The present study analyzed the change of Toll-like receptor 4 (TLR4) and nuclear factor-κB (NF-κB) expression in focal cerebral ischemia-reperfusion injury model. A sample of 36 Sprague-Dawley rats were randomly selected and divided into sham operation group (group S), control group (group C) and Chrysanthemum ester group (NF-κB inhibitor, group CE), each group consisted of 12 rats. The rat model of focal cerebral ischemia-reperfusion was established. The physiological indexes and neurological severity score of rats was recorded by a double-blind method. The cerebral infarction area was evaluated by triphenyltetrazole oxide (TTC) staining on brain slices. Apoptosis was evaluated by TUNEL staining. Semi-quantitative PCR and western blot analysis was used to measure the expression of TLR4 and NF-κB. The neurological severity score of rats in group C and CE were found to be significantly lower than group S (P<0.01). The TTC staining results showed that group C and CE had different levels of cerebral infarction but the area of infarction in group CE was significantly lower than group C (P<0.01). In addition, the number of TUNEL positive cells in group CE was significantly lower than group C (P<0.01). Semi-quantitative PCR and westernblot analysis results showed that the expression of NF-κB and TLR4 of group S was significantly lower than that of group C and group CE (P<0.01), the relative expression of NF-κB and TLR4 of group CE was significantly lower than that of group C (P<0.01). Moreover, the expression of NF-κB p65/p50 of group CE and group C was significantly higher than that of group S (P<0.01). This study concludes that the focal cerebral ischemia-reperfusion injury in rats can cause brain damage and cell apoptosis. This effect might be associated to the increased expression of NF-κB and TLR4, and the activation of TLR4/NF-κB signaling pathway.
Willed movement facilitates neurological rehabilitation in patients with stroke. Focal ischaemia is the hallmark of patients after stroke, though the detailed molecular mechanism by which willed movement affects neurological rehabilitation after stroke is not fully understood. The aim of the present study was to dissect the key factors of the hypoxia signaling pathway responsible for the willed movement‑improved rehabilitation. Sprague‑Dawley rats undergoing right middle cerebral artery occlusion (MCAO) surgery were randomly divided into four groups: MCAO alone, willed movement (WM), environmental modification (EM) and common rehabilitation (CR). The neurological behaviour score was assessed, and infarction areas were detected by TTC staining. In addition, angiogenesis‑associated genes (vascular epithelial growth factor, angiogenin‑1, matrix metalloproteinases‑2 and ‑9) and hypoxia inducible factor (HIF)‑1α expression was investigated in cells derived from MCAO, WM, EM and CR groups. Finally, the role of HIF‑1α using HIF‑1α knockdown in HUVECs under hypoxic conditions was evaluated. WM significantly improved neurological behaviour and rehabilitation by increasing the behaviour score and by decreasing the infarction area. In addition, CR, EM and WM raised the expression of angiogenesis‑associated genes and HIF‑1α, thereby promoting in vitro tube formation of primary endothelial cells. Knockdown of HIF‑1α in HUVECs restored the increased expression of angiogenesis‑associated genes to normal levels and inhibited in vitro tube formation of HUVECs. Willed movement most effectively improved the neurological rehabilitation of rats with focal ischaemia through upregulation of HIF‑1α. The present findings provide insight into willed movement‑facilitated rehabilitation and may help treat stroke‑triggered motor deficit and improve angiogenesis of cerebral endothelial cells.
Given that the role of Gelsemine in neuroinflammation has been demonstrated, this research aimed to investigate the effect of Gelsemine on neonatal hypoxic-ischemic (HI) brain injury. An in vivo HI brain injury neonatal mouse model and an in vitro oxygen–glucose deprivation (OGD) cell model were established and pretreated with Gelsemine. The brain infarct volume, neuronal loss and apoptosis, as well as spatial learning and memory were examined by TTC staining, Nissl’s staining, TUNEL staining and Morris water maze test. Immunohistochemical staining was applied to detect the microglia cells and astrocytes in the mouse brain tissue. The cell viability was analyzed by CCK-8 assay. The levels of malondialdehyde (MDA), superoxide dismutase (SOD), TNF-α, IL-1β, and IL-6 were determined via ELISA. The lactate dehydrogenase (LDH) release and reactive oxygen species (ROS) level in OGD-treated cells were detected by colorimetry and DCFH-DA staining. Nrf2, HO-1, and inflammation-related factors were analyzed by immunofluorescence, qRT-PCR, or western blot. Gelsemine reduced the infarct volume and neuronal loss and apoptosis, yet improved spatial learning and memory impairment of HI-injured mice. Gelsemine inhibited the elevated MDA, TNF-α, IL-1β, IL-6, LDH and ROS levels, promoted the reduced SOD level and viability, and strengthened the up-regulation of HO-1 and Nrf2 in brain tissues and OGD-treated cells. However, Nrf2 silencing reversed the effects of Gelsemine on the Nrf2/HO-1 pathway, inflammation, and oxidative stress in OGD-treated cells. Gelsemine produces neuroprotective effects on neonatal mice with HI brain injury by suppressing inflammation and oxidative stress via Nrf2/HO-1 pathway.
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