骨髓间充质干细胞移植治疗大鼠蛛网膜下腔出血后铁死亡的机制研究

Study on mechanism of iron death after bone mesenchymal stem cells(BMSCs)transplantation in rats with subarachnoid hemorrhage

目的 探讨骨髓间充质干细胞(BMSCs)能否通过调控核转录E2相关因子2(Nrf2)/谷胱甘肽过氧化物酶4(GPX4)信号通路减轻蛛网膜下腔出血(SAH)后脑组织的铁死亡。方法 100只雌性SD大鼠中随机取出25只作为假手术组(Sham组)(不刺破大脑前动脉与大脑中动脉分支,仅在感到穿刺线受阻时退出)。剩余75只大鼠通过血管穿刺法复制SAH大鼠模型,75只大鼠均造模成功,随机分为蛛网膜下腔出血组(SAH组)、蛛网膜下腔出血+BMSCs移植组(BMSCs组)和蛛网膜下腔出血+BMSCs移植+Nrf2特异性抑制剂ML385干预组(ML385组),每组25只。造模3 d后,进行大鼠神经功能学评分和脑组织含水量测定;普鲁士蓝染色检测神经细胞内铁沉积情况;分光光度计法测定神经脑细胞中铁含量、丙二醛(MDA)含量、谷胱甘肽(GSH)含量及谷胱甘肽过氧化物酶4(GPX4)活性;Western blot检测大鼠脑组织中Nrf2和GPX4蛋白表达水平。结果 BMSCs组大鼠神经功能学评分较SAH组显著增高,脑组织含水量明显低于SAH组大鼠(P均<0.05)。BMSCs组大鼠脑组织内铁沉积情况较SAH组明显减轻,铁含量、MDA含量明显减少,GSH含量和GPX4活性显著升高(P均<0.05)。与SAH组相比,BMSCs组大鼠脑组织中Nrf2、 GPX4表达水平显著增多(P均<0.05);与BMSCs组相比,ML385组大鼠脑组织中Nrf2、GPX4表达水平显著减少(P均<0.05)。结论 BMSCs可能通过Nrf2/GPX4信号通路减轻SAH后受损脑组织铁死亡,从而修复其受损的神经功能。

Objective To explore whether bone mesenchymal stem cells(BMSCs)can reduce ferroptosis in brain tissue of subarachnoid hemorrhage(SAH)rats by regulating Nrf2/GPX4 pathway.Methods 25 out of 100 female SD rats were randomly selected as sham surgery group(Sham group)(without puncturing the branches of the anterior and middle cerebral arteries,only exiting when the puncture line was obstructed).The remaining 75 rats replicated the SAH rat model through vascular puncture,and all 75 rats were successfully modeled.They were randomly divided into subarachnoid hemorrhage group(SAH group),subarachnoid hemorrhage+BMSCs transplantation group(BMSCs group),and subarachnoid hemorrhage+BMSCs transplantation+Nrf2 specific inhibitor ML385 intervention group(ML385 group),with 25 rats in each group.After 3 days of modeling,the neurological score and brain tissue water content of the rats were measured,Prussian blue staining was used to detect ferroptosis in nerve cells.Iron content,malondi-aldehyde(MDA)content,glutathione(GSH)content and glutathione peroxidase 4(GPX4)activity in cerebral cortex were determined by spectrophotometry.The expression levels of Nrf2 and GPX4 were de-tected by Western blot.Results The neurological function score in the BMSCs group was significantly higher than that in SAH group,and the brain water content was significantly lower than that in SAH group(both P<0.05).The iron deposition in brain tissue of BMSCs group was significantly reduced com-pared with SAH group,the iron content and MDA content in brain tissue were significantly decreased,and the GSH content and GPX4 activity were significantly increased(P<0.05).Compared with SAH group,the expression levels of Nrf2 and GPX4 in brain tissue of BMSCs group were significantly increased(P<0.05).Compared with BMSCs group,the expression levels of Nrf2 and GPX4 in brain tissue of ML385 group were significantly decreased(P<0.05).Conclusion BMSCs may reduce ferroptosis in damaged brain tissue after SAH through Nrf2/GPX4 signaling pathway,so as to repair the damaged nerve function.