Salvianolic Acid D Alleviates Cerebral Ischemia-Reperfusion Injury by Suppressing the Cytoplasmic Translocation and Release of HMGB1-Triggered NF-κB Activation to Inhibit Inflammatory Response

Abstract

Inflammatory response participates in the overall pathophysiological process of stroke. It is a promising strategy to develop antistroke drugs targeting inflammation. This study is aimed at investigating the therapeutic effect and anti-inflammatory mechanism of salvianolic acid D (SalD) against cerebral ischemia/reperfusion (I/R) injury. A rat middle cerebral artery occlusion/reperfusion (MCAO/R) injury model was established, and an oxygen-glucose deprivation/reoxygenation (OGD/R) injury model was established in PC12 cells. Neurological deficit score, cerebral infarction, and edema were studied in vivo. Cell viability was achieved using the MTT method in vitro. The Bax, Bcl-2, cytochrome c, HMGB1, TLR4, TRAF6, NF-κB p65, p-NF-κB p65, and cleaved caspase-3 and -9 were tested via the Western blot method. Cytokines and cytokine mRNA, including TNF-α, IL-1β, and IL-6, were studied via ELISA and PCR methods. The translocation of HMGB1 and NF-κB were studied by immunofluorescence assay. The HMGB1/NeuN, HMGB1/GFAP, and HMGB1/Iba1 double staining was carried out to observe the localization of HMGB1 in different cells. Results showed that SalD alleviated neurological impairment, decreased cerebral infarction, and reduced edema in I/R rats. SalD improved OGD/R-downregulated PC12 cell viability. SalD also promoted Bcl-2 expression and suppressed Bax, cytochrome c, and cleaved caspase-3 and -9 expression. SalD decreased the intensity of TLR4, MyD88, and TRAF6 proteins both in vivo and in vitro, and significantly inhibited the NF-κB nuclear translocation induced by I/R and OGD/R. What’s more, SalD inhibited HMGB1 cytoplasmic translocation in neurons, astrocytes, and microglia in both the cortex and hippocampus regions of I/R rats. In conclusion, SalD can alleviate I/R-induced cerebral injury in rats and increase the PC12 cell viability affected by OGD/R. The anti-inflammatory mechanism of SalD might result from the decreased nuclear-to-cytoplasmic translocation of HMGB1 and the inhibition on its downstream TLR4/MyD88/NF-κB signaling.

Transplantation of umbilical cord blood mononuclear cells attenuates the expression of IL-1β via the TLR4/NF-κB pathway in hypoxic-ischemic neonatal rats

“…However, whether the increase in TLR4 protein expression could alleviate the following inflammation injury remains uncertain. Studies showed that NF-κB could be activated by the upstream molecule TLR4, phosphorylated, and transferred to nuclei, resulting in inflammation injury [13]. In the present work, the expression of pNF-κB in the nuclei increased significantly in the HIBD model 7 days after HIBD, indicating that hypoxia-ischemia activated NF-κB.…”

Physcion Protects Rats Against Cerebral Ischemia-Reperfusion Injury via Inhibition of TLR4/NF-kB Signaling Pathway

“… 29 , 30 It has been shown that TLR4 contributes to I/R-induced inflammatory response. 31 Our data indicated that the expression of TLR4 was increased in SH-SY5Y cells exposed to OGD/R and in brain tissues of rats subjected to I/R injury. In addition, NF-κB is an essential transcription factor that plays an important role in cell growth and inflammation.…”

“… 9 , 32 NF-κB exists in cytoplasm as an inactive dimeric form binding to IκB. 31 TLR4/MyD88 signaling can phosphorylate IκB, and then release the active form of NF-κB (NF-κB translocation from the cytoplasm to the nucleus). 33 NF-κB participates in the pathological process of IS, which can be triggered by ROS, and several inflammatory cytokines.…”

Alpinia oxyphylla Miq extract reduces cerebral infarction by downregulating JNK-mediated TLR4/T3JAM- and ASK1-related inflammatory signaling in the acute phase of transient focal cerebral ischemia in rats

“…Post-ischemic inflammation, one of the main pathological features in the early stage of cerebral I/R injury, contributes to the release of pro-inflammatory mediators, which exacerbate cerebral infarction [ 39 ]. Studies have reported that TLR4 expressed on microglia and astrocytes plays a crucial role in the generation of pro-inflammatory cytokines in the initial stage of cerebral I/R injury, whereas pharmacological interventions alleviate cerebral infarction by inhibiting TLR4-mediated microglial and astrocytic activation in the ischemic area in the acute phase of transient MCAo [ 40 ]. In the present study, the TTC-stained brain sections revealed that cerebral infarction was predominantly distributed in the right cerebral hemisphere involving the cortex and striatum at 1 day after 90 min of MCAo.…”