IngentaConnect Fast Track Article: Activation of TLR4/NF-κB sig...ry after subarachnoid hemorrhage

Authors: Chun-Xiao Ma, Wei-Ning Yin, Bo-Wen Cai, Min He, Jian Wu, Jun-Yi Wang, Yi Zeng, Jun-Li Ding, Chao You

Abstract:

Objectives: To detect the expression of TLR4 in the brain and to clarify the activation of the Toll-like receptor 4/nuclear factor-kappa B (TLR4/NF-κB) signaling in the rat brain following subarachnoid hemorrhage (SAH). Methods: Male Sprague-Dawley rats were divided into four different experimental groups: sham, SAH, SAH+vehicle and SAH+E5564. TLR4 antagonist E5564 or vehicle was timely injected intracisternally after the induced SAH. The rats were decapitated and their brains were removed at different time points after a single injection of blood into the pre-chiasmatic cistern. The mRNA expression of TLR4 was measured with TaqMan real-time RT-PCR, and protein expression with immunohistochemistry and Western blot analysis. NF-κB activity was assessed by Western blot assay and ELISA. Concentrations of tumor necrosis factoralpha (TNF-α), interleukin-1beta (IL-1β) and interleukin-6 (IL-6) were determined by ELISA. In addition, neurological scores were assessed for each group. Results: A biphasic change in TLR4 expression was observed at the levels of both mRNA and protein: an initial peak (2–6 hours) and a sustained elevation (12–48 hours). The inducible expression of TLR4-like immunoreactions was detectable predominantly in glial cells and vascular endothelium. A similar biphasic change in the nuclear translocation and DNA-binding of NF-κB subunit p65 as well as the production of NFκB-regulated proinflammatory cytokines (TNF-α, IL-1β and IL-6) were observed. Administration of E5564 prevented the up-regulated NF-κB activity and cytokines levels, as well as reduced neurological deficits of the rats to a certain extent. Discussion: These data suggest that experimental SAH induces significant up-regulation of TLR4 expression and the NF-κB signaling in early brain injury. Activation of the TLR4/NF-κB signaling may regulate the inflammatory responses after SAH.

Activation of TLR4/NF-κB signaling pathway in early brain injury after subarachnoid hemorrhage