Fig. 4: Pluripotency-related features of ALDH⁺CD44⁺CXCR4⁺CD24⁺-CWR22 cells.

a As shown by Western blotting analysis, ALDH⁺ CD44⁺ CXCR4⁺ CD24⁺-CWR22 cells exhibit more robust expression of major pluripotency regulators such as Nanog, OCT4, Sox2 and FoxD3, as compared to ALDH⁺ CD44⁺-CWR22 cells, ALDH⁻ CD44⁻ CXCR4⁻ CD24⁻-CWR22 cells and bulk culture. b Quantitative RT-PCR analysis demonstrated that ALDH⁺ CD44⁺ CXCR4⁺ CD24⁺-PCa cells have much higher expression level of NanogP8 mRNA, which was thought to be a pluripotency marker gene in cancer stem cells, as compared to controls run in parallel. c Cells labelled with reporter gene NanogP8-GFP, 1 × 10³ ALDH⁺CD44⁺CXCR4⁺CD24⁺ -CWR22 cells could be differentiated into a mixed bulk culture in a “salt and pepper” pattern in vitro; whereas ALDH^-CD44^-CXCR4^-CD24^--PCa cells exhibited no NanogP8 expression under routine culture conditions. Scale bar, 10 µm. d The telomerase activity was measured by quantitative qRT-PCR. e Telomere fluorescence intensities and telomere length were measured by flow-fluorescence in situ hybridisation. Right panel, quantification of the left panel.