Biological characteristics and effect of TNFª²¦Á binding peptide and TNFR blocking peptide in vitro

    YIN Bingª²jiao, YU Mingª²xia, WANG Jing, JIANG Xiaoª²dan, LI Zhuoª²ya*

    Department of Immunology, Tongji Medical College of Huazhong University of Science and Technology, Wuhan 430030, China

    [Abstract] AIM: To investigate the biological characteristics and effect of TNFª²¦Á binding peptide (TBP) and TNFR blocking peptide (TRBP) in vitro. METHODS: The binding specificity of TBP or TRBP was tested by competition experiment using GFPª²TNF fusion protein and detected by FCM and fluorescent microscope. The interaction between TBP and TRBP was determined by nonª²denatured PAGE and the inhibitory effect of TBP and TRBP on TNFª²¦Á cytotoxicity against U937 was carried out by MTT colorimetry. The effects of TBP and TRBP on the functions of human monocytes activated by TNFª²¦Á and IFNª²¦Ã in vitro were detected by nitrotetrazoium blue chlorid (NBT) reduction assay for evaluating respiratory burst and by RTª²PCR for evaluating ILª²1¦Â and ILª²8 mRNA transcription. RESULTS: It was showed that TBP and TRBP was able to block the GFPª²TNF binding to the TNFR on the surface of cells and no binding interaction took place between TBP and TRBP. Both TBP and TRBP were able to inhibit the TNFª²¦Á cytotoxicity against U937 and this inhibitiory effect was doseª²dependent and the combination of TBP and TRBP (pep.38 +X4) was able to inhibit the TNFª²¦Á cytotoxicity more efficiently than the individual peptide at all tested concentrations. The combination of TBP and TRBP was able to obviously inhibit both respiratory burst of monocytes induced by TNFª²¦Á and IFNª²¦Ã and transcription level of ILª²1¦Â and ILª²8 mRNA induced by TNFª²¦Á. CONCLUSION: TBP or TRBP can bind with their ligands specifically and don¡¯t interact with each other. They can also block the biological activity of the TNFª²¦Á in vitro, and the combination of TBP and TRBP is able to inhibit the biological activity of the TNFª²¦Á more efficiently. This research will provide an experimental basis for the clinical treatment of the inflammatory disease. ......