Figure 2.
Inhibition of ALDH Activity Blocks the Formation and Proliferation of Spheroid Cells (EMN24 cells)
(A) Correlation between ALDH activity and ALDH1A1 mRNA levels on culture day 10.
(B) Immunofluorescence staining with anti-ALDH1A1 antibody and DAPI in ALDH-high and ALDH-low cells after sorting. Scale bars, 20 μm.
(C) Western blot analyses of spheroids cells after disulfiram treatment for 24 h.
(D) FACS analyses of ALDH activity in the presence or absence of disulfiram (24 h after treatment).
(E) Time course of cell growth in spheroid cells treated with the indicated amounts of disulfiram. n = 4 independent experiments, p < 0.001, Student's t test.
(F) Bright-phase images of spheroids (7 days after disulfiram treatment). Scale bars, 100 μm.
(G) Responses of ALDH-high and ALDH-low cells to different concentrations of disulfiram after treatment for 7 days. n = 4 independent experiments, p < 0.001, Student's t test.
(H) Tumor volumes (mean ± SEM) of xenograft tumors from 1 × 105 spheroid cells subcutaneously injected. Disulfiram (40 mg/kg) was intraperitoneally injected into mice in the disulfiram-treatment group, and vehicle (DMSO) was intraperitoneally injected into mice in the control group. n = 12 independent experiments, p < 0.001, Student's t test.
(I) Images of whole resected tumor xenografts excised on day 17. Scale bar, 10 mm.
(J) ALDH activity of the cancer cells derived from xenograft tumors.
(K) H&E staining and immunostaining of xenograft tumors, 17 days after transplantation. Scale bars, 100 μm.
(L) Western blot analyses after infection with the indicated lentiviruses.
(M) ALDH activity in the infected cells after ALDEFLUOR staining.
(N) Time course of proliferation of infected cells. n = 4 independent experiments, p < 0.01, Student's t test.
(O) Bright-phase images of the infected cells on day 7. Scale bars, 100 μm.
∗∗p < 0.01; ∗∗∗p < 0.001. See also Figure S2.