The expression of CXCL9,CXCL10,and CXCL11 in renal tubular epithelial cells induced by IFN-γ and TNF-α

This study aimed to investigate the effects of TNF-α and TNF-α combined with IFN-γ on the release of CXCL9,CXCL10,and CXCL11 in renal tubular epithelial cells(HK-2).After stimulation of various time of TNF-α and TNF-α combined with IFN-γ,the HK-2 cells were analyzed by Real-time PCR to detect the expression of CXCL9,CXCL10,and CXCL11 mRNA,and the supernatants were analyzed by ELISA to quantify the release of CXCL9,CXCL10,and CXCL11 from cells.Furthermore,the supernatant was used to activate lymphocytes,which were then analyzed by flow cytometry to evaluate the CXCR3 expression,and determined by chemotaxis assay to assess chemotactic activity of the culture supernatants of HK-2 cells.The mRNA level of CXCL9,CXCL10 and CXCL11 of HK-2 cells were significantly increased 12 h after TNF-α stimulation,and reached maximal response at 48 h,24 h and 24 h,respectively,while the protein level of CXCL9,CXCL10,and CXCL11 reached the peak at 48 h,48 h and 72 h,respectively.Compared with IFN-γ alone or TNF-α alone,the combination treatment could significantly enhance the mRNA expression and protein secretion of CXCL9,CXCL10 and CXCL11(P 0.05).The supernatant-activated lymphocytes expressed more CXCR3,as compared with freshly isolated lymphocytes.The activated lymphocytes can be recruited by the culture supernatants of HK-2 cells treated with TNF-α,IFN-γ,and combination of TNF-α and IFN-γ,but pretreatment of CXCR3 antibody may inhibited this chemotactic effect.These results indicated that TNF-α and IFN-γ can significantly up-regulate the mRNA expression and protein excretion of chemokines such as CXCL9,CXCL10 and CXCL11,in which IFN-γ exerted synergic effect.