Song JiaLi, Qiao HuiYing, Dong ShunLi, Tang JingMei, Li Bai, Zhou Xi, Lv Shan, Lv Rong
Department of Geriatrics, Suzhou Ninth People's Hospital, Suzhou, Jiangsu, China.
Department of Pharmacology, College of Pharmaceutical Sciences, Soochow University, Suzhou, Jiangsu, China.
Front Pharmacol. 2025 Jul 8;16:1566724. doi: 10.3389/fphar.2025.1566724. eCollection 2025.
This study aims to assess LCD's neuroprotective pharmacological effects against SBI post-ICH and identify its ferroptosis-inhibiting targets.
Animal models of ICH and cellular models of SBI were established. Subsequently, gradient concentrations of LCD were administered at both the animal and cellular/molecular levels. The extent of ICH injury was evaluated using a range of methods, including CCK8 assay, Flow Cytometry, quantification of CAT and MDA, CI staining, Western blot, and HE staining. The SWISS TARGET prediction tool and molecular docking were utilized to confirm LCD's target pathway and its binding site on COX2. Quantification of ferroptosis-executing proteins, BODIPY ROS staining, quantification of PGE2, MDA, and CAT were observed to assess the pharmacological effects, trends in ferroptosis influence, and to elucidate the underlying pathway mechanism.
Pretreatment with LCD can improve the state of SBI before the induction of an ICH model. Compound target prediction analysis revealed 102 differentially expressed genes ( < 0.05) associated with the drug target of LCD, with COX2 exhibiting the most significant expression. Furthermore, we found that LCD intervention suppressed COX2 expression, and pretreatment with COX2 overexpression in the ICH model group negated the pharmacological effects, of LCD on neuronal cell ferroptosis and SBI. It is proposed that by targeting COX2 through early LCD administration in ICH, ferroptosis in nerve cells can be reduced and SBI outcomes can be improved. To further elucidate the mechanism of targeting COX2, we found that PGE2, a downstream metabolite of COX2, is also regulated by LCD. By screening its impacts on the EP receptor family (EP1, EP2, EP3, EP4), it was found that COX2 is specifically targeted and suppressed by LCD pretreatment prior to ICH modeling, which further inhibits the PGE2/EP1 pathway, thereby reducing ferroptosis-specific lipid peroxidation.
LCD pretreatment reduces ferroptosis in neurons and alleviates SBI after ICH by blocking the COX2/PGE2/EP1 pathway. Early LCD use may improve SBI, highlighting its potential as a pharmacological option for ICH outcomes.
本研究旨在评估连钱草对脑出血后继发性脑损伤的神经保护药理作用，并确定其抑制铁死亡的靶点。
建立脑出血动物模型和继发性脑损伤细胞模型。随后，在动物及细胞/分子水平给予不同梯度浓度的连钱草。采用多种方法评估脑出血损伤程度，包括CCK8法、流式细胞术、CAT和MDA定量、CI染色、蛋白质印迹法及苏木精-伊红染色。利用SWISS TARGET预测工具和分子对接技术确定连钱草的作用靶点途径及其在COX2上的结合位点。通过观察铁死亡执行蛋白的定量、BODIPY活性氧染色、PGE2、MDA和CAT的定量，评估其药理作用、铁死亡影响趋势，并阐明潜在的途径机制。
连钱草预处理可改善脑出血模型诱导前的继发性脑损伤状态。复合靶点预测分析显示，与连钱草药物靶点相关的差异表达基因有102个（<0.05），其中COX2表达最为显著。此外，我们发现连钱草干预可抑制COX2表达，在脑出血模型组中过表达COX2进行预处理可消除连钱草对神经元细胞铁死亡和继发性脑损伤的药理作用。研究表明，在脑出血中通过早期给予连钱草靶向COX2，可减少神经细胞铁死亡并改善继发性脑损伤结局。为进一步阐明靶向COX2的机制，我们发现COX2的下游代谢产物PGE2也受连钱草调节。通过筛选其对EP受体家族（EP1、EP2、EP3、EP4）的影响，发现连钱草预处理在脑出血建模前可特异性靶向并抑制COX2，进而抑制PGE2/EP1途径，从而减少铁死亡特异性脂质过氧化。
连钱草预处理通过阻断COX2/PGE2/EP1途径减少神经元铁死亡并减轻脑出血后的继发性脑损伤。早期使用连钱草可能改善继发性脑损伤，突出了其作为改善脑出血结局的药理选择的潜力。
