Fig 1: DPP-4 inhibition-induced epithelial mesenchymal transition (EMT) impacted ABC transporters during chemotherapy treatment of breast cancer. (A) Western blot analysis of E-cadherin and α-SMA in 4T1 cells pretreated with KR62436 (50 μmol/L) for 48 h and then treated with or without DOX (0.425 μmol/L) for another 48 h. (B) Western blot analysis of E-cadherin and α-SMA in control and DPP-4-kd 4T1 cells treated with or without DOX (0.425 μmol/L) for 48 h. (C) Immunofluorescence analysis of E-cadherin and α-SMA expression in DPP-4-kd and control primary tumors treated with or without DOX. For each group, representative images of six different fields of view at 200× magnification were evaluated. The scale bar indicates 50 μm in each panel. (D) Western blot analysis of P-gp and ABCG2 in snail siRNA knockdown (snail siRNA) and control siRNA (100 nmol/L) 4T1 cells pretreated with KR62436 (50 μmol/L) for 24 h and then treated with or without DOX (0.425 μmol/L) for another 48 h. (E) Western blot analysis of phosphorylated smad3 (P-smad3), P-gp and ABCG2 in E-cadherin siRNA knockdown (E-cadherin siRNA) and control siRNA (100 nmol/L) 4T1 cells pretreated with KR62436 (50 μmol/L) for 24 h and then treated with or without DOX (0.425 μmol/L) for another 48 h. All densitometric quantification relative to β-actin levels and P-smad3 protein expression relative to smad3 levels (n = 3 per group) were performed by using ImageJ.
Fig 2: Reverse transcription-qPCR analysis of URAT1, GLUT9, OAT4 and ABCG2 in the kidney cortex tissues. Data are presented as mean ± SEM, n = 5/group.
Fig 3: DLGAP1-AS2 upregulated CD151 by interacting with E2F1 to regulate the radiation resistance of rectal cancer stem cells.(A–B) Western blot for measuring the protein levels of CD133, MDR1, BCRP1 and γ-H2AX and AKT/mTOR/cyclinD1 signaling. *P<0.05, **P<0.01, ***P<0.001. The error bar represents the mean ± SD. Each experiment was repeated three times.
Fig 4: The ALDH+CD44+CXCR4+CD24+ subpopulation in human PCa tissues.a Western blot analysis was performed to determine the protein expressions of PSA, HOXB9, ALDH, CD44, CXCR4 and CD24 in the controls (PCa tissue with Gleason score 6), para-carcinoma (2 mm away from PCa tissue), initial PCa tissue (derived from PCa at first diagnosis via radical prostatectomy) and refractory PCa tissue (derived after recurrence), respectively. ß-actin was used as an internal control. b Quantification of (a). *P < 0.05 vs. initial PCa tissues. (n = 6). c Human PCa tissue was subcutaneously implanted into NOD-SCID mice to establish a patient-derived xenograft (PDX) model. Subsets of cells (as indicated) were derived from the PDX model and seeded in 96-well plates (1 × 104 cells/well) and treated with different anti-androgens (as indicated) and chemotherapeutic agents, with 0.2% DMSO and 0.5% H2O2 were used as negative and positive controls, respectively. After 48 h of treatment, cells were incubated with alamarBlue solution for 4 h, and cell viability was measured with excitation wavelength at 530–560 nm and emission wavelength at 590 nm using a TECAN Infinite 200 PRO microplate reader. d Subsets of cells (as indicated) were derived from the PDX model and seeded in 96-well plates (1 × 104 cells/well). Cells were treated with different chemotherapeutic drugs, as indicated, for 72 h. Then, a WST-1 proliferation assay was performed. The absorbance was measured at 450 nm using a microplate reader. #P < 0.05 vs. bulk cells, as well as ALDH–CD44–CXCR4–CD24– -cells (n = 12); ?P < 0.05 vs. ALDH+CD44+CXCR4+CD24+-cells (n = 12). e ALDH+ CD44+ CXCR4+ CD24+-cells were isolated from PDX tumours at 8 weeks post implantation, before they were subject to HOXB9 knockdown. Then cells at 1 × 106 (together with 1 × 106 HS-5 cells to facilitate tumour formation) were subcutaneously injected into the NOD/SCID mouse. Derived tumours at 12 weeks post injection were subject to Western blotting analysis of the expression of APLN, HIF-1a, MSH6, GSTT2, metallothionein, ABCG2 and Bcl-2). GAPDH was used as an internal control. f Western blotting analysis of the expression of epithelial–mesenchymal transition-associated genes (Slug, Vimentin and E-cadherin). ß-actin was used as an internal control. (G-I) CD44+-, CD44+ a2ß1+-, ALDH+ CD44+ a2ß1+- and ALDH+ CD44+ CXCR4+ CD24+-PCa cells were obtained from orthotopic CWR22 tumours by FACS using the respective antibodies, whereas HOXB9-silenced ALDH+ CD44+ CXCR4+ CD24+-PCa cells were derived from ALDH+ CD44+ CXCR4+ CD24+-cells. Orthotopic tumour models were established using these subsets of cells, respectively. Mice were sacrificed at week 14 after inoculation. The time for developing a palpable tumour (g), tumour weights (h) and the number of metastatic foci (i) were recorded (n = 12). ?P < 0.05 vs. ALDH+ CD44+ CXCR4+ CD24+ cells-based implantation.
Fig 5: Inhibiting AKT/mTOR/cyclinD1 signaling weakened the promotive effect of overexpressed CD151 on the radioresistance of rectal cancer stem cells.Protein levels of CD133, MDR1, BCRP1 and ?-H2AX were detected with Western blot. *P<0.05, **P<0.01, ***P<0.001. The error bar represents the mean ± SD. Each experiment was repeated three times.
Supplier Page from Abcam for Anti-BCRP/ABCG2 antibody [EPR20080]