In this study, we report that the intra-hydrogel culture system mitigates the transformation of mesenchymal stem cells (MSCs) induced by two-dimensional (2D) expansion. MSCs expanded in monolayer culture prior to encapsulation in collagen hydrogels (group eMSCs-CH) featured impaired stemness in chondrogenesis, comparing with the freshly isolated bone marrow mononuclear cells seeded directly in collagen hydrogels (group fMSCs-CH). The molecular mechanism of the in vitro expansion-triggered damage to MSCs was detected through genome-wide microarray analysis. Results indicated that pathways such as proteoglycans in cancer and pathways in cancer expansion were highly enriched in eMSCs-CH. And multiple up-regulated oncoma-associated genes were verified in eMSCs-CH compared with fMSCs-CH, indicating that expansion in vitro triggered cellular transformation was associated with signaling pathways related to tumorigenicity. Besides, focal adhesion (FA) and mitogen-activated protein kinase (MAPK) signaling pathways were also involved in in vitro expansion, indicating restructuring of the cell architecture. Thus, monolayer expansion in vitro may contribute to vulnerability of MSCs through the regulation of FA and MAPK. This study indicates that intra-hydrogel culture can mitigate the monolayer expansion induced transformation of MSCs and maintain the uniformity of the stem cells, which is a viable in vitro culture system for stem cell therapy.

中文翻译：

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水凝胶内培养可防止单层扩增诱导的间充质干细胞转化

在这项研究中，我们报告水凝胶内培养系统减轻了由二维（2D）扩展诱导的间充质干细胞（MSCs）的转化。与直接植入胶原水凝胶（fMSCs-CH组）中的新鲜分离的骨髓单核细胞相比，MSCs在囊封在胶原水凝胶（eMSCs-CH组）中之前在单层培养中扩增，其特点是软骨形成中的干性受损。通过全基因组微阵列分析检测了体外扩增触发的MSC损伤的分子机制。结果表明，eMSCs-CH中高度富集了癌症中的蛋白聚糖等途径和癌症扩展中的途径。并且与fMSCs-CH相比，在eMSCs-CH中验证了多个上调的与昏迷相关的基因，这表明扩增体外触发的细胞转化与致瘤性相关的信号通路相关。此外，在体外扩增中还涉及粘着粘附（FA）和有丝分裂原激活的蛋白激酶（MAPK）信号传导途径，表明细胞结构发生了重组。因此，体外单层扩增可能通过调节FA和MAPK促进MSC的易损性。这项研究表明，水凝胶内培养可以减轻单层扩增诱导的MSCs转化并保持干细胞的均匀性，这是一种可行的干细胞治疗体外培养系统。