Zhang Junming, Liu Li, Li Yuexia, Wu Jinglei, Lou Xiangxin
College of Biological Science and Medical Engineering, Donghua University, Shanghai, China.
Cell J. 2023 Jul 25;25(7):447-454. doi: 10.22074/cellj.2023.1989426.1228.
Previous reports showed that mouse embryonic fibroblasts (MEFs) could support pluripotent stem cell selfrenewal and maintain their pluripotency. The goal of this study was to reveal whether the decellularized extracellular matrix derived from MEFs (MEF-ECM) is beneficial to promote the proliferation of inner ear-derived cells.
In this experimental study, we prepared a cell-free MEF-ECM through decellularization. Scanning electron microscope (SEM) and immunofluorescent staining were conducted for phenotype characterization. Organs of Corti were dissected from postnatal day 2 and the inner ear-derived cells were obtained. The identification of inner ear-derived cells was conducted by using reverse transcription-polymerase chain reaction (RT-PCR). Cell counting kit-8 (CCK-8) was used to evaluate the proliferation capability of inner ear-derived cells cultured on the MEFECM and tissue culture plate (TCP).
The MEF-ECM was clearly observed after decellularization via SEM, and the immunofluorescence staining results revealed that MEF-ECM was composed of three proteins, including collagen I, fibronectin and laminin. Most importantly, the results of CCK-8 showed that compared with TCP, MEF-ECM could effectively facilitate the proliferation of inner ear-derived cells.
The discovery of the potential of MEF-ECM in promoting inner ear-derived cell proliferation indicates that the decellularized matrix microenvironment may play a vital role in keeping proliferation ability of these cells. Our findings indicate that the use of MEF-ECM may serve as a novel approach for expanding inner ear-derived cells and potentially facilitating the clinical application of inner ear-derived cells for hearing loss in the future.
既往报道显示，小鼠胚胎成纤维细胞（MEFs）可支持多能干细胞自我更新并维持其多能性。本研究的目的是揭示源自MEFs的脱细胞细胞外基质（MEF-ECM）是否有利于促进内耳来源细胞的增殖。
在本实验研究中，我们通过脱细胞制备了无细胞的MEF-ECM。进行扫描电子显微镜（SEM）和免疫荧光染色以进行表型表征。从出生后第2天的小鼠中分离出柯蒂氏器并获得内耳来源的细胞。使用逆转录-聚合酶链反应（RT-PCR）对内耳来源的细胞进行鉴定。使用细胞计数试剂盒-8（CCK-8）评估在内耳来源的细胞在MEF-ECM和组织培养板（TCP）上培养时的增殖能力。
通过SEM在脱细胞后清晰观察到MEF-ECM，免疫荧光染色结果显示MEF-ECM由三种蛋白质组成，包括胶原蛋白I、纤连蛋白和层粘连蛋白。最重要的是，CCK-8结果显示，与TCP相比，MEF-ECM可有效促进内耳来源细胞的增殖。
MEF-ECM在促进内耳来源细胞增殖方面潜力的发现表明，脱细胞基质微环境可能在维持这些细胞的增殖能力中起关键作用。我们的研究结果表明，使用MEF-ECM可能是一种扩大内耳来源细胞的新方法，并有可能在未来促进内耳来源细胞用于听力损失的临床应用。
