Figure 4. Vinpocetine suppresses S. pneumoniae-induced inflammatory response via upregulating CYLD.

(A) HMEEC transfected with siCON or siCYLD were pre-treated with vinpocetine (10 μM) for 1 h, followed by S. pneumoniae stimulation for 6 h, and IL-8 and TNF-α mRNA expression was measured by real-time Q-PCR analysis. (B-D) WT and Cyld KO mice were inoculated transbullarly with S. pneumoniae (2 × 10⁶ CFU per mouse), and post-treated intraperitoneally with vinpocetine (10 mg/kg) 2 h after S. pneumoniae inoculation. The treatment was repeated at a dose of 10 mg/kg/day during the experiment. The inoculated mice were then sacrificed 3 days after S. pneumoniae inoculation. (B) 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. (C) H&E staining of the middle ear tissues from mice was performed (Magnification, ×400; Scale bar, 20 μm) for histological analysis, and (D) the thickness of middle ear mucosa was measured from 40 middle ear sections per experimental group. Data in (A) are mean ± SD (n = 3). Data in (B) and (D) are mean ± SEM (B, n = 4; D, n = 40). *p<0.05, **p<0.01, ***p<0.001. n.s., not significant. Statistical analysis was performed using Student’s t-test. Pictures of the H&E-stained middle ear tissues from one representative experiment are shown in (C) (n = 16). Data are representative of three independent experiments. CON, control. Sp, S. pneumoniae. Vinp, vinpocetine. siCYLD, CYLD siRNA. WT, wild-type. KO, knock-out.