Bone marrow mesenchymal stromal cell-derived exosomes (BMSC-Exos) can recruit stem cells for bone repair, with neovessels serving as the main migratory channel for stem cells to the injury site. However, existing exosome (Exo) delivery strategies cannot reach the angiogenesis phase following bone injury. To that end, an enzyme-sensitive Exo delivery material that responds to neovessel formation during the angiogenesis phase was designed in the present study to achieve spatiotemporally controlled Exo release. Herein, matrix metalloproteinase-1 (MMP1) was found to be highly expressed in neovascularized bone; as a result, we proposed an injectable MMP1-sensitive hydrogel microspheres (KGE) made using a microfluidic chip prepared by mixing self-assembling peptide (KLDL-MMP1), GelMA, and BMSC-Exos. The results revealed that KGE microspheres had a uniform diameter of 50-70 µm, ideal for minimally invasive injection and could release exosomes in response to MMP1 expression. In vitro experiments demonstrated that KGE had less cytotoxicity and could promote the migration and osteodifferentiation of BMSCs. Furthermore, in vivo experiments confirmed that KGE could promote bone repair during angiogenesis by recruiting CD90⁺ stem cells via neovessels. Collectively, our results suggest that injectable enzyme-responsive KGE microspheres could be a promising Exo-secreting material for accelerating neovascularized bone healing. STATEMENT OF SIGNIFICANCE: Exosomes can spread through blood vessels and activate stem cells to participate in bone repair, but under normal circumstances, exosomes lacking sustained-release delivery materials cannot be maintained until the angiogenesis phase. In this study, we found that MMP1 was highly expressed in neovascularized bone, then we proposed an MMP1-sensitive injectable microsphere that carries exosomes and responds temporally and spatially to neovascularization, which maximizes the ability of exosomes to recruit stem cells. Different from previous strategies that focus on promoting angiogenesis to accelerate bone healing, this is a brand new delivery strategy that is stimuli-responsive to neovessel formation. In addition, the preparation of self-assembled peptide microspheres by a microfluidic chip is also proposed for the first time.

Diabetic foot ulcer (DFU) is a common and severe complication of diabetes mellitus, the etiology of which remains insufficiently understood, particularly regarding the involvement of extracellular vesicles (EVs). In this study, nanoflow cytometry to detect EVs in DFU skin tissues is used and found a significant increase in the Translocase of Outer Mitochondrial Membrane 20 (TOM20)⁺ mitochondrial‐derived vesicles (MDVs). The role of MDVs in DFU is yet to be reported. Using single‐cell datasets, it is discovered that the increase in MDVs may be regulated by Sorting Nexin 9 (SNX9). In vitro experiments revealed that MDVs secreted by fibroblasts cultured in high glucose medium exhibited similar composition and protein enrichment results to those in DFU tissues, suggesting their potential as an ideal in vitro surrogate. These MDVs promoted apoptosis and intracellular oxidative stress, disrupted mitochondrial structure, and reduced aerobic metabolism in target cells. In vivo experiments also showed that MDV drops hindered wound healing in diabetic mice; however, this effect is rescued by SNX9 inhibitors, restoring mitochondrial dynamics and balance. Under high glucose conditions, MDVs significantly upregulated oxidative stress levels and induced mitochondrial dysfunction. This study proposes targeting MDVs as a potential therapeutic strategy for DFU.

A schematic overview of electrospinning technology and the structure of electrospun nanofibers.

To effectively treat osteoarthritis (OA), the existing inflammation must be reduced before the cartilage damage can be repaired; this cannot be achieved with a single type of extracellular vesicles (EVs). Here, a hydrogel complex with logic‐gates function is proposed that can spatiotemporally controlled release two types of EVs: interleukin 10 (IL‐10)⁺ EVs to promote M2 polarization of macrophage, and SRY‐box transcription factor 9 (SOX9)⁺ EVs to increase cartilage matrix synthesis. Following dose‐of‐action screening, the dual EVs are loaded into a matrix metalloporoteinase 13 (MMP13)‐sensitive self‐assembled peptide hydrogel (KM13E) and polyethylene glycol diacrylate/gelatin methacryloyl‐hydrogel microspheres (PGE), respectively. These materials are mixed to form a “microspheres‐in‐gel” KM13E@PGE system. In vitro, KM13E@PGE abruptly released IL‐10⁺ EVs after 3 days and slowly released SOX9⁺ EVs for more than 30 days. In vivo, KM13E@PGE increased the CD206⁺ M2 macrophage proportion in the synovial tissue and decreased the tumor necrosis factor‐α and IL‐1β levels. The aggrecan and SOX9 expressions in the cartilage tissues are significantly elevated following inflammation subsidence. This performance is not achieved using anti‐inflammatory or cartilage repair therapy alone. The present study provides an injectable, integrated delivery system with spatiotemporal control release of dual EVs, and may inspire logic‐gates strategies for OA treatment.

Current Ti‐6Al‐4V bone implants lack trabecular structure and pro‑angiogenic cues, both essential for regeneration. Herein, a dual biomimetic strategy is devised that integrates a 3D‐printed biomimetic trabecular porous Ti‐6Al‐4V scaffold (BTPS) with exosome‐loaded PEGDA/GelMA hydrogel microspheres (PGHExo) designed for sustained release. BTPS is designed using Voronoi algorithms and imaging data, and replicates the geometry and mechanical properties of natural bone. Hypoxia‐induced human umbilical vein endothelial cell (HUVEC) derived exosomes (HExo) are encapsulated in PGHExo microspheres via microfluidic technology, enabling controlled release of HExo, and anchored onto BTPS using polydopamine (pDA) modification (BTPS&pDA@PGHExo). BTPS exhibited an elastic modulus of ≈3.2 GPa and a permeability of 11.52 × 10 ⁻⁸ mm ² , mimicking natural bone. In vitro assays demonstrated that BTPS&pDA@PGHExo significantly enhanced osteogenesis and angiogenesis. mRNA‐Seq analysis suggested that BTPS&pDA@PGHExo regulates osteogenic and angiogenic gene expression through the activation of pathways including MAPK, mTOR, HIF‐1, and VEGF. In vivo, BTPS&pDA@PGHExo improved bone volume, density, and neovascularization in a rabbit model. This dual biomimetic strategy offers a promising clinical solution, addressing the limitations of conventional Ti‐6Al‐4V scaffolds and providing an innovative approach for personalized bone defect repair.

The slow proliferation rate and poor osteodifferentiation ability of inflammatory periodontal membrane stem cells extracted from periodontitis tissues (i-PDLSCs) account for poor efficiency in treating inflammatory bone loss. Exosomes reportedly have inducible and relatively stable components, allowing them to promote inflammatory bone repair, but obtaining i-PDLSCs exosomes with the ability to promote osteodifferentiation is challenging. In the present study, i-PDLSCs were extracted from periodontal membrane tissues of patients with severe periodontitis, and in vitro induction with gallic acid (GA) significantly promoted the proliferative activity of i-PDLSCs at a concentration of 10 mM, with TC₀ of 11.057 mM and TC₅₀ of 67.56 mM for i-PDLSCs. After mRNA sequencing, we found that GA could alleviate oxidative stress in i-PDLSCs and increase its mitochondrial membrane potential and glucose aerobic metabolism level, thus promoting the osteodifferentiation of i-PDLSCs. After exosomes of i-PDLSCs after GA induction (i-EXO-GA) were isolated by differential centrifugation, we found that 200 ug/mL of i-EXO-GA could remarkably promote the osteodifferentiation of i-PDLSCs. Overall, our results suggest that GA induction can enhance the proliferation and osteodifferentiation in primary cultures of i-PDLSCs in vitro, mediated by alleviating oxidative stress and glycometabolism levels in cells, which further influences the osteodifferentiation-promoting ability of i-EXO-GA. Overall, we provide a viable cell and exosome induction culture method for treating inflammatory alveolar defects associated with periodontitis.