Department of Neurosurgery, Tangshan Gongren Hospital, Tangshan, Hebei 063000, P.R. China.
Department of Neurosurgery, Tangshan Gongren Hospital, Tangshan, Hebei 063000, P.R. China.
Mol Med Rep. 2020 Dec;22(6):5399-5411. doi: 10.3892/mmr.2020.11618. Epub 2020 Oct 20.
Reperfusion is a critical therapeutic intervention used following acute ischemic stroke; however, it may cause cerebral ischemia/reperfusion injury (CIRI) and aggravate brain damage. Piceatannol (Pic), a hydroxylated analog of resveratrol, has been reported to exhibit anti‑inflammatory effects. However, the detailed molecular mechanisms and its effects on CIRI have not been sufficiently assessed, and, to the best of our knowledge, current methods of prevention of CIRI are limited. The aim of the present study was to investigate the effects of Pic on improving neurological function in a mouse model of CIRI. For the animal experiments, 8‑week‑old C57BL/6 mice were raised and randomly grouped, and an in vivo model of CIRI was established. Mice were administered a low (10 mg/kg/day) or high‑dose (20 mg/kg/d) of Pic 1 h after CIRI orally and once daily for the next 6 days. Neurological dysfunction was assessed using a modified neurological severity score and a rotarod test 1 week after CIRI establishment, and the cognitive status of the mice was assessed using a Morris water maze. Hematoxylin and eosin staining was used to evaluate the histopathological changes. The expression levels of sirtuin 1 (Sirt1), FoxO1, cleaved caspase‑3 (CC‑3), Bax and Bcl‑2 were measured using western blotting. Intracellular reactive oxygen species (ROS) generation, antioxidant enzymes [superoxide dismutase, glutathione (GSH) peroxidase and catalase] and non‑enzymatic antioxidants (GSH) were also detected using spectrophotometry. After inhibition of the Sirt1/FoxO1 pathway, a TUNEL assay was used for the detection of apoptotic cells in vitro and in vivo. The co‑localization of neuron‑specific nuclear protein and CC‑3 was assessing using immunofluorescent staining. Pic improved neurological functions and ameliorated hippocampal neuronal pathology following CIRI. In addition, the expression levels of CC‑3 and Bax and intracellular ROS levels were increased, while levels of antioxidant and non‑enzymatic enzymes were decreased in the mouse model of CIRI. Low and high doses of Pic significantly decreased ROS production and the expression levels of apoptosis‑related proteins, but increased antioxidant enzyme levels. However, a high‑dose of Pic did not result in increased levels of non‑enzymatic enzymes. Furthermore, low and high doses of Pic treatment significantly activated the Sirt1/FoxO1 pathway. Following inhibition of the Sirt1/FoxO1 pathway, the percentage of TUNEL‑positive cells and expression of CC‑3 were increased, and CC‑3 was enriched in neurons. The antioxidant effects of Pic were blocked by inhibition of Sirt1 in vitro and in vivo. In conclusion, these results suggested that Pic may exert a neuroprotective effect against in hippocampal neurons via the Sirt1/FoxO1 pathway.
再灌注是急性缺血性脑卒中后使用的一种关键治疗干预措施；然而，它可能导致脑缺血/再灌注损伤（CIRI）并加重脑损伤。白皮杉醇（Pic）是白藜芦醇的一种羟基化类似物，据报道具有抗炎作用。然而，其详细的分子机制及其对 CIRI 的影响尚未得到充分评估，而且据我们所知，目前预防 CIRI 的方法有限。本研究旨在探讨 Pic 对改善 CIRI 小鼠模型神经功能的影响。在动物实验中，饲养 8 周龄 C57BL/6 小鼠并进行随机分组，建立 CIRI 的体内模型。在 CIRI 后 1 小时，通过口服给予小鼠低（10mg/kg/天）或高（20mg/kg/d）剂量的 Pic，每天一次，连续 6 天。在 CIRI 建立后 1 周，使用改良神经严重程度评分和旋转棒试验评估神经功能障碍，使用 Morris 水迷宫评估小鼠的认知状态。苏木精和伊红染色用于评估组织病理学变化。使用蛋白质印迹法测量沉默调节蛋白 1（Sirt1）、叉头框蛋白 O1（FoxO1）、裂解的半胱天冬酶-3（CC-3）、Bax 和 Bcl-2 的表达水平。使用分光光度法还检测了细胞内活性氧（ROS）的产生、抗氧化酶[超氧化物歧化酶、谷胱甘肽（GSH）过氧化物酶和过氧化氢酶]和非酶抗氧化剂（GSH）。抑制 Sirt1/FoxO1 通路后，使用 TUNEL 测定法在体外和体内检测凋亡细胞。使用免疫荧光染色评估神经元特异性核蛋白和 CC-3 的共定位。Pic 改善了 CIRI 后神经功能并改善了海马神经元病理学。此外，在 CIRI 小鼠模型中，CC-3 和 Bax 的表达水平以及细胞内 ROS 水平增加，而抗氧化和非酶酶的水平降低。低剂量和高剂量的 Pic 可显著降低 ROS 产生和凋亡相关蛋白的表达水平，但可增加抗氧化酶水平。然而，高剂量的 Pic 并没有导致非酶酶水平的增加。此外，低剂量和高剂量的 Pic 治疗可显著激活 Sirt1/FoxO1 通路。抑制 Sirt1/FoxO1 通路后，TUNEL 阳性细胞的百分比和 CC-3 的表达增加，CC-3 在神经元中富集。体外和体内抑制 Sirt1 可阻断 Pic 的抗氧化作用。综上所述，这些结果表明，Pic 可能通过 Sirt1/FoxO1 通路在海马神经元中发挥神经保护作用。
