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SYSTEMATIC REVIEW
Year : 2022 | Volume
: 13
| Issue : 4 | Page : 111-116
Efficacy of Application of Periodontal Ligament Stem Cells in Bone Regeneration: A Systematic Review of Animal Studies
Pardis Aghandeh1, Farnaz Kouhestani2, Fatemeh Isamorad1, Solmaz Akbari3, Behrad Tanbakuchi4, Saeed Reza Motamedian5
1 School of Dentistry, Tehran University of Medical Sciences, Tehran, Iran
2 Department of Periodontics, School of Dentistry, Tehran University of Medical Sciences, Tehran, Iran
3 Department of Periodontics, School of Dentistry, Tehran University of Medical Sciences, Tehran; Dental Implant Research Center, Dentistry Research Institute, Tehran, Iran
4 Department of Orthodontics, School of Dentistry, Tehran University of Medical Sciences, Tehran, Iran
5 Dentofacial Deformities Research Center, Research Institute of Dental Sciences, Shahid Beheshti University of Medical Sciences, Tehran; Department of Orthodontics, School of Dentistry, Shahid Beheshti University of Medical Sciences, Tehran, Iran
Correspondence Address:
Farnaz Kouhestani
Tehran University of Medical Sciences Faculty of Dentistry, North Karegar St, Tehran
Iran
Source of Support: None, Conflict of Interest: None
CheckDOI: 10.4103/denthyp.denthyp_136_22
Introduction: We aimed to systematically review the animal studies that have investigated the efficacy of periodontal ligament stem cells (PDLSCs) in bone regeneration. Methods: Web of science, Scopus, PubMed, Embase, Cochrane Library, and ProQuest were searched to September 2022 for animal studies investigating bone regeneration using PDLSCs. Results: Twenty studies were included. Calvaria and alveolar defects were treated with stem cells. The cells were mainly carried with hydrogel, hydroxyapatite, and collagen scaffolds. PDLSCs promoted osteogenesis compared with cell-free scaffolds, except in one study where biphasic calcium phosphate block scaffolds alone or with Bone morphogenic protein 2 had superior outcomes in calvaria defects. Controversial results were reported when comparing the osteogenic potential of PDLSCs and bone marrow stem cells. Two studies reported higher potential in BMSCs, and two others reported higher bone formation and more bone quality in PDLSCs. Conclusion: With the limitations of this study, PDLSCs might have promising potential to accelerate bone regeneration in artificial defects; however, due to high heterogeneity in the outcomes of the present studies, before moving forward to human experiments, further preclinical in vivo studies are needed.
Keywords: Animal study, bone regeneration, periodontal ligament, stem cells tissue engineering, systematic review
Bone is a multi-functional organ that protects the body and provides structure and mechanical support to the body.[1] Bone defects in the oral cavity are very diverse in terms of extent and etiology. These defects range from localized alveolar bone resorption due to periodontal disease to extensive bone atrophy resulting from various syndromes, traumatic injuries, and surgical removal of benign or malignant bone lesions of the oral cavity.[2] Bone repair and regeneration are biological events that include several cell types and signaling pathways in a temporal and spatial sequence. Although the exact mechanisms that regulate bone regeneration at the deepest biomolecular levels are not yet understood, various methods for bone regeneration have been proposed.[1] Autografts have been commonly used for bone regeneration. In addition, studies have shown the successful implementation of other sources, including allografts and xenografts.[3] Various problems have been reported with these treatments. Limited access to cell sources, donor site morbidity, and high bone resorption probability are problems associated with autografts.[3] On the other hand, lack of sufficient cell population, reduced osteoinductive properties, and extended healing time are drawbacks of allografts and xenografts.[4]
In tissue engineering, stem cells are desirable to researchers because of their unique properties.[5] When stimulated by internal and external signals, these undifferentiated cells can proliferate and differentiate into other cells.[4] Mesenchymal stem cells (MSCs) have osteogenic differentiation ability and high proliferative and immunomodulatory capabilities, making them an excellent source for bone regeneration and repair.[6] The cell-based approach to repairing bone lesions involves using MSCs, which can come from a wide range of tissues such as bone marrow, adipose tissue, umbilical cord, lungs, or tooth tissue.[7] Bone marrow-derived mesenchymal stem cells (BMMSCs) are considered the gold standard for bone-regenerating cell-based therapies.[8] However, because of the difficulty in accessing sufficient cell numbers and the pain and complications of bone marrow removal, researchers are looking for other sources for MSCs.[9] One potential identified source is oral tissues such as dental pulp.[10] Multi-potential stem cells, first extracted from periodontal ligament (PDL) by Seo et al.[11] in 2004, have similar properties to BMMSCs and can differentiate into adipocytes, osteoblasts, and chondrocytes under certain conditions.
Recently, some studies have evaluated bone regeneration based on PDLSCs. In 2011, Wang et al.[12] reported that BMSCs and PDLSCs have similar mineralized tissue formation in 6-mm calvaria defects in nude mice. However, subcutaneous transplantation of BMSCs caused more bone formation than PDLSCs.[13] Although previous systematic reviews have reported that the use of these cells in periodontal bone regeneration was associated with promising results,[14],[15] none have evaluated their bone regenerative potentials. Considering the controversy and uncertainty of studies using PDLSCs for bone regeneration, the current study was performed to systematically review studies that have investigated the use of PDLSCs in treating bone lesions in animals.
MethodsProtocol
This study aimed to systematically review the existing animal studies related to the use of PDLSCs for bone regeneration. The study is reported based on Preferred Reporting Items for Systematic Reviews and Meta-Analayses (PRISMA-ScR) guideline.[16]
Eligibility criteria
This systematic review answers the following PICO: (P) critical-sized defects in any animal, (I) bone regeneration using periodontal ligament stem cells, (C) with a control group (including untreated defect or empty scaffold defect), and (O) bone regeneration. Animal studies that had examined the use and effectiveness of PDLSCs for bone regeneration and were published in English were included. No time limitation was applied. Narrative reviews, commentaries and opinions, in vitro studies, non-PDLSC studies, and studies that did not report bone regeneration outcomes were excluded. Also, studies that did not report flow cytometry results to differentiate PDLSCs were excluded.
Information sources and search
Searches were performed in Web of science, Scopus, PubMed, Embase, Cochrane Library, as well as ProQuest for thesis and Gray Literature, in English, without any time limit. The final search was executed in September 2022. The bibliographies of all included papers were manually searched for relevant studies. Search terms were tailored for each database. Supplementary Table 1[17] lists the search terms for each database.
Selection of sources of evidence
The article selection process was completed by two calibrated authors (P.A. and F.K.) independently. The initial screening was done based on the titles and abstracts of the studies. Then, the articles’ full texts were received and carefully evaluated according to the inclusion and exclusion criteria. Imported papers’ references were also carefully searched for relevant studies. At each step, any disagreement was resolved by discussion using brainstorming method.
Data items
Relevant data were extracted from the studies according to the objectives of this study, independently by two reviewers (P.A. and F.I.). Data items included author and information of the study, demographic characteristics of samples, type and location of the bone lesion, treatment of intervention group, control group treatments, scaffold type or growth factor or membrane, follow-up period, tests performed, results in bone regeneration, and complications.
Risk of bias (RoB)
Included studies were evaluated using the modified Systematic Review Center for Laboratory Animal Experimentation[18] RoB tool, blindly and independently by two reviewers (P.A. and F.K.). The expressions yes, no, and unspecified were used for low risk, high risk, and lack of sufficient data to judge regarding 10 various domains. The assessment of each study was done independently.
Synthesis of results
Final data were reported qualitatively. Due to the heterogeneity of samples, assessment methods, and reported outcomes, meta-analysis was not applicable.
Statistical methods
Inter-rater agreement assessed via linear weighted Cohen’s Kappa using R software (R Foundation for Statistical Computing, Vienna, Austria).
ResultsSelection of sources of evidence
Electronic search in Web of science (482), Scopus (637), PubMed (1507), Embase (964), Cochrane Library (37), and ProQuest (1075) led to a total of 4702 articles. After duplicates removal, 2688 articles remained. Then, 2515 articles with titles and abstracts unrelated to the study were removed. Finally, by reviewing the full text of the 173 remaining articles, 20 articles were included in our study [Figure 1]. Data extracted from the selected studies are presented in Supplementary Table 2.[17]
Figure 1 Flow diagram of studies selection according to PRISMA guidelines.Characteristics of sources of evidence
In the included studies, the interventions were on mice (n = 5), rats (n = 10), minipig (n = 2), rabbit (n = 1), and beagle dogs (n = 2). Human extracted teeth (n = 14), beagle dogs (n = 2), minipigs (n = 2), rats (n = 1), and rabbits (n = 1) were used for harvesting PDLSCs. Defects in the studies were categorized as subcutaneous (n = 1), alveolar defects (n = 5), and calvarium defects (n = 14). The included studies mostly used hydrogel, hydroxyapatite, and collagen as carriers. Other carriers were: TGF-β3/RHC/CS freeze-dried sponge, Matrigel, zein/gelatin, polyethyleneimine, Evo, and Furamatrix, each used by only one study. The follow-up period in the studies was from 2 to 17 weeks. Regarding the measurement tools, micro-CT (n = 3), immunohistochemical staining (IHC) (n = 5), western blotting (n = 2), and a combination of micro-CT and IHC (n = 10) were used.
Critical appraisal within sources of evidence
Most of the included studies had unclear risks in the domains of blinding against performance and detection bias. All studies had low risks regarding incomplete outcome data and selective outcome reporting. Unclear risks of bias in the domain of allocation concealment were observed in more than 50% of the included studies. [Figure 2] illustrates the critical appraisal of the included studies. Linear weighted Kappa value showed very good agreement between reviewers regarding assessment of risk of bias (k = 0.92, 95% confidence interval: 0.87–0.97).
Results of individual sources of evidence
Among the studies that evaluated the calvarium defects, except in Yi et al. study,[19] PDLSC-riched groups improved bone regeneration compared to their controls (including no intervention and scaffold-only treatments). These advantages were in various criteria including new bone formation,[20],[21],[22],[23],[24],[25],[26],[27] osteocalcin expression,[21],[22] Runx2 expression,[21] collagen-1 expression,[26] bone morphogenetic protein 2 (BMP-2) expression,[28],[29] osteoblast-like cells arrangement,[30] total bone value,[23] bone/tissue volume ratio.[24],[31] However, Yi et al.[19] showed that the treatment of critical-size bone defect biphasic calcium phosphate block scaffolds alone or in conjugation with BMP-2 had better outcomes compared with PDLSC-treated scaffolds. When PDLSCs compared with BMMSCs, the results were controversial. Kadkhoda et al.[27] and Moshaverinia et al.[21] reported that the new bone formation of BMMSCs groups was more than PDLSCs groups. However, Yu et al.[24] reported that PDLSC-treated Bio-Oss has better outcomes regarding bone regeneration.
Regarding alveolar defects studies, the results were almost similar. New bone formation[32],[33],[34],[35] and bone/tissue volume ratio[32],[36],[37] increased using PDLSC. Lee et al.[35] reported that adding BMP-2 to the scaffold can increase the regeneration capacity of the stem cells, Kim et al.[34] reported that in comparison to PDLSCs, the regenerated bone quality was better when BMMSCs were used.
DiscussionPDLSCs exhibit MSC-like features and have the potential to develop into osteoblasts, chondrocytes, adipocytes, and other lineages such as neurons.[38] Availability, culture adaptability, and strong osteogenesis capacity make PDLSCs proper candidates for regenerative bone treatments.[22] The present study was conducted to systematically review the studies outcomes on the bone formation efficacy of PDLSCs in artificial animal defects. Our results indicated that despite the limitations, PDLSCs might be the ideal means of achieving bone regeneration, especially in conjunction with suitable carrier and growth factors.
PDLSCs were isolated from lower third molars for the first time. Their bone regenerative potential has been investigated in several in vitro and in vivo studies.[39] Scaffolds, stem cells, and growth factors are the three main substructures for bone tissue engineering. The osteogenic differentiation capacity of stem cells, including PDLSCs, were affected by several factors, such as donor age and genetic variability, isolation and storage method, culturing conditions, and medium components.[14] Different scaffolds had shown acceptable efficacy in maintaining the viability and osteogenic capacity of PDLSCs during cell transplantation, including HA/TCP, HA/ECM, nano-HA-collagen-polylactic acid, bovine bone grafts, and gelatin sponges. VEGF, FGF-2, IGF-I, and recombinant human IGF binding protein five were reported as influential growth factors, enhancing osteogenic differentiation, and bone formation of PDLSCs.[39] In 2016, PDLSCs were used to treat an intra-bony defect in combination with Bio-Oss. This was the first clinical trial that used a dental stem cell for bone regeneration. The results did not significantly differ from the cell-free group.[40] Afterward, in 2020, Sanchez et al. used autologous PSLSC in combination with xenogeneic bone substitutes to treat human intra-bony periodontal defects. No clinical differences were reported among groups with or without PDLSCs.[41]
The first identified dental stem cells with mesenchymal characteristics were dental pulp stem cells.[42] These cells have higher proliferation capacity and colony-forming units than BMSCs; however, ectopic expansion of DPSCs without osteogenic induction materials leads to the formation of dentin-pulp-like tissue rather than bone.[39] Although successful in vivo bone regeneration using DPSCs have been reported, their regenerative capacity was lower than PDLSCs.[14]
In addition to oral tissue, other cell sources for bone regeneration have been found in bone marrow, fat, and placental tissues.[43] With the advantages of high stability in culture and osteoblastic differentiation affinity, bone marrow is still the main source of adult stem cells. Previous studies have shown more bone formation potential in BMSCs compared to PDLSCs. However, its harvesting methods are invasive, painful, and expensive, with the risk of donor infection.[14] Compared to other sources of MSCs, Similar bone regenerative potential has been reported in PDLSCs and umbilical cord MSCs.[14]In essence, bone regeneration involves forming and mineralizing an extracellular matrix, which should be Bio-Oss mimicked by scaffolds.[25] Growth factors can promote stem cells’ differentiation into osteoprogenitor cells, thus driving them toward osteogenic differentiation.[36] Differentiation of osteoprogenitor cells to osteoblast is the first step of the osteogenesis process. Following the related signals, the proliferation of progenitors, secretion, and mineralization of the extracellular matrix and then embedding of the proliferated cells in the mineralized matrix take place, respectively. During differentiation, osteocytic-related genes are expressed and transcripted to produce the essential proteins needed for bone formation.[27]
Limiting the search database to English studies could have possibly resulted in missing some studies. We tried to reduce publication bias by extending our search to various search engines. High inconsistency between the included studies in terms of cell types, scaffolds, animal models, follow-up and healing period, defect size, and location. The lack of possibility of doing the quantitative analysis was another limitation of the present study.
Despite the lack of consistency among the collected data of the present study, it seems that the presence of PDLSCs, along with sufficient scaffold and growth factors, might facilitate bone regeneration in both calvaria and alveolar defects. As a head-to-head comparison between the stem cells, scaffolds and growth factors were not possible by the present data; further preclinical animal studies are needed to reveal the proper combination for clinical regenerative treatments.
Financial support and sponsorship
Nil.
Conflicts of interest
The authors report no conflicts of interest.
References
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