HIF-1α siRNA inhibited ERK1/2, JNK, and Akt activation by suppressing BCR-ABL1 and Met expression in K562 and K562/IR cells. (A-L) K562 and K562/IR cells were treated with 10 nM HIF-1α or negative siRNA for three days. (A, G) RNA was extracted, and HIF-1α levels were examined using real-time PCR. The results were standardized using GAPDH values and then expressed as a test:control ratio. These results are the average of four independent experiments. *P < 0.01 vs. untreated K562 or K562/IR cells. (B, H) Cell lysates were evaluated by immunoblotting using the antibodies. The content of HIF-1α, is normalized to the content of β-actin. The results are exemplary of three independent experiments. *P < 0.01 vs. untreated K562 or K562/IR cells. (C, I) RNA was extracted, and BCR-ABL1 or Met levels were examined using real-time PCR. The results were standardized using GAPDH values and then expressed as a test:control ratio. These results are the average of four independent experiments. *P < 0.01 vs. untreated K562 or K562/IR cells. (D, J) Cell lysates were evaluated by immunoblotting using the antibodies. The content of BCR-ABL1 or Met, is normalized to the content of β-actin. The results are exemplary of three independent experiments. *P < 0.01 vs. untreated K562 or K562/IR cells. (E, K) Cell lysates were evaluated by immunoblotting using the indicated antibodies. The content of phospho-ERK1/2, phospho-JNK, and phospho-Akt, normalized to the content of ERK1/2, JNK, and Akt. The results are exemplary of three independent experiments. *P < 0.01 vs. untreated K562 or K562/IR cells. (F, L) The cell survival rate of HIF-1α siRNA, was determined by the trypan blue staining assay. The results are exemplary of five independent experiments. *P < 0.01 vs. untreated K562 or K562/IR cells. (M, N) RNA was extracted, and VEGF, GATA1, or CA9 levels were assessed using real-time PCR. GAPDH was used as the endogenous control to normalize the gene expression data. The results are the average of four independent experiments. *P < 0.01 vs. untreated K562 or K562/IR cells.
