
DPSCs	(HA/TCP) ceramic scaffolds	In vivo	Subcutaneous pockets were created by blunt dissection. Cell-loaded and control implants were placed subcutaneously in 10-week-old immunocompromised mice	The limited hard tissue regeneration ability of dental pulp stromal cells questions their practical application for complete tooth regeneration. Repeated cell passaging may explain the reduction of the osteogenic ability of both bone- and dentinal-derived stem cells	[74]
DPSCs	Granular deproteinized bovine bone with 10% porcine collagen and granular β;-tricalcium phosphate	In vivo	Critical calvarial defects in SCID Beige nude mice	The results showed that tissue-engineered constructs did not significantly improve bone-induced regeneration process when compared with the effect of scaffolds alone. In addition, the data also showed that the regeneration induced by β;-tricalcium phosphate alone was higher after 8 weeks than that of scaffold seeded with the 2 stem cell lines	[75]
DPSCs	3D 45S5 Bioglass scaffolds	In vivo	Intraperitoneal implantation in male immunecompromised nude mice	The study has demonstrated the potential of using a combination of HDPSCs with 3D 45S5 Bioglass scaffolds to promote bone-like tissue formation in vitro and in vivo	[76]
SHED	(HA/TCP) ceramic scaffolds	In vivo	Transplanted into the 2.7 mm diameter defect created by a trephine bur on the calvaria of immunocompromised mice	SHED are capable of repairing critical-size parietal defects in immunocompromised mice. However, SHED-mediated bone lacks hematopoietic marrow elements, unlike the bone/marrow organ-like structure generated by bone marrow mesenchymal stem cells	[102]
SHED	Hydroxyapatite tricalcium phosphate (HA/TCP)	In vivo	Subcutaneously transplanted into beige nude/nude Xid (III) mice	SHED possess similar stem cell properties as those seen in BMMSCs, including osteo/odontogenic and adipogenic differentiation in vitro, forming mineralized tissue in vivo, and expression of extensive mesenchymal stem cell markers	[103]
SHED	B-TCP	In vivo	Critical-size bone defects generated in swine mandible models	Stem cells from miniature pig deciduous teeth, an autologous and easily accessible stem cell source, were able to engraft and regenerate bone to repair critical-size mandibular defects at 6 months post-surgical reconstruction	[104]
SHED	PLGA-10% bioactive glass composite scaffold	In vivo	Male adult Sprague–Dawley rats aged 7 weeks were used. Cleft-mimicking lesions were induced by extracting the right permanent maxillary first molar	PLGA-10% bioactive glass composite scaffold enhanced osteogenic differentiation of SHED in vitro and in vivo	[105]
SHED	Ceramic bovine bone (CBB)	In vivo	Transplanted subcutaneously in immunocompromised mice	The results of the in vivo transplantation suggest that SHED have a higher capability of mineralization than the DPSCs	[106]
Fresh and cryoprese-rved human FDCs	DBM	In vivo	Mandibular defects of in miniature pigs and subcutaneous tissues of mice	hDFCs possess immunomodulatory properties that involved inhibition of the adaptive immune response mediated by CD4 and MHC II, which highlights the usefulness of hDFCs in tissue engineering	[108]
DFSCs, PDLSCs, and BMSCs	Collagen 1/ polystyrene dishes (Pol-d)	In vivo	A curvilinear sagittal incision was made in the calvarium scalp of anesthetized, immunocompromised rats	All three cell types showed equivalent osteogenic capacity in vivo at 4 weeks postoperatively. There were no statistically significant differences among the cell populations (BMSCs, DFSCs, PDLSCs) with respect to capacity for bone formation	[109]
DFSCs	Polycaprolactone (PCL) scaffold	In vivo	Critical-size defects were created on the skulls of 5-month-old immunocompetent rats, and the cell-scaffold constructs were transplanted into the defects	This study demonstrated that transplantation of DFSCs seeded into PCL scaffolds can be used to repair craniofacial defects	[110]
PDLSCs	HA/TCP	In vivo	Transplanted subcutaneously into the dorsal surfaces of 10‐week‐old immunocompromised beige mice	The PDLSC sheets showed relatively loose fibre arrangement with obvious directionality of fibers, which was similar to DPSC sheet, although there was no obvious vascular regeneration	[112]
PDLSCs	Nano-hydroxyapatite/collagen/poly(Llactide) (nHAC/PLA)	In vivo	Implanted into the subcutaneous pocket of 2-year-old beagle dogs	The amount of newly formed bone in the nHAC/PLA group was higher than that in the HA/TCP group (P < 0.05). This study suggests that nHAC/PLA can be used as a potent scaffold for alveolar bone regeneration	[113]
PDLSCs	nanohydroxyapatite-coated genipin-chitosan conjunction scaffold (HGCCS)	In vivo	Full-thickness defects measuring 5 mm in diameter were created bilaterally in the parietal bones in 8-week-old adult male Sprague–Dawley rats	The bone defects were filled with newly formed dense tissue in the HGCCS + PDLSC group. HGCCS promoted bone repair in vivo when implanted with PDLSCs in a critical size defect model. This study demonstrates that PDLSC-seeded HGCCS constructs are promising for bone tissue repair	[114]
PDLSCs	OsteoBiol® Dual-Block (Tecnoss® Dental, Coazze (TO), Italy). It is a collagenated porcine block constituted by natural cancellous and cortical bone	In vivo	Implanted in the mouse calvaria	This scaffold guarantees, due to its rigid consistency, that the original volume of the grafting site can be preserved. Our results suggest consideration of DB as a biocompatible, osteoinductive and osteoconductive biomaterial	[115]
PDLSCs	Gelfoam (Absorbable gelatin sponge)	In vivo	Periodontal alveolar bone defect model in rats	Defects treated with PDLSCs showed significantly greater percentage bone fill and length of new bone bridge compared with the untreated group or the group treated with Gelfoam alone	[116]
cMSCs, DTSCs, and DPSCs	PRP	In vivo	Parent Canine Mandible	All three cell types showed bone regeneration.DTSC showed slightly less bone regeneration than DPSC/cMSC	[65]
cBMSC, cDPSC, and pDTSC	PRP + HA-coated osseointegrated dental implants	In vivo	Mandible of adult hybrid dogs with a mean age of 2 years	The cBMMSCs/PRP, cDPSCs/PRP, and pDTSCs/PRP groups had well-formed mature bone and neovascularization	[92]
cDPSC, cBMSC, and cPC	PRP  + dental implants	In vivo	Mandible of dogs	DPSCs showed the highest bone regeneration.Both DPSCs and BMSCs had much higher bone regeneration compared to PCs	[93]
DPSCs	3D-Printed Polycaprolactone Scaffolds Coated with Freeze-Dried Platelet-Rich Plasma	Both	Calvarial defects in adult male Sprague–Dawley rats	In vitro: freeze dried PRP increased expression of osteogenic genes and increased ALP activity.Freeze dried PRP-PLC had more cell attachment, but no difference between freeze dried PRP-PLC and traditional PRP-PLC in cell migration and proliferation In vivo: freeze dried PRP-PLC showed highest bone regeneration compared to the other groups	[134]
DPSCs	Cell sheets	Both	Mouse Tibia Fracture	Osteogenic medium (OM) + TH showed highest bone regeneration	[95]
hDPSCs, BMSCs	Cell sheets	Both	Calvarial defects in mice	In vitro: TH + Regular medium (RM) showed no bone regeneration.TH + Osteogenic medium (OM) showed bone regeneration even in short term culture (faster than other factors) TH might be more effective at promoting maturation of DPSCs In vivo: new bone tissue was formed using TH + OM without the use of scaffolds or growth factors	[96]
BMSCs, and DPSCs	Bio-Oss	In vivo	Adult male New Zealand white rabbits calvarial Model	DPSCs and BMSCs seeded on Bio-Oss showed similar bone regeneration efficacy	[64]
hDPSCs	Bonelike	In vivo	Femur defects in healthy skeletally-mature Merino sheep (Ovis aries)	hDPSCs + (Bonelike) showed increased bone regeneration compared to Bonelike alone	[135]
hDPSCs + HUVECs	E1001(1 K)/β-TCP	In vivo	Mandibular defects on New Zealand White Rabbits	Osteodentin -like mineralized jawbone that is similar to natural rabbit bone was formed by the seeded scaffold	[71]
hDPSCs	E1001(1 K)/β-TCP	In vivo	Mandibular defects in female nude rats	Both seeded scaffolds and scaffolds supplemented with BMP2 showed significant bone regeneration but the group with BMP2 was higher	[98]
hBMSCs,hDPSCs, and SHED	polylactic-coglycolic acid barrier membrane (PLGA)	In vivo	Calvarial defects in immunodeficient mice	All 3 cell types with scaffolds induced similar bone regeneration.SHED produced more osteoid and collagen fibers	[100]
DPSCs	hydroxyapatite matrix and polylactic polyglycolic acid (HA/PLGA)	In vivo	New Zealand rabbits with bilateral mandibular critical-sized defects	DPSCs with scaffold caused increased radiodensity percentage compared to control DPSCs with scaffolds also caused the highest lesion size reduction (highest regeneration), and increased osteogenic marker expression	[70]
DPSCs	1- (3D-PLGA/TCP) 2-(3D-TCP) 3- C-TCP	In vitro	–	All scaffolds showed high DPSCs viability, although TCP scaffolds had higher viability compared to 3D-PLGA/TCP All DPSCs showed increased proliferation in scaffolds. However, 3D scaffolds had higher DPSC proliferation than C-TCP, and 3D-TCP scaffolds showed significantly higher regeneration than other scaffolds	[101]
DPSC	Biomimetic chitosan/gelatin Scaffolds 1- CS/Gel-0.1 2- CS/Gel-1	Both	In vivo: dorsum of immunocompromised mice	In vitro: Both scaffolds supported viability and proliferation, but the CS/Gel-0.1 scaffold showed a better response In vivo: abundant matrix mineralization was seen, with the BMP-2 group having a denser matrix and more osteoid than groups without BMP-2	[77]
hDPSCs	1- HNTs (halloysite nanotubes) incorporated hydrogel 2- (gelatin methacrylate (GelMA) and HNTs)	Both	Calvarial defect model of Sprague−Dawley rats	In vitro: HNTs/GelMA enhanced hDPSCs proliferation and osteogenesis In vivo: Adding HNTs increased expression of osteogenic genes and increased bone regeneration significantly	[136]
DPSCs	1- gelatin-HA-TCP scaffolds 2-gelatin-only scaffold	In vitro	–	Gelatin -HA-TCP scaffolds were better than control and gelatin only scaffolds in inducing ALP activity, proliferation, and osteogenic differentiation	[83]
hDPSCs + HUVECs	5% gelatin methacrylate (GelMA) hydrogel	In vivo	Implanted subcutaneously in nude rats	GelMA with hDPSCs and HUVECs formed robust pulp-like tissues along with increased vascularity	[84]
hDPSCs	Scaffold-free 3D Construct	In vitro	–	Cells in the 3D construct showed increased expression of bone related genes	[79]
SHED	chitosan	In vitro	–	SHED + scaffold + TGFb1 induced higher proliferation rates, higher mineralization rate, higher adhesion, and increased osteogenic differentiation compared to scaffold alone or growth factor alone	[137]
DPSCs	1-Coll/HA/PLCL 2-PLCL	In vitro	–	Coll/HA/PLCL scaffolds had high adhesion and viability, with DPSCs having higher affinity to it compared to PLCL Coll/HA/PLCL induced more osteoblast growth and differentiation, ALP activity, calcium deposition, and mineralization rate	[86]

Clinical trials

Periodontal disease (PD)

Ferrarotti et al. used DPSCs in a randomized clinical trial to treat deep intrabony defects caused by chronic periodontitis, researchers used dental pulp micrografts in collagen spongy scaffold and compared it to collagen scaffold alone [122]. The results of the clinical trial were significant, as it showed the importance of minimally invasive surgery to provide stability for this kind of procedures. However, Periodontal Ligament Stem Cells (PDLSCs) with Bio-Oss—a collagen-based scaffold—were used for the treatment of periodontal intrabony defects as well. While this study demonstrated the safety of this treatment, there was no significant difference in bone regeneration between the (scaffold and PDLSCs) group and the (scaffold only) group; they both were significant in comparison with the control group [137].

In a case report by Hernández-Monjaraz et al. deciduous DPSCs in lyophilized collagen-polyvinylpyrrolidone sponge scaffold were used to treat the premolar area of a patient with periodontitis. After follow up, the patient showed decreased tooth mobility, periodontal pocket depth and bone defect area as the bone mineral density had increased [123].

Recently, deciduous DPSCs were used in a QUASI randomized clinical trial to treat periodontal disease, cells were delivered in a collagen scaffold. The experimental group showed significant increase in both bone mineral density (BMD) and salivary superoxide dismutase (SOD) levels which indicates an increase in bone regeneration. It also showed decreased levels of IL1β [124].

Pulp necrosis

In a randomized clinical trial, autologous Deciduous Pulp Stem Cells (DPSCs) were used without a scaffold – instead researchers used aggregates containing cells and extracellular matrix—to treat traumatic dental injuries with pulp necrosis, the results of this clinical trial led to the conclusion that human DPSCs can regenerate whole dental pulp which makes it a better treatment than conventional apexification [125].

Post extraction complications

The first time autologous DPSCs were used to treat a clinical defect secondary to wisdom teeth (third molar) removal surgery was by d'Aquino et al. The operation produces an alveolar bone defect in the mandible which compromises the second molar teeth and may lead to its loss in the future. The researchers performed a study design known as Split-Mouth clinical trial where the patient had a test site on one side and a control site on the other, they used a collagen bioscaffold to deliver the stem cells to the test site and it was used alone in the control site. The results of this study showed complete bone regeneration in the test site. More importantly, this study was the first to use DPSCSs in treating bone defects [126].

A case report was published by Giuliani et al. as a follow up [127] to the patients of the previous study after three years to investigate the type of the regenerated bone using histological tests. The study showed the formation of a compact bone which is uniformly vascularized in the test site from the previous study in 2009. Although the formed bone is different from the physiologic bone (spongy bone) that is normally found in the mandible; the study suggests it may have positive clinical impacts like increased stability and resistance for different agents [127].

Although the previous studies demonstrated the ability of autologous DPSCs to reduce bone resorption and increase bone density, a split-mouth clinical trial to assess the efficacy of autologous DPSCs in bone resorption after inferior third molar teeth extraction found no significant difference between the control and experimental group [128].

Another split-mouth case report used autologous dental ligament stem cells (DLSC) micrograft delivered to the test site in a collagen sponge. They followed up the patient for six months and mineralization was higher in the test site compared to collagen sponge alone [129].

Atrophic maxilla

Autologous DPSCs were used for sinus lifting in a patient with atrophic maxilla that needed a prosthetic implant, in a case report by Brunelli et al. The researchers measured bone density using CT imaging and compared the newly formed bone to the native bone. The results of this study showed that density of treated bone was double of the native bone, but the study had multiple limitations and no conclusion could be drawn from it [130].

Cleft lip and palate

In a prospective cohort study, Tanikawa et al. used deciduous Dental pulp stem cells for the first time in maxillary alveolar repair in Cleft Lip and Palate (CLP) patients. Researchers delivered stem cells in a hydroxyapatite collagen (Bio-Oss) and compared them to a historical control group which were treated for the same condition with different approaches. The results of this study were astonishing as there were completion of defect 6 months after the operation and new tooth eruption in the defected area was observed in two thirds of the patients [131]. This study shows the potential of Dental stem cells in treating more complicated facial bony defects.

Table 2 summarizes all clinical studies related to the use of biomaterials combined with dental stem cells.

Table 2.

Summary of human clinical studies

Cell type	Scaffold	In vivo/in vitro	Defect	Study type	References
DPSCs	collagen sponge bio-complex	Human clinical study	Periodontal disease (PD)	Randomized clinical trial Significantly positive	[122]
PDLSC	Bio-Oss	Human clinical study	Periodontal disease (PD)	Randomized clinical trial	[138]
DPSCs	lyophilized collagen-polyvinyl-pyrrolidone sponge	Human clinical study	Periodontal disease (PD)	Case report	[124]
DPSCs	Collagen	Human clinical study	Periodontal disease (PD)	QUASI randomized clinical trial	[139]
DPSCs	No scaffold	Human clinical study	Pulp Necrosis	Randomized clinical trial	[126]
DPSCs	Collagen sponge bio-scaffold	Human clinical study	Post extraction complications	Split-mouth randomized clinical trial	[127]
Follow up for 6	Follow up for paper 6	Human clinical study	Post extraction complications	Case report	[128]
DPSCs	Collagen	Human clinical study	Post extraction complications	Split-mouth randomized clinical trial	[129]
DLSCs	Collagen sponge	Human clinical study	Post extraction complications	Split-mouth Case report	[130]
DPSCs	Collagen sponge	Human clinical study	Atrophic maxilla	Case report	[131]
DPSCs	Hydroxyapatite collagen (bio-oss)	Human clinical study	Cleft lip and palate (CLP)	prospective cohort study Significantly positive	[132]

Conclusion

Stem cell-mediated therapeutic interventions have gained a lot of attention recently and have significantly advanced the treatment of diseases, particularly those that are resistant to conventional therapies such as periodontitis and bone defects. Dental stem cells (DSCs) are emerging as the most feasible source of stem cells that can be used for the treatment of bone defects and periodontitis. Different combinations of DScs in conjunction with biomaterials have been investigated and many clinical trials are showing promising results for the treatment of maxillofacial defects in the field of tissue engineering. Therefore, more research must be done to improve DSCs' capacity to offer better novel treatment approaches, that can be applied clinically.

Authors’ contribution

DA: who had the idea for the article. LST, MBO, YIH, JAA, Nazneen Aslam, who performed the literature search and DA, NA, HJ and AA who drafted and/or critically revised the work.

Declarations

Conflict of interest

The authors have no conflict of interest to declare.

Ethical statement

There are no experiments carried out for this article.