ABSTRACT
Objective To investigate the effects of mannan-binding lectin(MBL) on TNF-α production induced by peptidoglycan (PGN) and its mechanism in human THP-1/CD14 monocytes.Methods The THP-1/CD14 cells were stimulated for 24 h with PGN at the indicated ratios after pretreated with human natural MBL at concentrations ranging from 1 to 20 mg/L for 2 h.The content of TNF-α and IL-6 in culture supernatants were detected by ELISA,and the levels of TNF-α and IL-6 mRNA expressions in these cells were determined by RT-PCR.FACS was used to investigate the interaction of MBL with THP-1/CD14 cells and the impact of MBL on PGN binding to THP-1/CD14 cells.Western blot was used to detect PGN-induced NF-κB translocation in THP-1/CD14 cells.Results ELISA showed that secretion of TNF-α and IL-6 from THP-1/CD14 cells could be induced by PGN ;The productions of TNF-α and IL-6 by THP-1/CD14 cells induced with PGN were profoundly inhibited by MBL at higher concentrations (10-20 mg/L) but not MBL at lower concentrations (1 mg/L).RT-PCR analysis also indicated that the mRNA expressions of TNF-α and IL-6 in THP-1/CD14 cells were decreased by MBL at higher concentration,compared to the corresponding THP-1/CD14 cells stimulated with PGN only.FACS showed that the binding of MBL to THP-1/CD14 cells was evident in a Ca2+-dependent manner.PGN could competitively inhibit the binding of MBL to THP-1/CD14 cells.MBL could competitively inhibit the binding of PGN to THP-1/CD14 cells by binding to THP-1/CD14 cells directly.Similarly,MBL at higher concentration (20 mg/L) decreased the NF-κB translocation in THP-1/CD14 cells.Conclusion MBL may inhibit TNF-α and IL-6 production induced by PGN in THP-1/CD14 cells through NF-κB signaling pathways,suggesting that MBL can play some roles in the regulation of PGN-induced inflammatory response.
