Vascular Pericyte-Derived Exosomes Inhibit Bone Resorption via Traf3.

Purpose: Blood vessels distribute cells, oxygen, and nutrients throughout the body to support tissue growth and balance. Pericytes and endothelial cells form the inner wall of blood vessels, crucial for organ development and tissue homeostasis by producing paracrine signaling molecules. In the skeletal system, pericyte-derived vascular factors along with angiogenic factors released by bone cells regulate angiogenesis and bone formation. Although the involvement of angiogenic factors and skeletal blood vessels in bone homeostasis is relatively clear, the role of pericytes and the underlying mechanisms remain unknown. Here, our objective was to elucidate the significance of pericytes in regulating osteoclast differentiation. Methods: We used tissue staining to detect the coverage of pericytes and osteoclasts in femoral tissues of osteoporotic mice and mice of different ages, analyzing their correlation. We developed mice with conditionally deleted pericytes, observing changes in bone mass and osteoclast activity using micro-computer tomography and tissue staining to detect the regulatory effect of pericytes on osteoclasts. Pericytes-derived exosomes (PC-EVs) were collected and co-cultured with monocytes that induce osteoclast differentiation to detect the effect of the former on the exosomes. Finally, the specific mechanism of PC-EVs regulating osteoclast differentiation was verified using RNA sequencing and Western blotting. Results: Our study indicates a significant correlation between pericytes and age-related bone resorption. Conditional deletion of pericytes activated bone resorption and led to osteopenia in vivo. We discovered that PC-EVs inhibited the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathway, which is mediated by tumor necrosis factor receptor-associated factor 3 (Traf3), negatively regulating osteoclast development and bone resorption. Silencing Traf3 in PC-EVs canceled their inhibitory effect on osteoclast differentiation. Conclusion: Our study provides a novel perspective into the regulatory role of pericytes on bone resorption and may provide potential strategies for developing novel anti-bone resorption therapies.

Bone-Targeted Extracellular Vesicles from Mesenchymal Stem Cells for Osteoporosis Therapy.

BACKGROUND: Current drugs used for osteoporosis therapy show strong adverse effects. Stem cell-derived extracellular vesicles (EVs) provide another choice for osteoporosis therapy. Mouse mesenchymal stem cells (mMSCs)-derived EVs promote bone regeneration; however, their clinical application is limited due to non-specific tissue targeting. Alendronate specifically targets bone tissue via hydroxyapatite. Therefore, EVs were combined with alendronate to generate Ale-EVs by "click chemistry" to facilitate EVs targeting bone via alendronate/hydroxyapatite binding. METHODS: Ale-EVs were characterized based on size using dynamic light scattering analysis and morphology was visualized by transmission electron microscopy. Hydroxyapatite affinity of Ale-EVs was detected by flow cytometry. Bone targeting of Ale-EVs was tested by ex vivo fluorescent imaging. Cell viability was assessed by using WST-8 reduction assay kit for testing the ability of Ale-EVs to promote mMSCs proliferation. Alkaline phosphatase experiment was used to detect ability of Ale-EVs to promote differentiation of mouse mesenchymal stem cells in vitro. Western blotting and Q-PCR assay were used to detect the early marker of osteogenic differentiation. Antiosteoporotic effects of Ale-EVs were detected in ovariectomy (OVX)-induced osteoporosis rat model. The safety of the Ale-EVs in vivo was measured by H&E staining and serum markers assay. RESULTS: In vitro, Ale-EVs had high affinity with hydroxyapatite. Also, ex vivo data indicated that Ale-EVs-DiD treatment of mice induced strong fluorescece in bone tissues compared with EVs-DiD group. Furthermore, results suggested that Ale-EVs promoted the growth and differentiation of mouse MSCs. They also protected against osteoporosis in ovariectomy (OVX)-induced osteoporotic rats. Ale-EVs were well tolerated and no side effects were found, indicating that Ale-EVs specifically target bone and can be used as a new therapeutic in osteoporosis treatment. CONCLUSION: We used the Ale-N3 to modify mouse mesenchymal stem cells-derived extracellular vesicles by copper-free "click chemistry" to generate a Ale-EVs system. The Ale-EVs had a high affinity for bone and have great potential for clinical applications in osteoporosis therapy with low systemic toxicity.

Study of a new bone-targeting titanium implant-bone interface.

New strategies involving bone-targeting titanium (Ti) implant-bone interface are required to enhance bone regeneration and osseointegration for orthopedic and dental implants, especially in osteoporotic subjects. In this study, a new dual-controlled, local, bone-targeting delivery system was successfully constructed by loading tetracycline-grafted simvastatin (SV)-loaded polymeric micelles in titania nanotube (TNT) arrays, and a bone-targeting Ti implant-bone interface was also successfully constructed by implanting the delivery system in vivo. The biological effects were evaluated both in vitro and in vivo. The results showed that Ti surfaces with TNT-bone-targeting micelles could promote cytoskeletal spreading, early adhesion, alkaline phosphatase activity, and extracellular osteocalcin concentrations of rat osteoblasts, with concomitant enhanced protein expression of bone morphogenetic protein (BMP)-2. A single-wall bone-defect implant model was established in normal and ovariectomized rats as postmenopausal osteoporosis models. Microcomputed tomography imaging and BMP-2 expression in vivo demonstrated that the implant with a TNT-targeting micelle surface was able to promote bone regeneration and osseointegration in both animal models. Therefore, beneficial biological effects were demonstrated both in vitro and in vivo, which indicated that the bone-targeting effects of micelles greatly enhance the bioavailability of SV on the implant-bone interface, and the provision of SV-loaded targeting micelles alone exhibits the potential for extensive application in improving local bone regeneration and osseointegration, especially in osteoporotic subjects.