Figure 3. CYLD negatively regulates S. pneumoniae-induced inflammation in vitro and in vivo.

(A-B) HMEEC transfected with (A) CYLD WT or (B) siCYLD were stimulated with S. pneumoniae for 6 h, and IL-1β, IL-6, IL-8 and TNF-α mRNA expression was measured by real-time Q-PCR analysis. (C-F) WT and Cyld KO mice were inoculated transtympanically with S. pneumoniae (5 × 10⁶ CFU per mouse) for 9 h. (C) IL-1β, IL-6, MIP-2 and TNF-α mRNA expression in the middle ear of mice was measured by real-time Q-PCR analysis. (D) Tympanic cavity of the mouse ear was observed and recorded under the video-otoscope. (E) H&E staining of the middle ear tissues from mice was performed (Magnification, ×400; Scale bar, 20 μm) for histological analysis, and (F) the thickness of middle ear mucosa was measured from 16 middle ear sections per experimental group. Data in (A)-(C) and (F) are mean ± SD (A-C, n = 3; F, n = 16). *p<0.05, **p<0.01, ***p<0.001. Statistical analysis was performed using Student’s t-test. Pictures of the video otoscopy and H&E-stained middle ear tissues from one representative experiment are shown in (D) and (E) (D, n = 6; E, n = 16). Data are representative of three or more independent experiments. Sp, S. pneumoniae. WT, wild-type. KO, knock-out. siCYLD, CYLD siRNA.