The TLR2 blocking antibody inhibition of NTHi induced HBD-2 mRNA expression in HMEEC-1 cells and in TLR2 knock out mice. (A) Pretreatment of HMEEC-1 with human TLR2 blocking antibody for 30 minutes (10 μg/ml, clone TL2.1, eBioscience) reduced the HBD-2 mRNA expression dramatically in response to NTHi lysate stimulation (5 μg/ml). In contrast, human TLR4 blocking antibody did not show any significant effect suggesting that TLR2 is one of the major signaling receptors for inducing human β-defensin 2 expression in response to NTHi stimulation in HMEEC-1. The control group was treated with an isogenic antibody. (B) NTHi WCL-induced expression of murine β-defensin 2 mRNA (mBD-2) in wild type (C57 BL/6) and TLR2-/- mice was compared. Mice were injected with 10 μl of NTHi WCL transtympanically in the left ear. The right ear served as a PBS injected control. Four hours post-injection, the total RNA was harvested and Defb2 expression measured by quantitative PCR (n = 3). Values are given as mean ± standard deviation. N = 3. *: p < 0.05.