Retinal endothelial cells (RECs) are the primary target cells for diabetes-induced vascular damage. The P2X7/NLRP3 pathway plays an essential role in amplifying inflammation via an ATP feedback loop, promoting the inflammatory response, pyroptosis, and apoptosis of RECs in the early stages of diabetic retinopathy induced by hyperglycemia and inflammation. 3TC, a type of nucleoside reverse transcriptase inhibitor, is effective against inflammation, as it can targeting formation of the P2X7 large pore formation. Hence, our aim was to evaluated the anti-inflammatory effects and potential mechanisms of action of 3TC in vitro in retinal microvascular endothelial cells treated with high-glucose (HG) and lipopolysaccharide (LPS), as well as in vivo in the retinas of C57BL/6J male mice with streptozotocin-induced diabetes. The expression of inflammasome-related proteins P2X7 and NLRP3, and apoptosis in the retinas of 3TC-treated diabetic mice were compared to those of untreated diabetic mice. Furthermore, the anti-inflammatory, anti-apoptotic, and anti-pyroptotic effects of 3TC were evaluated in vitro in cultured mice retinal endothelial cells. Co-application of HG and LPS significantly increased the secretion of IL-6, IL-1β, and TNF-α, and ATP levels, whereas 3TC decreased cell inflammation, apoptosis, and pyroptosis. Inhibition of P2X7R and NLRP3 inflammasome activation decreased NLRP3 inflammasome-mediated injury. 3TC prevented cytokine and ATP release following co-application of HG and LPS/BzATP. Our findings provide new insights regarding the mechanisms of action of 3TC in diabetic environment-induced retinal injury, including apoptosis and pyroptosis.

To explore the relevant RNA-binding proteins (RBPs) and alternative splicing events (ASEs) in diabetic retinopathy (DR). We devised a comprehensive work to integrate analyses of the differentially expressed genes, including differential RBPs, and variable splicing characteristics related to DR in human retinal endothelial cells induced by low glucose and high glucose in dataset GSE117238. A total of 2320 differentially expressed genes (DEGs) were identified, including 1228 upregulated genes and 1092 downregulated genes. Further analysis screened out 232 RBP genes, and 42 AS genes overlapped DEGs. We selected high expression and consistency six RBP genes (FUS, HNRNPA2B1, CANX, EIF1, CALR, and POLR2A) for coexpression analysis. Through analysis, we found eight RASGs (MDM2, GOLGA2P7, NFE2L1, KDM4A, FAM111A, CIRBP, IDH1, and MCM7) that could be regulated by RBP. The coexpression network was conducted to further elucidate the regulatory and interaction relationship between RBPs and AS. Apoptotic progress, protein phosphorylation, and NF-kappaB cascade revealed by the functional enrichment analysis of RASGs regulated by RBPs were closely related to diabetic retinopathy. Furthermore, the expression of differentially expressed RBPs was validated by qRT-PCR in mouse retinal microvascular endothelial cells and retinas from the streptozotocin mouse model. The results showed that Fus, Hnrnpa2b1, Canx, Calr, and Polr2a were remarkedly difference in high-glucose-treated retinal microvascular endothelial cells and Fus, Hnrnpa2b1, Canx, and Calr were remarkedly difference in retinas from streptozotocin-induced diabetic mice compared to control. The regulatory network between identified RBPs and RASGs suggests the presence of several signaling pathways possibly involved in the pathogenesis of DR. The verified RBPs should be further addressed by future studies investigating associations between RBPs and the downstream of AS, as they could serve as potential biomarkers and targets for DR.