Cancer Stem Cell Sorting from Colorectal Cancer Cell Lines by Sedimentation Field Flow Fractionation

Abstract

Recently, cancer stem cells (CSCs) have been identified in many types of cancers, such as colorectal cancer (CRC). CSCs seem to be involved in initiation, growth, and tumor metastasis, as well as in radio- and chemotherapy failures. CSCs appears as new biological targets for cancer therapy, requiring the development of noninvasive cell sorting methods. In this study, we used sedimentation field flow fractionation (SdFFF) to prepare enriched populations of CSCs from eight cell lines corresponding to different CRC grades. On the basis of phenotypic and functional characterizations, "hyperlayer" elution resulted in a fraction overexpressing CSC markers (CD44, CD166, EpCAM) for all cell lines. CSCs were eluted in the last fraction for seven out of eight cell lines, but in the first for HCT116. These results suggest, according to the literature, that two different pools of CSCs exist, quiescent and activated, which can both be sorted by SdFFF. Moreover, according to CSC properties, enriched fractions are able to form colonies.

New ex-ovo colorectal-cancer models from different SdFFF-sorted tumor-initiating cells

“…We then performed a pre-amplification step by in-vitro culturing cells in a 3D matrix: Matrigel 3D (tumoral mice extracellular matrix). According to self-renewal properties of TICs presented in cell lines [25], naked-eye-visible colonies appeared after five weeks culture of 500 cells/well, for both cell lines (data not shown). Colonies were recovered from Matrigel and deposited on the CAM.…”

“…The SdFFF separation device used in this study was the same as that described elsewhere, equipped with a 175 μm thick channel [25]. Optimum SdFFF elution conditions, enabling hyperlayer mode, have been defined elsewhere [25] and were 20g and 0.8 mL min −1 as external field strength and flow rate, respectively, for HCT-116 cells and 8g and 0.8 mL min −1 , respectively, for WiDr.…”

“…Optimum SdFFF elution conditions, enabling hyperlayer mode, have been defined elsewhere [25] and were 20g and 0.8 mL min −1 as external field strength and flow rate, respectively, for HCT-116 cells and 8g and 0.8 mL min −1 , respectively, for WiDr. Elution of CRC cell suspensions (100 μL, 2.5×10 6 cells per mL) resulted in the separation of three cell fractions (Fig.…”

“…Recently, we revealed the ability of SdFFF to generate enriched fractions with TIC characteristics [25,26]. SdFFF had successfully sorted colorectal TIC phenotypes on the basis of CD44 + , EpCAM high , and CD166 + expression in eight cell lines.…”

Cancer Stem-Like Cells in Glioblastoma

“…Usually, the largest and less dense cells (the most differentiated) are eluted first, while the smallest and the most dense cells (low nucleo-cytoplasmic ratio corresponding to stems cells) are eluted last. This elution order also depends on the ploidy or the position in the cell cycle; the cells eluted in the last fraction are usually in the G0/G1 phase (56,57).…”