Figure 1

Toll-like receptor 4 (TLR4) immunofluorescence and protein expression was increased in the corpus callosum and cerebellum in neonatal rats following hypoxic exposure. Confocal images showing the distribution of lectin/OX42 (green) and TLR4 (red) immunoreactive cells in the corpus callosum and cerebellum at 3 days after hypoxic exposure and the corresponding control (A-C). Colocalized expression of TLR4 and lectin/OX42 immunoreactive cells (arrows) in corpus callosum (Ac, Af; Bc, Bf) and cerebellum (Cc, Cf) can be seen. Note the upregulated expression of TLR4 in some lectin/OX42-positive microglial cells after hypoxia (Af, Bf, Cf). Western blotting of TLR4 protein expression in the corpus callosum (3 h, and 1, 3, 7 and 14 days) and cerebellum (3 h, and 1 and 3 days) of rats after hypoxic exposure and their corresponding controls is shown. The upper panels show specific bands of TLR4 (95 kDa) and β-actin (43 kDa). The lower panels are bar graphs showing significant changes in the optical density following hypoxic exposure (h) (normalized with β-actin, shown as fold change of control in 3 h (c)). TLR4 protein expression in the corpus callosum is significantly increased at 3 h, and 1 and 3 days after hypoxic exposure when compared with the matching control. At 7 and 14 days, TLR4 protein expression level was declined in comparison with the comparing control (D). TLR4 protein expression level in the cerebellum also increased significantly following hypoxia at 3 h, and 1 and 3 days when compared with the matching control (E). Significant differences in protein levels between hypoxic and control rats are expressed as *P <0.05 and **P <0.01. Fold-change values are calculated from triplicates and represented as mean ± SD. Scale bars in A-C = 50 μm. TLR4, Toll-like receptor 4.