Figure 4. PODXL expression correlates with oncosphere formation.

(A): Results of the limiting dilution assay using PODXL knockdown with TRCN0000117018 compared to control virus in JHU-0879. The 0.37 intercept was 64 cells/well for the control cells and 374 cells/well for the PODXL knockdown cells. Inset shows a control well, left, and a PODXL knockdown well, right (40×). Quantitatively similar results were obtained with two additional knockdown experiments. (B): Results of the limiting dilution assay using PODXL knockdown with V2THS_36258 compared to control virus in JHU-0879. The 0.37 intercept was 38 cells/well for the control cells and 125 cells/well for the PODXL knockdown cells. Quantitatively similar results were obtained with two additional knockdown experiments. (C): qRT-PCR results of Musashi1, SOX2, and BMI1 gene expression in JHU-0879 cells transduced with either PODXL shRNA or control virus. Expression was normalized to GAPDH expression. ns: p>0.05; *: p≤0.01. (D): Results of the limiting dilution assay using PODXL-positive cells compared to PODXL-negative cells obtained using FACS on 020913. The 0.37 intercept was 37 cells/well for the PODXL-positive cells and 632 cells/well for the PODXL-negative cells. Inset shows a PODXL-positive well, left, and a PODXL-negative well, right (40×). Quantitatively similar results were obtained with two additional FACS experiments using 020913. (E): Results of the limiting dilution assay using PODXL-positive cells compared to PODXL-negative cells obtained using FACS on JHU-0879. The 0.37 intercept was 53 cells/well for the PODXL-positive cells and 356 cells/well for the PODXL-negative cells. Quantitatively similar results were obtained with two additional FACS experiments using JHU-0879. (F): qRT-PCR results of Musashi1, SOX2, and BMI1 gene expression comparing FACS-sorted PODXL-positive and PODXL-negative populations from 020913. Expression was normalized to GAPDH expression. *: p≤0.01; **: p≤0.001.