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The oral and craniofacial region consists of various types of hard and soft tissues with the intricate organization. With the high prevalence of tissue defects in this specific region, it is highly desirable to enhance tissue regeneration through the development and use of engineered biomaterials. Collagen, the major component of tissue extracellular matrix, has come into the limelight in regenerative medicine. Although collagen has been widely used as an essential component in biomaterial engineering owing to its low immunogenicity, high biocompatibility, and convenient extraction procedures, there is a limited number of reviews on this specific clinic sector. The need for mechanical enhancement and functional engineering drives intensive efforts in collagen-based biomaterials concentrating on therapeutical outcomes and clinical translation in oral and craniofacial tissue regeneration. Herein, we highlighted the status quo of the design and applications of collagen-based biomaterials in oral and craniofacial tissue reconstruction. The discussion expanded on the inspiration from the leather tanning process on modifications of collagen-based biomaterials and the prospects of multi-tissue reconstruction in this particular dynamic microenvironment. The existing findings will lay a new foundation for the optimization of current collagen-based biomaterials for rebuilding oral and craniofacial tissues in the future. Graphical Abstract

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PDF file - 121K, Determination of M1 and M2 macrophages differentiated from bone marrow-derived macrophages (BMDMs) from Zoledronate (Zol)-treated C57BL/6 mice.

PDF file - 72K, Systemic infusion of human serum from BRONJ patients promotes BRONJ-like lesion development in C57BL/6 mice.

PDF file - 97K, Jaw bone marrows of mice exhibit elevated level of Th-17/IL-17 than long bone marrows.

Purpose: Osteonecrosis of the jaw (ONJ) is emerging as one of the important complications in cancer patients treated with antiresorptive agents. This study explored the potential role of interleukin (IL)-17–mediated M1/M2 macrophage alterations in the pathogenesis of bisphosphonate-related osteonecrosis of the jaw (BRONJ).Experimental Design: The expression of IL-17 and M1 and M2 macrophage markers at the local mucosal site of human BRONJ lesions was examined by immunofluorescence studies. BRONJ-like disease was induced in C57BL/6 mice and multiple myeloma-burdened mice by intravenous injection of zoledronate to evaluate the correlation of elevated IL-17 levels with changes in M1 and M2 macrophage phenotypes and the therapeutic effects of blocking IL-17 on pathogenesis of BRONJ-like disease.Results: Increased T-helper (TH)17 cells and IL-17 cytokine correlate with an increase in M1/M2 macrophages ratio at the local mucosal site of both murine and human BRONJ lesion. Convincingly, in mice burdened with multiple myeloma, a combination of elevated suprabasal level and drug-induced IL-17 activity augmented the incidence of BRONJ; both systemic increase of IL-17 and disease severity could be reversed by adoptive transfer of ex vivo expanded M2 macrophages. Targeting IL-17 via specific neutralizing antibodies or a small inhibitory molecule, laquinimod, significantly decreased M1/M2 ratio and concomitantly suppressed BRONJ-like condition in mice. Mechanistically, IL-17 enhanced IFN-γ–induced M1 polarization through augmenting STAT-1 phosphorylation while suppressing IL-4–mediated M2 conversion via inhibiting STAT-6 activation.Conclusions: These findings have established a compelling linkage between activated IL-17–mediated polarization of M1 macrophages and the development of BRONJ-like conditions in both human disease and murine models. Clin Cancer Res; 19(12); 3176–88. ©2013 AACR.

PDF file - 116K, The potential positive feedback loop between Th17/IL-17 and M1 macrophages involved in bisphosphonate-related osteonecrosis of the jaws (BRONJ).

PDF file - 444K, Representative microCT of BRONJ-like lesions and X-radiographs of normal healing of tibia in BgXID (III) nu/nu mice.

Mesenchymal stem cells (MSCs) influence T cells in health, disease and therapy through messengers of intercellular communication including extracellular vesicles (EVs). Apoptosis is a mode of cell death that tends to promote immune tolerance, and a large number of apoptotic vesicles (apoVs) are generated from MSCs during apoptosis. In an effort to characterize these apoVs and explore their immunomodulatory potential, here we show that after replenishing them systemically, the apoV deficiency in Fas mutant mice and pathological lymphoproliferation were rescued, leading to the amelioration of inflammation and lupus activity. ApoVs directly interacted with CD4(+) T cells and inhibited CD25 expression and IL-2 production in a dose-dependent manner. A broad range of Th1/2/17 subsets and cytokines including IFNγ, IL17A and IL-10 were suppressed while Foxp3(+) cells were maintained. Mechanistically, exposed phosphatidylserine (PtdSer/PS) on apoVs mediated the interaction with T cells to disrupt proximal T cell receptor signaling transduction. Remarkably, administration of apoVs prevented Th17 differentiation and memory formation, and ameliorated inflammation and joint erosion in murine arthritis. Collectively, our findings unveil a previously unrecognized crosstalk between MSC apoVs and CD4(+) T cells and suggest a promising therapeutic use of apoVs for autoimmune diseases.

Diabetes is a major public health issue because of its widely epidemic nature and lack of cure. Here, we show that pancreas-derived mesenchymal stem cells (PMSCs) are capable of regenerating exocrine pancreas when implanted into the kidney capsule of mice with streptozotocin (STZ)–induced diabetes. Mechanistically, we found that the regenerated exocrine pancreas elevated interleukin-6 (IL-6) in PMSC implants, which transiently activated tumor necrosis factor–α (TNF-α) and interferon-γ (IFN-γ) to inhibit IL-17, thereby rescuing damaged exocrine pancreas and islet β cells. In addition, we used knockout mouse models to show that global lack of IL-6, TNF-α, or IFN-γ resulted in increased severity of STZ-induced diabetes and resistance to PMSC implantation therapy, confirming the roles of these factors in safeguarding pancreatic β cells. Furthermore, removal of the kidney capsule PMSC implants at 28 days after implantation did not affect the PMSC-initiated therapeutic effect on diabetic mice. This study reveals a previously unknown role of exocrine pancreas regeneration in safeguarding β cells and demonstrates a “soil-rescues-seed” strategy for type 1 diabetes therapy.

Soft tissue reconstruction has remained a major clinical challenge in dentistry and regenerative medicine. Although current methods have shown partial success, there are several disadvantages associated with these approaches. Gingival mesenchymal stem cells (GMSCs) can be simply obtained in the oral cavity for soft tissue augmentation. Regenerative potential of mesenchymal stem cells (MSCs) encapsulated in hydrogels is well documented. Here, an alginate-gelatin methacrylate (GelMA) hydrogel formulation is developed encapsulating GMSCs within the developed hydrogel. The results confirm that the encapsulated MSCs remain viable within the hydrogel with enhanced collagen deposition. An excisional wound model in mice is utilized to evaluate the in vivo functionality of the GMSC-hydrogel construct for wound healing and soft tissue regeneration. The histology and immunofluorescence analyses confirm the effectiveness of the GMSC-hydrogel in expediting wound healing via enhancing angiogenesis and suppressing local proinflammatory cytokines. Altogether, the findings demonstrate that GMSCs encapsulated in an engineered hydrogel sheet based on alginate and GelMA can be used to expedite wound healing and soft tissue regeneration, with potential applications in plastic and reconstructive surgery as well as dentistry.

Gene regulatory divergence is thought to play a central role in determining human-specific traits. However, our ability to link divergent regulation to divergent phenotypes is limited. Here, we utilized human-chimpanzee hybrid induced pluripotent stem cells to study gene expression separating these species. The tetraploid hybrid cells allowed us to separate cis- from trans-regulatory effects, and to control for non-genetic confounding factors. We differentiated these cells into cranial neural crest cells (CNCCs), the primary cell type giving rise to the face. We discovered evidence of lineage-specific selection on the hedgehog signaling pathway, including a human-specific 6-fold down-regulation of EVC2 (LIMBIN), a key hedgehog gene. Inducing a similar down-regulation of EVC2 substantially reduced hedgehog signaling output. Mice and humans lacking functional EVC2 show striking phenotypic parallels to human-chimpanzee craniofacial differences, suggesting that the regulatory divergence of hedgehog signaling may have contributed to the unique craniofacial morphology of humans.

Sensorineural hearing loss in mammals occurs due to irreversible damage to the sensory epithelia of the inner ear and has very limited treatment options. The ability to regenerate the auditory progenitor cells is a promising approach for the treatment of sensorineural hearing loss; therefore, finding an appropriate and easily accessible stem cell source for restoring the sense of hearing would be of great interest. Here, we proposed a novel easy-to-access source of cells with the ability to recover auditory progenitor cells. In this study, gingival mesenchymal stem cells (GMSCs) were utilized, as these cells have high self-renewal and multipotent differentiation capacity and can be obtained easily from the oral cavity or discarded tissue samples at dental clinics. To manipulate the biophysical properties of the cellular microenvironment for promoting GMSC differentiation toward the target cells, we also tried to propose a candidate biomaterial. GMSCs in combination with an appropriate scaffold material can, therefore, present advantageous therapeutic options for a number of conditions. Here, we report the potential of GMSCs to differentiate into auditory progenitor cells while supporting them with an optimized three-dimensional scaffold and certain growth factors. A hybrid hydrogel scaffold based on peptide modified alginate and Matrigel was used here in addition to the presence of fibroblast growth factor-basic (bFGF), insulin-like growth factor (IGF), and epidermal growth factor (EGF). Our in vitro and in vivo studies confirmed the auditory differentiation potential of GMSCs within the engineered microenvironment.

Osteoconductive hydrogels can be fabricated by incorporating necessary growth factors and bioactive particles or simply utilizing the ability of the hydrogel itself to induce bone regeneration. The osteogenic inductive potential of the bioactive glass microparticles (BG MPs) has been well-studied. However, the role of the hydrogel embedding the BG MPs on the osteogenic differentiation of the encapsulated stem cells has not been well established. Moreover, it has been reported that the dental pulp stem cell (DPSC) has the capability of regenerating the craniofacial bone tissue when receiving the necessary osteogenic signals from the microenvironment. In the current study, we have fabricated a composite hydrogel based on alginate and Matrigel containing BG MPs and evaluated the role of the BG MPs and Matrigel on osteogenic differentiation of the encapsulated DPSCs. Our results confirmed that presence of Matrigel enhances the osteogenic differentiation of the DPSCs regardless of the decrease in elasticity of the hydrogel in presence of the BG MPs. This strategy of modifying the microenvironment can be a promising approach for craniofacial bone tissue engineering.

Over 300 billion of cells die every day in the human body, producing a large number of endogenous apoptotic extracellular vesicles (apoEVs). Also, allogenic stem cell transplantation, a commonly used therapeutic approach in current clinical practice, generates exogenous apoEVs. It is well known that phagocytic cells engulf and digest apoEVs to maintain the body's homeostasis. In this study, we show that a fraction of exogenous apoEVs is metabolized in the integumentary skin and hair follicles. Mechanistically, apoEVs activate the Wnt/β-catenin pathway to facilitate their metabolism in a wave-like pattern. The migration of apoEVs is enhanced by treadmill exercise and inhibited by tail suspension, which is associated with the mechanical force-regulated expression of DKK1 in circulation. Furthermore, we show that exogenous apoEVs promote wound healing and hair growth

Growth-factor-free bone regeneration remains a challenge in craniofacial engineering. Here, we engineered an osteogenic niche composed of a commercially modified alginate hydrogel and whitlockite microparticles (WHMPs), which impart tunable physicochemical properties that can direct osteogenesis of human gingival mesenchymal stem cells (GMSCs). Our in vitro studies demonstrate that WHMPs induce osteogenesis of GMSCs more effectively than previously demonstrated hydroxyapatite microparticles (HApMPs). Alginate-WHMP hydrogels showed higher elasticity without any adverse effects on the viability of the encapsulated GMSCs. Moreover, the alginate-WHMP hydrogels upregulate the mitogen-activated protein kinase (MAPK) pathway, which in turn orchestrates several osteogenic markers, such as RUNX2 and OCN, in the encapsulated GMSCs. Concurrent coculture studies with human osteoclasts demonstrate that GMSCs encapsulated in alginate-WHMP hydrogels downregulate osteoclastic activity, potentially due to release of Mg2+ ions from the WHMPs along with secretion of osteoprotegerin from the GMSCs. In vivo studies demonstrated that the GMSCs encapsulated in our osteogenic niche were able to promote bone repair in calvarial defects in murine models. Altogether, our results confirmed the development of a promising treatment modality for craniofacial bone regeneration based on an injectable growth-factor-free hydrogel delivery system.

Type 1 diabetes (T1D) imposes a significant health burden by negatively affecting tissue regeneration during wound healing. The adverse effect of diabetes is attributed to high levels of inflammation, but the cellular mechanisms responsible remain elusive. In this study, we show that intrinsic skeletal stem cells (SSCs), a subset of mesenchymal stem cells, are essential for resolution of inflammation to occur during osseous healing by using genetic approaches to selectively ablate SSCs. T1D caused aberrant nuclear factor-κB (NF-κB) activation in SSCs and substantially enhanced inflammation in vivo. Constitutive or tamoxifen-induced inhibition of NF-κB in SSCs rescued the impact of diabetes on inflammation, SSC expansion, and tissue formation. In contrast, NF-κB inhibition in chondrocytes failed to reverse the effect of T1D. Mechanistically, diabetes caused defective proresolving macrophage (M2) polarization by reducing TGF-β1 expression by SSCs, which was recovered by NF-κB inhibition or exogenous TGF-β1 treatment. These data identify an underlying mechanism for altered healing in T1D and demonstrate that diabetes induces NF-κB hyperactivation in SSCs to disrupt their ability to modulate M2 polarization and resolve inflammation.

Long bone development involves the embryonic formation of a primary ossification center (POC) in the incipient diaphysis followed by postnatal development of a secondary ossification center (SOC) at each epiphysis. Studies have elucidated major basic mechanisms of POC development, but relatively little is known about SOC development. To gain insights into SOC formation, we used Col2-Cre Rosa-tdTomato (Col2/Tomato) reporter mice and found that their periarticular region contained numerous Tomato-positive lineage cells expressing much higher Tomato fluorescence (termed TomatoH) than underlying epiphyseal chondrocytes (termed TomatoL). With time, the TomatoH cells became evident at the SOC invagination site and cartilage canal, increased in number in the expanding SOC, and were present as mesenchymal lineage cells in the subchondral bone. These data were verified in two mouse lineage tracing models, Col2-CreER Rosa-tdTomato and Gli1-CreER Rosa-tdTomato. In vitro tests showed that the periarticular TomatoH cells from Col2/Tomato mice contained mesenchymal progenitors with multidifferentiation abilities. During canal initiation, the cells expressed vascular endothelial growth factor (VEGF) and migrated into epiphyseal cartilage ahead of individual or clusters of endothelial cells, suggesting a unique role in promoting vasculogenesis. Later during SOC expansion, chondrocytes in epiphyseal cartilage expressed VEGF, and angiogenic blood vessels preceded TomatoH cells. Gene expression analyses of microdissected samples revealed upregulation of MMPs in periarticular cells at the invagination site and suggested potential roles for novel kinase and growth factor signaling pathways in regulating SOC canal initiation. In summary, our data indicate that the periarticular region surrounding epiphyseal cartilage contains mesenchymal progenitors that initiate SOC development and form subchondral bone. Stem Cells 2019;37:677–689

Mesenchymal stem cell (MSC)-derived exosome plays a central role in the cell-free therapeutics involving MSCs and the contents can be customized under disease-associated microenvironments. However, optimal MSC-preconditioning to enhance its therapeutic potential is largely unknown. Here, we show that preconditioning of gingival tissue-derived MSCs (GMSCs) with tumor necrosis factor-alpha (TNF-α) is ideal for the treatment of periodontitis. TNF-α stimulation not only increased the amount of exosome secreted from GMSCs, but also enhanced the exosomal expression of CD73, thereby inducing anti-inflammatory M2 macrophage polarization. The effect of GMSC-derived exosomes on inflammatory bone loss were examined by ligature-induced periodontitis model in mice. Local injection of GMSC-derived exosomes significantly reduced periodontal bone resorption and the number of tartrate-resistant acid phosphatase (TRAP)-positive osteoclasts, and these effects were further enhanced by preconditioning of GMSCs with TNF-α. Thus, GMSC-derived exosomes also exhibited anti-osteoclastogenic activity. Receptor activator of NF-κB ligand (RANKL) expression was regulated by Wnt5a in periodontal ligament cells (PDLCs), and exosomal miR-1260b was found to target Wnt5a-mediated RANKL pathway and inhibit its osteoclastogenic activity. These results indicate that significant ability of the TNF-α-preconditioned GMSC-derived exosomes to regulate inflammation and osteoclastogenesis paves the way for establishment of a therapeutic approach for periodontitis.

Oral mucositis (OM), a common debilitating toxicity associated with chemo- and radiation therapies, is a significant unmet clinical need for head and neck cancer patients. The biological complexities of chemoradiotherapy-induced OM involve interactions among disrupted tissue structures, inflammatory infiltrations, and oral microbiome, whereby several master inflammatory pathways constitute the complicated regulatory networks. Oral mucosal damages triggered by chemoradiotherapy-induced cell apoptosis were further exacerbated by the amplified inflammatory cascades dominantly governed by the innate immune responses. The coexistence of microbiome and innate immune components in oral mucosal barriers indicates that a signaling hub coordinates the interaction between environmental cues and host cells during tissue and immune homeostasis. Dysbiotic shifts in oral microbiota caused by cytotoxic cancer therapies may also contribute to the progression and severity of chemoradiotherapy-induced OM. In this review, we have updated the mechanisms involving innate immunity-governed inflammatory cascades in the pathobiology of chemoradiotherapy-induced OM and the development of new interventional targets for the management of this severe morbidity in head and neck cancer patients.

Bone marrow mesenchymal lineage cells are a heterogeneous cell population involved in bone homeostasis and diseases such as osteoporosis. While it is long postulated that they originate from mesenchymal stem cells, the true identity of progenitors and their in vivo bifurcated differentiation routes into osteoblasts and adipocytes remain poorly understood. Here, by employing large scale single cell transcriptome analysis, we computationally defined mesenchymal progenitors at different stages and delineated their bi-lineage differentiation paths in young, adult and aging mice. One identified subpopulation is a unique cell type that expresses adipocyte markers but contains no lipid droplets. As non-proliferative precursors for adipocytes, they exist abundantly as pericytes and stromal cells that form a ubiquitous 3D network inside the marrow cavity. Functionally they play critical roles in maintaining marrow vasculature and suppressing bone formation. Therefore, we name them marrow adipogenic lineage precursors (MALPs) and conclude that they are a newly identified component of marrow adipose tissue.

Mesenchymal stem cells (MSCs) closely interact with the immune system, and they are known to secrete inflammatory cytokines in response to stress stimuli. The biological function of MSC-derived inflammatory cytokines remains elusive. Here, we reveal that even under physiological conditions, MSCs produce and release a low level of tumor necrosis factor alpha (TNFα), which is unexpectedly required for preserving the self-renewal and differentiation of MSCs via autocrine/paracrine signaling. Furthermore, TNFα critically maintains MSC function in vivo during bone homeostasis. Mechanistically, we unexpectedly discovered that physiological levels of TNFα safeguard MSC homeostasis in a receptor-independent manner through mechanical force-driven endocytosis and that endocytosed TNFα binds to mammalian target of rapamycin (mTOR) complex 2 and restricts mTOR signaling. Importantly, inhibition of mTOR signaling by rapamycin serves as an effective osteoanabolic therapeutic strategy to protect against TNFα deficiency and mechanical unloading. Collectively, these findings unravel the physiological framework of the dynamic TNFα shuttle-based mTOR equilibrium that governs MSC and bone homeostasis.

Background Immunosuppression risks are a major concern with vascularized composite allotransplantation (VCA). As an emerging strategy, the antirejection role played by mesenchymal stem cells (MSCs) is receiving attention. However, the current literature reports are inconclusive regarding the robustness of the MSC monotherapy. Using a rat forelimb VCA model, this study tested the robustness of the immunomodulation efficacy of gingival-derived MSCs (GMSCs) and bone marrow–derived MSCs (BMMSCs). Methods Forelimbs were transplanted on pairs of major histocompatibility complex–incompatible rats (Wistar-Kyoto donor, Lewis [LEW] recipient). Twenty-four LEW rats were randomly divided into four groups, including control (no treatment) and three treatment groups: rapamycin (2 mg/kg/day for 28 days, postoperatively), BMMSC and GMSC, both of which received donor-derived stem cells administered intravenously on postoperative days (PODs) 0, 3, 7, and 14. Rejection was considered as 80% skin necrosis of the allograft. Microcomputed tomography (µCT) was performed to evaluate healing at osteosynthesis site. On POD 14, limbs from each group underwent histological analysis and rejection grading using the Banff system. Results Both BMMSC (15.0 days) and GMSC (14.7 days) treatment failed to prolong VCA survival in comparison with the control group (13.8 days; p > 0.050), while the rapamycin significantly delayed acute VCA rejection (24.5 days; p = 0.003). Micro-CT imaging revealed no gross visual difference across all groups. Histology revealed that the control group was most severely affected (grades III and IV) followed by MSC (grade II) and rapamycin (grade I). Conclusion MSC monotherapy, both BMMSC and GMSC, did not inhibit rejection in our VCA model. Skin immunogenicity is an important issue in promoting rejection, and a concomitant immunosuppression regimen should be considered to prolong allograft survival.

Mesenchymal stem cell (MSC)–derived exosome plays a central role in the cell-free therapeutics involving MSCs and the contents can be customized under disease-associated microenvironments. However, optimal MSC-preconditioning to enhance its therapeutic potential is largely unknown. Here, we show that preconditioning of gingival tissue-derived MSC (GMSC) with tumor necrosis factor-alpha (TNF-α) is ideal for the treatment of periodontitis. TNF-α stimulation not only increased the amount of exosome secreted from GMSC, but also enhanced the exosomal expression of CD73, thereby inducing anti-inflammatory M2 macrophage polarization. The effect of GMSC-derived exosomes on inflammatory bone loss were examined by ligature-induced periodontitis model in mice. Local injection of GMSCs-derived exosomes significantly reduced periodontal bone resorption and the number of tartrate-resistant acid phosphatase (TRAP)-positive osteoclasts, and these effects were further enhanced by preconditioning of GMSCs with TNF-α. Thus, GMSCs-derived exosomes also exhibited anti-osteoclastogenic activity. Receptor activator of NF-κB ligand (RANKL) expression was regulated by Wnt5a in periodontal ligament cells (PDLCs), and exosomal miR-1260b was found to target Wnt5a-mediated RANKL pathway and inhibit its osteoclastogenic activity. These results indicate that significant ability of the TNF-α-preconditioned GMSC-derived exosomes to regulate inflammation and osteoclastogenesis paves the way for establishment of a new therapeutic approach for periodontitis.

The preservation of vital dental pulp with vasculature and nerve components remains one of the most significant challenges in modern dentistry. Due to the immense potential for neurovascularization, mesenchymal stem cell (MSC) transplantation has shown emerging promise in regenerative medicine and dental translational practice. Actually, pulp mesenchymal stem cells, including postnatal dental pulp stem cells (from permanent teeth) and stem cells from human exfoliated deciduous teeth, possess unique properties based on their origins from neural crest or glial cells. Furthermore, they reside in a neurovascular niche and have the potential for neurogenesis, angiogenesis, and neurovascular inductive activity. According to current pulp regeneration strategies, pulp stem cell–mediated approaches to regeneration have demonstrated convincing evidence that they can rebuild the complex histologic structure of native pulp in situ with highly organized physiologic patterns or even achieve de novo regeneration of complete dental pulp tissues. More importantly, recent clinical studies emphasized in situ neurovascularization outcomes in successful regeneration of vitalized pulp via pulp stem cell transplantation. In this review, we summarize recent breakthroughs in pulp stem cell–mediated pulp regeneration, emphasizing the crucial achievement of neurovascularization. This functional pulp regeneration represents an innovative and promising approach for future regenerative endodontics.

Ten-eleven translocation (Tet) family-mediated DNA oxidation represents an epigenetic modification capable of converting 5-methylcytosine (5-mC) to 5-hydroxymethylcytosine (5-hmC), which regulates various biological processes. However, it is unknown whether Tet family affects mesenchymal stem cells (MSCs) or the skeletal system. Here we show that depletion of Tet1 and Tet2 results in impaired self-renewal and differentiation of bone marrow MSCs (BMMSCs) and a significant osteopenia phenotype. Tet1 and Tet2 deficiency reduces demethylation of the P2rX7 promoter and downregulates exosome release, leading to intracellular accumulation of miR-297a-5p, miR-297b-5p, and miR-297c-5p. These miRNAs inhibit Runx2 signaling to impair BMMSC function. We show that overexpression of P2rX7 rescues the impaired BMMSCs and osteoporotic phenotype in Tet1 and Tet2 double knockout mice. These results indicate that Tet1 and Tet2 play a critical role in maintaining BMMSC and bone homeostasis through demethylation of P2rX7 to control exosome and miRNA release. This Tet/P2rX7/Runx2 cascade may serve as a target for the development of novel therapies for osteopenia disorders., Tet-mediated DNA oxidation converts 5-methylcytosine (5-mC) to 5-hydroxymethylcytosine (5-hmC), which is essential to regulate different biological processes. Here the authors show that Tet1 and Tet2 regulate mesenchymal stem cell and bone homeostasis through demethylation of P2rX7 promoter.