Haiyan Weng

Antiangiogenesis with bevacizumab, an antibody against vascular endothelial growth factor (VEGF), has been used for devascularization to limit the growth of malignant glioma. However, the benefits are transient due to elusive mechanisms underlying resistance to the antiangiogenic therapy. Glioma stem cells (GSCs) are capable of forming vasculogenic mimicry (VM), an alternative microvascular circulation independent of VEGF-driven angiogenesis. Herein, we report that the formation of VM was promoted by bevacizumab-induced macroautophagy/autophagy in GSCs, which was associated with tumor resistance to antiangiogenic therapy. We established a 3-dimensional collagen scaffold to examine the formation of VM and autophagy by GSCs, and found that rapamycin increased the number of VM and enhanced KDR/VEGFR-2 phosphorylation. Treatment with chloroquine, or knockdown of the autophagy gene ATG5, inhibited the formation of VM and KDR phosphorylation in GSCs. Notably, neutralization of GSCs-produced VEGF with bevacizumab failed to recapitulate the effect of chloroquine treatment and ATG5 knockdown, suggesting that autophagy-promoted formation of VM was independent of tumor cell-derived VEGF. ROS was elevated when autophagy was induced in GSCs and activated KDR phosphorylation through the phosphoinositide 3-kinase (PI3K)-AKT pathway. A ROS inhibitor, N-acetylcysteine, abolished KDR phosphorylation and the formation of VM by GSCs. By examination of the specimens from 95 patients with glioblastoma, we found that ATG5 and p-KDR expression was strongly associated with the density of VM in tumors and poor clinical outcome. Our results thus demonstrate a crucial role of autophagy in the formation of VM by GSCs, which may serve as a therapeutic target in drug-resistant glioma.

INTRODUCTION: Human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) and their small extracellular vesicles (hUC-MSC-sEVs) have shown attractive prospects applying in regenerative medicine. This study aimed to compare the therapeutic effects of two agents on osteoarthritis (OA) and investigate underlying mechanism using proteomics. METHODS: ) or hUC-MSC-sEVs (30 μg/time) were injected into the knee joints of anterior cruciate ligament transection-induced OA model. Hematoxylin and eosin, Safranin O/Fast Green staining were used to observe cartilage degeneration. The levels of cartilage matrix metabolic molecules (Collagen II, MMP13 and ADAMTS5) and macrophage polarization markers (CD14, IL-1β, IL-10 and CD206) were assessed by immunohistochemistry. Finally, proteomics analysis was performed to characterize the proteinaceous contents of two agents. RESULTS: In vitro data showed that hUC-MSC-sEVs were taken up by chondrocytes. A total of 15 μg/mL of sEVs show the greatest proliferative and migratory capacities among all groups. In the animal study, hUC-MSCs and hUC-MSC-sEVs alleviated cartilage damage. This effect was mediated via maintaining cartilage homeostasis, as was confirmed by upregulation of the COL II and downregulation of the MMP13 and ADAMTS5. Moreover, the M1 macrophage markers (CD14) were significantly reduced, while the M2 macrophage markers (CD206 and IL-10) were increased in the hUC-MSCs and hUC-MSC-sEVs relative to the untreated group. Mechanistically, we found that many proteins connected to cartilage repair were more abundant in sEVs. Notably, compared to hUC-MSCs, the upregulated proteins in sEVs were mostly involved in the regulation of immune effector process, extracellular matrix organization, PI3K-AKT signaling pathways, and Rap1 signaling pathway. CONCLUSION: Our study indicated that hUC-MSC-sEVs protect cartilage from damage and many cartilage repair-related proteins are probably involved in the restoration process. These data suggest the promising potential of hUC-MSC-sEVs as a therapeutic agent for OA.

Abstract The highly heterogeneous characteristics of Wharton's jelly mesenchymal stem cells (WJ‐MSCs) may be responsible for the poor clinical outcomes and poor reproducibility of treatments based on WJ‐MSCs. Exploration of WJ‐MSC heterogeneity with multimodal single‐cell technologies will aid in establishing accurate MSC subtyping and developing screening protocols for dominant functional subpopulations. Here, the characteristics of WJ‐MSCs are systematically analyzed by single cell and spatial transcriptome sequencing. Single‐cell transcriptomics analysis identifies four WJ‐MSC subpopulations, namely proliferative_MSCs, niche‐supporting_MSCs, metabolism‐related_MSCs and biofunctional‐type_MSCs. Furthermore, the transcriptome, cellular heterogeneity, and cell‐state trajectories of these subpopulations are characterized. Intriguingly, the biofunctional‐type MSCs (marked by S100A9, CD29, and CD142) selected in this study exhibit promising wound repair properties in vitro and in vivo. Finally, by integrating omics data, it has been found that the S100A9 + CD29 + CD142 + subpopulation is more enriched in the fetal segment of the umbilical cord, suggesting that this subpopulation deriving from the fetal segment may have potential for developing into an ideal therapeutic agent for wound healing. Overall, the presented study comprehensively maps the heterogeneity of WJ‐MSCs and provides an essential resource for future development of WJ‐MSC‐based drugs.

off-target detection pipeline for three generations of artificial nucleases provided useful tools for genome engineering-based gene therapy.