Mesenchymal stem cells in craniofacial reconstruction: a comprehensive review.

Craniofacial reconstruction faces many challenges, including high complexity, strong specificity, severe injury, irregular and complex wounds, and high risk of bleeding. Traditionally, the "gold standard" for treating craniofacial bone defects has been tissue transplantation, which involves the transplantation of bone, cartilage, skin, and other tissues from other parts of the body. However, the shape of craniofacial bone and cartilage structures varies greatly and is distinctly different from ordinary long bones. Craniofacial bones originate from the neural crest, while long bones originate from the mesoderm. These factors contribute to the poor effectiveness of tissue transplantation in repairing craniofacial defects. Autologous mesenchymal stem cell transplantation exhibits excellent pluripotency, low immunogenicity, and minimally invasive properties, and is considered a potential alternative to tissue transplantation for treating craniofacial defects. Researchers have found that both craniofacial-specific mesenchymal stem cells and mesenchymal stem cells from other parts of the body have significant effects on the restoration and reconstruction of craniofacial bones, cartilage, wounds, and adipose tissue. In addition, the continuous development and application of tissue engineering technology provide new ideas for craniofacial repair. With the continuous exploration of mesenchymal stem cells by researchers and the continuous development of tissue engineering technology, the use of autologous mesenchymal stem cell transplantation for craniofacial reconstruction has gradually been accepted and promoted. This article will review the applications of various types of mesenchymal stem cells and related tissue engineering in craniofacial repair and reconstruction.

Gingival mesenchymal stem cell therapy, immune cells, and immunoinflammatory application.

MSC-based therapeutic strategies have proven to be incredibly effective. Robust self-renewal, multilineage differentiation, and potential for tissue regeneration and disease treatments are all features of MSCs isolated from oral tissue. Human exfoliated deciduous teeth, dental follicles, dental pulp, apical papilla SCs, and alveolar bone are the primary sources of oral MSC production. The early immunoinflammatory response is the first stage of the healing process. Oral MSCs can interact with various cells, such as immune cells, revealing potential immunomodulatory regulators. They also have strong differentiation and regeneration potential. Therefore, a ground-breaking strategy would be to research novel immunomodulatory approaches for treating disease and tissue regeneration that depend on the immunomodulatory activities of oral MSCs during tissue regeneration.

Resveratrol May Reduce the Degree of Periodontitis by Regulating ERK Pathway in Gingival-Derived MSCs.

Gingival-derived mesenchymal stem cells (GMSCs) have strong self-renewal, multilineage differentiation, and immunomodulatory properties and are expected to be applied in anti-inflammatory and tissue regeneration. However, achieving the goal of using endogenous stem cells to treat diseases and even regenerate tissues remains a challenge. Resveratrol is a natural compound with multiple biological activities that can regulate stem cell immunomodulation when acting on them. This study found that resveratrol can reduce inflammation in human gingival tissue and upregulate the stemness of GMSCs in human gingiva. In cell experiments, it was found that resveratrol can reduce the expression of TLR4, TNFα, and NFκB and activate ERK/Wnt crosstalk, thereby alleviating inflammation, promoting the proliferation and osteogenic differentiation ability of GMSCs, and enhancing their immunomodulation. These results provide a new theoretical basis for the application of resveratrol to activate endogenous stem cells in the treatment of diseases in the future.

miR-1260b inhibits periodontal bone loss by targeting ATF6ß mediated regulation of ER stress.

The expression profiles of exosomal microRNAs (miRNAs) are regulated by the microenvironment, and appropriate priming with mesenchymal stem cells (MSCs) is one of the strategies to enhance the paracrine potency of MSCs. Our previous work demonstrated that exosomes from tumor necrosis factor (TNF)-α-primed human gingiva-derived MSCs (GMSCs) could be a therapeutic tool against periodontitis, and that TNFα-inducible exosomal miR-1260b is essential for the inhibition of alveolar bone loss. However, the precise molecular mechanism underlying miR-1260b-mediated inhibition of osteoclastogenesis is not yet fully understood. Here, we found that the activating transcription factor (ATF)-6ß, a novel miR-1260b-targeting gene, is critical for the regulation of osteoclastogenesis under endoplasmic reticulum (ER) stress. An experimental periodontal mouse model demonstrated that induction of ER stress was accompanied by enhanced ATF6ß expression, and local administration of miR-1260b and ATF6ß siRNA using polyethylenimine nanoparticles (PEI-NPs) significantly suppressed the periodontal bone resorption. In periodontal ligament (PDL) cells, the ER stress inducer, tunicamycin, enhanced the expression of the receptor activator of NF-κB ligand (RANKL), while miR-1260b-mediated downregulation of ATF6ß caused RANKL inhibition. Furthermore, the secretome from miR-1260b/ATF6ß-axis-activated PDL cells inhibited osteoclastogenesis in human CD14+ peripheral blood-derived monocytes. These results indicate that the miR-1260b/ATF6ß axis mediates the regulation of ER stress, which may be used as a novel therapeutic strategy to treat periodontal disease.

Supramolecular Hydrogel-Wrapped Gingival Mesenchymal Stem Cells in Cutaneous Radiation Injury.

Radiation-induced skin wound/dermatitis is one of the common side effects of radiotherapy or interventional radiobiology. Gingiva-derived mesenchymal stem cells (GMSCs) were indicated to have therapeutic potentials in skin diseases. However, stem cells are prone to spread and difficult to stay in the skin for a long time, limiting their curative effects and application. This study investigated the therapeutic efficacy of Nap-GDFDFpDY (pY-Gel) self-assembled peptide hydrogel-encapsulated GMSCs to treat 137Cs γ-radiation-induced skin wounds in mice. The effects were evaluated by skin damage score, hind limb extension measurement and histological and immunohistochemical analysis. In vivo studies showed that pY-Gel self-assembled peptide hydrogel-encapsulated GMSCs could effectively improve wound healing in irradiated skin tissues. In addition, it was found that GMSCs conditioned medium (CM) could promote the proliferation, migration and DNA damage repair ability of skin cells after irradiation in human keratinocyte cell line HaCaT and normal human dermal fibroblasts (HFF). Mechanistically, GMSCs-CM can promote the expression of epidermal growth factor receptor (EGFR), signal transducers and activators of transcription 3 (STAT3) and matrix metalloproteinases (MMPs), suggesting that activation of the EGFR/STAT3 signaling pathway may be involved in the repair of skin cells after exposure to radiations. In conclusion, pY-Gel self-assembled peptide hydrogel-encapsulated GMSCs have a beneficial therapeutic effect on radiation-induced cutaneous injury and may serve as a basis of novel cells therapeutic approach.

Comparative proteomics analysis of human stem cells from dental gingival and periodontal ligament.

Dental stem cells isolated from oral tissues have been shown to provide with high proliferation ability and multilineage differentiation potential. Gingival mesenchymal stem cells (GMSCs) and periodontal ligament stem cells (PDLSCs), kinds of dental stem cells, can be used as substitutes for tissue repair materials because of their similar regenerative functions. In this study, we aim to explore the similarities and differences between the protein profiles of GMSCs and PDLSCs through quantitative proteomics. A total of 2821 proteins were identified and retrieved, of which 271 were upregulated and 57 were downregulated in GMSCs compared to PDLSCs. Gene Ontology (GO) analysis demonstrated that the 328 differentially abundant proteins (DAPs) were involved in the regulation of gene expression, metabolism, and signal transduction in biological process, mainly distributed in organelles related to vesicle transport, and involved in the molecular function of binding protein. And Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that the DAPs were committed to regulating the synthesis of proteasome and spliceosome. Real-time quantitative polymerase chain reaction (RT-qPCR) results showed that ARPC1B, PDAP1, and SEC61B can be used as special markers to distinguish GMSCs from PDLSCs. This research contributes to explaining the molecular mechanism and promoting the clinical application of tissue regeneration of GMSCs and PDLSCs.

Growth and Osteogenic Differentiation of Discarded Gingiva-Derived Mesenchymal Stem Cells on a Commercial Scaffold.

BACKGROUND: In periodontal patients with jawbone resorption, the autologous bone graft is considered a "gold standard" procedure for the placing of dental prosthesis; however, this procedure is a costly intervention and poses the risk of clinical complications. Thanks to the use of adult mesenchymal stem cells, smart biomaterials, and active biomolecules, regenerative medicine and bone tissue engineering represent a valid alternative to the traditional procedures. AIMS: In the past, mesenchymal stem cells isolated from periodontally compromised gingiva were considered a biological waste and discarded during surgical procedures. This study aims to test the osteoconductive activity of FISIOGRAFT Bone Granular® and Matriderm® collagen scaffolds on mesenchymal stem cells isolated from periodontally compromised gingiva as a low-cost and painless strategy of autologous bone tissue regeneration. MATERIALS AND METHODS: We isolated human mesenchymal stem cells from 22 healthy and 26 periodontally compromised gingival biopsy tissues and confirmed the stem cell phenotype by doubling time assay, colony-forming unit assay, and expression of surface and nuclear mesenchymal stem cell markers, respectively by cytofluorimetry and real-time quantitative PCR. Healthy and periodontally compromised gingival mesenchymal stem cells were seeded on FISIOGRAFT Bone Granular® and Matriderm® scaffolds, and in vitro cell viability and bone differentiation were then evaluated. RESULTS: Even though preliminary, the results demonstrate that FISIOGRAFT Bone Granular® is not suitable for in vitro growth and osteogenic differentiation of healthy and periodontally compromised mesenchymal stem cells, which, instead, are able to grow, homogeneously distribute, and bone differentiate in the Matriderm® collagen scaffold. CONCLUSION: Matriderm® represents a biocompatible scaffold able to support the in vitro cell growth and osteodifferentiation ability of gingival mesenchymal stem cells isolated from waste gingiva, and could be employed to develop low-cost and painless strategy of autologous bone tissue regeneration.

Human gingiva-derived mesenchymal stem cells are therapeutic in lupus nephritis through targeting of CD39-CD73 signaling pathway.

Cell specific and cytokine targeted therapeutics have underperformed in systemic lupus erythematosus (SLE). Mesenchymal stem cells (MSCs) have emerged as a novel therapy to address the dysregulation in autoimmune diseases but also have limitations. Human gingiva derived MSCs (GMSCs) are superior in regulating immune responses. Here, we demonstrate that the adoptive transfer of GMSCs homes to and maintains in the kidney and has a robust therapeutic effect in a spontaneous lupus nephritis model. Specifically, GMSCs limits the development of autoantibodies as well as proteinuria, decreases the frequency of plasma cells and lupus nephritis histopathological scores by directly suppressing B cells activation, proliferation and differentiation. The blockage of CD39-CD73 pathway dramatically abrogates the suppressive capacities of GMSCs in vitro and in vivo and highlights the significance of this signaling pathway in SLE. Collectively, manipulation of GMSCs provides a promising strategy for the treatment of patients with SLE and other autoimmune diseases.

Gingiva-derived Mesenchymal Stem Cells and Their Potential Applications in Oral and Maxillofacial Diseases.

BACKGROUND: Stem cells are undifferentiated cells with multilineage differentiation potential. They can be collected from bone marrow, fat, amniotic fluid, and teeth. Stem cell-based therapies have been widely used to treat multiple diseases, such as cardiac disease, and hematological disorders. The cells may also be beneficial for controlling the disease course and promoting tissue regeneration in oral and maxillofacial diseases. Oral-derived gingival mesenchymal stem cells are easy to access and the donor sites heal rapidly without a scar. Such characteristics demonstrate the beneficial role of GMSCs in oral and maxillofacial diseases. OBJECTIVE: We summarize the features of GMSCs, including their self-renewal, multipotent differentiation, immunomodulation, and anti-inflammation properties. We also discuss their applications in oral and maxillofacial disease treatment and tissue regeneration. CONCLUSION: GMSCs are easily harvestable adult stem cells with outstanding proliferation, differentiation, and immunomodulation characteristics. A growing body of evidence indicates that GMSCs have strong potential use in accelerating wound healing and promoting the regeneration of bone defects, periodontium, oral neoplasms, salivary glands, peri-implantitis, and nerves. Moreover, alginate, polylactic acid and polycaprolactone can be used as biodegradable scaffolds for GMSC encapsulation. Various growth factors can be applied to the corresponding scaffolds to obtain the desired GMSC differentiation and phenotypes. Three-dimensional spheroid culture systems could optimize GMSC properties and improve the performance of the cells in tissue engineering. The immunomodulatory property of GMSCs in controlling oral and maxillofacial inflammation needs further research.

Muscone Promotes The Adipogenic Differentiation Of Human Gingival Mesenchymal Stem Cells By Inhibiting The Wnt/ß-Catenin Signaling Pathway.

OBJECTIVES: This study was performed to evaluate the effects of muscone on the proliferation, migration and differentiation of human gingival mesenchymal stem cells (GMSCs) and to explore the relevant mechanisms. MATERIALS AND METHODS: We performed studies to determine the effects and mechanisms of muscone on GMSC proliferation, migration and differentiation. We conducted CCK-8, colony formation, transwell chamber, scratch wound, alkaline phosphatase (ALP) staining and activity, and alizarin red and oil red O staining assays, as well as real-time quantitative polymerase chain reaction (qRT-PCR), to ascertain the effects of muscone on GMSC proliferation, migration and differentiation in vitro. The mechanism by which muscone influences the osteogenic and adipogenic differentiation of GMSCs was elucidated by qRT-PCR and Western blotting. RESULTS: We found that muscone significantly promoted GMSC proliferation, chemotaxis, wound healing and fat droplet formation and inhibited ALP activity and mineral deposition. Notably, we observed that the Wnt/ß-catenin pathway was closely related to the ability of muscone to inhibit the osteogenic differentiation and promote the adipogenic differentiation of GMSCs. The effect of muscone on the multidirectional differentiation capacity of GMSCs was significantly reversed by the agonist lithium chloride through the Wnt/ß-catenin signaling pathway. CONCLUSION: Muscone effectively increased the proliferation and migration, promoted the adipogenic differentiation and inhibited the osteogenic differentiation of GMSCs by inhibiting the Wnt/ß-catenin signaling pathway. These results may provide a theoretical basis for the application of GMSCs and muscone in tissue engineering and regenerative medicine.

A preclinical study-systemic evaluation of safety on mesenchymal stem cells derived from human gingiva tissue.

BACKGROUND: Mounting evidence has shown that a novel subset of mesenchymal stem cells (MSCs) derived from human gingiva referred to as gingival mesenchymal stem cells (GMSCs) displays a greater immunotherapeutic potential and regenerative repair expression than MSCs obtained from other tissues. However, the safety of the use of transplanted GMSCs in humans remains unclear. METHODS: In this study, we evaluated the safety of GMSCs transplanted into mouse, rat, rabbit, beagle dog, and monkey as well as two animal models of autoimmune diseases. RESULTS: In short- and long-term toxicity tests, infused GMSCs had no remarkable adverse effects on hematologic and biochemical indexes, particularly on the major organs such as heart, liver, spleen, and kidney in recipient animals. It was also shown that GMSCs were well tolerated in other assays including hemolysis, vascular, and muscular stimulation, as well as systemic anaphylaxis and passive skin Arthus reaction in animal models. GSMC infusion did not cause any notable side effects on animal models of either autoimmune arthritis or lupus. Significantly, GMSCs most likely play no role in genotoxicity and tumorigenesis. The biological features remained stable for an extended period after cell transfer. CONCLUSIONS: GMSCs are safe in various animal models of autoimmunity, even during active disease episodes, especially in monkeys. This study paves a solid road for future clinical trials of GMSCs in patients with autoimmune and inflammatory diseases.

Small molecule inhibitor of TGF-ß signaling enables robust osteogenesis of autologous GMSCs to successfully repair minipig severe maxillofacial bone defects.

BACKGROUND: Clinically, for stem cell-based therapy (SCBT), autologous stem cells are considered better than allogenic stem cells because of little immune rejection and no risk of communicable disease infection. However, severe maxillofacial bone defects restoration needs sufficient autologous stem cells, and this remains a challenge worldwide. Human gingival mesenchymal stem cells (hGMSCs) derived from clinically discarded, easily obtainable, and self-healing autologous gingival tissues, have higher proliferation rate compared with autologous bone marrow mesenchymal stem cells (hBMSCs). But for clinical bone regeneration purpose, GMSCs have inferior osteogenic differentiation capability. In this study, a TGF-ß signaling inhibitor SB431542 was used to enhance GMSCs osteogenesis in vitro and to repair minipig severe maxillofacial bone defects. METHODS: hGMSCs were isolated and cultured from clinically discarded gingival tissues. The effects of SB431542 on proliferation, apoptosis, and osteogenic differentiation of hGMSCs were analyzed in vitro, and then, SB431542-treated hGMSCs composited with Bio-Oss® were transplanted into immunocompromised mice subcutaneously to explore osteogenic differentiation in vivo. After that, SB431542-treated autologous pig GMSCs (pGMSCs) composited with Bio-Oss® were transplanted into circular confined defects (5 mm × 12 mm) in minipigs maxillary to investigate severe bone defect regeneration. Minipigs were sacrificed at 2 months and nude mice at 8 weeks to retrieve specimens for histological or micro-CT or CBCT analysis. Effects of SB431542 on TGF-ß and BMP signaling in hGMSCs were investigated by Western Blot or qRT-PCR. RESULTS: One micromolar of SB431542 treatment induced a robust osteogenesis of hGMSCs in vitro, without adverse effect on apoptosis and growth. In vivo, 1 µM SB431542 treatment also enabled striking osteogenesis of hGMSCs subcutaneously in nude mice and advanced new bone formation of pGMSCs in minipig maxillary bone defect model. In addition, SB431542-treated hGMSCs markedly increased bone-related proteins expression, and BMP2 and BMP4 gene expression. Conversely, SMAD3 protein-dependent TGF-ß signal pathway phosphorylation was decreased. CONCLUSIONS: Our study show that osteogenic differentiation of GMSCs treated with TGF-ß signaling inhibitor SB431542 was increased, and SB431542-treated autologous pig GMSCs could successfully repair minipig severe maxillofacial bone defects. This preclinical study brings about a promising large bone regeneration therapeutic potential of autologous GMSCs induced by SB431542 in clinic settings.

Small molecules enhance neurogenic differentiation of dental-derived adult stem cells.

OBJECTIVE: Dental-derived stem cells originate from the embryonic neural crest, and exhibit high neurogenic potential. This study aimed to investigate whether a cocktail of eight small molecules (Valproic acid, CHIR99021, Repsox, Forskolin, SP600125, GO6983, Y-27632 and Dorsomorphin) can enhance the in vitro neurogenic differentiation of dental pulp stem cells (DPSCs), stem cells from apical papilla (SCAPs) and gingival mesenchymal stem cells (GMSCs), as a preliminary step towards clinical applications. MATERIALS AND METHODS: Neural induction was carried out with a small molecule cocktail based two-step culture protocol, over a total duration of 14 days. At the 8 and 14 day timepoints, the cells were analyzed for expression of neural markers with immunocytochemistry, qRT-PCR and Western Blot. The Fluo 4-AM calcium flux assay was also performed after a further 14 days of neural maturation. RESULTS: More pronounced morphological changes characteristic of the neural lineage (i.e. neuritogenesis) were observed in all three cell types treated with small molecules, as compared to the untreated controls. This was corroborated by the immunocytochemistry, qRT-PCR and western blot data, which showed upregulated expression of several early and mature neural markers in all three cell types treated with small molecules, versus the corresponding untreated controls. Finally, the Fluo-4 AM calcium flux assay showed consistently higher calcium transient (F/Fo) peaks for the small molecule-treated versus untreated control groups. CONCLUSIONS: Small molecules can enhance the neurogenic differentiation of DPSCs, SCAPs and GMSCs, which offer much potential for therapeutic applications.

Time-Course Transcriptome Analysis of Gingiva-Derived Mesenchymal Stem Cells Reveals That Fusobacterium nucleatum Triggers Oncogene Expression in the Process of Cell Differentiation.

Fusobacterium nucleatum has pathogenic effects on oral squamous cell carcinoma and colon cancer, while the effects of continuously altered gene expression in normal human cells, as induced by persistent exposure to F. nucleatum, remain unclear. In this study, a microarray Significant Profiles (maSigPro) analysis was used to obtain the transcriptome profile of gingiva-derived mesenchymal stem cells (GMSCs) stimulated by F. nucleatum for 3, 7, 14, and 21 day, and the results revealed 790 (nine clusters) differentially expressed genes (DEGs), which were significantly enriched in cell adherens junctions and cancer-related pathways. On the basis of a short time-series expression miner (STEM) analysis, all the expressed genes in the GMSCs were grouped into 50 clusters according to dynamic gene expression patterns, and the expression levels of three gene clusters in the F. nucleatum-treated GMSCs were significantly different than the predicted values. Among the 790 DEGs, 50 tumor-associated genes (TAGs; such as L3MBTL4, CD163, CCCND2, CADM1, BCL7A, and IGF1) and five core dynamic DEGs (PLCG2, CHI3L2, L3MBTL4, SH2D2A, and NLRP3) were identified during F. nucleatum stimulation. Results from a GeneMANIA database analysis showed that PLCG2, CHI3L2, SH2D2A, and NLRP3 and 20 other proteins formed a complex network of which 12 genes were enriched in cancer-related pathways. Based on the five core dynamic DEGs, the related microRNAs (miRNAs) and transcription factors (TFs) were obtained from public resources, and an integrated network composed of the related TFs, miRNAs, and mRNAs was constructed. The results indicated that these genes were regulated by several miRNAs, such as miR-372-3p, miR-603, and miR-495-3p, and several TFs, including CREB3, GATA2, and SOX4. Our study suggests that long-term stimulation by F. nucleatum may trigger the expression of cancer-related genes in normal gingiva-derived stem cells.

Gingiva-Derived Mesenchymal Stem Cell-Extracellular Vesicles Activate Schwann Cell Repair Phenotype and Promote Nerve Regeneration.

IMPACT STATEMENT: Peripheral nerve injuries (PNIs) are common and debilitating, usually resulting in considerable long-term disability and remaining an unmet clinical need. Even though the combination of mesenchymal stem cells (MSCs) and the state-of-the-art tissue engineering technologies has shown promising therapeutic potentials for PNI, there is still not a single licensed stem cell-based product for peripheral nerve repair/regeneration. Emerging evidence indicates that MSC-derived extracellular vesicles (EVs) are comparably effective as MSCs in the therapy of a variety of disease models or pathological conditions. This report shows that local delivery of gingiva-derived mesenchymal stem cell (GMSC)-derived EVs could obviously promote axonal regeneration and functional recovery of injured mice sciatic nerves. Importantly, the findings suggest that GMSC-derived EVs promoted the expression of Schwann cell dedifferentiation/repair phenotype-related genes in vitro, particularly c-JUN, a key transcription factor that drives the activation of repair phenotype of Schwann cells during PNI and regeneration.

SIS-ECM Laden with GMSC-Derived Exosomes Promote Taste Bud Regeneration.

Oral cancer has a high annual incidence rate all over the world, and the tongue is the most frequently affected anatomic structure. The current standard care is ablative surgery of malignant neoplasm, followed by tongue reconstruction with free flap. However, such reconstructive modalities with postsurgery radiotherapy or chemotherapy can hardly support the functional recovery of the tongue-particularly, functional taste bud regeneration-in reconstructed areas, thus seriously affecting patients' prognosis and life quality. Using a critical-sized tongue defect model in rats, we show that combinatory transplantation of small intestinal submucosa-extracellular matrix (SIS-ECM) with gingival mesenchymal stem cells (GMSCs) or their derivative exosomes promoted tongue lingual papillae recovery and taste bud regeneration as evidenced by increased expression of CK14, CK8, and markers for type I, II, and III taste bud cells (NTPdase 2, PLC-ß2, and AADC, respectively). In addition, our results indicate that GMSCs or their derivative exosomes could increase BDNF expression, a growth factor that plays an important role in the proliferation and differentiation of epithelial basal progenitor cells into taste bud cells. Meanwhile, we showed an elevated expression level of Shh-which is essential for development, homeostasis, and maintenance of the taste bud organ-in wounded areas of the tongue among animals treated with GMSC/SIS-ECM or exosome/SIS-ECM as compared with SIS-ECM control. Moreover, our data show that GMSCs or their derivative exosomes promoted innervation of regenerated taste buds, as evidenced by elevated expressions of neurofilament and P2X3 at the injury areas. Together, our findings indicate that GMSC/SIS-ECM and exosome/SIS-ECM constructs can facilitate taste bud regeneration and reinnervation with promising potential application in postsurgery tongue reconstruction of patients with tongue cancer.

MicroRNA-21 promotes wound healing via the Smad7-Smad2/3-Elastin pathway.

Wound healing is regulated by a complex network of cells, molecules, and cytokines, as well as microRNAs (miRNAs). miRNAs were confirmed to influence the wound healing process, and miR-21, an important member of the miRNA family, was also shown to regulate wound healing. The aim of the present study was to investigate the role of miR-21 in the wound healing process and the possible underlying cell signaling pathways. We isolated GMSCs from WT and miR-21-KO mouse gingiva. Flow cytometric analysis and immunocytofluorescense staining were used to identify the GMSCs acquired from WT and miR-21-KO mice. RT-PCR, western blot analysis and immunohistofluorescence staining were performed to examine the expression of extracellular matrix components and key proteins of cell signaling pathways. TargetScan and pmiR-RB-REPORT vectors were used to verify that Smad7 was a direct target of miR-21. Compared to WT mice, miR-21-KO mice showed slower wound healing. RT-PCR and western blot analysis indicated that Elastin expression was downregulated in miR-21-deficient samples. We confirmed that Smad7 was a direct target of miR-21. miR-21 knockout resulted in increased expression of Smad7 and impaired phosphorylation of the Smad2/3 complex. The expression of the Smad7-Smad2/3-Elastin axis in palate tissues sections acquired from WT and miR-21-KO mice showed the same trend. Based on all these results, we demonstrated that miR-21 promoted the wound healing process via the Smad7-Smad2/3-Elastin pathway.