Despite the therapeutic promise of stem cell aggregates in bone regeneration, clinical outcomes remain unstable due to the complex and incompletely understood fate of transplanted cells. Here, we investigated the role of programmed cell death in this process, moving beyond the established paradigm of apoptosis. Our study demonstrates that stem cell aggregates (CAs) from human exfoliated deciduous teeth (SHEDs) implanted into a murine femoral defect model undergo necroptosis in a time-dependent manner. Pre-induction of necroptosis in CAs prior to transplantation profoundly impaired bone repair, significantly reducing new bone formation, osteogenic differentiation, and vascularization at the defect site. Conversely, pharmacological inhibition of necroptosis using Necrostatin-1 rescued the regenerative capacity. Crucially, we identified that this inhibitory effect is primarily mediated by extracellular vesicles released during necroptosis (Nec-EVs). Inhibition of Nec-EV biogenesis with GW4869 restored bone healing. Characterization confirmed the successful isolation of phosphorylated mixed lineage kinase domain-like protein (pMLKL)-enriched Nec-EVs. Proteomic analysis revealed a distinct cargo profile in Nec-EVs, notably enriched in proteins involved in translation regulation and RNA metabolism. Functionally, Nec-EVs directly suppressed the proliferation and osteogenic potential of bone marrow stem cells (BMSCs) in vitro, and their local application in vivo was sufficient to recapitulate the impaired bone regeneration phenotype. Collectively, this work establishes necroptosis and its associated vesicular signaling as novel detrimental axes in cell-based bone regeneration, providing mechanistic insights and potential therapeutic targets for enhancing the reliability of regenerative therapies.

Healthy aging is a common goal for humanity and society, and one key to achieving it is the rejuvenation of senescent resident stem cells and empowerment of aging organ regeneration. However, the mechanistic understandings of stem cell senescence and the potential strategies to counteract it remain elusive. Here, we reveal that the aging bone microenvironment impairs the Golgi apparatus thus diminishing mesenchymal stem cell (MSC) function and regeneration. Interestingly, replenishment of cell aggregates-derived extracellular vesicles (CA-EVs) rescues Golgi dysfunction and empowers senescent MSCs through the Golgi regulatory protein Syntaxin 5. Importantly, in vivo administration of CA-EVs significantly enhanced the bone defect repair rate and improved bone mass in aging mice, suggesting their therapeutic value for treating age-related osteoporosis and promoting bone regeneration. Collectively, our findings provide insights into Golgi regulation in stem cell senescence and bone aging, which further highlight CA-EVs as a potential rejuvenative approach for aging bone regeneration.

Infection control and bone regeneration remain critical challenges in bone defect treatment. We developed a 3D-printed scaffold incorporating copper-based metal–organic framework-74 (Cu-MOF-74) within a polycaprolactone/hydroxyapatite composite. The synthesized Cu-MOF-74 exhibited a well-defined crystalline structure and rod-like morphology, as confirmed by TEM, EDS, FTIR, and XRD analyses. The scaffolds exhibited hierarchical pores (100–200 μm) and demonstrated tunable hydrophilicity, as evidenced by the water contact angles decreasing from 103.3 ± 2.02° (0% Cu-MOF-74) to 63.60 ± 1.93° (1% Cu-MOF-74). A biphasic Cu2+ release profile was observed from the scaffolds, reaching cumulative concentrations of 98.97 ± 3.10 ppm by day 28. Antimicrobial assays showed concentration-dependent efficacy, with 1% Cu-MOF-74 scaffolds achieving 90.07 ± 1.94% and 80.03 ± 2.17% inhibition against Staphylococcus aureus and Escherichia coli, respectively. Biocompatibility assessments using bone marrow-derived mesenchymal stem cells revealed enhanced cell proliferation at Cu-MOF-74 concentrations ≤ 0.2%, while concentrations ≥ 0.5% induced cytotoxicity. Osteogenic differentiation studies highlighted elevated alkaline phosphatase activity and mineralization in scaffolds with 0.05–0.2% Cu-MOF-74 scaffolds, particularly at 0.05% Cu-MOF-74 scaffolds, which exhibited the highest calcium deposition and upregulation of bone sialoprotein and osteopontin expression. These findings demonstrate the dual functional efficacy of Cu-MOF-74/PCL/HAp scaffolds in promoting both infection control and bone regeneration. These optimized Cu-MOF-74 concentrations (0.05–0.2%) effectively balance antimicrobial and osteogenic properties, presenting a promising strategy for bone defect repair in clinical applications.

Extracellular Vesicles In article number 2300019, Bing-Dong Sui, Yan Jin, Fang Jin, and co-workers discover mesenchymal stem cell aggregates secrete angiogenic extracellular vesicles, which exert high efficacy in boosting CD31+EMCN+ vessels with adjacent cell proliferation and provide an effective therapy to alleviate non-healing diabetic wounds.

The blood vessel system is essential for skin homeostasis and regeneration. While the heterogeneity of vascular endothelial cells has been emergingly revealed, whether a regeneration-relevant vessel subtype exists in skin remains unknown. Herein, a specialized vasculature in skin featured by simultaneous CD31 and EMCN expression contributing to the regeneration process is identified, the decline of which functionally underlies the impaired angiogenesis of diabetic nonhealing wounds. Moreover, enlightened by the developmental process that mesenchymal condensation induces angiogenesis, it is demonstrated that mesenchymal stem/stromal cell aggregates (CAs) provide an efficacious therapy to enhance regrowth of CD31+ EMCN+ vessels in diabetic wounds, which is surprisingly suppressed by pharmacological inhibition of extracellular vesicle (EV) release. It is further shown that CAs promote secretion of angiogenic protein-enriched EVs by proteomic analysis, which directly exert high efficacy in boosting CD31+ EMCN+ vessels and treating nonhealing diabetic wounds. These results add to the current knowledge on skin vasculature and help establish feasible strategies to benefit wound healing under diabetic condition.

Mesenchymal stem cells (MSCs), characterized by their self-renewal ability and multilineage differentiation potential, can be derived from various sources and are emerging as promising candidates for regenerative medicine, especially for regeneration of the tooth, bone, cartilage, and skin. The self-assembled approach of MSC aggregation, which notably constructs cell clusters mimicking the developing mesenchymal condensation, allows high-density stem cell delivery along with preserved cell-cell interactions and extracellular matrix (ECM) as the microenvironment niche. This method has been shown to enable efficient cell engraftment and survival, thus promoting the optimized application of exogenous MSCs in tissue engineering and safeguarding clinical organ regeneration. This paper provides a detailed protocol for the construction and characterization of self-assembled aggregates based on umbilical cord mesenchymal stem cells (UCMSCs), as well as an example of the cranial bone regenerative application. The implementation of this procedure will help guide the establishment of an efficient MSC transplantation strategy for tissue engineering and regenerative medicine.

Stem cells play a crucial role in re-establishing homeostasis in the body, and the search for mechanisms by which they interact with the host to exert their therapeutic effects remains a key question currently being addressed. Considering their significant regenerative/therapeutic potential, research on mesenchymal stem cells (MSCs) has experienced an unprecedented advance in recent years, becoming the focus of extensive works worldwide to develop cell-based approaches for a variety of diseases. Initial evidence for the effectiveness of MSCs therapy comes from the restoration of dynamic microenvironmental homeostasis and endogenous stem cell function in recipient tissues by systemically delivered MSCs. The specific mechanisms by which the effects are exerted remain to be investigated in depth. Importantly, the profound cell-host interplay leaves persistent therapeutic benefits that remain detectable long after the disappearance of transplanted MSCs. In this review, we summarize recent advances on the role of MSCs in multiple disease models, provide insights into the mechanisms by which MSCs interact with endogenous stem cells to exert therapeutic effects, and refine the interconnections between MSCs and cells fused to damaged sites or differentiated into functional cells early in therapy.

Immunotherapy has changed the treatment landscape for multiple cancer types. In the recent decade, great progress has been made in immunotherapy, including immune checkpoint inhibitors, adoptive T-cell therapy, and cancer vaccines. ICIs work by reversing tumor-induced immunosuppression, resulting in robust activation of the immune system and lasting immune responses. Whereas, their clinical use faces several challenges, especially the low response rate in most patients. As an increasing number of studies have focused on membrane immune checkpoint protein trafficking and degradation, which interferes with response to immunotherapy, it is necessary to summarize the mechanism regulating those transmembrane domain proteins translocated into the cytoplasm and degraded via lysosome. In addition, other immune-related transmembrane domain proteins such as T-cell receptor and major histocompatibility are associated with neoantigen presentation. The endosomal-lysosomal system can also regulate TCR and neoantigen-MHC complexes on the membrane to affect the efficacy of adoptive T-cell therapy and cancer vaccines. In conclusion, we discuss the process of surface delivery, internalization, recycling, and degradation of immune checkpoint proteins, TCR, and neoantigen-MHC complexes on the endosomal-lysosomal system in biology for optimizing cancer immunotherapy.