PANCREATIC ELASTASE INDUCED TNF-α SECRETION AND CD11b EXPRESSION ARE MEDIATED BY TLR4 RECEPTOR ON CULTURED HUMAN MYELOID CELLS.

Abstract

Introduction: Pancreatic elastase has been implicated in the pathophysiology of severe acute pancreatitis characterized by systemic inflammatory response, distant organ failure and high mortality. Elastase induces NF-κB activation and TNF-α secretion from inflammatory cells in culture. Aims and Methods: Our aim was to study the signal transduction of pancreatic elastase-induced effects in inflammatory cells. Human myeloid cells (THP-1) were cultured in RPMI-1640 supplemented with 10% FCS and antibiotics. Pancreatic elastase (1 U/ml) was used to stimulate cells. TLR4 receptor was blocked by a monoclonal neutralizing anti-TLR4-antibody (Serotec). MG-132 (25μM) and PGA1 (100μM) were used to inhibit NF-κB activation. Transcription factor activation in THP-1 cells was measured by Transfactor assay (BD Biosciences). TNF-α secretion of cells was measured from culture media by using a commercial enzyme-linked immunosorbent assay (ELISA) kit. CD11b expression was studied by flow cytometry. Results: Pancreatic elastase induced activation of NF-κB and AP-1 in THP-1 cells. NF-κB activation was blocked by anti-TLR4 antibodies as well as by MG-132 and PGA1. Anti-TLR4-antibodies inhibited elastase-induced TNF-α secretion and increased expression of CD11b in THP-1 cells while MG-132 secretion from THP-1 cells. and PGA1 prevented elastase-induced TNF-α Conclusion: Our findings suggest that the proinflammatory effects of pancreatic elastase, such as NF-κB activation, TNF-α secretion and increased expression of CD11b are mediated by TLR4.