Figure 2

In vivo tumor formation assay of ALDH1^high cells in the presence or absence of fibroblasts. (a) Schema of cell transplantation into NOD/SCID mice. 1 × 10⁴ ALDH1^high cells with or without a mixture of 1 × 10⁴ human normal skin fibroblasts (N25-5) were suspended in 100 μl of PBS matrigel mixture and inoculated into NOD/SCID mice. (b) Tumor formation assay. Fibroblast-mixed ALDH1^high cells developed earlier and significantly larger tumors than did ALDH1^high only cells. Data represent means±s.d. Differences between fibroblast-mixed ALDH1^high tumor and ALDH1^high only tumor were examined for statistical significance using Student’s t-test. *P-values. (c) HE staining of ALDH1^high only tumor. Representative picture of hematoxylin and eosin staining of tumor tissue. × 200 magnification. (d) HE staining and immunostaining of fibroblast-mixed ALDH1^high tumor. Upper panel: Representative picture of hematoxylin and eosin staining of the tumor tissue. Abundant CAFs are induced in tumor tissue. × 100 magnification. Middle panel: Immunostaining with anti-HLA class I antibody. Tumor cells are positive for HLA, but stromal cells are negative. × 100 magnification. Lower panel: Immunostaining with anti-α-SMA antibody. Stromal fibroblasts are positive for α-SMA. × 100 magnification.