Purpose. Retinal ischemia–reperfusion injury (RIRI) is the basis of the pathology that leads to many retinal diseases and induces necroptosis and apoptosis. Tumor necrosis factor-α (TNF-α) is critically involved in necroptosis and apoptosis. Delta-opioid receptor (DOR) activation inhibits TNF-α release in our previous studies, it might prevent necroptosis and apoptosis by inhibiting the release of TNF-α. However, the role of TNF-α and DOR in necroptosis and apoptosis of retinal pigment epithelial (RPE) cells remains largely unknown. Here, we explored the mechanisms of TNF-α and DOR in necroptosis and apoptosis using an oxygen-glucose deprivation/reoxygenation (OGD/R) model of adult retinal pigment epithelial cell line-19 (ARPE19) cells. Materials and Methods. ARPE19 cells were exposed to OGD/R conditions to mimic RIRI in vitro. Cell viability was quantified using the Cell Counting Kit-8 (CCK-8) assay. Morphological changes were observed by inverted microscopy. TNF-α protein levels in cell lysates were measured by enzyme-linked immunosorbent assay (ELISA). The DOR agonist TAN-67 and antagonist naltrindole (NTI) were used to pretreat cells for 1 or 2 hours before OGD24/R36 administration. Calcein acetoxymethylester/propidium iodide (Calcein-AM/PI) and Terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) staining were used to detect necroptotic and apoptotic ARPE19 cells, respectively. The protein expression of DOR, p-RIP1 (RIP1), p-RIP3 (RIP3), p-MLKL (MLKL), and cleaved Caspase3 (Caspase3) was measured by western blotting. Results. OGD severely damaged ARPE19 cells. Prolonged reoxygenation significantly increased TNF-α level and decreased DOR expression in ARPE19 cells. Pretreatment with the DOR agonist TAN-67 (10 µM) significantly improved ARPE19 cell viability after OGD24/R36 by reducing the number of necroptotic and apoptotic cells. Furthermore, DOR activation significantly inhibited TNF-α release and suppressed the expression of proteins related to necroptosis and apoptosis, including p-RIP1, p-RIP3, p-MLKL, and cleaved Caspase3, after OGD24/R36. This effect was reversed by the DOR antagonist NTI. Conclusion. These results strongly suggest that DOR activation inhibits necroptosis and apoptosis by decreasing TNF-α release, leading to the prevention of OGD/R-induced injury in ARPE19 cells. This study provides an innovative idea for clinical treatment strategies for retinal damage and vision loss due to RIRI.

Objectives: Retinal ischemia-reperfusion injury (RIRI) is the common pathological basis of many ophthalmic diseases in the later stages, and inflammation is the primary damage mechanism of RIRI. Our study aimed to assess whether electroacupuncture (EA) has a protective effect against RIRI and to elucidate its related mechanisms.Methods: A high-intraocular pressure (HIOP) model was used to simulate RIRI in Wistar rats. EA was applied to the EA1 group [Jingming (BL1) + Shuigou (GV26)] and the EA2 group [Jingming (BL1) + Hegu (LI4)] respectively for 30 min starting immediately after the onset of reperfusion and repeated (30 min/time) at 12 h and then every 24 h until days 7 after reperfusion. The pathological changes in the retina were observed by H and E staining after HIOP. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining was utilized to observe retinal cell apoptosis. The mRNA expression of IL1-β, TNF-α, IL-4, IL-10, δ-opioid receptor (DOR), brain-derived neurotrophic factor (BDNF), and tropomyosin-related kinase B (TrkB) in the retina was measured by quantitative real-time PCR.Results: HIOP caused structural disorders of the retina, decreased RGCs, and increased retinal cell apoptosis. At 1 and 3 days of RIRI, retinal apoptotic cells in the EA group were significantly reduced, while there was no distinct difference in the EA group compared with the HIOP group at 7 days of RIRI. Compared with that in the HIOP group, the expression of anti-inflammatory factors, DOR and TrkB was increased, and the expression of pro-inflammatory factors was decreased in the EA group. In contrast, HIOP had no appreciable effect on BDNF expression.Conclusion: EA at Jingming (BL1) and Shuigou (GV26) or at Jingming (BL1) and Hegu (LI4) may inhibit RIRI induced inflammation through activating the DOR-BDNF/TrkB pathway to protect the retina, especially the pair of Jingming (BL1) and Shuigou (GV26) has better inhibitory effects on inflammation.