Inhibition of DC maturation in response to different TLR ligands. (A) Human monocyte–derived DCs were cultured under normoxic or hypoxic conditions for 18 hours and analyzed for mRNA expression of TLR members by real-time PCR. Data are means plus or minus SD of 3 independent experiments. TLR mRNA levels in normoxic conditions () were set to 1.0 arbitrary unit. (B) Surface expression (flow cytometric analysis) of selected chemokine receptors and the DC maturation marker CD83 on cells exposed to different TLR agonsists. DCs were cultured under normoxic or hypoxic conditions for 18 hours in the presence or absence of different TLRs agonists as indicated: 10 ng/mL LPS (TLR2/4; Sigma-Aldrich); 10 ng/mL LPS (TLR4; Alexis Biochemicals); 2 μg/mL PAM₃CSK₄ (TLR2); 10 μg/mL poly (I:C) (TLR3); or 3 μg/mL R-848 (TLR7/8). Data are means plus or minus SD of 3 independent experiments. *P < .05 versus normoxic counterparts. (C) Supernatants were collected 18 hours after treatments and analyzed for cytokine production by ELISA after exposure to different TLR agonists. Results are means plus or minus SD of 4 experiments. *P < .05 versus DCs matured in normoxia.