Daphnetin Attenuates Inflammatory and Apoptotic Responses Induced by High Glucose/High Free Fatty Acids and Lipopolysaccharide in an HK-2 Cell-Neutrophil Co-Culture System.

OBJECTIVES: The present study aimed to investigate the protective effect of daphnetin (dap) against high glucose and high free fatty acids (HGHF) and lipopolysaccharide(LPS)-induced injury in the aforementioned co-culture system under normal control conditions (normal glucose and no exogenous FFA supplementation), and to explore its potential molecular mechanisms. METHODS: Neutrophils were isolated from the peripheral blood of healthy individuals. Subsequently, HK-2 cells and a neutrophil suspension were co-cultured at 1x105 cells/mL to establish the experimental model. The non-cytotoxic concentration of dap for subsequent experiments was determined by Cell Counting Kit-8 (CCK-8) assay. Heparin-binding protein (HBP) concentration was measured using dry quantitative immunofluorescence assay. Inflammatory cytokines were quantified via cytometric bead array, and cell apoptotic rate was assessed by Annexin V-FITC-based flow cytometry. RESULTS: Both HGHF treatment and LPS exposure significantly elevated the levels of the inflammatory mediator HBP, cytokines (TNF-α, IFN-γ, IL-1β, IL-2, IL-6 and IL-10) and cellular apoptosis, whereas dap administration markedly attenuated these inflammatory and apoptotic alterations induced by HGHF or LPS (all P<0.05). CONCLUSION: In this in vitro model, dap attenuated HGHF- and LPS-induced inflammatory and apoptotic responses. Further studies are needed to assess its relevance to diabetic kidney disease.