The E3 ubiquitin ligase complex CDC20-activated anaphase-promoting complex/Cyclosome (APC/C ^CDC20 ) plays a critical role in governing mitotic progression by targeting key cell cycle regulators for degradation. Cell division cycle protein 20 homolog (CDC20), the co-activator of APC/C, is required for full ubiquitin ligase activity. In addition to its well-known cell cycle-related functions, we demonstrate that CDC20 plays an essential role in osteogenic commitment of bone marrow mesenchymal stromal/stem cells (BMSCs). Cdc20 conditional knockout mice exhibit decreased bone formation and impaired bone regeneration after injury. Mechanistically, we discovered a functional interaction between the WD40 domain of CDC20 and the DNA-binding domain of p65. Moreover, CDC20 promotes the ubiquitination and degradation of p65 in an APC11-dependent manner. More importantly, knockdown of p65 rescues the bone loss in Cdc20 conditional knockout mice. Our current work reveals a cell cycle-independent function of CDC20, establishes APC11 ^CDC20 as a pivotal regulator for bone formation by governing the ubiquitination and degradation of p65, and may pave the way for treatment of bone-related diseases.

Bone marrow stem cells (BMSCs) are a promising source of seed cells in bone tissue engineering, which needs a great quantity of cells. Cell senescence occurs as they are passaged, which could affect the therapeutic effects of cells. Therefore, this study aims to explore the transcriptomic differences among the uncultured and passaged cells, finding a practical target gene for anti-aging. We sorted PαS (PDGFR-α⁺SCA-1⁺CD45^-TER119^-) cells as BMSCs by flow cytometry analysis. The changes in cellular senescence phenotype (Counting Kit-8 (CCK-8) assay, reactive oxygen species (ROS) test, senescence-associated β-galactosidase (SA-β-Gal) activity staining, expression of aging-related genes, telomere-related changes and in vivo differentiation potential) and associated transcriptional alterations during three important cell culture processes (in vivo, first adherence in vitro, first passage, and serial passage in vitro) were studied. Overexpression plasmids of potential target genes were made and examed. Gelatin methacryloyl (GelMA) was applied to explore the anti-aging effects combined with the target gene. Aging-related genes and ROS levels increased, telomerase activity and average telomere length decreased, and SA-β-Gal activities increased as cells were passaged. RNA-seq offered that imprinted zinc-finger gene 1 (Zim1) played a critical role in anti-aging during cell culture. Further, Zim1 combined with GelMA reduced the expression of P16/P53 and ROS levels with doubled telomerase activities. Few SA-β-Gal positive cells were found in the above state. These effects are achieved at least by the activation of Wnt/β-catenin signaling through the regulation of Wnt2. The combined application of Zim1 and hydrogel could inhibit the senescence of BMSCs during in vitro expansion, which may benefit clinical application.

Primary bone mesenchymal stem cells (BMSCs) gradually lose stemness during in vitro expansion, which significantly affects the cell therapeutic effects. Here, we chose murine PαS (SCA-1⁺PDGFRα⁺CD45⁻TER119⁻) cells as representative of BMSCs and aimed to explore the premium culture conditions for PαS cells. Freshly isolated (fresh) PαS cells were obtained from the limbs of C57/6N mice by fluorescence-activated cell sorting (FACS). We investigated the differences in the stemness of PαS cells by proliferation, differentiation, and stemness markers in vitro and by ectopic osteogenesis and chondrogenesis ability in vivo, as well as the changes in the stemness of PαS cells during expansion in vitro. Gain- and loss-of-function experiments were applied to investigate the critical role and underlying mechanism of the basic helix–loop–helix family member E40 (BHLHE40) in maintaining the stemness of PαS cells. The stemness of fresh PαS cells representative in vivo was superior to that of passage 0 (P0) PαS cells in vitro. The stemness of PαS cells in vitro decreased gradually from P0 to passage 4 (P4). Moreover, BHLHE40 plays a critical role in regulating the stemness of PαS cells during in vitro expansion. Mechanically, BHLHE40 regulates the stemness of PαS cells by targeting Zbp1 through the Wnt/β-catenin signaling pathway. This work confirms that BHLHE40 is a critical factor for regulating the stemness of PαS cells during expansion in vitro and may provide significant indications in the exploration of premium culture conditions for PαS cells.

High-translucency zirconia (HTZ) has superior esthetic properties, but its unreliable resin bonding limits minimally invasive anterior restorations. An in situ surface modification was developed to synthesize CaZrO₃ particulates on pre-sintered HTZ for enhanced bonding durability. HTZ specimens were randomized into control (Zr-c) and calcium-modified (Zr-Ca) groups; Zr-Ca was treated with NaF/HCl mixture, calcium chloride glycerol solution, NaOH incubation (80 °C, 2 h), and sintering. Surface characteristics were characterized by SEM/EDS, AFM, X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), and FTIR. Flexural strength was tested via three-point bending; shear bond strength (SBS) was evaluated immediately and after 5000 thermocycles with resin cements (with/without 10-MDP). Zr-Ca showed uniform surface particulates, increased roughness, enhanced wettability, and surface Ca; XRD/FTIR/XPS confirmed CaZrO₃ and Ca-O-P species (after MDP). Zr-Ca with 10-MDP-containing resin adhesive had significantly higher SBS before/after aging (predominantly mixed failures), with flexural strength within clinical limits. In situ CaZrO₃ formation on HTZ strengthens MDP-mediated resin bonding and thermocycling resistance while preserving mechanical integrity, providing a feasible strategy for durable adhesion.

Glioblastoma (GBM) is an aggressive brain malignancy with a median survival of only 15 months despite current therapeutic interventions. A major obstacle to effective treatment is the intricate tumor microenvironment (TME), which drives tumor progression and therapy resistance. However, existing in vitro models lack micrometer‐scale spatial control over multiple cellular components and fail to preserve their dynamic interactions, limiting our understanding of tumor biology and treatment responses. Here, we present a spatially controlled GBM‐on‐a‐chip (scGoC) platform that integrates a novel organoid positioning strategy, enabling the reconstruction of the TME within a 3D matrix at micron‐level precision. This platform allows real‐time monitoring of dynamic tumor‐immune‐vascular interactions, providing refined insights into tumor progression and drug responses. Using the scGoC, we demonstrate that endothelial cells not only promote microglial migration toward tumor organoids but also drive their polarization toward an immunosuppressive phenotype, faithfully recapitulating hallmark features of GBM progression. Importantly, the platform reveals a pro‐tumorigenic adaptive response to bevacizumab monotherapy, driven by the upregulation of HIF‐1 signaling and metabolic reprogramming pathways, providing mechanistic insights into GBM treatment. Collectively, the scGoC establishes a versatile and standardized tool for decoding complex cellular communications and investigating mechanisms of treatment response with broad applicability across cancer research.