Huang Yu, Xia Xin, Li DanYang, Lyu Yi, Zhou Pai, Cui Hongda, Zhou YaSha, Yang YiJing, Peng QingHua
Hunan University of Chinese Medicine, Changsha, Hunan, China; Hunan University of Chinese Medicine, First College of Clinical Chinese Medicine, Changsha, Hunan, China.
Fujian University of Traditional Chinese Medicine, College of Traditional Chinese Medicine, Fuzhou, Fujian, China.
J Ethnopharmacol. 2025 Sep 18;355(Pt A):120576. doi: 10.1016/j.jep.2025.120576.
Glaucomatous optic neuropathy (GON), characterized by the progressive degeneration of retinal ganglion cells (RGCs), is the primary pathological basis of irreversible visual impairment in glaucoma. Qingguang'An II decoction (QGAII), a traditional Chinese medicine formula with decades of clinical application, has been shown to promote optic nerve repair and preserve the visual field in patients with GON. Nevertheless, the precise mechanisms underlying the neuroprotective effects of QGAII remain incompletely understood.
This study aimed to investigate the specific molecular mechanisms by which QGAII mitigates RGC pyroptosis through the targeted modulation of the TRPV4/NF-κB signaling pathway.
The neuroprotective effects of QGAII were evaluated both in vivo using a DBA/2J mouse model of chronic ocular hypertension and in vitro using a novel cell model of glaucomatous injury. In vivo, the effect of QGAII intervention was assessed by hematoxylin and eosin (H&E) staining and transmission electron microscopy (TEM). The mechanism of action was further investigated by immunofluorescence, flow cytometry, Western blot (WB), qPCR, and ELISA. In vitro, a novel glaucomatous cell injury model was established by combining continuous hydrostatic pressure (CHP) and oxygen-glucose deprivation (OGD). Liposome transfection was employed to explore the roles of TRPV4, NF-κB, and GSDMD. Subsequently, QGAII was administered to this injury model, and its effects were analyzed using a cell counting kit-8 (CCK-8), flow cytometry, scanning electron microscopy (SEM), immunofluorescence, WB, qPCR, and ELISA to clarify its role in attenuating pyroptosis via the TRPV4/NF-κB signaling pathway.
In vivo, H&E staining and TEM showed that QGAII exerted a protective effect on RGCs in the retina of DBA/2J mice, and this effect was linked to a reduction in RGC pyroptosis. Further analysis showed that QGAII attenuated RGC pyroptosis in DBA/2J mice, potentially through the TRPV4/NF-κB signaling pathway. In vitro, we first established that the cell injury model, induced by CHP and OGD, exhibited a GSDMD-dependent pyroptotic phenotype. This process was confirmed to be regulated by the TRPV4/NF-κB signaling pathway. Subsequent QGAII intervention reduced the incidence of RGC pyroptosis. This protective effect was mediated by the inhibition of the TRPV4/NF-κB signaling pathway, as confirmed by immunofluorescence, WB, qPCR, and ELISA.
QGAII suppresses RGC pyroptosis through the targeted modulation of the TRPV4/NF-κB signaling pathway, providing effective optic neuroprotection for the therapeutic management of GON.
青光眼性视神经病变（GON）以视网膜神经节细胞（RGCs）的进行性变性为特征，是青光眼不可逆视力损害的主要病理基础。青光安II号方（QGAII）是一种有着数十年临床应用历史的中药方剂，已被证明可促进视神经修复并保护GON患者的视野。然而，QGAII神经保护作用的确切机制仍未完全明确。
本研究旨在探讨QGAII通过靶向调节TRPV4/NF-κB信号通路减轻RGC焦亡的具体分子机制。
在体内使用慢性高眼压DBA/2J小鼠模型，在体外使用新型青光眼损伤细胞模型，评估QGAII的神经保护作用。在体内，通过苏木精-伊红（H&E）染色和透射电子显微镜（TEM）评估QGAII干预的效果。通过免疫荧光、流式细胞术、蛋白质免疫印迹（WB）、定量聚合酶链反应（qPCR）和酶联免疫吸附测定（ELISA）进一步研究其作用机制。在体外，通过联合持续静水压力（CHP）和氧糖剥夺（OGD）建立新型青光眼细胞损伤模型。采用脂质体转染法探讨瞬时受体电位香草酸亚型4（TRPV4）、核因子κB（NF-κB）和Gasdermin D（GSDMD）的作用。随后，将QGAII应用于该损伤模型，并使用细胞计数试剂盒-8（CCK-8）法、流式细胞术、扫描电子显微镜（SEM）、免疫荧光、WB、qPCR和ELISA分析其效果，以阐明其通过TRPV4/NF-κB信号通路减轻焦亡的作用。
在体内，H&E染色和TEM显示QGAII对DBA/2J小鼠视网膜中的RGCs具有保护作用，且这种作用与RGC焦亡的减少有关。进一步分析表明，QGAII可能通过TRPV4/NF-κB信号通路减轻DBA/2J小鼠的RGC焦亡。在体外，我们首先确定由CHP和OGD诱导的细胞损伤模型表现出依赖GSDMD的焦亡表型。该过程被证实受TRPV4/NF-κB信号通路调控。随后的QGAII干预降低了RGC焦亡的发生率。免疫荧光、WB、qPCR和ELISA证实，这种保护作用是通过抑制TRPV4/NF-κB信号通路介导的。
QGAII通过靶向调节TRPV4/NF-κB信号通路抑制RGC焦亡，为GON的治疗管理提供了有效的视神经保护作用。
