Skip to main content

Strategies of cell and cell-free therapies for periodontal regeneration: the state of the art



Periodontitis often causes irrevocable destruction of tooth-supporting tissues and eventually leads to tooth loss. Currently, stem cell-based tissue engineering has achieved a favorable result in regenerating periodontal tissues. Moreover, cell-free therapies that aim to facilitate the recruitment of resident repair cell populations to injured sites by promoting cell mobilization and homing have become alternative options to cell therapy.

Main text

Cell aggregates (e.g., cell sheets) retain a large amount of extracellular matrix which can improve cell viability and survival rates after implantation in vivo. Electrostatic spinning and 3D bioprinting through fabricating specific alignments and interactions scaffold structures have made promising outcomes in the construction of a microenvironment conducive to periodontal regeneration. Cell-free therapies with adding biological agents (growth factors, exosomes and conditioned media) to promote endogenous regeneration have somewhat addressed the limitations of cell therapy.


Hence, this article reviews the progress of stem cell-based tissue engineering and advanced strategies for endogenous regeneration based on stem cell derivatives in periodontal regeneration.


The periodontium is an intercalated structure of hard and soft tissues surrounding and supporting the teeth and is capable of resisting and dispersing masticatory forces in a highly hierarchical manner. The periodontium consists of four different tissues: the gingiva, cementum, periodontal ligament (PDL), and alveolar bone [1, 2]. Periodontitis is the most common reason for the destruction of periodontal tissue, leading to the loss of teeth in serious situations, and it significantly decreases mastication function and quality of life [2, 3]. Conventional treatments include supragingival cleaning, subgingival scaling, and root planning to remove pathogenic factors [4], which only halt the further development of inflammation and do not work to restore damaged periodontium. In recent decades, bone grafting and guided tissue/bone regeneration techniques have been widely used in the clinic and can achieve some periodontal regeneration; however, the results are unpredictable and limited.

Tissue engineering plays a significant role in regenerative medicine and consists of three basic elements: cells, scaffolds, and growth factors. Of these, stem cells are crucial given their ability to self-renew and differentiate into a variety of cell types. Many animal models and clinical studies [5, 6] have confirmed that stem cell-based tissue engineering has achieved a favorable result in regenerating periodontal tissues. Nevertheless, its clinical application is hampered by various issues, such as stem cell acquisition, in vitro expansion, immunogenicity, and ethics. The phenomenon of the native wound healing cascade hints that it is possible to harness the intrinsic regenerative potential of endogenous tissues for tissue repair and regeneration without the aid of exogenous stem cells [7]. However, naturally endogenous regenerative processes are generally limited and unable to successfully regenerate many tissues. Therefore, current strategies for endogenous regenerative medicine aim to facilitate the recruitment of resident repair cell populations to injured sites by promoting cell mobilization and homing and have become alternative options to cell therapy.

Hence, this paper will review the progress of cell-based tissue engineering and cell-free endogenous regeneration applied to periodontal regeneration (a schematic of periodontal regeneration is shown in Fig. 1) and discuss the most likely means of achieving functional periodontal tissue regeneration.

Fig. 1
figure 1

Schematic of periodontal regeneration. Cell-based tissue engineering and cell-free endogenous regeneration in periodontal regeneration. Created with

Tissue engineering and periodontal regeneration

Mesenchymal stem cells (MSCs) originate from the mesoderm and possess immunomodulatory and anti-inflammatory functions as well as the ability to regulate intercellular communication and function via paracrine secretion [8], which makes them the most commonly used stem cells in current research for the study and treatment of human refractory diseases [9]. Greater than 1300 clinical trials have utilized MSCs, and these trials are currently listed at

Advances in MSCs-based tissue engineering have given rise to novel therapeutic strategies for tooth and periodontal repair and regeneration. Stem cells applied to periodontal regeneration can be classified as dental stem cells and nondental stem cells. Dental stem cells include dental pulp stem cells (DPSCs), PDL stem cells (PDLSCs), dental follicle cells, stem cells from human exfoliated deciduous teeth (SHED), stem cells from the apical papilla, and gingival MSCs (GMSCs) [10, 11]. Among these, PDLSCs [12,13,14] are the most commonly used in periodontal tissue regeneration given their multilineage differentiation potency, anti-inflammation properties, and immunomodulation properties [15]. Nondental stem cells include embryonic stem cells, induced pluripotent stem cells, bone marrow MSCs (BMMSCs), and adipose-derived stem cells. BMMSCs [16,17,18] have been reported to differentiate into PDL, have a strong osteogenic differentiation ability and are readily available, allowing bone marrow to become one of the main stem cell sources for regenerating the periodontium.

Cell aggregates and periodontal regeneration

Transplanting an adequate number of seed cells with optimal viability into the defects and guaranteeing that they remain functional is the first step toward achieving clinical efficacy for cell therapies. Therefore, it is essential to choose an appropriate method of cell transplantation. When administered as a suspension into the area of periodontal defects, there is little time for the cells to adhere, which results in low cell survival and carries the risk of cell migration to other parts of the body [19].These deficiencies are ameliorated by cell aggregates [20, 21], which are a cluster of cells bound to each other through cell proliferation and/or cell aggregation and are valuable for studying tissue development [22]. Various studies have shown enhanced cellular function in cell aggregates compared to that in single-cell suspensions, such as immunomodulatory potency, multilineage differentiation of MSCs aggregates, and improved cell viability and survival rates after implantation in vivo [23]. Cell aggregates can be produced by a variety of methods; among them, the cell sheet technique is the most frequently used method in periodontal regeneration. Cell sheets [24] retain a large amount of extracellular matrix (ECM), which provides a microenvironment for cell survival, assists in maintaining cell stemness, enhances cell viability, promotes cell adhesion to damaged areas, and stimulates host cell homing. The most popular approach to harvesting cell sheets is via temperature-responsive intelligent polymer cell culture dishes. After lowering the temperature below 32 °C, the polymer dish surfaces become hydrophilic; as a result, an intact cell sheet can be separated from the dish surface without enzymatic treatment [25].

PDLSCs sheets could form in these temperature-responsive culture dishes and achieve ectopic regeneration of cementum-like and PDL-like tissues after transplantation [26]. However, only a thin layer of cell sheet was insufficient for the regeneration of large periodontal defects. One study added 50 ug/mL ascorbic acid to the cell culture medium, prolonged the culture of cells for 14 days, and reported an increase in secreted ECM proteins, such as type I collagen, laminin and fibronectin [27]. Raju et al. [28] fabricated a three-dimensional (3D) composite cell sheets containing 10 layers of cells. Only the complex cell sheet, combined with the functional linkage of PDL-like fibers to the tooth root, was shown to physically renew the bone-ligament structure in ectopic and orthotopic transplantation. Each cell type derived from a different tissue has a characteristic ability to differentiate, so the combination of stem cells from distinct tissue sources could produce enhanced periodontal regeneration outcomes. Zhang et al. [29] combined PDLSCs with BMMSCs, significantly enhancing the expression of bone- and ECM-related genes and proteins. To address problems, such as necrosis and inflammation of the transplanted tissue, a previous study mixed PDLSCs and human umbilical vein endothelial cells (HUVECs) for transplantation, which led to the formation of PDL-like tissues in vivo and higher numbers of blood vessel lumens [30]. In addition, numerous studies have further improved regeneration by adding bioactive molecules or bone graft materials to enhance the properties and mechanical strength of the cell sheets.

Scaffold materials

For sophisticated and variable periodontal regeneration microenvironments and large-scale periodontal defects, in addition to the use of cell sheets, researchers are turning their attention to well-designed scaffold materials that mimic periodontal structures to promote 3D periodontal regeneration. There are different types of scaffold materials (Summarized in Table 1) [31]. Natural biomaterials, such as collagen and chitosan, possess good biocompatibility and low immunogenicity, and chitosan [32] also has favorable osteoinductive, antibacterial, and anti-inflammatory properties. Bioceramic materials [33], such as hydroxyapatite (HA), β-tricalcium phosphate (β-TCP), and bioactive glass, have been extensively used to bolster the healing of alveolar bone in the periodontium. Bioceramic scaffolds typically provide high mechanical stability with outstanding osteoconductive and osteoinductive properties and are suitable for alveolar bone regeneration. Synthetic polyester-based polymers [31], such as polylactic acid, polyglycolic acid, poly(lactic-co-glycolic acid) (PLGA), and polycaprolactone (PCL), have many unique advantages, including highly tunable physicochemical properties, controlled biodegradation rates, and a simple and straightforward manufacturing process that allows for mass production. They are therefore commonly used as periodontal scaffold materials. It should be noted that the degradation byproducts of polyesters can be toxic but are considered safe for this application due to the minimal number of residual particles being released at a very slow rate. A scaffold, as a delivery vehicle for cells and bioactive molecules and as a means to supply space for tissue growth, can be fabricated into diverse structural forms by different manufacturing techniques. Hydrogels, microspheres, micropatterns, and monolayer or multilayer scaffolds are each manufactured differently and have their own unique merits. Generally, previous studies focused on promoting tissue generation by combining different scaffolding materials or by adding active factors. However, currently, the fabrication strategy of scaffolds has moved from [34] to multiphasic constructs [35,36,37,38,39] with biomimetic periodontal tissue to address the highly hierarchical structure of the native periodontium, in which electrospinning and 3D bioprinting technologies show tremendous potential.

Table 1 A summary of the diverse types of scaffold materials

Electrospinning technology and periodontal regeneration

Electrospinning technology [40, 41] increases the protein content through controlled fiber diameter, porosity, and ideal morphology. Better surface properties may activate some related acellular signaling and specific gene expression, stimulate cellular responses and are appropriate for the architecture of orderly aligned periodontal fibers and random mineralized matrix accumulation to form alveolar bone. With rat PDLSCs, Shang et al. [41] created electrospun scaffolds with two distinct orientations as an example of in vitro electrospun contact guiding. Contrary to randomly arranged fibers and a porosity control, where the cells took on polygonal or random geometries, the parallel and cross-aligned scaffolds showed elongated morphologies along the direction of fiber alignment. By inserting highly aligned biodegradable poly(-caprolactone)-poly(ethylene glycol) (PCE) copolymer electrospun nanofibrous mats into porous chitosan, Jiang et al. created a 3D multilayered scaffold. The aligned nanofiber-embedded scaffold guided arrangement and enhanced cell behavior in vitro. In vivo, it showed a more orderly arrangement of regenerated PDL, a more extensive formation of mature collagen fibers, and an increase in periostin expression compared to those of the other two groups.

To increase osteoconductivity, improve cellular and tissue interactions, and improve vascular growth, a biphasic scaffold was built by attaching a fused deposition modeled bone compartment coated with a calcium phosphate (CaP) layer to a melt electrospun periodontal compartment comprising larger pores. In vivo, increased bone formation, more pronounced vascularization, and greater periodontal attachment on the dentin surface were observed [43]. Furthermore, some bioactive components, such as chitosan [32], bioceramics [44] and functional polymers [45], can be easily blended into the electrospun nanofiber matrix to modulate its physical, chemical, and biological properties and its regenerative capacity. Although the histologic sections provided in Jiang et al.’s study showed good attachment and orientation of the newly formed PDL, the presence of slowly degrading PCE electrospinning material was not evident [42]. This finding may indicate that the scaffold material was moved during implantation or was only partially located at the periphery of the defect and was not necessarily placed exactly in its central portion. Limitations of electrospinning include that it is slow, costly, difficult to control, and does not yield a wide range of scalable pattern geometries. In contrast, 3D printing, with rapid production and low material wastage, has been demonstrated to be an effective approach to reconstructing periodontal tissue.

3D printing and periodontal regeneration

3D printing [46] is a method of additive manufacturing where a 3D object can be created by adding material layer by layer. It has been applied to reconstruct and regenerate organs and tissues. For irregular periodontal defects caused by periodontitis, 3D bioprinting has the benefit of being able to realize structures with specific alignments and interactions [47, 48]. Utilizing dental stem cells, Lee et al. [36] developed a region-specific scaffold with three stages of microstructures that was preoptimized for the regeneration of cementum, PDL, and alveolar bone. In 2015, Rasperini et al. [49] evaluated the first clinical case of applying a 3D-printed scaffold to a patient with a periodontal defect. Specifically, 3D-printed PCL scaffolds containing recombinant human platelet-derived growth factor-BB (rhPDGF-BB) were transplanted in single-walled bone defects in patients. The transplanted scaffolds were then exposed intraorally at 13 months. A 3-mm increase in adhesion and 75.9% stent retention were noted at 14 months with predominantly connective tissue healing and minimal evidence of bone repair. Although clinical outcomes need further improvement, the use of 3D-printed scaffolds with different spaces for PDL and alveolar bone is a worthwhile endeavor. To boost the degradation of PCL, Peng et al. [50] mixed PCL with PLGA in different ratios and found that the increase in the PLGA ratio significantly accelerated the degradation of the hybrid scaffold, smoothed the surface, and increased wettability. Furthermore, human PDLSCs (hPDLSCs) showed significantly increased adhesion, proliferation, and osteogenic capacity at 0.5 PCL/0.5 PLGA.

Advances in material technology have made 3D bioprinting possible, allowing cells to be blended into bioink and positioned in certain locations in the printed tissue, increasing the accuracy of the structural details of the inner and outer layers [51,52,53]. Several studies have been conducted to explore various bioprinting materials and optimize printing conditions using hPDLSCs [51, 52]. These studies revealed that adjusting the composition and concentration of the materials helped to improve cell viability and diffusion and to increase cell-material interactions, which promoted tissue regeneration [51, 52]. Tian et al. [51] printed sodium alginate (SA)/gelatin (Gel)/nano-HA (na-HA) composite bioscaffolds by extrusion 3D bioprinting technology and subsequently used hPDLSCs and composite hydrogels as bioinks to print SA/Gel/na-HA/hPDLSC cellular bioscaffolds. The results showed that the SA/Gel/na-HA composite hydrogels exhibited good rheological properties and a high swelling rate suitable for the printed scaffolds, which indicated that they had sufficient porosity. The compression properties of the composite bioscaffolds were also improved by the addition of na-HA. In another study, Li et al. [54] mixed hPDLSCs into bioink (1 × 107 cells/mL) and printed PDL layers on the surface of 3D-printed titanium scaffolds. Scanning electron microscopy revealed that the hPDLSCs were neatly arranged and aligned in the cell printing group compared to the cell seeding group. On Day 7 of culture, Cementum protein (CEMP1) expression in the cell printing groups was significantly higher than that in the cell seeding groups. This finding indicates that the cell printing method is more reliable than cell seeding. HE staining at 6 weeks showed new bone formation in the porous scaffold of the cell seeding group, whereas the printed group showed well-organized connective tissue between the rat cranial bone and the scaffold [54]. As a fibrous connective tissue connecting the tooth root to the alveolar bone, the PDL prevents infection and bone resorption associated with mechanical stress [55] and exists to allow dynamic action even in functionally strong osseointegrated implants [56]. 3D bioprinting produces bioimplants that produce structures similar to the PDL of natural teeth, which shows the value of 3D bioprinting in reconstructing periodontal tissue and treating tooth loss. However, current research using 3D bioprinting is still in the initial stage. The technology necessary to resolve microstructures deserves further exploration, and the issues of printing accuracy, cytocompatibility, and blood supply still need to be solved.

Tissue engineering in periodontal regeneration has evolved greatly in recent years from using only one type of cell to combinations of different cell types and from single scaffold structures to the emergence of techniques that mimic periodontal tissue structures that all hold promise for achieving periodontal regeneration in the clinic (An overview of these studies is given in Table 2). Although a single-center, randomized trial that used autologous PDLSCs-based treatment for periodontal intrabony defects showed it was safe, it still needs more rigorous clinical trials to evaluate the efficacy of this therapy [57]. However, several issues still need to be addressed for stem cell transplantation: safe and effective in vitro expansion; effective transplantation methods; long-term storage of engineered tissues; standardized procedures for sterilization, purification, and the quantitative production of cell products; age limitations for autologous cell transplantation; and the potential risks and availability of allogeneic and xenogeneic cells. Further research and understanding of stem cell physiology may facilitate the development of novel and powerful therapeutic approaches and will help realize the tremendous impact of stem cell therapies on the future of health care. A prerequisite for functional periodontal tissue regeneration is novel biomaterial technology aimed at mimicking the structure of periodontal tissue closely at the micron and nanoscale. Technologies, such as electrostatic spinning and 3D bioprinting, hold promise, but more research is needed to address the need for improved cell viability, printing accuracy, etc.

Table 2 An overview of these studies on cell-based tissue engineering

Endogenous regeneration and periodontal regeneration

Although cell therapy has shown regenerative potential in the treatment of periodontal defects, the translation of cell therapies has been hampered by challenges in production, storage, delivery to patients, optimal cell viability, stem cell immunogenicity, ethics, and safety. Some studies [58, 59] found that in periodontal regeneration, only a small percentage of green fluorescent protein-labeled transplanted cells existed in defect areas, suggesting that the transplanted stem cells may be less engaged in differentiation to repair damaged tissue. According to previous stem cell transplantation studies, transplanted cells are believed to promote tissue regeneration through two possible pathways: participating in the formation of tissues through their own proliferation and differentiation (direct contribution) or inducing the formation of new tissues in host cells through the secretion of cytokines/growth factors (indirect contribution). A new view has emerged that MSCs may affect tissue repair primarily through their paracrine factors and the stimulation of host cells, rather than through cell replacement [60, 61].

MSCs secretome derivatives, such as conditioned medium (CM), include different proteins, cytokines, growth factors, enzymes, extracellular vesicles (EVs) [62] with various proteins, such as cargo, coding and noncoding RNA, small RNAs, autophagosomes, and mitophagosomes. EVs can be further subdivided into apoptotic bodies, microparticles, and exosomes. Subsequent studies [63] have revealed that exosomes participate in cell communication, immunomodulation, promotion of cell proliferation, angiogenesis, and neuroprotection, which largely explains the extremely broad therapeutic effects previously attributed to MSCs. Secretome derivatives are gaining popularity due to their safety, low immunogenicity, and the reduced number of ethical issues involved. Furthermore, the ECM [64] is an intricate and dynamic bioenvironment with precisely regulated mechanical and biochemical properties that provides the appropriate microenvironment for cells and acts as a powerful tool for endogenous regeneration without cellular involvement. Drugs targeting signaling molecules involved in periodontitis progression, modulating osteoimmunity and promoting tissue repair and regeneration to drive endogenous regeneration may also be used as a tool for cell-free therapy.

Strategies, such as adding biological agents (growth factors, exosomes, and CM), adding well-designed or natural biomaterials (ECM) to induce cell homing and promote cell residence and differentiation, and using signaling molecules to drive endogenous regeneration, are all based on the concept of endogenous regeneration. Endogenous regeneration [65] involves creating a microenvironment suitable for the promotion of cell proliferation and differentiation and mobilizing chemokines and cytokines to promote the continuous migration and aggregation of the organism's internal stem cells to the injured region. This section describes the current state of various strategies for endogenous regeneration (Studies of this section is summarized in Table 3).

Table 3 Various strategies for endogenous regeneration in periodontal regeneration

Scaffold-based cell-free therapy

Several events need to occur in order to repair periodontal defects caused by periodontitis or other causes, including cell proliferation, migration differentiation and angiogenesis. Particular factors participating in these processes will also be needed, such as SDF-1, BMP, insulin-like growth factor (IGF-1), and tumor necrosis factor β1 (TGF-β1) [66,67,68]. Bioactive agents can be added into biomaterials by noncovalent methods such as physical capture, surface adsorption, and ion complexation [67].

However, a single growth factor has a limited ability to regenerate tissue, and an overdose of a single growth factor can cause severe inflammatory reactions [69]. Several studies revealed that the use of both angiogenic and BMP cytokines was more effective than the use of one single growth factor [70, 71]. Cytokines are mainly large molecular proteins with a short biological half-life, poor stability, and high cost, but these deficits can be overcome by modifying and packaging cytokines with scaffold materials for spatiotemporal control. Chitosan/alginate/PLGA hybrid scaffolds were designed by Duruel et al. for the coordinated and sequential administration of IGF-1 and BMP-6 [72]. Cell culture studies showed that the scaffolds induce the proliferation and osteoblastic differentiation of cementoblasts more effectively than IGF-1- and BMP-6-free chitosan scaffolds. Tan et al. [68] designed a hydrogel called NapFFY that allows the release of two bioactive factors, SDF-1 and BMP-2, from the hydrogel in a controlled, sequential and sustained manner as intended. These conditions were created by using various materials in combination with growth factors to create favorable alveolar bone, PDL, and alveolar bone growth conditions. Cellular studies showed that the scaffold aided in the differentiation of human DFCs into cementogenic, fibrous, and osteogenic tissues, and in vivo findings further supported the formation of new cementum, fibrous PDL, and alveolar bone with distinct bone trabeculae. [39].

Several growth factors are effective for tissue reconstruction in specific conditions but are not sufficient for periodontal tissue regeneration processes that involve multiple growth factors and in which diverse types and volumes of factors are involved at different times. In this way, MSCs-CM or EVs offer new possibilities.

MSCs-conditioned medium and exosomes

In recent years, MSCs-CM and exosomes have been widely used in the field of skin, cardiovascular, liver and kidney tissue regeneration due to their angiogenic, anti-inflammatory, immunomodulatory, and anti-apoptotic activities as well as their cell growth support and chemoattraction [73, 74]. Their use in periodontal tissue regeneration is gradually increasing. MSCs-CM [61] contains anti-inflammatory TGF-β1, IL-10, IL-12, IL-17E, IL-27, and proinflammatory cytokines, such as IL-1b, IL-6, IL-8, and IL-9. The final outcome can depend on how these proinflammatory and antiinflammatory cytokines are balanced. Reduced salivary CD9/CD81 exosome levels were linked to the pathogenesis of periodontitis, according to a research article [75]. In periodontitis, programmed death-ligand 1 (PD-L1) expression positively correlated with inflammation, whereas PD-L1 mRNA in salivary exosomes was enriched [76]. This finding suggests that exosomes may be involved in inflammatory signaling and periodontal disease progression. Nagata et al. [77] showed that transplantation of CM obtained from cultured PDLSCs resulted in decreased TNF-α mRNA levels in healing periodontal tissues and suppressed TNF-α mRNA levels in the monocyte/macrophage cell line RAW stimulated with IFN-γ. Qiu et al. [78] revealed that CM of both GMSCs and PDLSCs promoted periodontal regeneration in a rat periodontal defect model. Immunostaining results showed that the CM transplants decreased the expression levels of IL-1 and TNF-α and increased the levels of IL-10. Another study [79] found that TNF-α isolated from pretreated GMSCs exosomes induced polarization of anti-inflammatory M2 macrophages through the improved secretion of GMSCs exosomes and increased exosomal expression of CD73. This finding suggests that CM from PDLSCs and GMSCs has the potential to improve the microenvironment of periodontal regeneration by participating in inflammatory regulatory responses.

CM and exosomes act as carriers of growth factors and cytokines, including IGF-1, TGF-β1, VEGF, and hypoxia inducible factor-1α. These growth factors and cytokines also contribute to the recruitment of bone and vascular precursor cells, stimulate their proliferation and differentiation into osteoblasts and enhance angiogenesis, which are all key components of periodontal regeneration [80]. This notion has been validated by some studies. Human MSCs exosome-loaded collagen sponges could increase migration and proliferation through CD73-mediated adenosine receptor activation of prosurvival AKT and ERK signaling, which leads to the regeneration of periodontal tissues [81]. A study [82] indicated that conditioned SHED exosome-enhanced PDLSCs osteogenic differentiation was partly due to it carrying Wnt3a and BMP-2. Exosomes released by human SHED, according to a study from Wu et al., support the healing of alveolar bone defects by controlling angiogenesis and osteogenesis [83]. In other fields of tissue regeneration, MSCs-derived exosomes have been found to act mainly through the horizontal transfer of mRNA, miRNA, and proteins, which then alter the activity of target cells through various mechanisms [84]. The mechanisms by which exosomes promote tissue regeneration are diverse and complex, and this feature may be partly due to the large number of components they contain.

Investigating the mechanisms by which exosomes promote tissue regeneration and repair would contribute to a better understanding and use of MSCs therapy. Although a substantial number of studies have reported that exosomes from MSCs are as effective as MSCs in preserving tissue and/or promoting functional recovery after injury, it is difficult to compare efficacy due to the lack of appropriate controls, differing doses, the use of different disease endpoints, and the frequency and duration of doses. There is also the question regarding whether exosome number, instability, and freezing/thawing affect exosome potency. In addition, the status of MSCs-derived exosomes depends on the cellular origin and physiological state with different cells and different states having different effects. Finally, the standardization of production and storage of the product also requires further investigation.

Extracellular matrix

Natural tissues are a mixture of cells, growth factors, and ECM. Unlike CM or exosomes, the ECM [85] is an intricate and dynamic bioenvironment with precisely regulated mechanical and biochemical properties, providing the appropriate microenvironment for cells, influencing stem cell behavior, and participating in mechanical signaling through mechanoreceptors (e.g., integrins). ECM extraction from natural tissues is accomplished by a decellularization process that uses chemical, enzymatic, and/or physical means to remove cellular and genetic material from ECM that would cause immunogenicity while maintaining its structure, including proteins and polysaccharides. Thereafter, the decellularized ECM can be populated with the patient's cells to produce individualized tissue [64]. Decellularized ECM have been used to recreate skeletal muscle, spinal cord, bone, kidney, liver, lung, and heart tissue [86].

Tissue-derived ECM have some complex structures, some of which are difficult to separate, so incomplete decellularization occurs and creates the risk of immune rejection and disease transmission. Mixing different cell types to create tissue scaffolds can address some of the limitations of tissue-derived ECM. Selecting appropriate cells to secrete and integrate new components to form an ECM allows for their further modification (e.g., genetic) and exposes them to specific stimuli, thus enabling the creation of an ECM with desired properties. Combining porcine dental epithelial cells with human DPSCs and HUVECs and implanting them into extraction sockets of adult piglets led to decellularized tooth bud-ECM structures, which were found to consistently guide the formation of organized bioengineered teeth comparable in size to natural human teeth [87]. Decellularized PDLSCs sheets have been coupled with PCL scaffolds to promote angiogenesis and increase periodontal attachment formation [88]. Another study used decellularized PDLSCs sheets with PCL/GE nanofibers as carriers of 15-deoxy-delta(12,14)-prostaglandin J(2) nanoparticles. These nanoparticles were transplanted into periodontal defects in rats, leading to bone, dental bone, and periodontal ligament formation after 4 weeks [89]. The decellularized ECM scaffold was approved recently by the US FDA for human therapeutic applications, making ECM scaffolds a promising therapy for human tooth regeneration.

The most important aspects of the ECM are its intrinsic properties, including both biochemical and biomechanical properties. However, current methods are still some distance from this goal to achieve a complete intrinsic vital structure, but the process of producing ECM scaffolds can be modified by adding growth factors and bioactive molecules to the ECM. Combinations of different techniques for different tissues can improve the efficiency of decellularization and reduce the negative effects caused by using a single technique. Optimizing decellularization methods to utilize scaffolds from human and animal tissues and organs can help address the increasing needs of patients for clinical transplantation.

Drug therapy

Periodontitis is a disease caused by the destruction of periodontal tissue when the inflammatory response, wherein plaque serves as the initiating factor, exceeds the host's immune regulatory capacity. The conventional approach to inflammation management is surgical and nonsurgical treatment coupled with antibiotics as an adjunct to therapy, but there are limitations in drug resistance and effectiveness. Alternative therapies that target the signaling molecules that participate in the progression of periodontitis, modulate osteoimmunity and promote tissue repair and regeneration have emerged.

Teriparatide (FORTEO) [90] is an anabolic drug that consists of the bioactive portion of parathyroid hormone which activates a signaling cascade to stimulate bone formation. Clinical studies involving the systematic administration of teriparatide for periodontal tissue regeneration showed that it had a significant bone formation effect [91], which is consistent with the results of preclinical studies [92].

Sclerostin and Dickkopf-related protein 1 (DKK-1) are both antagonistic regulators of the Wnt signaling pathway that limit bone growth [93]. These proteins at higher levels in patients with periodontitis than in patients without periodontitis [94] and are expressed at significantly higher levels in rats with experimental periodontitis [95]. Romosozumab, blosozumab and BPS-804 (setrusumab) inhibit sclerostin, of which romosozumab has been approved for marketing by the FDA. Previous studies showed that a Scl-Ab increased bone mass and bone density and decreased bone resorption [96]. Taut et al. [97] discovered that 6 weeks of Scl-Ab significantly improved maxillary bone healing, as measured by linear alveolar bone loss, bone volume fraction, and tissue mineral density. Another study [98] showed that scl-Ab combined with DKK-1 antibody more dramatically enhanced alveolar process healing. As mentioned before, these studies were undertaken by means of systemic administration, which suffers from safety and unclear tissue localization. These issues can be solved by local administration. In a study comparing the effectiveness of systemically and locally administered low-dose Scl-Ab using PLGA microspheres in rats with periodontitis, it was found that systemic Scl-Ab significantly enhanced alveolar bone and cementum regeneration in comparison to locally administered Scl-Ab [99]. This finding can be attributed to the difference in dosage and characteristics of the PLGA microspheres. Consequently, for local drug administration, various factors, such as the form of the drug, the mode of transport, and the spatiotemporal controlled release of the drug, need to be addressed in a multidisciplinary manner.

Currently, the complement system is considered a pivotal immune locus for inducing and regulating various immune and inflammatory functions [100]. Studies have demonstrated that the complement system is overactivated during periodontitis and that AMY-101 (C3 inhibitor) targets its complement component (C3) with therapeutic benefit and no adverse toxicity [101]. Preclinical trials [102] and a phase IIa clinical study that was randomized, placebo-controlled, and double-blind (NCT03694444) confirmed that C3-targeted inhibition prevented gingival inflammation in patients with periodontitis. C3-targeted intervention may represent an innovative host-modulated therapy in the treatment of periodontitis.

Combination of MSCs secretome derivatives with MSCs

The ability of MSCs to multi-differentiate is key to their role, but maintaining the integrity of MSCs in the microenvironment of the transplanted region is a great challenge. Exosomes and ECM, which contain a variety of bioactive molecules, regulate functions such as inflammatory response, signal transduction, and maintenance of cell function and structure [62]. Many studies have been conducted on the application of exosomes in periodontal regeneration, but the use of cell derivatives in combination with MSCs for periodontal regeneration has not been reported. However, there are some reports of other tissue regeneration. After rabbit cartilage progenitor cell-alginate constructs were implanted subcutaneously in nude mice, the injection of exosomes derived from chondrocytes weekly might imitate the chondrogenic niche, which maintenance of cartilage stability [103].

The method for creating chondrocytes folded within hBMSCs-ECM requires significantly fewer cells than the chondrocyte cell sheet technology, and it also exhibited chondrocyte proliferation, maintenance of chondrocytic phenotype, and promotion of chondrocyte redifferentiation after expansion both in vitro and in vivo [104].The ECM derived from adipose tissue has basic components of the gradient tendon-to-bone-interface (TBI)structure, after adding the HA gradient to the scaffold, which could provide umbilical cord derived MSCs with a favorable microenvironment to regenerate the structure of the TBI [105]. The multilayered interconnected structure of periodontal tissue is similar to that of TBI, and the microenvironment provided by the extracellular matrix that favors MSCs in combination with dental stem cells may also help to reconstruct the ordered structure of the periodontium.

LL37, an antimicrobial peptide, plays a central role in wound healing, angiogenesis, and arteriogenesis and acts as an immune adjuvant [106]. One study discovered that the combination of LL37 and BMP2-modified MSCs promoted MSC-mediated increaseed the number of new blood vessel and ectopic bone formation in the LPS-induced osteolytic defects in mouse calvaria. This hints to us that LL37 can be a potential candidate drug for periodontal regeneration because of its ability to promote osteogenesis and for inhibiting bacterial growth [107].

Combining MSCs derivatives with MSCs is more effective and maximizes the potential of MSCs, but the difficulty of establishing standardized procedures for exosome and MSC isolation, purification, storage, and standardization increases accordingly. Clinical treatment plans are not static due to the diversity of the periodontal foundation of the patients, and cell and cell free therapy each have their own advantages and limitations, and either alone or in combination are applied to specific type. Further studies on the identification, signal transduction, and mechanisms of action of key active molecules in MSCs, exosomes, and ECM are needed.

Conclusion and future perspectives challenge

MSCs-based periodontal tissue engineering applications, especially the construction of a microenvironment that is conducive to periodontal regeneration through well-designed guided scaffold structures that are biomimetic of periodontal multilayer structures, have made promising progress. However, the application of MSCs still faces numerous challenges, including the safety of cell expansion in vitro, standardized production procedures for cellular products, inherent immunogenicity, precision of scaffold materials, cytocompatibility, and material degradation. Cell-free therapy with the addition of biological agents (growth factors, exosomes, and CM) to promote endogenous regeneration in organisms has partially addressed the limitations of cell therapy and contributed to understanding the mechanisms behind MSCs functions. However, the environment and state of exosome-derived cell culture strongly influence the properties of exosomes. Defining exosome activity, dose, and potency as well as effective methods of extraction and storage require additional studies. Finally, drug therapy targeting signaling molecules aids in functional periodontal regeneration.

Availability of data and materials

Not applicable.



Periodontal ligament


Mesenchymal stem cells


Dental pulp stem cells


Periodontal ligament stem cells


Stem cells from human exfoliated deciduous teeth


Gingival MSCs


Bone marrow MSCs


Extracellular matrix


Human umbilical vein endothelial cells




Poly(lactic-co-glycolic acid)






β-Tricalcium phosphate


Poly(caprolactone)-poly(ethylene glycol)


Calcium phosphate


Recombinant human platelet-derived growth factor-BB


Human PDLSCs






Sodium alginate


Cementum protein1


Tumor necrosis factor-α






Human dental pulp stem cells


Human jaw bone marrow mesenchymal stem cells


Human alveolar bone stem cells


Bone morphogenetic factor


Fibroblast growth factor


Rat bone marrow mesenchymal stem cells


Conditioned medium


Extracellular vesicles


Stromal cell derived factor-1


Insulin-like growth factor-1


Tumor necrosis factor β1


Human bone marrow mesenchymal stem cells


Programmed death-ligand 1


Dickkopf-related protein1


Sclerostin-neutralizing monoclonal antibody


Sclerostin antibody


DKK1 antibody




  1. Bartold PM, Shi ST, Gronthos S. Stem cells and periodontal regeneration. Periodontol 2000. 2006;40:164–72.

    Article  Google Scholar 

  2. Kinane DF, Stathopoulou PG, Papapanou PN. Periodontal diseases. Nat Rev Dis Primers. 2017;3:17038.

    Article  Google Scholar 

  3. Pihlstrom BL, Michalowicz BS, Johnson NW. Periodontal diseases. Lancet. 2005;366(9499):1809–20.

    Article  Google Scholar 

  4. Deas DE, Moritz AJ, Sagun RS Jr, Gruwell SF, Powell CA. Scaling and root planing vs. conservative surgery in the treatment of chronic periodontitis. Periodontol 2000. 2016;71(1):128–39.

    Article  Google Scholar 

  5. Bartold PM, Gronthos S, Ivanovski S, Fisher A, Hutmacher DW. Tissue engineered periodontal products. J Periodontal Res. 2016;51(1):1–15.

    Article  CAS  Google Scholar 

  6. Trounson A, McDonald C. Stem cell therapies in clinical trials: progress and challenges. Cell Stem Cell. 2015;17(1):11–22.

    Article  CAS  Google Scholar 

  7. Wu R-X, Xu X-Y, Wang J, He X-T, Sun H-H, Chen F-M. Biomaterials for endogenous regenerative medicine: coaxing stem cell homing and beyond. Appl Mater Today. 2018;11:144–65.

    Article  Google Scholar 

  8. Pittenger MF, Discher DE, Peault BM, Phinney DG, Hare JM, Caplan AI. Mesenchymal stem cell perspective: cell biology to clinical progress. NPJ Regen Med. 2019;4:22.

    Article  Google Scholar 

  9. Zhuang WZ, Lin YH, Su LJ, Wu MS, Jeng HY, Chang HC, et al. Mesenchymal stem/stromal cell-based therapy: mechanism, systemic safety and biodistribution for precision clinical applications. J Biomed Sci. 2021;28(1):28.

    Article  CAS  Google Scholar 

  10. Zhai Q, Dong Z, Wang W, Li B, Jin Y. Dental stem cell and dental tissue regeneration. Front Med. 2019;13(2):152–9.

    Article  Google Scholar 

  11. Magalhaes FD, Sarra G, Carvalho GL, Pedroni ACF, Marques MM, Chambrone L, et al. Dental tissue-derived stem cell sheet biotechnology for periodontal tissue regeneration: A systematic review. Arch Oral Biol. 2021;129: 105182.

    Article  CAS  Google Scholar 

  12. Ding G, Liu Y, Wang W, Wei F, Liu D, Fan Z, et al. Allogeneic periodontal ligament stem cell therapy for periodontitis in swine. Stem Cells. 2010;28(10):1829–38.

    Article  CAS  Google Scholar 

  13. Zhu W, Liang M. Periodontal ligament stem cells: current status, concerns, and future prospects. Stem Cells Int. 2015;2015: 972313.

    Article  Google Scholar 

  14. Seo BM, Miura M, Gronthos S, Bartold PM, Batouli S, Brahim J, et al. Investigation of multipotent postnatal stem cells from human periodontal ligament. Lancet. 2004;364(9429):149–55.

    Article  CAS  Google Scholar 

  15. Liu D, Xu J, Liu O, Fan Z, Liu Y, Wang F, et al. Mesenchymal stem cells derived from inflamed periodontal ligaments exhibit impaired immunomodulation. J Clin Periodontol. 2012;39(12):1174–82.

    Article  CAS  Google Scholar 

  16. Ishikawa I, Iwata T, Washio K, Okano T, Nagasawa T, Iwasaki K, et al. Cell sheet engineering and other novel cell-based approaches to periodontal regeneration. Periodontol 2000. 2009;51:220–38.

    Article  Google Scholar 

  17. Du J, Shan Z, Ma P, Wang S, Fan Z. Allogeneic bone marrow mesenchymal stem cell transplantation for periodontal regeneration. J Dent Res. 2014;93(2):183–8.

    Article  CAS  Google Scholar 

  18. Kawaguchi H, Hirachi A, Hasegawa N, Iwata T, Hamaguchi H, Shiba H, et al. Enhancement of periodontal tissue regeneration by transplantation of bone marrow mesenchymal stem cells. J Periodontol. 2004;75(9):1281–7.

    Article  Google Scholar 

  19. Mooney DJ, Vandenburgh H. Cell delivery mechanisms for tissue repair. Cell Stem Cell. 2008;2(3):205–13.

    Article  CAS  Google Scholar 

  20. Hu J, Cao Y, Xie Y, Wang H, Fan Z, Wang J, et al. Periodontal regeneration in swine after cell injection and cell sheet transplantation of human dental pulp stem cells following good manufacturing practice. Stem Cell Res Ther. 2016;7(1):130.

    Article  Google Scholar 

  21. Guo R, Wan F, Morimatsu M, Xu Q, Feng T, Yang H, et al. Cell sheet formation enhances the therapeutic effects of human umbilical cord mesenchymal stem cells on myocardial infarction as a bioactive material. Bioact Mater. 2021;6(9):2999–3012.

    Article  CAS  Google Scholar 

  22. Zhang J, Yun S, Du Y, Zannettino A, Zhang H. Hydrogel-based preparation of cell aggregates for biomedical applications. Appl Mater Today. 2020;20: 100747.

    Article  Google Scholar 

  23. Zhang J, Chan HF, Wang H, Shao D, Tao Y, Li M. Stem cell therapy and tissue engineering strategies using cell aggregates and decellularized scaffolds for the rescue of liver failure. J Tissue Eng. 2021;12:2041731420986711.

    Article  Google Scholar 

  24. Wang J, Zhang R, Shen Y, Xu C, Qi S, Lu L, et al. Recent advances in cell sheet technology for periodontal regeneration. Curr Stem Cell Res Ther. 2014;9(3):162–73.

    Article  CAS  Google Scholar 

  25. Iwata T, Washio K, Yoshida T, Ishikawa I, Ando T, Yamato M, et al. Cell sheet engineering and its application for periodontal regeneration. J Tissue Eng Regen Med. 2015;9(4):343–56.

    Article  CAS  Google Scholar 

  26. Washio K, Iwata T, Mizutani M, Ando T, Yamato M, Okano T, et al. Assessment of cell sheets derived from human periodontal ligament cells: a pre-clinical study. Cell Tissue Res. 2010;341(3):397–404.

    Article  Google Scholar 

  27. Yang Z, Jin F, Zhang X, Ma D, Han C, Huo N, et al. Tissue engineering of cementum/periodontal-ligament complex using a novel three-dimensional pellet cultivation system for human periodontal ligament stem cells. Tissue Eng Part C Methods. 2009;15(4):571–81.

    Article  CAS  Google Scholar 

  28. Raju R, Oshima M, Inoue M, Morita T, Huijiao Y, Waskitho A, et al. Three-dimensional periodontal tissue regeneration using a bone-ligament complex cell sheet. Sci Rep. 2020;10(1):1656.

    Article  CAS  Google Scholar 

  29. Zhang H, Liu S, Zhu B, Xu Q, Ding Y, Jin Y. Composite cell sheet for periodontal regeneration: crosstalk between different types of MSCs in cell sheet facilitates complex periodontal-like tissue regeneration. Stem Cell Res Ther. 2016;7(1):168.

    Article  Google Scholar 

  30. Panduwawala CP, Zhan X, Dissanayaka WL, Samaranayake LP, Jin L, Zhang C. In vivo periodontal tissue regeneration by periodontal ligament stem cells and endothelial cells in three-dimensional cell sheet constructs. J Periodontal Res. 2017;52(3):408–18.

    Article  CAS  Google Scholar 

  31. d’Avanzo N, Bruno MC, Giudice A, Mancuso A, Gaetano F, Cristiano MC, et al. Influence of materials properties on bio-physical features and effectiveness of 3D-scaffolds for periodontal regeneration. Molecules. 2021;26(6):1643.

    Article  Google Scholar 

  32. Niu XL, Wang LF, Xu MJ, Qin M, Zhao LQ, Wei Y, et al. Electrospun polyamide-6/chitosan nanofibers reinforced nano-hydroxyapatite/polyamide-6 composite bilayered membranes for guided bone regeneration. Carbohyd Polym. 2021;260: 117769.

    Article  CAS  Google Scholar 

  33. Woo HN, Cho YJ, Tarafder S, Lee CH. The recent advances in scaffolds for integrated periodontal regeneration. Bioact Mater. 2021;6(10):3328–42.

    Article  CAS  Google Scholar 

  34. Dan HX, Vaquette C, Fisher AG, Hamlet SM, Xiao Y, Hutmacher DW, et al. The influence of cellular source on periodontal regeneration using calcium phosphate coated polycaprolactone scaffold supported cell sheets. Biomaterials. 2014;35(1):113–22.

    Article  CAS  Google Scholar 

  35. Puppi D, Migone C, Grassi L, Pirosa A, Maisetta G, Batoni G, et al. Integrated three-dimensional fiber/hydrogel biphasic scaffolds for periodontal bone tissue engineering. Polym Int. 2016;65(6):631–40.

    Article  CAS  Google Scholar 

  36. Lee CH, Hajibandeh J, Suzuki T, Fan A, Shang P, Mao JJ. Three-dimensional printed multiphase scaffolds for regeneration of periodontium complex. Tissue Eng Pt A. 2014;20(7–8):1342–51.

    Article  CAS  Google Scholar 

  37. Vaquette C, Fan W, Xiao Y, Hamlet S, Hutmacher DW, Ivanovski S. A biphasic scaffold design combined with cell sheet technology for simultaneous regeneration of alveolar bone/periodontal ligament complex. Biomaterials. 2012;33(22):5560–73.

    Article  CAS  Google Scholar 

  38. Varoni EM, Vijayakumar S, Canciani E, Cochis A, De Nardo L, Lodi G, et al. Chitosan-based trilayer scaffold for multitissue periodontal regeneration. J Dent Res. 2018;97(3):303–11.

    Article  CAS  Google Scholar 

  39. Sowmya S, Mony U, Jayachandran P, Reshma S, Kumar RA, Arzate H, et al. Tri-layered nanocomposite hydrogel scaffold for the concurrent regeneration of cementum, periodontal ligament, and alveolar bone. Adv Healthc Mater. 2017;6(7):1601251.

    Article  Google Scholar 

  40. Zhuang Y, Lin K, Yu H. Advance of nano-composite electrospun fibers in periodontal regeneration. Front Chem. 2019;7:495.

    Article  CAS  Google Scholar 

  41. Shang SH, Yang F, Cheng XR, Walboomers XF, Jansen JA. The effect of electrospun fibre alignment on the behaviour of rat periodontal ligament cells. Eur Cells Mater. 2010;19:180–92.

    Article  CAS  Google Scholar 

  42. Jiang WL, Li L, Zhang D, Huang SS, Jing Z, Wu YK, et al. Incorporation of aligned PCL-PEG nanofibers into porous chitosan scaffolds improved the orientation of collagen fibers in regenerated periodontium. Acta Biomater. 2015;25:240–52.

    Article  CAS  Google Scholar 

  43. Vaquette C, Ivanovski S, Hamlet SM, Hutmacher DW. Effect of culture conditions and calcium phosphate coating on ectopic bone formation. Biomaterials. 2013;34(22):5538–51.

    Article  CAS  Google Scholar 

  44. Wang YF, Liu YN, Zhang XP, Liu N, Yu XX, Gao MH, et al. Engineering electrospun nanofibers for the treatment of oral diseases. Front Chem. 2021;9: 797523.

    Article  CAS  Google Scholar 

  45. Qian YZ, Zhou XF, Zhang FM, Diekwisch TGH, Luan XH, Yang JX. Triple PLGA/PCL scaffold modification including silver impregnation, collagen coating, and electrospinning significantly improve biocompatibility, antimicrobial, and osteogenic properties for orofacial tissue regeneration. ACS Appl Mater Inter. 2019;11(41):37381–96.

    Article  CAS  Google Scholar 

  46. Ngo TD, Kashani A, Imbalzano G, Nguyen KTQ, Hui D. Additive manufacturing (3D printing): a review of materials, methods, applications and challenges. Compos Part B-Eng. 2018;143:172–96.

    Article  CAS  Google Scholar 

  47. Park CH, Rios HF, Jin QM, Bland ME, Flanagan CL, Hollister SJ, et al. Biomimetic hybrid scaffolds for engineering human tooth-ligament interfaces. Biomaterials. 2010;31(23):5945–52.

    Article  CAS  Google Scholar 

  48. Park CH, Rios HF, Jin QM, Sugai JV, Padial-Molina M, Taut AD, et al. Tissue engineering bone-ligament complexes using fiber-guiding scaffolds. Biomaterials. 2012;33(1):137–45.

    Article  CAS  Google Scholar 

  49. Rasperini G, Pilipchuk SP, Flanagan CL, Park CH, Pagni G, Hollister SJ, et al. 3D-printed bioresorbable scaffold for periodontal repair. J Dent Res. 2015;94(9):153s-s157.

    Article  CAS  Google Scholar 

  50. Peng CX, Zheng JX, Chen DR, Zhang XQ, Deng LD, Chen ZY, et al. Response of hPDLSCs on 3D printed PCL/PLGA composite scaffolds in vitro. Mol Med Rep. 2018;18(2):1335–44.

    CAS  Google Scholar 

  51. Tian Y, Liu M, Liu Y, Shi C, Wang Y, Liu T, et al. The performance of 3D bioscaffolding based on a human periodontal ligament stem cell printing technique. J Biomed Mater Res A. 2021;109(7):1209–19.

    Article  CAS  Google Scholar 

  52. Thattaruparambil Raveendran N, Vaquette C, Meinert C, Samuel Ipe D, Ivanovski S. Optimization of 3D bioprinting of periodontal ligament cells. Dent Mater. 2019;35(12):1683–94.

    Article  CAS  Google Scholar 

  53. Ma Y, Ji Y, Huang G, Ling K, Zhang X, Xu F. Bioprinting 3D cell-laden hydrogel microarray for screening human periodontal ligament stem cell response to extracellular matrix. Biofabrication. 2015;7(4): 044105.

    Article  Google Scholar 

  54. Lee UL, Yun S, Cao HL, Ahn G, Shim JH, Woo SH, et al. Bioprinting on 3D printed titanium scaffolds for periodontal ligament regeneration. Cells-Basel. 2021;10(6):1337.

    Article  CAS  Google Scholar 

  55. Nakajima K, Oshima M, Yamamoto N, Tanaka C, Koitabashi R, Inoue T, et al. Development of a functional biohybrid implant formed from periodontal tissue utilizing bioengineering technology. Tissue Eng Part A. 2016;22(17–18):1108–15.

    Article  Google Scholar 

  56. Giannobile WV. Getting to the root of dental implant tissue engineering. J Clin Periodontol. 2010;37(8):747–9.

    Google Scholar 

  57. Chen FM, Gao LN, Tian BM, Zhang XY, Zhang YJ, Dong GY, et al. Treatment of periodontal intrabony defects using autologous periodontal ligament stem cells: a randomized clinical trial. Stem Cell Res Ther. 2016;7:33.

    Article  Google Scholar 

  58. Hasegawa N, Kawaguchi H, Hirachi A, Takeda K, Mizuno N, Nishimura M, et al. Behavior of transplanted bone marrow-derived mesenchymal stem cells in periodontal defects. J Periodontol. 2006;77(6):1003–7.

    Article  Google Scholar 

  59. Yu N, Bronckers AL, Oortgiesen DA, Yan X, Jansen JA, Yang F, et al. Periodontal cell implantation contributes to the regeneration of the periodontium in an indirect way. Tissue Eng Part A. 2015;21(1–2):166–73.

    Article  CAS  Google Scholar 

  60. Phinney DG, Pittenger MF. Concise review: MSC-derived exosomes for cell-free therapy (vol 35, pg 851, 2017). Stem Cells. 2017;35(9):2103.

    Article  Google Scholar 

  61. Vizoso FJ, Eiro N, Cid S, Schneider J, Perez-Fernandez R. Mesenchymal stem cell secretome: toward cell-free therapeutic strategies in regenerative medicine. Int J Mol Sci. 2017;18(9):1852.

    Article  Google Scholar 

  62. Bjorge IM, Kim SY, Mano JF, Kalionis B, Chrzanowski W. Extracellular vesicles, exosomes and shedding vesicles in regenerative medicine—a new paradigm for tissue repair. Biomater Sci. 2017;6(1):60–78.

    Article  CAS  Google Scholar 

  63. Skotland T, Sagini K, Sandvig K, Llorente A. An emerging focus on lipids in extracellular vesicles. Adv Drug Deliv Rev. 2020;159:308–21.

    Article  CAS  Google Scholar 

  64. Gilpin A, Yang Y. Decellularization strategies for regenerative medicine: from processing techniques to applications. Biomed Res Int. 2017;2017:9831534.

    Article  Google Scholar 

  65. Yin Y, Li X, He XT, Wu RX, Sun HH, Chen FM. Leveraging stem cell homing for therapeutic regeneration. J Dent Res. 2017;96(6):601–9.

    Article  CAS  Google Scholar 

  66. Carreira AC, Lojudice FH, Halcsik E, Navarro RD, Sogayar MC, Granjeiro JM. Bone morphogenetic proteins: facts, challenges, and future perspectives. J Dent Res. 2014;93(4):335–45.

    Article  CAS  Google Scholar 

  67. Chen FM, Shelton RM, Jin Y, Chapple IL. Localized delivery of growth factors for periodontal tissue regeneration: role, strategies, and perspectives. Med Res Rev. 2009;29(3):472–513.

    Article  CAS  Google Scholar 

  68. Tan J, Zhang M, Hai Z, Wu C, Lin J, Kuang W, et al. Sustained release of two bioactive factors from supramolecular hydrogel promotes periodontal bone regeneration. ACS Nano. 2019;13(5):5616–22.

    Article  CAS  Google Scholar 

  69. Perri B, Cooper M, Lauryssen C, Anand N. Adverse swelling associated with use of rh-BMP-2 in anterior cervical discectomy and fusion: a case study. Spine J. 2007;7(2):235–9.

    Article  Google Scholar 

  70. Patel ZS, Young S, Tabata Y, Jansen JA, Wong ME, Mikos AG. Dual delivery of an angiogenic and an osteogenic growth factor for bone regeneration in a critical size defect model. Bone. 2008;43(5):931–40.

    Article  CAS  Google Scholar 

  71. Kuttappan S, Mathew D, Jo JI, Tanaka R, Menon D, Ishimoto T, et al. Dual release of growth factor from nanocomposite fibrous scaffold promotes vascularisation and bone regeneration in rat critical sized calvarial defect. Acta Biomater. 2018;78:36–47.

    Article  CAS  Google Scholar 

  72. Duruel T, Cakmak AS, Akman A, Nohutcu RM, Gumusderelioglu M. Sequential IGF-1 and BMP-6 releasing chitosan/alginate/PLGA hybrid scaffolds for periodontal regeneration. Int J Biol Macromol. 2017;104(Pt A):232–41.

    Article  CAS  Google Scholar 

  73. Madrigal M, Rao KS, Riordan NH. A review of therapeutic effects of mesenchymal stem cell secretions and induction of secretory modification by different culture methods. J Transl Med. 2014;12:260.

    Article  Google Scholar 

  74. Ratajczak MZ, Kucia M, Jadczyk T, Greco NJ, Wojakowski W, Tendera M, et al. Pivotal role of paracrine effects in stem cell therapies in regenerative medicine: can we translate stem cell-secreted paracrine factors and microvesicles into better therapeutic strategies? Leukemia. 2012;26(6):1166–73.

    Article  CAS  Google Scholar 

  75. Tobon-Arroyave SI, Celis-Mejia N, Cordoba-Hidalgo MP, Isaza-Guzman DM. Decreased salivary concentration of CD9 and CD81 exosome-related tetraspanins may be associated with the periodontal clinical status. J Clin Periodontol. 2019;46(4):470–80.

    Article  CAS  Google Scholar 

  76. Yu J, Lin Y, Xiong X, Li K, Yao Z, Dong H, et al. Detection of exosomal PD-L1 RNA in saliva of patients with periodontitis. Front Genet. 2019;10:202.

    Article  CAS  Google Scholar 

  77. Nagata M, Iwasaki K, Akazawa K, Komaki M, Yokoyama N, Izumi Y, et al. Conditioned medium from periodontal ligament stem cells enhances periodontal regeneration. Tissue Eng Part A. 2017;23(9–10):367–77.

    Article  CAS  Google Scholar 

  78. Qiu J, Wang X, Zhou H, Zhang C, Wang Y, Huang J, et al. Enhancement of periodontal tissue regeneration by conditioned media from gingiva-derived or periodontal ligament-derived mesenchymal stem cells: a comparative study in rats. Stem Cell Res Ther. 2020;11(1):42.

    Article  CAS  Google Scholar 

  79. Nakao Y, Fukuda T, Zhang Q, Sanui T, Shinjo T, Kou X, et al. Exosomes from TNF-alpha-treated human gingiva-derived MSCs enhance M2 macrophage polarization and inhibit periodontal bone loss. Acta Biomater. 2021;122:306–24.

    Article  CAS  Google Scholar 

  80. Diomede F, Marconi GD, Fonticoli L, Pizzicanella J, Merciaro I, Bramanti P, et al. Functional relationship between osteogenesis and angiogenesis in tissue regeneration. Int J Mol Sci. 2020;21(9):3242.

    Article  CAS  Google Scholar 

  81. Chew JRJ, Chuah SJ, Teo KYW, Zhang S, Lai RC, Fu JH, et al. Mesenchymal stem cell exosomes enhance periodontal ligament cell functions and promote periodontal regeneration. Acta Biomater. 2019;89:252–64.

    Article  CAS  Google Scholar 

  82. Wang M, Li J, Ye Y, He S, Song J. SHED-derived conditioned exosomes enhance the osteogenic differentiation of PDLSCs via Wnt and BMP signaling in vitro. Differentiation. 2020;111:1–11.

    Article  CAS  Google Scholar 

  83. Wu J, Chen L, Wang R, Song Z, Shen Z, Zhao Y, et al. Exosomes secreted by stem cells from human exfoliated deciduous teeth promote alveolar bone defect repair through the regulation of angiogenesis and osteogenesis. ACS Biomater Sci Eng. 2019;5(7):3561–71.

    Article  CAS  Google Scholar 

  84. Thery C. Exosomes: secreted vesicles and intercellular communications. F1000 Biol Rep. 2011;3:15.

    Article  Google Scholar 

  85. Taylor DA, Sampaio LC, Ferdous Z, Gobin AS, Taite LJ. Decellularized matrices in regenerative medicine. Acta Biomater. 2018;74:74–89.

    Article  CAS  Google Scholar 

  86. Saldin LT, Cramer MC, Velankar SS, White LJ, Badylak SF. Extracellular matrix hydrogels from decellularized tissues: structure and function. Acta Biomater. 2017;49:1–15.

    Article  CAS  Google Scholar 

  87. Zhang W, Vazquez B, Oreadi D, Yelick PC. Decellularized tooth bud scaffolds for tooth regeneration. J Dent Res. 2017;96(5):516–23.

    Article  CAS  Google Scholar 

  88. Farag A, Hashimi SM, Vaquette C, Bartold PM, Hutmacher DW, Ivanovski S. The effect of decellularized tissue engineered constructs on periodontal regeneration. J Clin Periodontol. 2018;45(5):586–96.

    Article  CAS  Google Scholar 

  89. Jiang Y, Liu JM, Huang JP, Lu KX, Sun WL, Tan JY, et al. Regeneration potential of decellularized periodontal ligament cell sheets combined with 15-deoxy-Delta(12,14)-prostaglandin J2 nanoparticles in a rat periodontal defect. Biomed Mater. 2021;16(4): 045008.

    Article  CAS  Google Scholar 

  90. Fujita T, Meguro T, Fukuyama R, Nakamuta H, Koida M. New signaling pathway for parathyroid hormone and cyclic AMP action on extracellular-regulated kinase and cell proliferation in bone cells. Checkpoint of modulation by cyclic AMP. J Biol Chem. 2002;277(25):22191–200.

    Article  CAS  Google Scholar 

  91. Bashutski JD, Eber RM, Kinney JS, Benavides E, Maitra S, Braun TM, et al. Teriparatide and osseous regeneration in the oral cavity. N Engl J Med. 2010;363(25):2396–405.

    Article  CAS  Google Scholar 

  92. Zou Z, Wang L, Zhou Z, Sun Q, Liu D, Chen Y, et al. Simultaneous incorporation of PTH(1–34) and nano-hydroxyapatite into Chitosan/Alginate Hydrogels for efficient bone regeneration. Bioact Mater. 2021;6(6):1839–51.

    Article  CAS  Google Scholar 

  93. Kim JH, Lee DE, Woo GH, Cha JH, Bak EJ, Yoo YJ. Osteocytic sclerostin expression in alveolar bone in rats with diabetes mellitus and ligature-induced periodontitis. J Periodontol. 2015;86(8):1005–11.

    Article  Google Scholar 

  94. Napimoga MH, Nametala C, da Silva FL, Miranda TS, Bossonaro JP, Demasi AP, et al. Involvement of the Wnt-beta-catenin signalling antagonists, sclerostin and dickkopf-related protein 1, in chronic periodontitis. J Clin Periodontol. 2014;41(6):550–7.

    Article  CAS  Google Scholar 

  95. Duan P, Bonewald LF. The role of the wnt/beta-catenin signaling pathway in formation and maintenance of bone and teeth. Int J Biochem Cell Biol. 2016;77(Pt A):23–9.

    Article  CAS  Google Scholar 

  96. Maeda K, Kobayashi Y, Koide M, Uehara S, Okamoto M, Ishihara A, et al. The regulation of bone metabolism and disorders by Wnt signaling. Int J Mol Sci. 2019;20(22):5525.

    Article  CAS  Google Scholar 

  97. Taut AD, Jin Q, Chung JH, Galindo-Moreno P, Yi ES, Sugai JV, et al. Sclerostin antibody stimulates bone regeneration after experimental periodontitis. J Bone Miner Res. 2013;28(11):2347–56.

    Article  CAS  Google Scholar 

  98. Liu M, Kurimoto P, Zhang J, Niu QT, Stolina M, Dechow PC, et al. Sclerostin and DKK1 inhibition preserves and augments alveolar bone volume and architecture in rats with alveolar bone loss. J Dent Res. 2018;97(9):1031–8.

    Article  CAS  Google Scholar 

  99. Yao Y, Kauffmann F, Maekawa S, Sarment LV, Sugai JV, Schmiedeler CA, et al. Sclerostin antibody stimulates periodontal regeneration in large alveolar bone defects. Sci Rep. 2020;10(1):16217.

    Article  CAS  Google Scholar 

  100. Hajishengallis G, Reis ES, Mastellos DC, Ricklin D, Lambris JD. Novel mechanisms and functions of complement. Nat Immunol. 2017;18(12):1288–98.

    Article  CAS  Google Scholar 

  101. Hajishengallis G, Kajikawa T, Hajishengallis E, Maekawa T, Reis ES, Mastellos DC, et al. Complement-dependent mechanisms and interventions in periodontal disease. Front Immunol. 2019;10:406.

    Article  CAS  Google Scholar 

  102. Maekawa T, Abe T, Hajishengallis E, Hosur KB, DeAngelis RA, Ricklin D, et al. Genetic and intervention studies implicating complement C3 as a major target for the treatment of periodontitis. J Immunol. 2014;192(12):6020–7.

    Article  CAS  Google Scholar 

  103. Chen Y, Xue K, Zhang X, Zheng Z, Liu K. Exosomes derived from mature chondrocytes facilitate subcutaneous stable ectopic chondrogenesis of cartilage progenitor cells. Stem Cell Res Ther. 2018;9(1):318.

    Article  CAS  Google Scholar 

  104. Yang Y, Lin H, Shen H, Wang B, Lei G, Tuan RS. Mesenchymal stem cell-derived extracellular matrix enhances chondrogenic phenotype of and cartilage formation by encapsulated chondrocytes in vitro and in vivo. Acta Biomater. 2018;69:71–82.

    Article  CAS  Google Scholar 

  105. Yea JH, Bae TS, Kim BJ, Cho YW, Jo CH. Regeneration of the rotator cuff tendon-to-bone interface using umbilical cord-derived mesenchymal stem cells and gradient extracellular matrix scaffolds from adipose tissue in a rat model. Acta Biomater. 2020;114:104–16.

    Article  CAS  Google Scholar 

  106. Ramos R, Silva JP, Rodrigues AC, Costa R, Guardao L, Schmitt F, et al. Wound healing activity of the human antimicrobial peptide LL37. Peptides. 2011;32(7):1469–76.

    Article  CAS  Google Scholar 

  107. Liu Z, Yuan X, Liu M, Fernandes G, Zhang Y, Yang S, et al. Antimicrobial peptide combined with BMP2-modified mesenchymal stem cells promotes calvarial repair in an osteolytic model. Mol Ther. 2018;26(1):199–207.

    Article  CAS  Google Scholar 

Download references


The authors thank American Journal Experts (AJE) for English language editing.


This work was supported by the financial support of National Key Research and Development Program of China (2017YFA0104800) and Natural Science Foundation of Sichuan Province (2022NSFSC1540).

Author information

Authors and Affiliations



XW wrote the manuscript and designed the figure; XW, JC and WT reviewed edited the manuscript. All authors read and approved the final manuscript.

Corresponding authors

Correspondence to Jinlong Chen or Weidong Tian.

Ethics declarations

Ethics approval and consent to participate

Not applicable.

Consent for publication

Not applicable.

Competing interests

The authors declare that they have no conflict of interest.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit The Creative Commons Public Domain Dedication waiver ( applies to the data made available in this article, unless otherwise stated in a credit line to the data.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Wang, X., Chen, J. & Tian, W. Strategies of cell and cell-free therapies for periodontal regeneration: the state of the art. Stem Cell Res Ther 13, 536 (2022).

Download citation

  • Received:

  • Accepted:

  • Published:

  • DOI: