The Expression Of CXCL9/10/11 In Human Renal Tubular Epithelial Cells And The Modulatory Effect Of 15D-PGJ2

Objective:To study the effect and mechanism of tumor necrosis factor-alpha(TNF-a) combined with interferon-gamma (IFN-y) on the expression of CXCL9、CXCL10 and CXCL11 in human renal tubular epithelial cells (HK-2), and to investigate the effect and mechanism of peroxisome proliferator-activated receptor-y (PPAR-y) agonist 15-Deoxy-Δ12，14-prostaglandin J2 (15d-PGJ2) on CXCL9、CXCL10、CXCL11 expression in HK-2 cells, and provide some evidences for the new ways to treat kidney transplant rejection.Methods:HK-2 cells were cultured and divided into four groups:the control group, TNF-a combined with IFN-y treatment group,15d-PGJ2 treatment group and pyrrolidine dithiocarbamate (PDTC) treatment group. The expression of CXCL9, CXCL10 and CXCL11 mRNA were measured by real-time quantitative PCR and the levels of CXCL9, CXCL10 and CXCL11 in culture supernatant were tested by enzyme-linked immunosorbent assay (ELISA). The expression of p65, IκB-α and their phosphorylated protein p-p65, p-IκBα were measured by western blot.Results:1、The mRNA expression in HK-2 cells were up-regulated after TNF-a and IFN-y stimulation, the real-time PCR suggests the expression of CXCL9, CXCL10, CXCL11 mRNA was up-regulated compared with the control group (P<0.01), and ELISA indicates the releases of CXCL9, CXCL10, CXCL11 in supernatant were also up-regulated compared with the control group (P<0.05); 2、The CXCL9, CXCL10, CXCL11 were down-regulated by 85.44%、45.94%、45.03% at mRNA level and 60.87%、47.59%、53.42% at protein level when preteated with PDTC for 2h and then incubated with TNF-a combined with IFN-y, respectively (P<0.05); 3、The phosphorylation levels of p65 and IκBa were significantly up-regulated in TNF-a combined with IFN-y treated HK-2 cells for 5min (P<0.01); 4、The CXCL9, CXCL10 and CXCL11 were down-regulated by 93.87%,92.4% and 86.81% at mRNA level and 60.87%,47.59% and 53.42% at protein level when HK-2 were pretreated with 15d-PGJ2 for 0.5h and then incubated with TNF-a combined with IFN-y, respectively (P <0.01); 5、The phosphorylation levels of p65 and IκBa were down-regulated when HK-2 were pretreated with 15d-PGJ2 for 0.5h and then incubated with TNF-a combined with IFN-y (P<0.01).Conclusion:1、TNF-a combine with IFN-y could up-regulate the expression of CXCL9, CXCL10 and CXCL11 through NF-κB signal pathway; 2、15d-PGJ2 could inhibits the expression of CXCL9, CXCL10 and CXCL11 through NF-κB signal pathway. Given that CXCL9, CXCL10 and CXCL11 play important roles in kidney transplant rejection and together with our results in this study, these results suggest that the modulatory effect of 15d-PGJ2 on the expression of CXCL9, CXCL10 and CXCL11 in renal tubular epithelial cells may provide a novel strategy in kidney transplant rejection treatment.