Initiate Tumors with Single Cell Spheres Formed in Serum-Containing Medium

Abstract

Background: It is difficult to prospectively identify and maintain putative tumor-initiating cells (TICs). Spheres that formed in serum-free media contained more TICs while spheres formed in serum-containing media were not used in tumor-initiating.Methods: Soft-agar was used to isolate colonies. A continuous, static suspension culture using serum-containing media was modified from liquid overlay technique and tumor cell spheres could be maintained by this method for >90 days. Tumor-initiating capacity of these spheres was tested in nude mice. The nuclear staining of OCT4 in sphere cells and the expression profile of hepatic cell lineage related genes were examined.Results: Soft-agar derived HepG2 colonies indicated different properties from their parental cells. The suspension cells of A549 and MCF7 could initiate tumors at 104 cells level. The growth proportions of individual A549, MCF7 and HepG2 spheres with diameter of 101-150 µm were significantly higher than smaller spheres. After suspension culture for 15-27 days, HepG2 and MCF7 spheres could initiate tumors with diameter up to 200 µm; the estimated TIC frequency was 1/1500-1/400. The HepG2 and MCF7 spheres retain tumor-initiating potential for at least 52 days.Conclusion: After 15 days' serum-containing suspension culture small HepG2 and MCF7 cell spheres (diameter ~200 µm) could initiate tumors in nude mice.

MiRNA Transcriptome Profiling of Spheroid-Enriched Cells with Cancer Stem Cell Properties in Human Breast MCF-7 Cell Line

“…As such, it is generally accepted that clonal and aggregates of cells might coexist and resulted in a heterogeneous spheroid 29 . Therefore, several studies characterized CSCs across the different types of cancers including breast, prostate, colon and melanoma have relied on cell aggregation method 30 - 33 . The models were not just biologically spherical in shape, but shared many features with that of solid tumors, and were not observed in traditional 2D monolayer cultures 34 - 37 .…”

Three dimensional tumor models for cancer studies

“…Recent developments in tissue culture technology made it possible to culture patient derived cells to obtain tumor spheroids without losing the original tumor's properties in terms of genotype and phenotype (82)(83)(84). These developments may soon replace the traditional 2D cell culture protocols and will establish themselves as standard techniques for culturing cancer cells.…”

“…For example, traditional cytotoxicity assays are developed and optimized for 2D cell culture models and may not serve the purpose in 3D models. 3D cell culture is currently used in anticancer studies, cytotoxicity studies, drug discovery experiments and biosensor/bioassay applications (84)(85)(86). 3D cell culture models are superior models and better mimic the actual tumor microenvironment and pathological conditions.…”