Abstract
OBJECTIVES: Glaucoma is the leading cause of irreversible blindness worldwide. Reducing intraocular pressure is currently one of the most effective treatment strategies; however, it cannot completely prevent retinal ganglion cells (RGCs) death and the resulting vision loss. Traditional Chinese medicine has been widely investigated in glaucoma treatment. This study aims to determine whether geniposide can effectively inhibit retinal cell death and to explore its potential role in glaucoma therapy. METHODS: Cell experiments: Retinal R28 cells were divided into a control (CTL) group, an oxygen-glucose deprivation/reoxygenation (OGD/R) group, an OGD/R+geniposide (OGD/R+ Gen) group, and an OGD/R+Gen+heat shock protein family A member 1A (Hspa1a) small interfering RNA (OGD/R+Gen+H-KD) group. Proteomics analysis was first performed to identify key molecules altered after geniposide intervention. Quantitative real-time PCR (qPCR) and Western blotting were used to detect molecular changes after geniposide and H-KD interventions. Cell viability and death were assessed using cell counting kit-8 (CCK-8) and lactate dehydrogenase (LDH) assays. Flow cytometry, reactive oxygen species (ROS) detection, and terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) staining were used to evaluate apoptosis. Animal experiments: SD rats were divided into a sham control (CTL) group, an acute high intraocular pressure (aHIOP) group, an aHIOP+Gen group, and an aHIOP+Gen+Hspa1a inhibitor apoptozole (aHIOP+Gen+Apo) group. Hematoxylin-eosin (HE) staining, TUNEL staining, and flash electroretinogram (fERG) were used to evaluate retinal structure, apoptosis, and functional changes. RESULTS: 1) Proteomics combined with qPCR and Western blotting showed that Hspa1a expression was significantly increased after geniposide treatment. 2) Compared with the OGD/R group, the OGD/R+Gen group showed increased cell viability (P<0.05), fewer dead cells (P<0.05), decreased ROS levels (P<0.001), and reduced apoptosis (P<0.05), indicating that geniposide alleviated OGD/R-induced retinal R28 cell injury and death. 3) Compared with the OGD/R+Gen group, the OGD/R+Gen+H-KD group showed decreased phosphorylated protein kinase B (Akt, p-Akt) expression (P<0.01), reduced cell viability (P<0.05), increased cell death (P<0.05), elevated ROS levels (P<0.01), and increased apoptosis (P<0.05), suggesting that Hspa1a regulates the Akt pathway and mediates the protective effect of geniposide on retinal R28 cells. 4) Compared with the CTL group, the aHIOP group showed reduced retinal thickness (P<0.01), increased apoptosis (P<0.001), and decreased fERG b-wave amplitude (P<0.001). Compared with the aHIOP group, the aHIOP+Gen group showed increased retinal thickness (P<0.05), reduced apoptosis (P<0.05), and increased fERG b-wave amplitude (P<0.05). Compared with the aHIOP+Gen group, the aHIOP+Gen+Apo group showed decreased retinal thickness (P<0.05), increased apoptosis (P<0.05), and reduced fERG b-wave amplitude (P<0.05), indicating that geniposide alleviates aHIOP-induced retinal injury through the Hspa1a pathway. CONCLUSIONS: Geniposide inhibits OGD/R- and aHIOP-induced retinal cell apoptosis and tissue injury through the Hspa1a-Akt pathway, providing a potential therapeutic target for glaucoma treatment.