Abstract: Objective To explore the effect of contrast agent-induced macrophage-derived exosomes on renal tubular epithelial cells and the intervention effect of glycyrrhizin. Methods THP-1 cells were stimulated with PBS, iohexol and iohexol+glycyrrhizin, respectively, and three kinds of exosomes (NC-Exo, CIN-Exo, CIN-GL-Exo) were obtained. Transmission electron microscopy was used to observe the morphology of exosomes. Western blot assay was used to detect the expression levels of CD63, TSG101 and calnexin. Fluorescent labeling method was used to detect the uptake of exosomes. HK-2 cells were randomly divided into 4 groups: Con group, NC-Exo group, CIN-Exo group and CIN-GL-Exo group. Western blot assay was used to detect the expression levels of HMGB1 in three exosomes, the expression levels of apoptosis-related proteins and HMGB1/TLR4/NF-κB signaling pathway related proteins in 4 groups of cells. Annexin Ⅴ-FITC/PI double staining method was used to detect the apoptosis rate of cells. Quantitative real-time PCR was used to detect the mRNA expression of inflammatory factors ［interleukin (IL)-1β, IL-6, tumor necrosis factor (TNF)-α］ in 4 groups of cells. Results The exosomes derived from THP-1 cells were round or round-like small vesicles, which expressed the marker protein CD63 and TSG101, but not the endoplasmic reticulum protein calnexin. DiR-labeled exosomes could be taken up by HK-2 cells. The expression level of HMGB1 was higher in CIN-Exo than that of NC-Exo and CIN-GL-Exo (P＜0.05). The apoptosis rate and the expression levels of Bax and Cleaved caspase-3 were higher in CIN-Exo group than those in Con group, NC-Exo group and CIN-GL-Exo group, while the expression level of Bcl-2 was lower than that in the other three groups (P＜0.05). The mRNA levels of IL-1β, IL-6 and TNF-α, and the expression levels of HMGB1, TLR4 and NF-κB were higher in CIN-Exo group than those in the other three groups (P＜0.05). Conclusion Contrast agents can induce apoptosis and inflammation of renal tubular epithelial cells through macrophage-derived exosomes, which may be related to the upregulation of exosome HMGB1 levels. Glycyrrhizin can inhibit this pathway and exert a protective effect.

Key words: macrophages, exosomes, HMGB1 protein, apoptosis, renal tubular epithelial cells, contrast induced nephropathy, glycyrrhizin