Gao Shang, Qi Lijie, Xu Zhaoyao, Li Xin, Han Bingbing, Zhang Zhaoqi, Zhang Jie, Su Ming, Zhu Weiming, Cao Linya, Wu Tong, Li Nianhu, Liu Wei
The First Clinical Medical College of Shandong University of Traditional Chinese Medicine.
Affiliated Hospital of Shandong University of Traditional Chinese Medicine.
J Ethnopharmacol. 2025 Sep 5:120543. doi: 10.1016/j.jep.2025.120543.
Fujian Tablet (FJT), a traditional Chinese herbal compound formulation developed under the theoretical framework of "nourishing the liver and kidney, replenishing essence and marrow" , has been clinically applied for over two decades to treat post-stroke neurological deficits. Preliminary studies demonstrated its efficacy in improving motor function and promoting cervical spinal cord neuroaxonal growth in a middle cerebral artery occlusion (MCAO) rat model. Building upon these findings, this study integrates metabolomic evidence of Foxo3a-GPX4 axis activation to systematically elucidate Fujian Tablet's neurorestorative mechanisms through three interconnected pathways: regulation of ferroptosis, promotion of oligodendrocyte proliferation, and remyelination. By bridging traditional TCM therapeutic principles with contemporary molecular neuroscience, this investigation aims to provide mechanistic insights for optimizing Fujian Tablet's clinical application and advancing its global recognition in neurorehabilitation.
This study aims to verify the molecular mechanism by which FJT activates Forkhead box O3a (Foxo3a) to promote glutathione peroxidase 4 (GPX4) expression, thereby regulating oligodendrocyte survival and remyelination, based on the regulatory network of "traditional Chinese medicine compound - target - cellular function". This study provides experimental evidence for explaining the material basis and action targets of the compound's efficacy.
The middle cerebral artery embolization method was employed to establish a rat model of MCAO. Then, FJT was used for intervention via intragastric administration at doses of 0.72g/kg and 1.44g/kg, twice daily for 14 consecutive days.Behavioral observations of the rats were carried out using the Catwalk system and beam walking test (BWT). remyelination was assessed via luxol fast blue (LFB) staining and transmission electron microscopy. In addition, in vitro experiments with oligodendrocytes were conducted in combination. The expressions of myelin basic protein (MBP) was detected by immunofluorescence. The expressions of Ferritin, and GPX4 were detected by RT-qPCR and Western blot. The levels of ferroptosis - related metabolites were measured using flow cytometry and enzyme - linked immunosorbent assay (ELISA). The transcriptional regulatory effect of Foxo3a on GPX4 was examined by ChIP-qPCR.
Fujian Tablet dose-dependently improved the motor function of MCAO rats, and upregulated the expression of Foxo3a to enhance its binding activity to the GPX4 promoter, thereby increasing GPX4 expression. Knockdown of Foxo3a or treatment with the GPX4 inhibitor (RSL3) reversed the inhibitory effect of FJT on ferroptosis (decrease in lipid reactive oxygen species (ROS) and malondialdehyde (MDA), and increase in reduced glutathione (GSH)). This reversal led to a reduction in the expression of MBP and myelin oligodendrocyte glycoprotein (MOG), inhibited the level of remyelination (decrease in LFB optical density and increase in the electron microscopy G-ratio), and hindered the recovery of motor function. In vitro experiments confirmed that serum containing FJT inhibited ferroptosis through the Foxo3a-GPX4 axis, and promoted the proliferation of oligodendrocytes.
FJT exerts a neuroprotective effect by activating the Foxo3a - GPX4 axis to inhibit ferroptosis, promote the proliferation of oligodendrocytes and remyelination, and ultimately improve motor function. This study clarifies that Fujian Tablet exerts a neuroprotective effect through regulating the "ferroptosis - oligodendrocyte proliferation - remyelination" axis, providing a scientific basis for its clinical application in ethnic medicine.
福建片（FJT）是一种在“滋补肝肾、填精益髓”理论框架下研制的传统中药复方制剂，已临床应用二十余年，用于治疗中风后神经功能缺损。初步研究表明，在大脑中动脉闭塞（MCAO）大鼠模型中，它在改善运动功能和促进颈脊髓神经轴突生长方面具有疗效。基于这些发现，本研究整合了Foxo3a - GPX4轴激活的代谢组学证据，通过三条相互关联的途径系统地阐明福建片的神经修复机制：铁死亡调节、少突胶质细胞增殖促进和髓鞘再生。通过将传统中医治疗原则与当代分子神经科学相结合，本研究旨在为优化福建片的临床应用并提高其在神经康复领域的全球认可度提供机制性见解。
本研究旨在基于“中药复方 - 靶点 - 细胞功能”调控网络，验证福建片激活叉头框O3a（Foxo3a）以促进谷胱甘肽过氧化物酶4（GPX4）表达，从而调节少突胶质细胞存活和髓鞘再生的分子机制。本研究为解释该复方药效的物质基础和作用靶点提供实验依据。
采用大脑中动脉栓塞法建立MCAO大鼠模型。然后，以0.72g/kg和1.44g/kg的剂量通过灌胃给予福建片进行干预，每日两次，连续14天。使用Catwalk系统和平衡木行走试验（BWT）对大鼠进行行为观察。通过Luxol快速蓝（LFB）染色和透射电子显微镜评估髓鞘再生情况。此外，结合少突胶质细胞进行体外实验。通过免疫荧光检测髓鞘碱性蛋白（MBP）的表达。通过RT - qPCR和蛋白质免疫印迹法检测铁蛋白和GPX4的表达。使用流式细胞术和酶联免疫吸附测定（ELISA）测量铁死亡相关代谢物的水平。通过染色质免疫沉淀 - qPCR（ChIP - qPCR）检测Foxo3a对GPX4的转录调控作用。
福建片剂量依赖性地改善MCAO大鼠的运动功能，并上调Foxo3a的表达以增强其与GPX4启动子的结合活性，从而增加GPX4表达。敲低Foxo3a或用GPX4抑制剂（RSL3）处理可逆转福建片对铁死亡的抑制作用（脂质活性氧（ROS）和丙二醛（MDA）减少，还原型谷胱甘肽（GSH）增加）。这种逆转导致MBP和髓鞘少突胶质细胞糖蛋白（MOG）表达降低，抑制髓鞘再生水平（LFB光密度降低，电子显微镜下G比值增加），并阻碍运动功能恢复。体外实验证实，含福建片的血清通过Foxo3a - GPX4轴抑制铁死亡，并促进少突胶质细胞增殖。
福建片通过激活Foxo3a - GPX4轴发挥神经保护作用，抑制铁死亡，促进少突胶质细胞增殖和髓鞘再生，最终改善运动功能。本研究阐明福建片通过调节“铁死亡 - 少突胶质细胞增殖 - 髓鞘再生”轴发挥神经保护作用，为其在民族医学中的临床应用提供了科学依据。
