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A Novel 3D Culture Model of Human ASCs Reduces Cell Death in Spheroid Cores and Maintains Inner Cell Proliferation Compared With a Nonadherent 3D Culture.

ABSTRACT: 3D cell culture technologies have recently shown very valuable promise for applications in regenerative medicine, but the most common 3D culture methods for mesenchymal stem cells still have limitations for clinical application, mainly due to the slowdown of inner cell proliferation and increase in cell death rate. We previously developed a new 3D culture of adipose-derived mesenchymal stem cells (ASCs) based on its self-feeder layer, which solves the two issues of ASC 3D cell culture on ultra-low attachment (ULA) surface. In this study, we compared the 3D spheroids formed on the self-feeder layer (SLF-3D ASCs) with the spheroids formed by using ULA plates (ULA-3D ASCs). We discovered that the cells of SLF-3D spheroids still have a greater proliferation ability than ULA-3D ASCs, and the volume of these spheroids increases rather than shrinks, with more viable cells in 3D spheroids compared with the ULA-3D ASCs. Furthermore, it was discovered that the SLF-3D ASCs are likely to exhibit the abovementioned unique properties due to change in the expression level of ECM-related genes, like COL3A1, MMP3, HAS1, and FN1. These results indicate that the SLF-3D spheroid is a promising way forward for clinical application.

SUBMITTER: Luo L

PROVIDER: S-EPMC8632442 | biostudies-literature | 2021

REPOSITORIES: biostudies-literature

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A Novel 3D Culture Model of Human ASCs Reduces Cell Death in Spheroid Cores and Maintains Inner Cell Proliferation Compared With a Nonadherent 3D Culture.

Luo Liang L Zhang Wei W Wang Jing J Zhao Ming M Shen Kuo K Jia Yanhui Y Li Yan Y Zhang Jian J Cai Weixia W Xiao Dan D Bai Xiaozhi X Liu Kaituo K Wang Kejia K Zhang Yue Y Zhu Huayu H Zhou Qin Q Hu Dahai D

Frontiers in cell and developmental biology 20211111

3D cell culture technologies have recently shown very valuable promise for applications in regenerative medicine, but the most common 3D culture methods for mesenchymal stem cells still have limitations for clinical application, mainly due to the slowdown of inner cell proliferation and increase in cell death rate. We previously developed a new 3D culture of adipose-derived mesenchymal stem cells (ASCs) based on its self-feeder layer, which solves the two issues of ASC 3D cell culture on ultra-l ...[more]

PMID: 34858974

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Project description:BackgroundThree dimensional (3D) cell cultures have been an area of increasing interest and relevance across several research fields including drug discovery, developmental biology and stem cell-based therapies. However, handling 3D structures can be difficult. In particular, the replacement of liquid media and reagents in which liquid is removed using pipettes is difficult to perform as the 3D spheroids can be easily aspirated into the pipette tip.ResultsWe have developed the 3D-tip, a novel tool that facilitates media change and washing procedures of 3D-spheroid cultures. The 3D-tip contains a mesh with 40-μm pores allowing the aspiration of liquids including media, drugs, buffers and reagents, with the mesh acting as a barrier preventing the spheroids being aspirated into the pipette tip. After aspiration of liquids, the spheroids are gently deposited back into the culture vessel. Our results demonstrate that the 3D-tips offer superior handling of 3D-spheroid cultures in comparison to commonly used methods. We showed that the 3D-tips can easily be used on both fixed and unfixed spheroids and on cancer cell, stem cell and glial cell spheroids.In contrast with the 50/50 media exchange method, the 3D-tips allow a complete media change with minimal loss of spheroids and without damaging their morphology. Our results showed that 86.0% of spheroids remained in the chamber after changing the media using the 3D-tips. In contrast, only 45.0% of spheroids remained using the 50/50 media exchange strategy.In comparison with the centrifugation technique, the 3D-tips preserved spheroids whereas centrifugation led to the loss of spheroids and/or the alteration of the size and shape of the 3D cellular structures. We observed that 87.6 and 84.6% of the fixed and unfixed spheroids remained using the 3D-tip, respectively. In contrast, only 66.3% of the fixed spheroids and 36.4% of the unfixed spheroids were left using the centrifugation method. From a time perspective, the 3D-tips dramatically reduce the time taken for replacing media.ConclusionsThis novel pipette tip is suitable for high throughput screening and automation and will revolutionise the techniques used for the production and analysis of 3D spheroids.

Dataset Information

A Novel 3D Culture Model of Human ASCs Reduces Cell Death in Spheroid Cores and Maintains Inner Cell Proliferation Compared With a Nonadherent 3D Culture.

Publications

A Novel 3D Culture Model of Human ASCs Reduces Cell Death in Spheroid Cores and Maintains Inner Cell Proliferation Compared With a Nonadherent 3D Culture.

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