In vitro anti-inflammatory activity and molecular docking of Peperomia pellucida (L.) Kunth extract via the NF-κB and PPAR-γ signalling in human retinal pigment epithelial cells

This study aims to elucidate the anti-inflammatory mechanism of Peperomia pellucida (L.) Kunth in human retinal pigment epithelial cell line (ARPE-19) as stimulated by high glucose (34 mM and 68 mM), and advanced glycation end product (AGE) under different glucose (17 mM, 34 mM and 68 mM) environments via the nuclear factor kappa B (NF-κB) and peroxisome proliferator activated receptor gamma (PPAR-γ) signalling pathways. The cytotoxicity of P. pellucida in ARPE-19 cells was evaluated with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay. The genes and proteins expression of nine pro-inflammatory, angiogenic and antioxidant markers, including glutathione peroxidase (GPx), interleukin 8, matrix metalloproteinase 2, monocyte chemoattractant protein 1, NF-κB, PPAR-γ, receptor for AGE (RAGE), soluble RAGE (sRAGE), and vascular endothelial growth factor in P. pellucida-treated ARPE-19 cells were compared to non-treated control via real-time polymerase chain reaction and western blot. Both P. pellucida methanolic extract (1.5 mg/mL and 3 mg/mL) and ethyl acetate fraction (4 mg/mL) were non-toxic to ARPE-19 cells and demonstrated cytoprotective effect against the high glucose (34 mM) and AGE (17 mM glucose)-induced cellular stress. High glucose and AGE activated the pro-inflammatory signalling in ARPE-19 cells, as evidenced by the increased NF-κB p65 phosphorylation, up-regulation of pro-inflammatory and angiogenic mediators (p<0.05) but reduced GPx, PPAR-γ and sRAGE protein expression. Both P. pellucida methanolic extract (3 mg/mL) and ethyl acetate fraction (4 mg/mL) suppressed (p<0.05) the pro-inflammatory and angiogenic markers expression under high glucose and AGE environment. The main phytochemicals identified in P. pellucida were dillapiole, 2,4,5-trimethoxystyrene, 9-octadecenoic acid, and pheophorbide A-methyl ester which displayed relatively strong binding affinity towards NF-κB p65 and PPAR-γ proteins in molecular docking analysis. This study has demonstrated that P. pellucida is a potential alternative anti-inflammatory source for managing diabetic retinopathy via NF-κB and PPAR-γ signalling.