Human Gingiva-derived Mesenchymal Stem Cells Research Articles

Influence of Topography and Composition of Commercial Titanium Dental Implants on Cell Adhesion of Human Gingiva-Derived Mesenchymal Stem Cells: An In Vitro Study

The topography and composition of dental implant surfaces directly impact mesenchymal cell adhesion, proliferation, and differentiation, crucial aspects of achieving osseointegration. However, cell adhesion to biomaterials is considered a key step that drives cell proliferation and differentiation. The aim of this study was to characterize characterize the topography and composition of commercial titanium dental implants manufactured with different surface treatments (two sandblasted/acid-etched (SLA) (INNO Implants, Busan, Republic of Korea; BioHorizonsTM, Oceanside, CA, USA) and two calcium phosphate (CaP) treated (Biounite®, Berazategui, Argentina; Zimmer Biomet, Inc., Warsaw, IN, USA)) and to investigate their influence on the process of cell adhesion in vitro. A smooth surface implant (Zimmer Biomet, Inc.) was used as a control. For that, high-resolution methodologies such as scanning electron microscopy (SEM), X-ray dispersive spectroscopy (EDX), laser scanning confocal microscopy (LSCM), and atomic force microscopy (AFM) were employed. Protein adsorption and retromolar gingival mesenchymal stem cells (GMSCs) adhesion to the implant surfaces were evaluated after 48 h. The adherent cells were examined by SEM and LSCM for morphologic and quantitative analyses. ANOVA and Tukey tests (α = 0.05) were employed to determine statistical significance. SEM revealed that INNO, BioHorizonsTM, and Zimmer implants have an irregular surface, whereas Biounite® has a regular topography consisting of an ordered pattern. EDX confirmed a calcium and phosphate layer on the Biounite® and Zimmer surfaces, and AFM exhibited different roughness parameters. Protein adsorption and cell adhesion were detected on all the implant surfaces studied. However, the Biounite® implant with CaP and regular topography showed the highest protein adsorption capacity and density of adherent GMSCs. Although the Zimmer implant also had a CaP treatment, protein and cell adhesion levels were lower than those observed with Biounite®. Our findings indicated that the surface regularity of the implants is a more determinant factor in the cell adhesion process than the CaP treatment. A regular, nanostructured, hydrophilic, and moderately rough topography generates a higher protein adsorption capacity and thus promotes more efficient cell adhesion.

Insulin-producing cell clusters derived from human gingival mesenchymal stem cells as a model for diabetes research.

The human gingiva-derived mesenchymal stem cells (hGMSCs) possess a great potential to develop the cell-based therapy for diabetes due to its unscarred healing capacity and reparative potential. In this current study, we isolated, cultured and characterised the GMSCs and explored their potential to differentiate into Insulin Producing Cell Clusters (IPCCs). The cells derived from gingival tissues exhibited fibroblast-like morphology. The flow cytometric analysis revealed positive expression of CD73(97.43%), CD90(95.05%), and CD105(93.17%) and negative expression of CD34(0.05%), CD45(0.09%), and HLA-DR (0.025) surface markers. We then converted this adherent fibroblast-like GMSCs into floating IPCCs using a sequential three-step protocol containing a different combination of differentiating agents. Initially, the presence of insulin in IPCCs was confirmed by dithizone staining. Glucose-stimulated insulin secretion (GSIS) assay confirmed that IPCCs secrete insulin in response to glucose. Generated IPCCs express pancreatic markers such as insulin, pdx1, glucagon, GLUT4 and GLUT2 as evidenced by RT-PCR analysis. Our results unequivocally showed that IPCCs can be generated from gingiva which is a potential source of postnatal MSCs. Our results offer the IPCCs generated from hGMSCs a platform for screening anti-diabetic drugs and a new autologous source of tissue for islet transplantation for the treatment of diabetes. Our results unequivocally demonstrate for the first time that hGMSCs can be used as an attractive non-invasive tissue source for generating IPCCs, which can be employed in diabetes research for screening antidiabetic agents and also for transplantation in type 1 diabetic patients as autologous source without the need of immunosuppression.

Gingiva-Derived Mesenchymal Stem Cells Attenuate Imiquimod- (IMQ-) Induced Murine Psoriasis-Like Skin Inflammation.

Human gingiva-derived mesenchymal stem cells (GMSCs) are isolated from the gingival propria with promising regenerative, immunomodulatory, and anti-inflammatory properties. Recently, several studies, including ours, have found that GMSCs have the therapeutic potentials of nerve regeneration and skin disorders in various types such as the cell itself, cell-free conditioned medium, or extracellular vesicles (EVs). However, the mechanobiological behavior of GMSCs is closely related to the culture conditions. Therefore, the purpose of this study was to evaluate the function of human GMSCs on imiquimod- (IMQ-) induced murine psoriasis-like skin inflammation in two-dimensional (2D) and three-dimensional (3D) culture conditions. Here, we isolated and characterized GMSCs in 2D and 3D culture conditions and found that GMSCs in 2D and 3D infusion can significantly ameliorate the IMQ-induced murine psoriasis-like skin inflammation, reduce the levels of Th1- and Th17-related cytokines IFN-γ, TNF-α, IL-6, IL-17A, IL-17F, IL-21, and IL-22, and upregulate the percentage of spleen CD25+CD3+ T cells while downregulate the percentage of spleen IL-17+CD3+ T cells. In summary, our novel findings reveal that GMSCs in 2D and 3D infusion may possess therapeutic effects in the treatment of psoriasis.

Comparison of Cellular and Differentiation Characteristics of Mesenchymal Stem Cells Derived from Human Gingiva and Periodontal Ligament.

ABSTRACTObjectives:Dental tissues possess multipotent stem cells with varying biological properties. The present study was aimed to establish a primary culture of human gingiva-derived mesenchymal stem cells (GMSCs) and periodontal ligament-derived stem cells (PDLSCs) from periodontally healthy subjects and compare their biological characteristics.Materials and Methods:Gingival and periodontal ligament (PDL) tissues were collected from extracted premolar teeth of five healthy subjects and primary cultures were established. Basic biological characteristics, such as cell morphology, viability, proliferation capacity, and colony-forming units, and in vitro osteogenic and adipogenic differentiation potential were performed at passage 3 of GMSCs and PDLSCs. This was followed by immuno-phenotyping and flow cytometric analysis for identification of positive mesenchymal stem cell (MSC) markers, such as CD73, CD90, and CD105, and negative markers CD45 and CD34.Statistical Analysis Used:One-way analysis of variance (ANOVA).Results:Primary cultures of GMSCs and PDLSCs were successfully established. Cells exhibited a fibroblast-like morphology with a homogeneous population at passage 3. Cells derived from both tissues were highly viable (>95%), proliferative, and capable of forming colonies. Both cells did not exhibit any noticeable differences in cellular properties. Immunofluorescence and flow cytometric analyses showed positivity for MSC markers, CD73, CD90, and CD105, and negativity for CD34 and CD45. Furthermore, GMSCs and PDLSCs were capable of differentiating in vitro into osteocytes as evidenced by Alizarin red-S staining, and adipocytes as demonstrated by oil red O staining.Conclusions:The results of the present study indicate that both GMSCs and PDLSCs have similar cellular characteristics and mesenchymal differentiation potential. Therefore, they may serve as an equally potent source of stem cells for use in cell-based periodontal therapies.

Harnessing 3D collagen hydrogel-directed conversion of human GMSCs into SCP-like cells to generate functionalized nerve conduits

Achieving a satisfactory functional recovery after severe peripheral nerve injuries (PNI) remains one of the major clinical challenges despite advances in microsurgical techniques. Nerve autografting is currently the gold standard for the treatment of PNI, but there exist several major limitations. Accumulating evidence has shown that various types of nerve guidance conduits (NGCs) combined with post-natal stem cells as the supportive cells may represent a promising alternative to nerve autografts. In this study, gingiva-derived mesenchymal stem cells (GMSCs) under 3D-culture in soft collagen hydrogel showed significantly increased expression of a panel of genes related to development/differentiation of neural crest stem-like cells (NCSC) and/or Schwann cell precursor-like (SCP) cells and associated with NOTCH3 signaling pathway activation as compared to their 2D-cultured counterparts. The upregulation of NCSC-related genes induced by 3D-collagen hydrogel was abrogated by the presence of a specific NOTCH inhibitor. Further study showed that GMSCs encapsulated in 3D-collagen hydrogel were capable of transmigrating into multilayered extracellular matrix (ECM) wall of natural NGCs and integrating well with the aligned matrix structure, thus leading to biofabrication of functionalized NGCs. In vivo, implantation of functionalized NGCs laden with GMSC-derived NCSC/SCP-like cells (designated as GiSCs), significantly improved the functional recovery and axonal regeneration in the segmental facial nerve defect model in rats. Together, our study has identified an approach for rapid biofabrication of functionalized NGCs through harnessing 3D collagen hydrogel-directed conversion of GMSCs into GiSCs.

Microstructure and mechanical behaviors of tibia for collagen-induced arthritic mice treated with gingiva-derived mesenchymal stem cells

Rheumatoid arthritis (RA) is a systemic polyarticular arthritis that primarily affects the small joints but also causes bone erosion in large joints. None of the currently existing treatment approaches is curable. In this study, the effects of human gingiva-derived mesenchymal stem cells (GMSCs) on collagen-induced arthritis (CIA) mice are examined by experimentally assessing the microstructure and mechanical behaviors of tibia. Bone morphology and mineral density of mouse tibiae were assessed using micro-X-ray computed tomography (micro-CT). Compression testing was performed on mouse tibia to access its stiffness. The deformation and strain localized inside proximal tibia were mapped using mechanical testing coupled with micro-CT and digital volume correlation of micro-CT images. The results show that CIA disease caused bone erosion in epiphyseal cortical bone, which manifested into the adjacent epiphyseal trabecular bone, and also affected the metaphyseal cortical bone. CIA disease also weakened the load-bearing function of proximal tibia. GMSC treatment interfered with the progress of CIA, attenuated the bone erosion in epiphyseal and metaphyseal trabecular bone and resulted in improved load-bearing function of proximal tibia. GMSCs provide a promising potential treatment of autoimmune arthritis.

Role of ascorbic acid in the regulation of epigenetic processes induced by Porphyromonas gingivalis in endothelial-committed oral stem cells

Periodontitis is a common inflammatory disease that affects the teeth-supporting tissue and causes bone and tooth loss. Moreover, in a worldwide population, periodontal disease is often associated with cardiovascular diseases. Emerging studies have reported that one of the major pathogens related to periodontitis is Porphyromonas gingivalis (P. gingivalis), which triggers the inflammatory intracellular cascade. Here, we hypothesized a possible protective effect of ascorbic acid (AA) in the restoration of the physiological molecular pathway after exposure to lipopolysaccharide derived from P. gingivalis (LPS-G). In particular, human gingiva-derived mesenchymal stem cells (hGMSCs) and endothelial-differentiated hGMSCs (e-hGMSCs) exposed to LPS-G showed upregulation of p300 and downregulation of DNA methyltransferase 1 (DNMT1), proteins associated with DNA methylation and histone acetylation. The co-treatment of AA and LPS-G showed a physiological expression of p300 and DNMT1 in hGMSCs and e-hGMSCs. Moreover, the inflammatory process triggered by LPS-G was demonstrated by evaluation of reactive oxygen species (ROS) and their intracellular localization. AA exposure re-established the physiological ROS levels. Despite the limitations of in vitro study, these findings collectively expand our knowledge regarding the molecular pathways involved in periodontal disease, and suggest the involvement of epigenetic modifications in the development of periodontitis.

WITHDRAWN: Human gingiva-derived mesenchymal stem cells promote osteogenic differentiation through their immunosuppressive function

Human gingiva-derived mesenchymal stem cells (GMSCs) have emerged as a new MSC population exhibiting robust immune regulatory functions, multipotent differentiation potential, and regenerative ability. However, the effects of GMSCs on T cells remain unexplored. Herein, we aimed to evaluate whether GMSCs promote osteogenic differentiation by regulating immune cells. The GMSC phenotype was confirmed using the colony-forming assay, immunophenotyping, Oil red O staining, and Alizarin red staining. mRNA expression levels of proinflammatory molecules (interleukin-1β [IL-1β] and tumor necrosis factor-α [TNF-α]) and anti-inflammatory factors (IL-10) were measured by quantitative reverse-transcription PCR (qRT-PCR). Then, MC3T3-E1 cells were treated with the collected co-culture supernatant, followed by alkaline phosphatase (ALP) and immunofluorescence staining to evaluate osteogenic differentiation of MC3T3-E1 cells. qRT-PCR and western blotting were employed to analyze the expression levels of osteogenic differentiation proteins, including collagen type I (COL-1), ALP, osteopontin (OPN), and runt-related transcription factor 2 (RUNX2). GMSCs were successfully isolated and identified. We observed that GMSCs suppressed the activated T-cell function by downregulating IL-1β and TNF-α and upregulating IL-10. Simultaneously, the expression levels of osteogenesis-related genes (COL-1, ALP, OPN, and RUNX2) were markedly lower in the co-culture supernatant and Jurkat T cell supernatant groups than those in the normal culture medium group; however, expression levels were significantly increased in the co-culture supernatant group when compared with the Jurkat T cell supernatant group. Our findings indicate that GMSCs could promote the osteogenic differentiation of MC3T3-E1 cells by inhibiting the biological activity of activated T cells.

B7-H1 Promotes the Functional Effect of Human Gingiva-Derived Mesenchymal Stem Cells on Collagen-Induced Arthritis Murine Model

Recent studies found that mesenchymal stem cells (MSCs), by virtue of their tissue recovery and immunoregulatory properties, have shown a broad prospect for applications in various autoimmune and degenerative diseases. Although the potential therapeutic use of MSCs is considerable, studies and clinical treatment efficacy are preliminary due to the heterogeneity of MSCs. Herein, based on RNA-sequencing (RNA-seq) and single cell sequence properties, we demonstrated that B7-H1 plays an important role in the immunosuppressive function of human gingiva-derived mesenchymal stem cells (GMSCs) in a collagen-induced arthritis murine model that is dependent on STAT3 signaling. Our data offer convincing evidence that B7-H1 expression by GMSCs helps to identify a new subpopulation of MSCs with a greater immunosuppressive property. The approach provides a unique and additional strategy for stem cells-based therapies of autoimmune and other inflammatory diseases.

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.

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

Abstract 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 adoptively transferred GMSC home to and maintain in the kidney and have 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 abrogated 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.

Persistent Exposure to Fusobacterium nucleatum Triggers Chemokine/Cytokine Release and Inhibits the Proliferation and Osteogenic Differentiation Capabilities of Human Gingiva-Derived Mesenchymal Stem Cells.

Fusobacterium nucleatum is one of the most frequent pathogenic bacteria causing periodontitis. The direct effect of Fusobacterium nucleatum (F. nucleatum) on oral stem cells has rarely been reported. In this study, we aimed to evaluate how gingiva-derived mesenchymal stem cells (GMSCs) respond to a direct challenge with F. nucleatum. GMSCs were isolated by the limiting dilution method and exposed to F. nucleatum at various multiplicities of infection (MOIs; F. nucleatum:cell ratios of 10:1, 50:1, and 100:1) for 24 h to 4 weeks. Our results indicated that F. nucleatum significantly inhibited cell proliferation in a dose-dependent manner and promoted cell migration and the release of chemokines/cytokines, such as CCL2, CXCL1, and IL-6. Additionally, F. nucleatum inhibited GMSC osteogenic differentiation partly by decreasing alkaline phosphatase (ALP) activity, mineralized nodule formation, and osteogenesis-related gene and protein expression. RNA-sequencing analyses indicated that F. nucleatum time-dependently activated cellular signaling pathways during the process of osteogenic differentiation. A total of 64 cell differentiation-related genes were found to be differentially expressed between non-infected and F. nucleatum-infected GMSCs at 3, 7, 14, and 21 d. Intriguingly, we discovered that the 64 cell differentiation-related differentially expressed genes (DEGs) were significantly enriched in cancer-related pathways, such as bone cancer, osteosarcoma and bone marrow cancer, which provides new insight into tumorigenesis during the process of GMSC osteogenic differentiation. In conclusion, this study demonstrates that persistent exposure to F. nucleatum promotes cell migration and chemokine/cytokine release and inhibits the proliferation and osteogenic differentiation of GMSCs. Our study provides a novel and long-time bacteria-cell co-culture in vitro model and makes a foundation for the future mechanistic studies of GMSCs under F. nucleatum infection.

Systematically transplanted human gingiva-derived mesenchymal stem cells regulate lipid metabolism and inflammation in hyperlipidemic mice with periodontitis.

Gingiva-derived mesenchymal stem cells (GMSCs) have been the focus of extensive research due to their numerous distinct properties, including their homing to injury sites and their contribution to tissue regeneration. However, the role of transplanted GMSCs in the regulation of lipid metabolism and inflammation in hyperlipidemic mice with periodontitis has not been demonstrated. In the present study, apolipoprotein E-deficient (ApoE−/−) mice were used to establish a hyperlipidemia model with periodontitis and divided into two groups: Group B and Group C (n=20 per group), and wild-type C57BL/6J mice without any treatment were assigned to Group A (n=20). Animals in Group C were then injected with human GMSCs through the tail vein and animals in Group B were injected with α-MEM as control. Animals were sacrificed at indicated time points. Serum was collected to determine the lipids and inflammatory cytokines. Liver samples were collected to estimate lipid-associated gene expression. Morphometric and histological analyses were performed to maxillaries. The results demonstrated that the delivery of GMSCs led to a significant decrease in triglyceride (TG), total cholesterol (TC), low density lipoprotein cholesterol (LDL), interleukin (IL)-6, tumor necrosis factor (TNF)-α, alveolar bone loss (ABL), and sterol regulatory element binding protein-1c (SREBP-1c) mRNA, and a significant increase in high density lipoprotein cholesterol (HDL), IL-10 and peroxisome proliferator-activated receptor α (PPARα) mRNA in Group C compared to Group B. Histological examination showed increased formation of new bone and higher alveolar bone height in Group C. Systematically transplanted GFP-positive cells were detected through both fluorescence microscope observation and immunohistochemical staining in the periodontal tissues. Overall, systematically transplanted GMSCs attenuated the hyperlipidemia and inflammatory responses in hyperlipidemic mice with periodontitis, and improved periodontal tissue regeneration.

Bone morphogenetic protein‑7 upregulates genes associated with osteoblast differentiation, including collagen I, Sp7 and IBSP in gingiva‑derived stem cells

The present study was performed to evaluate the effects of short-term application of bone morphogenetic protein-7 (BMP-7) on human gingiva-derived mesenchymal stem cells with next-generation sequencing. Human gingiva-derived stem cells were treated with a final concentration of 100 ng/ml BMP-7 and the same concentration of a vehicle control. mRNA sequencing and data analysis were performed along using gene ontology and pathway analysis. RT-qPCR of mRNA of collagen I, Sp7, IBSP and western blot analysis of collagen I, osterix and bone sialoprotein was also performed. A total of 25,737 mRNAs were identified to be differentially expressed. Regarding osteoblast differentiation, 14 mRNAs were upregulated and 10 were downregulated when the results of the BMP-7 at 3 h were compared with the control at 3 h. The expression of collagen I was increased following the application of BMP-7 at 3 h, and this increase was also observed following western blot analysis. The effects of BMP-7 on stem cells were evaluated with mRNA sequencing, and the expression was validated with RT-qPCR and western blot analysis. The short-term application of BMP-7 produced an increased expression of collagen I, which was associated with target genes selected for osteoblast differentiation. This study may provide novel insights into the role of BMP-7 using mRNA sequencing.

Human gingiva-derived mesenchymal stem cells alleviate inflammatory bowel disease via IL-10 signalling-dependent modulation of immune cells.

Current evidence indicates that inflammatory bowel disease (IBD) is caused primarily by impaired mucosal immunity, resulting in an imbalance between epithelial barrier function and tissue inflammation. Human gingiva-derived mesenchymal stem cells (GMSCs) exhibit immunomodulatory and anti-inflammatory effects in a variety of immunity- and inflammation-associated diseases. However, the role of GMSCs in treating IBD has not been elucidated. Our study, therefore, examined the therapeutic effect and mechanism of GMSCs in a murine colitis model of IBD. Our results indicate that the infusion of GMSCs significantly prolonged survival and relieved symptoms. Phenotype analyses showed that the frequencies of NK1.1+ and CD11b+ cells, as well as CD4 T cells in the spleen, were suppressed in GMSC-treated mice compared with the PBS- or fibroblast-treated control groups. Additionally, GMSC treatment markedly increased the numbers of interleukin (IL)-10+ regulatory T cells, reduced the secretion of pro-inflammatory cytokines, and increased production of anti-inflammatory cytokines. A mechanistic study revealed that anti-IL-10R antibody abolished the protective effect of GMSCs compared with mice treated with anti-IgG antibody. Thus, our results indicate that GMSCs play a critical role in alleviating colitis by modulating inflammatory immune cells via IL-10 signalling.

Exosomes from Human Gingiva-Derived Mesenchymal Stem Cells Combined with Biodegradable Chitin Conduits Promote Rat Sciatic Nerve Regeneration.

At present, repair methods for peripheral nerve injury often fail to get satisfactory result. Although various strategies have been adopted to investigate the microenvironment after peripheral nerve injury, the underlying molecular mechanisms of neurite outgrowth remain unclear. In this study, we evaluate the effects of exosomes from gingival mesenchymal stem cells (GMSCs) combined with biodegradable chitin conduits on peripheral nerve regeneration. GMSCs were isolated from human gingival tissue and characterized by surface antigen analysis and in vitro multipotent differentiation. The cell supernatant was collected to isolate the exosomes. The exosomes were characterized by transmission electron microscopy, Western blot, and size distribution analysis. The effects of exosomes on peripheral nerve regeneration in vitro were evaluated by coculture with Schwann cells and DRGs. The chitin conduit was prepared and combined with the exosomes to repair rat sciatic nerve defect. Histology, electrophysiology, and gait analysis were used to test the effects of exosomes on sciatic nerve function recovery in vivo. We have successfully cultured GMSCs and isolated exosomes. The exosomes from GMSCs could significantly promote Schwann cell proliferation and DRG axon growth. The in vivo studies showed that chitin conduit combined with exosomes from GMSCs could significantly increase the number and diameter of nerve fibers and promote myelin formation. In addition, muscle function, nerve conduction function, and motor function were also obviously recovered. In summary, this study suggests that GMSC-derived exosomes combined with biodegradable chitin conduits are a useful and novel therapeutic intervention in peripheral nerve repair.

Dexamethasone Leads to Upregulation of BMP6 and ACHE Suppression of SMAD3 and ESR1 Genes in Human Mesenchymal Stem Cells

The purpose of this study is to evaluate the effects of dexamethasone on mesenchymal stem cells from human gingiva using next-generation sequencing. The verification and suggestion of possible mechanisms was performed. Human gingiva-derived stem cells were treated with a final concentration of 10-7M dexamethasone at 2 and 24 hours. Extraction of RNA, sequencing of mRNA, gene ontology and pathway analysis were performed. Quantification by real-time polymerase chain reaction was conducted for validation. A fold change of two was applied for this study, and a log2 normalized read count of 5 or greater was applied to minimize false counts. Expression of SMAD3 and ESR1 was decreased in dexamethasone at 24 hours. Increased expression of BMP6 and ACHE was noted in dexamethasone at 24 hours. TGF-β signaling was involved in the target genes chosen for osteoblast differentiation. It was clear that the application of dexamethasone produced reduced expression of SMAD3 and ESR1 and enhanced expression of ACHE and BMP6 of human gingiva-derived mesenchymal stem cells. An RNA sequencing strategy can provide new insights into the mechanism of dexamethasone in stem cells originating from dental areas.

Updates on GMSCs Treatment for Autoimmune Diseases.

Autoimmune disease is a refractory disease. Accumulating Evidence has revealed that the manipulation of mesenchymal stem cells may have the potential to control or even treat autoimmune diseases. Human gingiva-derived mesenchymal stem cells (GMSCs) are emerging as a new line of mesenchymal stem cells that have displayed some potential advantages in controlling and treating autoimmune diseases. In this review, we briefly update the current understanding on the biology of GMSCs and their effects on preventing and treating autoimmune diseases. The availability of gingival mesenchymal stem cells (GMSCs), together with their potent capacity of multi-directional differentiation and inflammatory modulation, making GMSCs an ideal subtype of MSCs in treating autoimmune disease. Our and other studies have launched the earliest appraisal on GMSCs and carried out a lot of biological researches. The clinical trial of GMSCs on patients with autoimmune diseases will further approve their therapeutic effects, as well as its cellular and molecular mechanisms.

Human GMSCs Treat Murine Model with Aplastic Anemia Through Regulating the Balance between Treg and Th17 Cells

Severe aplastic anemia (AA) is a rare autoimmune disease characterized by severe pancytopenia and bone marrow failure. The recent finding of an abnormal quantity and function of T helper (Th) and regulatory T (Treg) cells in peripheral blood of patients has emerged as the major cause of the condition. Accumulating evidence has revealed that human gingiva-derived mesenchymal stem cells (GMSCs) are emerging as a new line of mesenchymal stem cells may have the potential to control or even treat autoimmune diseases through correcting the balance between Th and Treg cells. Given that GMSCs have a robust immune regulatory function and regenerative ability, we investigated the effect of GMSCs on preventing T cell-mediated bone marrow failure (BMF) in an AA mouse model. We observed that infusion of GMSCs markedly attenuated histological BM damage, controlled immunologically-mediated disorders in AA mice, significantly improved AA mice survival by significantly reducing cell infiltration of both CD8 cells and CD4 T cells (including Th1 and Th17 cells) in lymph nodes and decreased pro-inflammatory cytokines levels in lymph nodes and plasma. Moreover, GMSCs up-regulated the levels of Tregs and increased Foxp3 expression in lymph nodes. Our results demonstrate that the predominant Th1 and Th17 cell infiltration in the AA model could be significantly reduced while Treg cells are increased by GMSCs treatment. Collectively, our data demonstrate that GMSCs can attenuate T cell-mediated bone marrow failure through targeting the balance of Th17 and Treg cells, implicating that the use of GMSCs many have potential clinical application in prevention and treatment of patients with AA. Funding Statement: This study was supported by the National Key R&D Program of China (2017YFA0105800), the National Natural Science Foundation of China (No. 81671611), the Program for Guangdong Introducing Innovative and Entrepreneurial Teams (2016ZT06S252), the National Natural Science Foundation of Guangdong Province (No. 2014A030313131) and NIH R61 AR073409. Declaration of Interests: All authors have no conflict of interest. Ethics Approval Statement: This study was carried out in accordance with the recommendations of the animal use protocol, which was approved by the Institutional Animal Care and Use Committee of Sun Yat-Sen University (Approve number: [2018]02-195-01). Human gingiva samples were collected following routine dental procedures at the Division of Dentistry in the Third Hospital at the Sun Yat-sen University, which were approved by the medical ethics committees of Institutional Review Boards (IRB) at the Third Hospital at the Sun Yat-sen University.

Human Gingiva-Derived Mesenchymal Stem Cells Modulate Monocytes/Macrophages and Alleviate Atherosclerosis.

Atherosclerosis is the major cause of cardiovascular diseases. Current evidences indicate that inflammation is involved in the pathogenesis of atherosclerosis. Human gingiva-derived mesenchymal stem cells (GMSC) have shown anti-inflammatory and immunomodulatory effects on autoimmune and inflammatory diseases. However, the function of GMSC in controlling atherosclerosis is far from clear. The present study is aimed to elucidate the role of GMSC in atherosclerosis, examining the inhibition of GMSC on macrophage foam cell formation, and further determining whether GMSC could affect the polarization and activation of macrophages under different conditions. The results show that infusion of GMSC to AopE−/− mice significantly reduced the frequency of inflammatory monocytes/macrophages and decreased the plaque size and lipid deposition. Additionally, GMSC treatment markedly inhibited macrophage foam cell formation and reduced inflammatory macrophage activation, converting inflammatory macrophages to anti-inflammatory macrophages in vitro. Thus, our study has revealed a significant role of GMSC on modulating inflammatory monocytes/macrophages and alleviating atherosclerosis.