Abstract
The culture of tumor cell lines in three-dimensional scaffolds is considered to more closely replicate the in vivo tumor microenvironment than the standard method of two-dimensional cell culture. We hypothesized that our method of encapsulating and maintaining viable and functional pancreatic islets in agarose-agarose macrobeads (diameter 6-8 mm) might provide a novel method for the culture of tumor cell lines. In this report we describe and characterize tumor colonies that form within macrobeads seeded with mouse renal adenocarcinoma cells. Approximately 1% of seeded tumor cells survive in the macrobead and over several months form discrete elliptical colonies appearing as tumor cell niches with increasing metabolic activity in parallel to colony size. The tumor colonies demonstrate ongoing cell turnover as shown by BrdU incorporation and activated caspase-3 and TUNEL staining. Genes upregulated in the tumor colonies of the macrobead are likely adaptations to this novel environment, as well as an amplification of G(1)/S cell-cycle checkpoints. The data presented, including SCA-1 and Oct4 positivity and the upregulation of stem cell-like genes such as those associated with the Wnt pathway, support the notion that the macrobead selects for a subpopulation of cells with cancer stem cell or cancer progenitor properties.
Highlights
The in vitro investigation of cancer began with the report of the first human tumor cell line (HeLa) in 1952 (1)
The formation of viable and metabolically active tumor colonies within the macrobead To test the hypothesis that neoplastic cells entrapped in an agarose matrix would survive during in vitro culture, mouse renal adenocarcinoma (RENCA), feline breast carcinoma (K12), as well as human breast (MCF7), prostate (ArCap[10] and DU145), bladder (J82), colon (HCT116) and choriocarcinoma (JEG-3) cell lines were all successfully encapsulated and maintained in 6- to 8-mm diameter agarose–agarose
Cells derived from epithelial tumors encapsulated in agarose–agarose macrobeads undergo a process that results in a selection of at least 2 subpopulations of cells
Summary
The in vitro investigation of cancer began with the report of the first human tumor cell line (HeLa) in 1952 (1). Use of this cell line has resulted in numerous advances in our understanding of cell and cancer biology which is largely attributable to its capacity to be cultured and grown in standard laboratory glassware or plasticware as 2-dimensional (2D) monolayers. Standard 2D cell culture, is limited in its ability to accurately recreate the in vivo tumor environment. The use of 3D culture systems are increasingly being employed to better mimic the in vivo tumor environment. The use of 3D tumor cell culture has been shown to more accurately predict the in vivo response to cytotoxic therapy (6, 7)
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