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. Author manuscript; available in PMC: 2020 Jul 1.
Published in final edited form as: Mol Cancer Ther. 2019 Sep 18;19(1):199–210. doi: 10.1158/1535-7163.MCT-19-0242

Figure 2.

Figure 2.

ALDH1A1 enhances resistance of EOC cells to olaparib. A, ALDHbr cells exhibit resistance to olaparib. ALDHdim and ALDHbr cells were sorted from PEO1 and Kuramochi cells, treated with olaparib at various doses for 7 days, cell viability was determined using methylene blue staining (IC50: PEO1-ALDHdim: 1.43 μM, PEO1-ALDHbr: 3.52 μM; Kura-ALDHdim: 0.82 μM, Kura-ALDHbr: 2.78 μM). N = 3, Bar: SD, **: P < 0.01 compared to the ALDHdim group. B and C, ALDH1A1 is the major ALDH family gene contributes to the ALDH activity in ALDHbr cells and olaparib-induced high ALDH activity in EOC cells. Expression of various ALDH family genes in ALDHdim and ALDHbr cells sorted from PEO1 cells were analyzed using qRT-PCR (B). PEO1 cells were treated with olaparib for 7 days, expression of various ALDH family genes in these cells were determined using qRT-PCR (C). N = 3, Bar: SD, *: P < 0.05; **: P < 0.01 compared to the ALDHdim group and the DMSO group, respectively. D and E, ALDH1A1 overexpression decreased the sensitivity of EOC cells to olaparib. PEO1 cells were transfected with ALDH1A1 expressing plasmids for 48 h, treated with olaparib at various doses for 7 days. Immunoblotting was conducted to determine the expression level of ALDH1A1 after 48 h of transfection (D). Cell viability after olaparib treatment was determined using methylene blue staining (IC50: EV: 1.45 μM, ALDH1A1: 1.9 μM) (E). N = 4, Bar: SD, **: P < 0.01 compared to the EV group. F-H, Knockdown of ALDH1A1 sensitizes EOC cells to olaparib. PEO1-R cells were transfected with ALDH1A1 siRNA for 48 h, treated with olaparib at various doses for 7 days. Immunoblotting was conducted to determine the expression level of ALDH1A1 after 48 h of transfection (F). The ALDEFLUOR assay was used to determine the ALDH activity in these cells after 48 h of transfection (G). Cell viability after olaparib treatment was determined using methylene blue staining (IC50: siCtrl: 65.3 μM, si1A1-pool: 12.5 μM, si1A1-1: 23.3 μM, si1A1-4: 20.8 μM) (H). N = 4, Bar: SD, **: P < 0.01 compared with the siCtrl group.