Protective effects of omega-3 fatty acids in a blood-brain barrier-on-chip model and on postoperative delirium-like behaviour in mice.

BACKGROUND: Peripheral surgical trauma can trigger neuroinflammation and ensuing neurological complications, such as delirium. The mechanisms whereby surgery contributes to postoperative neuroinflammation remain unclear and without effective therapies. Here, we developed a microfluidic-assisted blood-brain barrier (BBB) device and tested the effects of omega-3 fatty acids on neuroimmune interactions after orthopaedic surgery. METHODS: A microfluidic-assisted BBB device was established using primary human cells. Tight junction proteins, vascular cell adhesion molecule 1 (VCAM-1), BBB permeability, and astrocytic networks were assessed after stimulation with interleukin (IL)-1ß and in the presence or absence of a clinically available omega-3 fatty acid emulsion (Omegaven®; Fresenius Kabi, Bad Homburg, Germany). Mice were treated 1 h before orthopaedic surgery with 10 µl g-1 body weight of omega-3 fatty acid emulsion i.v. or equal volumes of saline. Changes in pericytes, perivascular macrophages, BBB opening, microglial activation, and inattention were evaluated. RESULTS: Omega-3 fatty acids protected barrier permeability, endothelial tight junctions, and VCAM-1 after exposure to IL-1ß in the BBB model. In vivo studies confirmed that omega-3 fatty acid treatment inhibited surgery-induced BBB impairment, microglial activation, and delirium-like behaviour. We identified a novel role for pericyte loss and perivascular macrophage activation in mice after surgery, which were rescued by prophylaxis with i.v. omega-3 fatty acids. CONCLUSIONS: We present a new approach to study neuroimmune interactions relevant to perioperative recovery using a microphysiological BBB platform. Changes in barrier function, including dysregulation of pericytes and perivascular macrophages, provide new targets to reduce postoperative delirium.

Molecularly Tailored Interface for Long-Term Xenogeneic Cell Transplantation.

Encapsulation of therapeutic cells in a semipermeable device can mitigate the need for systemic immune suppression following cell transplantation by providing local immunoprotection while being permeable to nutrients, oxygen, and different cell-secreted biomolecules. However, fibrotic tissue deposition around the device has been shown to compromise the long-term function of the transplanted cells. Herein, a macroencapsulation device design that improves long-term survival and function of the transplanted cells is reported. The device is comprised of a semipermeable chitosan pouch with a tunable reservoir and molecularly engineered interface. The chitosan pouch interface decorated with 1,12-dodecanedioic acid (DDA), limits the cell adhesion and vigorous foreign body response while maintaining the barrier properties amenable to cell encapsulation. The device provides long-term protection to the encapsulated human primary hepatocytes in the subcutaneous space of immunocompetent mice. The device supports the encapsulated cells for up to 6 months as evident from cell viability and presence of human specific albumin in circulation. Solutions that integrate biomaterials and interfacial engineering such as the one described here may advance development of easy-to manufacture and retrievable devices for the transplantation of therapeutic cells in the absence of immunosuppression.

In vivo RNA editing of point mutations via RNA-guided adenosine deaminases.

We present in vivo sequence-specific RNA base editing via adenosine deaminases acting on RNA (ADAR) enzymes with associated ADAR guide RNAs (adRNAs). To achieve this, we systematically engineered adRNAs to harness ADARs, and comprehensively evaluated the specificity and activity of the toolsets in vitro and in vivo via two mouse models of human disease. We anticipate that this platform will enable tunable and reversible engineering of cellular RNAs for diverse applications.

In vivo engineering of bone tissues with hematopoietic functions and mixed chimerism.

Synthetic biomimetic matrices with osteoconductivity and osteoinductivity have been developed to regenerate bone tissues. However, whether such systems harbor donor marrow in vivo and support mixed chimerism remains unknown. We devised a strategy to engineer bone tissues with a functional bone marrow (BM) compartment in vivo by using a synthetic biomaterial with spatially differing cues. Specifically, we have developed a synthetic matrix recapitulating the dual-compartment structures by modular assembly of mineralized and nonmineralized macroporous structures. Our results show that these matrices incorporated with BM cells or BM flush transplanted into recipient mice matured into functional bone displaying the cardinal features of both skeletal and hematopoietic compartments similar to native bone tissue. The hematopoietic function of bone tissues was demonstrated by its support for a higher percentage of mixed chimerism compared with i.v. injection and donor hematopoietic cell mobilization in the circulation of nonirradiated recipients. Furthermore, hematopoietic cells sorted from the engineered bone tissues reconstituted the hematopoietic system when transplanted into lethally irradiated secondary recipients. Such engineered bone tissues could potentially be used as ectopic BM surrogates for treatment of nonmalignant BM diseases and as a tool to study hematopoiesis, donor-host cell dynamics, tumor tropism, and hematopoietic cell transplantation.

Three-Dimensional Monolayer Stress Microscopy.

Many biological processes involve the collective generation and transmission of mechanical stresses across cell monolayers. In these processes, the monolayer undergoes lateral deformation and bending because of the tangential and normal components of the cell-generated stresses. Monolayer stress microscopy (MSM) methods have been developed to measure the intracellular stress distribution in cell monolayers. However, current methods assume plane monolayer geometry and neglect the contribution of bending to the intracellular stresses. This work introduces a three-dimensional (3D) MSM method that calculates monolayer stress from measurements of the 3D traction stresses exerted by the cells on a flexible substrate. The calculation is carried out by imposing equilibrium of forces and moments in the monolayer, subject to external loads given by the 3D traction stresses. The equilibrium equations are solved numerically, and the algorithm is validated for synthetic loads with known analytical solutions. We present 3D-MSM measurements of monolayer stress in micropatterned islands of endothelial cells of different sizes and shapes. These data indicate that intracellular stresses caused by lateral deformation emerge collectively over long distances; they increase with the distance from the island edge until they reach a constant value that is independent of island size. On the other hand, bending-induced intracellular stresses are more concentrated spatially and remain confined to within one to two cell lengths of bending sites. The magnitude of these bending stresses is highest at the edges of the cell islands, where they can exceed the intracellular stresses caused by lateral deformations. Our data from nonpatterned monolayers suggests that biomechanical perturbations far away from monolayer edges also cause significant localized alterations in bending tension. The localized effect of bending-induced stresses may be important in processes like cellular extravasation, which are accompanied by significant normal deflections of a cell monolayer (i.e., the endothelium) and require localized changes in monolayer permeability.

In Situ Gene Therapy via AAV-CRISPR-Cas9-Mediated Targeted Gene Regulation.

Development of efficacious in vivo delivery platforms for CRISPR-Cas9-based epigenome engineering will be critical to enable the ability to target human diseases without permanent modification of the genome. Toward this, we utilized split-Cas9 systems to develop a modular adeno-associated viral (AAV) vector platform for CRISPR-Cas9 delivery to enable the full spectrum of targeted in situ gene regulation functionalities, demonstrating robust transcriptional repression (up to 80%) and activation (up to 6-fold) of target genes in cell culture and mice. We also applied our platform for targeted in vivo gene-repression-mediated gene therapy for retinitis pigmentosa. Specifically, we engineered targeted repression of Nrl, a master regulator of rod photoreceptor determination, and demonstrated Nrl knockdown mediates in situ reprogramming of rod cells into cone-like cells that are resistant to retinitis pigmentosa-specific mutations, with concomitant prevention of secondary cone loss. Furthermore, we benchmarked our results from Nrl knockdown with those from in vivo Nrl knockout via gene editing. Taken together, our AAV-CRISPR-Cas9 platform for in vivo epigenome engineering enables a robust approach to target disease in a genomically scarless and potentially reversible manner.

Calcium phosphate-bearing matrices induce osteogenic differentiation of stem cells through adenosine signaling.

Synthetic matrices emulating the physicochemical properties of tissue-specific ECMs are being developed at a rapid pace to regulate stem cell fate. Biomaterials containing calcium phosphate (CaP) moieties have been shown to support osteogenic differentiation of stem and progenitor cells and bone tissue formation. By using a mineralized synthetic matrix mimicking a CaP-rich bone microenvironment, we examine a molecular mechanism through which CaP minerals induce osteogenesis of human mesenchymal stem cells with an emphasis on phosphate metabolism. Our studies show that extracellular phosphate uptake through solute carrier family 20 (phosphate transporter), member 1 (SLC20a1) supports osteogenic differentiation of human mesenchymal stem cells via adenosine, an ATP metabolite, which acts as an autocrine/paracrine signaling molecule through A2b adenosine receptor. Perturbation of SLC20a1 abrogates osteogenic differentiation by decreasing intramitochondrial phosphate and ATP synthesis. Collectively, this study offers the demonstration of a previously unknown mechanism for the beneficial role of CaP biomaterials in bone repair and the role of phosphate ions in bone physiology and regeneration. These findings also begin to shed light on the role of ATP metabolism in bone homeostasis, which may be exploited to treat bone metabolic diseases.

Biomineralized matrices dominate soluble cues to direct osteogenic differentiation of human mesenchymal stem cells through adenosine signaling.

Stem cell differentiation is determined by a repertoire of signals from its microenvironment, which includes the extracellular matrix (ECM) and soluble cues. The ability of mesenchymal stem cells (MSCs), a common precursor for the skeletal system, to differentiate into osteoblasts and adipocytes in response to their local cues plays an important role in skeletal tissue regeneration and homeostasis. In this study, we investigated whether a bone-specific calcium phosphate (CaP) mineral environment could induce osteogenic differentiation of human MSCs, while inhibiting their adipogenic differentiation, in the presence of adipogenic-inducing medium. We also examined the mechanism through which the mineralized matrix suppresses adipogenesis of hMSCs to promote their osteogenic differentiation. Our results show that hMSCs cultured on mineralized matrices underwent osteogenic differentiation despite being cultured in the presence of adipogenic medium, which indicates the dominance of matrix-based cues of the mineralized matrix in directing osteogenic commitment of stem cells. Furthermore, the mineralized matrix-driven attenuation of adipogenesis was reversed with the inhibition of A2b adenosine receptor (A2bR), implicating a role of adenosine signaling in mineralized environment-mediated inhibition of adipogenesis. Such synthetic matrices with an intrinsic ability to direct differentiation of multipotent adult stem cells toward a targeted phenotype while inhibiting their differentiation into other lineages not only will be a powerful tool in delineating the role of complex microenvironmental cues on stem cell commitment but also will contribute to functional tissue engineering and their translational applications.

Rapid self-healing hydrogels.

Synthetic materials that are capable of autonomous healing upon damage are being developed at a rapid pace because of their many potential applications. Despite these advancements, achieving self-healing in permanently cross-linked hydrogels has remained elusive because of the presence of water and irreversible cross-links. Here, we demonstrate that permanently cross-linked hydrogels can be engineered to exhibit self-healing in an aqueous environment. We achieve this feature by arming the hydrogel network with flexible-pendant side chains carrying an optimal balance of hydrophilic and hydrophobic moieties that allows the side chains to mediate hydrogen bonds across the hydrogel interfaces with minimal steric hindrance and hydrophobic collapse. The self-healing reported here is rapid, occurring within seconds of the insertion of a crack into the hydrogel or juxtaposition of two separate hydrogel pieces. The healing is reversible and can be switched on and off via changes in pH, allowing external control over the healing process. Moreover, the hydrogels can sustain multiple cycles of healing and separation without compromising their mechanical properties and healing kinetics. Beyond revealing how secondary interactions could be harnessed to introduce new functions to chemically cross-linked polymeric systems, we also demonstrate various potential applications of such easy-to-synthesize, smart, self-healing hydrogels.

3D traction stresses activate protease-dependent invasion of cancer cells.

Cell invasion and migration that occurs, for example, in cancer metastasis is rooted in the ability of cells to navigate through varying levels of physical constraint exerted by the extracellular matrix. Cancer cells can invade matrices in either a protease-independent or a protease-dependent manner. An emerging critical component that influences the mode of cell invasion is the traction stresses generated by the cells in response to the physicostructural properties of the extracellular matrix. In this study, we have developed a reference-free quantitative assay for measuring three-dimensional (3D) traction stresses generated by cells during the initial stages of invasion into matrices exerting varying levels of mechanical resistance. Our results show that as cells encounter higher mechanical resistance, a larger fraction of them shift to protease-mediated invasion, and this process begins at lower values of cell invasion depth. On the other hand, the compressive stress generated by the cells at the onset of protease-mediated invasion is found to be independent of matrix stiffness, suggesting that 3D traction stress is a key factor in triggering protease-mediated cancer cell invasion. At low 3D compressive traction stresses, cells utilize bleb formation to indent the matrix in a protease independent manner. However, at higher stress values, cells utilize invadopodia-like structures to mediate protease-dependent invasion into the 3D matrix. The critical value of compressive traction stress at the transition from a protease-independent to a protease-dependent mode of invasion is found to be ∼165 Pa.

Differential roles of normal and lung cancer-associated fibroblasts in microvascular network formation.

Perfusable microvascular networks offer promising three-dimensional in vitro models to study normal and compromised vascular tissues as well as phenomena such as cancer cell metastasis. Engineering of these microvascular networks generally involves the use of endothelial cells stabilized by fibroblasts to generate robust and stable vasculature. However, fibroblasts are highly heterogenous and may contribute variably to the microvascular structure. Here, we study the effect of normal and cancer-associated lung fibroblasts on the formation and function of perfusable microvascular networks. We examine the influence of cancer-associated fibroblasts on microvascular networks when cultured in direct (juxtacrine) and indirect (paracrine) contacts with endothelial cells, discovering a generative inhibition of microvasculature in juxtacrine co-cultures and a functional inhibition in paracrine co-cultures. Furthermore, we probed the secreted factors differential between cancer-associated fibroblasts and normal human lung fibroblasts, identifying several cytokines putatively influencing the resulting microvasculature morphology and functionality. These findings suggest the potential contribution of cancer-associated fibroblasts in aberrant microvasculature associated with tumors and the plausible application of such in vitro platforms in identifying new therapeutic targets and/or agents that can prevent formation of aberrant vascular structures.

Branched poly-l-lysine for cartilage penetrating carriers.

Joint diseases, such as osteoarthritis, often require delivery of drugs to chondrocytes residing within the cartilage. However, intra-articular delivery of drugs to cartilage remains a challenge due to their rapid clearance within the joint. This problem is further exacerbated by the dense and negatively charged cartilage extracellular matrix (ECM). Cationic nanocarriers that form reversible electrostatic interactions with the anionic ECM can be an effective approach to overcome the electrostatic barrier presented by cartilage tissue. For an effective therapeutic outcome, the nanocarriers need to penetrate, accumulate, and be retained within the cartilage tissue. Nanocarriers that adhere quickly to cartilage tissue after intra-articular administration, transport through cartilage, and remain within its full thickness are crucial to the therapeutic outcome. To this end, we used ring-opening polymerization to synthesize branched poly(l-lysine) (BPL) cationic nanocarriers with varying numbers of poly(lysine) branches, surface charge, and functional groups, while maintaining similar hydrodynamic diameters. Our results show that the multivalent BPL molecules, including those that are highly branched (i.e., generation two), can readily adhere and transport through the full thickness of cartilage, healthy and degenerated, with prolonged intra-cartilage retention. Intra-articular injection of the BPL molecules in mouse knee joint explants and rat knee joints showed their localization and retention. In summary, this study describes an approach to design nanocarriers with varying charge and abundant functional groups while maintaining similar hydrodynamic diameters to aid the delivery of macromolecules to negatively charged tissues.

Grafting of cationic molecules to hyaluronic acid improves adsorption and cartilage lubrication.

Synovial fluid lubricates articular joints by forming a hydrated layer between the cartilage surfaces. In degenerative joint diseases like osteoarthritis (OA), the synovial fluid is compromised, which leads to less effective innate lubrication and exacerbated cartilage degeneration. Studies over the years have led to the development of partially or fully synthetic biolubricants to reduce the coefficient of friction with cartilage in knee joints. Cartilage-adhering, hydrated lubricants are particularly important to provide cartilage lubrication and chondroprotection under high normal load and slow speed. Here, we report the development of a hyaluronic acid (HA)-based lubricant functionalized with cationic branched poly-L-lysine (BPL) molecules that bind to cartilage via electrostatic interactions. We surmised that the electrostatic interactions between the BPL-modified HA molecules (HA-BPL) and the cartilage facilitate localization of the HA molecules to the cartilage surface. The number of BPL molecules on the HA backbone was varied to determine the optimal grafting density for cartilage binding and HA localization. Collectively, our results show that our HA-BPL molecules adhered readily to cartilage and were effective as a lubricant in cartilage-on-cartilage shear measurements where the modified HA molecules significantly reduce the coefficient of friction compared to phosphate-buffered saline or HA alone. This proof-of-concept study shows how the incorporation of cartilage adhering moieties, such as cationic molecules, can be used to enhance cartilage binding and lubrication properties of HA.

Active immunotherapy for C5a-mediated inflammation using adjuvant-free self-assembled peptide nanofibers.

The terminal protein in the complement cascade C5a is a potent inflammatory molecule and chemoattractant that is involved in the pathology of multiple inflammatory diseases including sepsis and arthritis, making it a promising protein to target with immunotherapies. Active immunotherapies, in which patients are immunized against problematic self-molecules and generate therapeutic antibodies as a result, have received increasing interest as an alternative to traditional monoclonal antibody treatments. In previous work, we have designed supramolecular self-assembling peptide nanofibers as active immunotherapies with defined combinations of B- and T-cell epitopes. Herein, the self-assembling peptide Q11 platform was employed to generate a C5a-targeting active immunotherapy. Two of three predicted B-cell epitope peptides from C5a were found to be immunogenic when displayed within Q11 nanofibers, and the nanofibers were capable of reducing C5a serum concentrations following immunization. Contrastingly, C5a's precursor protein C5 maintained its original concentration, promising to minimize side effects heretofore associated with C5-targeted therapies. Immunization protected mice against an LPS-challenge model of sepsis, and it reduced clinical severity in a model of collagen-antibody induced arthritis. Together, this work indicates the potential for targeting terminal complement proteins with active immunotherapies by leveraging the immunogenicity of self-assembled peptide nanomaterials. STATEMENT OF SIGNIFICANCE: Chronic inflammatory diseases such as rheumatoid arthritis, psoriasis, and inflammatory bowel disease are currently treated primarily with monoclonal antibodies against key inflammatory mediators. While helpful for many patients, they have high non-response rates, are costly, and commonly fail as anti-drug antibodies are raised by the patient. The approach we describe here explores a fundamentally different treatment paradigm: raising therapeutic antibody responses with an active immunotherapy. We employ innovative supramolecular peptide nanomaterials to elicit neutralizing antibody responses against complement component C5a and demonstrate therapeutic efficacy in preclinical mouse models of sepsis and rheumatoid arthritis. The strategy reported may represent a potential alternative to monoclonal antibody therapies.

Gene Therapy and Spinal Fusion: Systematic Review and Meta-Analysis of the Available Data.

OBJECTIVE: To analyze the extant literature describing the application of gene therapy to spinal fusion. METHODS: A systematic review of the English-language literature was performed. The search query was designed to include all published studies examining gene therapy approaches to promote spinal fusion. Approaches were classified as ex vivo (delivery of genetically modified cells) or in vivo (delivery of growth factors via vectors). The primary endpoint was fusion rate. Random effects meta-analyses were performed to calculate the overall odds ratio (OR) of fusion using a gene therapy approach and overall fusion rate. Subgroup analyses of fusion rate were also performed for each gene therapy approach. RESULTS: Of 1179 results, 35 articles met criteria for inclusion (all preclinical), of which 26 utilized ex vivo approaches and 9 utilized in vivo approaches. Twenty-seven articles (431 animals) were included in the meta-analysis. Gene therapy use was associated with significantly higher fusion rates (OR 77; 95% confidence interval {CI}: [31, 192]; P < 0.001); ex vivo strategies had a greater effect (OR 136) relative to in vivo strategies (OR 18) (P = 0.017). The overall fusion rate using a gene therapy approach was 80% (95% CI: [62%, 93%]; P < 0.001); overall fusion rates were significantly higher in subjects treated with ex vivo compared to in vivo strategies (90% vs. 42%; P = 0.011). For both ex vivo and in vivo approaches, the effect of gene therapy on fusion was independent of animal model. CONCLUSIONS: Gene therapy may augment spinal fusion; however, future investigation in clinical populations is necessary.

UDP-6-glucose dehydrogenase in hormonally responsive breast cancers.

Survival for metastatic breast cancer is low and thus, continued efforts to treat and prevent metastatic progression are critical. Estrogen is shown to promote aggressive phenotypes in multiple cancer models irrespective of estrogen receptor (ER) status. Similarly, UDP-Glucose 6-dehydrogenase (UGDH) a ubiquitously expressed enzyme involved in extracellular matrix precursors, as well as hormone processing increases migratory and invasive properties in cancer models. While the role of UGDH in cellular migration is defined, how it intersects with and impacts hormone signaling pathways associated with tumor progression in metastatic breast cancer has not been explored. Here we demonstrate that UGDH knockdown blunts estrogen-induced tumorigenic phenotypes (migration and colony formation) in ER+ and ER- breast cancer in vitro. Knockdown of UGDH also inhibits extravasation of ER- breast cancer ex vivo, primary tumor growth and animal survival in vivo in both ER+ and ER- breast cancer. We also use single cell RNA-sequencing to demonstrate that our findings translate to a human breast cancer clinical specimen. Our findings support the role of estrogen and UGDH in breast cancer progression provide a foundation for future studies to evaluate the role of UGDH in therapeutic resistance to improve outcomes and survival for breast cancer patients.

Grand Challenges at the Interface of Engineering and Medicine.

Over the past two decades Biomedical Engineering has emerged as a major discipline that bridges societal needs of human health care with the development of novel technologies. Every medical institution is now equipped at varying degrees of sophistication with the ability to monitor human health in both non-invasive and invasive modes. The multiple scales at which human physiology can be interrogated provide a profound perspective on health and disease. We are at the nexus of creating "avatars" (herein defined as an extension of "digital twins") of human patho/physiology to serve as paradigms for interrogation and potential intervention. Motivated by the emergence of these new capabilities, the IEEE Engineering in Medicine and Biology Society, the Departments of Biomedical Engineering at Johns Hopkins University and Bioengineering at University of California at San Diego sponsored an interdisciplinary workshop to define the grand challenges that face biomedical engineering and the mechanisms to address these challenges. The Workshop identified five grand challenges with cross-cutting themes and provided a roadmap for new technologies, identified new training needs, and defined the types of interdisciplinary teams needed for addressing these challenges. The themes presented in this paper include: 1) accumedicine through creation of avatars of cells, tissues, organs and whole human; 2) development of smart and responsive devices for human function augmentation; 3) exocortical technologies to understand brain function and treat neuropathologies; 4) the development of approaches to harness the human immune system for health and wellness; and 5) new strategies to engineer genomes and cells.

Effect of scaffold microarchitecture on osteogenic differentiation of human mesenchymal stem cells.

Design of macroporous synthetic grafts that can promote infiltration of cells, their differentiation, and synthesis of bone-specific extracellular matrix is a key determinant for in vivo bone tissue regeneration and repair. In this study, we investigated the effect of the microarchitecture of the scaffold on osteogenic differentiation of human mesenchymal stem cells (hMSCs). Poly(ethylene glycol) diacrylate-co-N-acryloyl 6-aminocaproic acid cryogels were fabricated to have either a pore network consisting of cellular, randomly oriented pores (termed 'spongy') or a pore network consisting of lamellar columns (termed 'columnar'), with both cryogel types showing a similar porosity. Both spongy and columnar cryogels supported comparable levels of cell viability and proliferation of hMSCs in vitro. However, spongy cryogels promoted osteogenic differentiation to a greater extent than their columnar counterparts, as evidenced by increased alkaline phosphatase activity and osteoblastic gene expression over 21 days post culture. Leveraging upon our previous work, we further evaluated the ability of these synthetic scaffolds in conjunction with mineralisation to promote ectopic bone formation upon subcutaneous implantation in nude rats. Mineralised spongy and columnar cryogels, both in the presence and absence of exogenous hMSCs, promoted ectopic bone formation in vivo. No such bone formation was observed in acellular cryogels devoid of mineralisation, with extensive host cell infiltration and vascularisation in columnar cryogels, and negligible infiltration into spongy cryogels. Our results thus present a novel method to tune the microarchitecture of porous polymeric scaffolds, in addition to suggesting their efficacy as synthetic bone grafts.

Extracellular adenosine signaling in bone health and disease.

Purinergic signaling is a key molecular pathway in the maintenance of bone health and regeneration. P1 receptor signaling, which is activated by extracellular adenosine, has emerged as a key metabolic pathway that regulates bone tissue formation, function, and homeostasis. Extracellular adenosine is mainly produced by ectonucleotidases, and alterations in the function of these enzymes or compromised adenosine generation can result in bone disorders, such as osteoporosis and impaired fracture healing. This mini review discusses the key role played by adenosine in bone health and how its alterations contribute to bone diseases, as well as potential therapeutic applications of exogenous adenosine to combat bone diseases like osteoporosis and injury.