Effects of nourishing qi, activating blood circulation, and inducing resuscitation on nerve cell pyroptosis after cerebral ischemia-reperfusion

Lin-Quan Liu, Xiao-ping Huang, Yazhen Cai, Yan She, Chang-Qing Deng
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Abstract

Cerebral ischemia-reperfusion (CIR) is a serious complication often associated with cerebral ischemia. The purpose of this study was to explore the therapeutic effect of nourishing qi, activating blood circulation, and inducing resuscitation (Borneol with astragaloside IV and Panax notoginseng total saponins, BAP) on CIR. Neurological function score system was used to determine the neurological function. The survival of nerve cells was detected by Nissl staining. The levels of IL-1β, IL-18, IL-4, and IL-10 were detected by ELISA. The expression of GSDMD, GSDMD-N, Nrf2, and HO-1 proteins in hippocampus tissues was measured by immunohistochemistry (IHC). Western blot, RT-qPCR, or immunofluorescence (IF) were used to detect the NACHT, LRR, and PYD domains-containing protein 3 (NLRP3), pro-Caspase-1, Caspase-1, Nrf2, and HO-1 expression. Lactate dehydrogenase (LDH) level was analyzed by LDH release assay. Cell viability was determined by cell counting kit-8 (CCK8). Apoptosis was detected by flow cytometry. BAP significantly promoted the recovery of nerve function, the activity of nerve cells, and the expression of Nrf2, HO-1, IL-4, and IL-10 in rat hippocampus tissues after CIR. BAP has an obvious inhibitory effect on the expression of NLRP3, pro-Caspase-1, and Caspase-1 proteins, the release of IL-1β and IL-18 factors, and neuronal pyroptosis in hippocampal tissues. BAP also promoted IL-4 and IL-10 levels, and the activity of SH-SY5Y cells. The IL-1β, IL-18, NLRP3, pro-Caspase-1, Caspase-1, GSDMD, and GSDMD-N expressions were significantly inhibited by BAP in vitro, which was reversed by Nrf2 knockdown. This study confirmed that BAP alleviated rat CIR and inhibited the pyroptosis of SH-SY5Y cells by regulating the Nrf2/HO-1 signaling pathway. This study provided new directions and ideas for the treatment of CIR.
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益气活血复苏对脑缺血再灌注后神经细胞焦亡的影响
脑缺血再灌注(CIR)是脑缺血的严重并发症。本研究旨在探讨益气活血醒脑(黄芪甲苷冰片加三七总皂苷,BAP)对大鼠CIR的治疗作用,并采用神经功能评分系统对大鼠的神经功能进行评分。尼氏染色检测神经细胞的存活情况。ELISA法检测各组IL-1β、IL-18、IL-4、IL-10水平。免疫组化(IHC)法检测海马组织中GSDMD、GSDMD- n、Nrf2、HO-1蛋白的表达。采用Western blot、RT-qPCR或免疫荧光(IF)检测NACHT、LRR和PYD结构域蛋白3 (NLRP3)、pro-Caspase-1、Caspase-1、Nrf2和HO-1的表达。乳酸脱氢酶(LDH)释放法测定乳酸脱氢酶(LDH)水平。采用细胞计数试剂盒-8 (CCK8)检测细胞活力。流式细胞术检测细胞凋亡。BAP显著促进大鼠CIR后海马组织神经功能恢复、神经细胞活性恢复及Nrf2、HO-1、IL-4、IL-10的表达,对海马组织NLRP3、pro-Caspase-1、Caspase-1蛋白表达、IL-1β、IL-18因子释放、神经元凋亡有明显抑制作用。BAP还能提高SH-SY5Y细胞的IL-4、IL-10水平和活性。BAP在体外显著抑制IL-1β、IL-18、NLRP3、pro-Caspase-1、Caspase-1、GSDMD、GSDMD- n的表达,并通过下调Nrf2使其表达逆转。本研究证实BAP通过调节Nrf2/HO-1信号通路减轻大鼠CIR,抑制SH-SY5Y细胞的焦亡。本研究为CIR的治疗提供了新的方向和思路。
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