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Life Sci. 2012 Aug 21;91(3-4):143-50. doi: 10.1016/j.lfs.2012.06.030. Epub 2012 Jul 4.

Trypsin inhibits lipopolysaccharide signaling in macrophages via toll-like receptor 4 accessory molecules.

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  • 1Laboratory of Veterinary Pharmacology, Joint Faculty of Veterinary Medicine, Yamaguchi University, Japan.



To examine the role of trypsin in the immune response of macrophages and to determine whether protease-activated receptors (PARs) are involved in the effects of trypsin.


We used RAW264.7 cells and peritoneal macrophages isolated from C57BL/6 wild-type mice, PAR2 knockout mice, and ddY mice. Macrophages were stimulated with lipopolysaccharide (LPS) in the presence or absence of trypsin, thrombin, and PAR subtype-specific agonists (PARs-AP). Activation of macrophages was quantified by nitric oxide production and expression of inflammatory mediators, such as inducible nitric oxide synthase, interleukin-1β, and interleukin-6. To clarify the effect of trypsin on LPS receptors, we also investigated the expression of toll-like receptor 4 (TLR4), soluble MD-2 (sMD-2), membrane-bound MD-2 (mMD-2), soluble CD14 (sCD14), and membrane-bound CD14 (mCD14). To directly investigate the effect of trypsin on CD14 protein, we expressed recombinant CD14 protein.


Trypsin inhibited LPS-induced nitric oxide production and expression of inducible nitric oxide synthase, interleukin-1β, and interleukin-6. The same inhibitory effects of trypsin were observed in wild-type macrophages and in PAR2 knockout macrophages. Furthermore, the other PAR agonists, thrombin, PAR1-AP, PAR2-AP, and PAR4-AP, did not mimic the effect of trypsin. Although trypsin did not affect TLR4 or mMD-2 expression, sCD14, mCD14, and sMD-2 expressions were decreased by trypsin. Furthermore, trypsin also degraded recombinant CD14 protein.


Trypsin inhibited LPS signaling PAR-independently via degradation of TLR4 accessory molecules. This observation provides a better understanding of the complicated immune response in acute pancreatitis.

Copyright © 2012 Elsevier Inc. All rights reserved.

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