Intestine-derived exosomes regulates inflammatory response of HK-2 cells through miR-146a-regulated NF-κB pathway
Objective To investigate the role of intestine-derived exosomes in regulating the inflammatory response of HK-2 cells through the NF-κB pathway regulated by miR-146a.Methods Caco-2 cells were divided into control group,model group(constructed intestinal epithelial cell inflammation model by stimulating with 10 ng·mL-1 LPS),NC mimic group,and miR-146a mimic group.The exosomes(30 μg·mL-1)from each group were extracted and incubated with HK-2 cells,respectively,and named as control-Exos,LPS-Exos,NC-Exos,and miR-146a-Exos group.Transcriptome sequencing was used to detect differential genes between the control group and the model group;real-time fluorescence quantitative PCR(qRT-PCR)was used to detect miR-146a expression;ELISA was used to detect the levels of interleukin(IL)-1β,IL-6,and tumor necrosis factor-α(TNF-α)in the cell culture supernatant;Western blotting was used to detect the expressions of IL-1β,IL-6,TNF-α,NF-κB,p-NF-κB,and Toll-like receptor 4(TLR4)inflammatory signaling pathway proteins in the cells.Results Compared with the control group,the expressions of inflammatory factors IL-1β,IL-6,and TNF-α and TLR4,NF-κB,and p-NF-κB proteins in the model group were significantly increased(P＜0.05,0.01),the levels of IL-iβ,IL-6,and TNF-α in the cell culture supernatant were significantly increased(P＜0.05),and the expressions of miR-146a in the cells and exosomes were significantly increased(P＜0.05,0.01);the expressions of miR-146a in the cells and exosomes of the miR-146a mimic group were significantly increased(P＜0.01).Compared with the control-Exos group,the levels of inflammatory factors IL-1β,IL-6,and TNF-α in the culture supernatant of the LPS-Exos and miR-146a mimic groups were significantly increased(P＜0.05).Conclusion Intestine-derived exosomes regulate the inflammatory response of HK-2 cells through miR-146a-regulated NF-κb pathway.
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