Research Paper Volume 12, Issue 10 pp 9534—9548

Figure 4. HIF1α is required for regulating the anti-inflammatory effect of DEX on LPS-treated macrophages. (A) BMDMs were treated with 100 ng/ml LPS and 1 μM DEX for 4 h. The mRNA and protein levels of HIF1α were determined by RT-PCR and Western blotting, respectively. n = 3; mean ± SEM; ** P < 0.01, *** P < 0.001. (B) BMDMs were transfected with HIF1α siRNA or Negative control siRNA for twenty-four hours. The mRNA and protein levels of HIF1α were determined by real-time PCR and Western blotting, respectively. n = 3; mean ± SEM; *** P < 0.001. (C) BMDMs were transfected as in (B). Twenty-four hours after transfection, the cells were treated with 100 ng/ml LPS and 1 μM DEX for 4 h. Supernatants were collected, and the levels of glucose and lactate were measured. n = 3; mean ± SEM; * P < 0.05. (D) BMDMs were transfected as in (B). Twenty-four hours after transfection, the cells were treated with 100 ng/ml LPS and 1 μM DEX for 4 h. The mRNA levels of GLUT1, HK2 and PFKFB3 were determined by RT-PCR. n = 3; mean ± SEM; * P < 0.05. (E) BMDMs were transfected as in (B). Twenty-four hours after transfection, the cells were treated with 100 ng/ml LPS and/or 5 mM ATP and 1 μM DEX for 4 h. Levels of IL-1β, TNF-α and IL-6 were determined by ELISA. n = 3; mean ± SD; ** P < 0.01.