ATC monolayer-derived cells initiate tumors in serially transplanted immunodeficient mice.

<p>(A) Tumor growth curve generated by subcutaneous injection of THJ-11T parental monolayer-derived cells. The number of cells injected is indicated. (B) A representative subcutaneous tumor removed from a mouse xenograft. (C) Western blot analysis showing the expression of ALDH, CD44, and CXCR4 in cells derived from primary, secondary and tertiary xenografts. (D) H&E staining showed that tumors originating from primary, secondary and tertiary xenografts are composed of a highly heterogeneous population of cells with a large nucleus-to-cytoplasm ratio. (E) Immunohistochemical analysis of ALDH, CD44, CXCR4 and CD133 levels in mouse xenografts generated from primary xenografts of THJ-11T and THJ-16T. Note that mouse xenografts do not express CD133.</p

ATC cells retain self-renewal capacity <i>in vitro</i>.

<p>(A) Representative phase contrast microscopy analysis of colonies (left) and thyrospheres (right) after seven days of culture. Scale bar, 100 µm. (B) Percentage of primary and secondary thyrospheres in all four ATC cell lines. (C) Representative confocal microscopy images of THJ-21T thyrospheres indicated the expression of Nanog (red), Oct4 (green) and DAPI (blue). Scale bar, 10 µm (upper panel). Representative phase contrast images of THJ-21T parental monolayer cells indicated the expression of Nanog and Oct4 is unique to thyrospheres and is not seen in the parental monolayer cells (lower panel).</p

Characterization of ATC cells <i>in vitro</i>.

<p>(A) Phase contrast microscopy images of four ATC cell lines cultured as monolayers in RPMI medium. (B) Growth proliferation curve over 96 hours demonstrates the proliferative potential of these ATC cell lines. (C) qRT-PCT analysis of thyroid transcription factors <i>Pax8</i> and <i>TTF1</i> and thyroid differentiation markers <i>TSHR</i>, <i>TG</i>, <i>NIS</i>, and <i>TPO</i>. Human GAPDH was used as the housekeeping gene during the amplifications.</p

The effect of cisplatin on ATC cells.

<p>(A) Representative phase contrast microscopy images of THJ-11T parental monolayer-derived cells exposed to the indicated dose of cisplatin for 48 hours. (B) Dose-dependent inhibition of growth in all ATC cell lines. IC₅₀ values are as shown for each cell line. (C) Comparison of <i>in vitro</i> resistance to cisplatin of the parental monolayer cells and spheroid-forming cells. The THJ-11T and THJ-16T parental monolayer-derived cells and thyrosphere-derived cells were treated with 10 µM of cisplatin and the fraction of proliferating cells remaining was subsequently estimated based on the Alamar Blue cell proliferation assay. All experiments were performed in triplicate. ****, <i>P</i><0.0001; ***, <i>P</i><0.001; **, <i>P</i><0.05.</p

Limiting dilution analysis using human ATC cell lines in a subcutaneous mouse model of thyroid carcinoma.

<p>Limiting dilution analysis using human ATC cell lines in a subcutaneous mouse model of thyroid carcinoma.</p