醒脑开窍针刺法对出血性卒中大鼠神经细胞铁死亡的影响

Effect of Xingnao Kaiqiao Acupuncture on Ferroptosis of Nerve Cells in Rats with Hemorrhagic Stroke

[目的]探讨醒脑开窍针刺法对出血性卒中大鼠脑组织神经细胞铁死亡的影响,及其促进神经功能修复的铁死亡应激机制。[方法]将30只雄性SD大鼠随机分为假手术组、模型组、针刺组各10只,模型组、针刺组采用自体血注入法构建出血性卒中大鼠模型;假手术组采用与模型组相同的操作,以微量注射器于相同位置进针,注入等量生理盐水。针刺组造模成功后选取人中、内关、三阴交穴进行针刺,每日1次,共3d;模型组与假手术组不做处理。采用Zausinger六分法对大鼠神经功能进行评分,流式细胞术检测神经细胞凋亡情况,比色法检测铁含量,ELISA检测法测定丙二醛(MDA)、谷胱甘肽(GSH)的含量,免疫荧光检测谷胱甘肽过氧化物酶4(GPX4)表达。[结果]与假手术组比较,模型组大鼠神经功能评分降低(P<0.05),而干预后针刺组大鼠神经功能评分高于模型组(P<0.05);模型组大鼠早期凋亡细胞、晚期凋亡细胞及总凋亡细胞百分数比假手术组高(P<0.01),而针刺组大鼠各凋亡细胞百分数比模型组低(P<0.01);与假手术组比较,模型组大鼠GPX4、GSH水平降低,总铁含量、MDA水平升高(P<0.05),而针刺组大鼠GPX4、GSH水平较模型组高,铁蛋白含量、MDA水平较模型组低(P<0.05)。[结论]醒脑开窍针刺法可促进出血性卒中大鼠神经功能修复,可能与降低GPX4、GSH水平,从而提高细胞抗氧化能力,抑制神经细胞铁死亡的发生有关。

[Objective]To investigate the effect of Xingnao Kaiqiao acupuncture on ferroptosis in brain tissue of rats with hemorrhagic stroke and its mechanism of promoting nerve function repair。[Methods]Thirty male SD rats were randomly divided into sham operation group,model group and acupuncture group,with 10 rats in each group.The rat model of hemorrhagic stroke was constructed by autologous blood injection in model group and acupuncture group.The sham operation group was treated with the same operation as the model group,and the same amount of normal saline was injected into the same position with a micro syringe.After successful modeling,Renzhong,Neiguan and Sanyinjiao as acupoints were selected for acupuncture in the acupuncture group,once a day for 3 days.The rats in the model group and the sham operation group were only fixed without other treatment.The neurological function of rats was scored by Zausinger six-point method.The apoptosis of nerve cells was detected by flow cytometry.The content of ferritin was de⁃tected by ferrozine colorimetry.The contents of malondialdehyde(MDA)and glutathione(GSH)were determined by ELI⁃SA.The expression of glutathione peroxidase 4(GPX4)was detected by immunofluorescence.[Results]Compared with the sham operation group,the neurological function score of the model group was decreased(P<0.05),while the neuro⁃logical function score of the acupuncture group was higher than that of the model group(P<0.05).Compared with the sham operation group,the percentage of early apoptotic cells,late apoptotic cells and total apoptotic cells in the model group was significantly increased(P<0.01),while the percentage of apoptotic cells in the acupuncture group was signifi⁃cantly lower than that in the model group(P<0.01).Compared with the sham operation group,the levels of GPX4 and GSH in the model group decreased,and the levels of ferritin and MDA increased(P<0.05).The levels of GPX4 and GSH in the acupuncture group were higher than those in the model group,and the levels of ferritin and MDA were lower than those in the model group(P<0.05)[.Conclusion]Xingnao Kaiqiao acupuncture can promote the repair of neurolog⁃ical function in hemorrhagic stroke rats,which may be related to the reduction of GPX4 and GSH levels,thereby im⁃proving the antioxidant capacity of cells and inhibiting the occurrence of ferroptosis in nerve cells.