Figure 1.
Figure 1. Effects of K-11706 and K-13144 on GATA and HIF-1 binding activities. (A) Electrophoretic mobility shift assay (EMSA) was performed using 1.0 μg protein from Hep3B cells under normoxic conditions (lanes 2-4 and 6-8), and incubated with 100 nM K-11706 (lanes 3 and 7) and 100 nM K-13144 (lanes 4 and 8). The closed circle and triangle at the left indicate the positions of the GATA and HIF-1 transcription factors. The autoradiograph is representative of 3 different experiments, using different nuclear extracts, with similar results. Densitometric analyses of the bands expressed relative to the control are indicated by circles or triangles. * indicates significance compared with normoxic control, P < .005. (B) Effect of K-11706 on enhanced expression of GATA induced by IL-1β or TNF-α. EMSA was performed using 1.0 μg protein from Hep3B cells under hypoxic conditions (lanes 2-7), normoxic condition (lane 9), and incubated with 15 U/mL rhIL-1β (lane 3), 220 U/mL rhTNF-α (lane 4), 100 nM K-11706 (lane 5), 15 U/mL rhIL-1β plus 100 nM K-11706 (lane 6), and 220 U/mL rhTNF-α plus 100 nM K-11706 (lane 7) for 24 hours. The closed circle at the left indicates the position of the GATA transcription factors. The autoradiograph is representative of 3 different experiments, using different nuclear extracts, giving similar results. Densitometric analyses of the bands expressed relative to the control are indicated by the circles. * indicates significance compared with hypoxic control, P < .005; **, significance compared with IL-1β, P < .005; ***, significance compared with TNF-α, P < .005. (C) Effects of K-11706 on enhanced expression of HIF-1 induced by IL-1β or TNF-α. EMSA was performed using 1.0 μg protein from Hep3B cells under normoxic conditions (lanes 2-5, 7-11), hypoxic condition (lane 12), and incubated with 15 U/mL rhIL-1β (lane 3), 220 U/mL rhTNF-α (lane 8), 100 nM K-11706 (lanes 5 and 10), 15 U/mL rhIL-1β plus 100 nM K-11706 (lane 4), and 220 U/mL TNF-α plus 100 nM K-11706 (lane 9) for 24 hours. The closed triangle at the left indicates the position of the HIF-1 transcription factor. The autoradiograph is representative of 3 different experiments using different nuclear extracts, giving similar results. Densitometric analyses of the bands expressed relative to the control are indicated by the triangles. * indicates significance compared with normoxic control, P < .005; **, significance compared with IL-1β, P < .005; ***, significance compared with TNF-α, P < .005.

Effects of K-11706 and K-13144 on GATA and HIF-1 binding activities. (A) Electrophoretic mobility shift assay (EMSA) was performed using 1.0 μg protein from Hep3B cells under normoxic conditions (lanes 2-4 and 6-8), and incubated with 100 nM K-11706 (lanes 3 and 7) and 100 nM K-13144 (lanes 4 and 8). The closed circle and triangle at the left indicate the positions of the GATA and HIF-1 transcription factors. The autoradiograph is representative of 3 different experiments, using different nuclear extracts, with similar results. Densitometric analyses of the bands expressed relative to the control are indicated by circles or triangles. * indicates significance compared with normoxic control, P < .005. (B) Effect of K-11706 on enhanced expression of GATA induced by IL-1β or TNF-α. EMSA was performed using 1.0 μg protein from Hep3B cells under hypoxic conditions (lanes 2-7), normoxic condition (lane 9), and incubated with 15 U/mL rhIL-1β (lane 3), 220 U/mL rhTNF-α (lane 4), 100 nM K-11706 (lane 5), 15 U/mL rhIL-1β plus 100 nM K-11706 (lane 6), and 220 U/mL rhTNF-α plus 100 nM K-11706 (lane 7) for 24 hours. The closed circle at the left indicates the position of the GATA transcription factors. The autoradiograph is representative of 3 different experiments, using different nuclear extracts, giving similar results. Densitometric analyses of the bands expressed relative to the control are indicated by the circles. * indicates significance compared with hypoxic control, P < .005; **, significance compared with IL-1β, P < .005; ***, significance compared with TNF-α, P < .005. (C) Effects of K-11706 on enhanced expression of HIF-1 induced by IL-1β or TNF-α. EMSA was performed using 1.0 μg protein from Hep3B cells under normoxic conditions (lanes 2-5, 7-11), hypoxic condition (lane 12), and incubated with 15 U/mL rhIL-1β (lane 3), 220 U/mL rhTNF-α (lane 8), 100 nM K-11706 (lanes 5 and 10), 15 U/mL rhIL-1β plus 100 nM K-11706 (lane 4), and 220 U/mL TNF-α plus 100 nM K-11706 (lane 9) for 24 hours. The closed triangle at the left indicates the position of the HIF-1 transcription factor. The autoradiograph is representative of 3 different experiments using different nuclear extracts, giving similar results. Densitometric analyses of the bands expressed relative to the control are indicated by the triangles. * indicates significance compared with normoxic control, P < .005; **, significance compared with IL-1β, P < .005; ***, significance compared with TNF-α, P < .005.

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