Isolation of cancer cells with augmented spheroid-forming capability using a novel tool equipped with removable filter.
Isolation of cancer cells with augmented spheroid-forming capability using a novel tool equipped with removable filter.
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DOI：
10.18632/oncotarget.26092
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发表时间：
2018-09-21
期刊：
影响因子：
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通讯作者：
Nojima H
中科院分区：
文献类型：
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作者：
Fujibayashi E;Yabuta N;Nishikawa Y;Uchihashi T;Miura D;Kurioka K;Tanaka S;Kogo M;Nojima H
三维 (3D) 细胞培养系统已用于从癌细胞中获得多细胞球状细胞聚集体或球体。然而，从癌细胞中有效制备大型肿瘤来源的球体是很困难的。为了解决这个问题，我们在这里使用了一种配备去除膜的工具，称为 Spheroid Catch，用于从人鳞状细胞癌（SAS 细胞）中选择和富集大尺寸和/或尺寸匹配的球体，而不会损失恢复。经过五轮的选择和富集过程，我们成功分离出了具有增强球体形成能力的 SAS 细胞亚群，命名为 eSAS：球体形成效率为 28.5%（eSAS），而亲本 SAS 为 16.8%。值得注意的是，我们发现一些 eSAS 细胞在 3D 培养物中暴露于高剂量顺铂后存活下来。此外，与注射SAS细胞的裸鼠相比，通过将eSAS细胞注射到裸鼠舌头中的原位植入显示出存活率降低和肿瘤生长增加。这些结果表明，通过使用 Spheroid Catch 可以有效地收集表现出较高球体形成能力的球体。事实上，全基因组 cDNA 微阵列和蛋白质印迹分析表明，在 eSAS 细胞中，hedgehog 酰基转移酶 (HHAT) 的 mRNA 和蛋白水平高于 SAS 细胞，HHAT 通过催化 Hedgehog 蛋白的 N-棕榈酰化与细胞癌中的干细胞维持相关。我们建议 Spheroid Catch 作为其他类型细胞过滤器的替代方法，可用于基础科学和临床科学中的球体和潜在类器官的研究。
Three-dimensional (3D) cell culture systems have been used to obtain multicellular spheroidal cell aggregates, or spheroids, from cancer cells. However, it is difficult to efficiently prepare large tumor-derived spheroids from cancer cells. To circumvent this problem, we here used a tool equipped with removal membrane, called Spheroid Catch, for the selection and enrichment of large-sized and/or size-matched spheroids from human squamous cell carcinoma (SAS cells) without loss of recovery. After a five-round process of selection and enrichment, we successfully isolated a subpopulation of SAS cells with augmented spheroid-forming capability, named eSAS: the efficiency of spheroid formation is 28.5% (eSAS) vs 16.8% (parental SAS). Notably, we found that some of eSAS cells survived after exposure of high doses of cisplatin in 3D culture. Moreover, orthotopic implantation by injecting eSAS cells into the tongues of nude mice showed reduced survival rate and increased tumor growth compared with those of nude mice injected with SAS cells. These results suggest that spheroids exhibiting properties of higher spheroid forming capacity can be efficiently collected by using Spheroid Catch. Indeed, genome-wide cDNA microarray and western blot analyses demonstrated higher mRNA and protein levels of hedgehog acyltransferase (HHAT), which is associated with stem maintenance in cell carcinoma by catalysing the N-palmitoylation of Hedgehog proteins, in eSAS cells than in SAS cells. We propose that Spheroid Catch could be useful for the study of spheroids, and potentially organoids, in the basic and clinical sciences, as an alternative method to other type of cell strainers.