Protective Effects Of 4-octyl Itaconate Against Inflammatory Response In Angiotensin Ⅱ-induced Oxidative Stress In Human Primary Retinal Pigment Epithelium

PURPOSE: To investigate the role of 4-octyl itaconate(OI),a cell-permeable derivative of itaconate,in the amelioration of angiotensin Ⅱ(Ang Ⅱ)-induced inflammatory response and oxidative stress in human primary retinal pigment epithelium(hRPE)cells as well as its underlying mechanisms.METHEDS: The RPE-specific antibody,anti-human RPE65 was used to confirm the hRPE.Cell counting kit-8(CCK-8)was used to detect the cell viability of hRPE.The gene expression was detected by real-time PCR,and protein levels were determined by Western blotting or ELISA.The changes of reactive oxygen species(ROS),malondialdehyde(MDA)and superoxide dismutase(SOD)were tested by detection kits.The differential expression profiles of long non-coding RNAs(lnc RNAs)and m RNAs were tested using microarray analysis.Gene ontology(GO)and Kyoto Encyclopaedia of Genes and Genomes(KEGG)were performed to identify the related biological modules and pathways of the differentially expressed genes.Coding-non-coding gene co-expression networks(CNC network)were built to identify the correlations between lnc RNAs and m RNAs.RESULTS: The cultured hRPE cells were identified as RPE cells.CCK-8showed that hRPE cells were pre-incubated(or not)with 200 μM OI for 30 min and then stimulated(or not)with 20 μM Ang Ⅱ for 24 h.OI inhibited the changes of pro-inflammatory cytokines(MCP-1,IL-8 and IL-6),ROS,MDA and SOD via activation of Nrf2 signaling in Ang Ⅱ-treated hRPE cells.A total of 645 altered expressed lnc RNAs and 455 m RNAs were identified.Ten lnc RNAs were analyzed using the CNC network and KEGG analysis,revealed that many differentially expressed lnc RNAs were enriched in the immune response-related pathways,such as TNF,IL-17 and NF-κB signaling pathways.CONCLUSIONS: Our study revealed OI inhibits Ang Ⅱ-induced inflammatory response and oxidative stress via activating the Nrf2 signaling in hRPE cells.We also provided a novel perspective on the role of lnc RNAs in these protective effects of OI.