Cot/tpl2 (MAP3K8) Mediates Myeloperoxidase Activity and Hypernociception following Peripheral Inflammation*

Irene Soria-Castro,Agnieszka Krzyzanowska,Marta López Pelaéz,Javier Regadera,Gema Ferrer,Lluis Montoliu,Rosario Rodríguez-Ramos,Margarita Fernández,Susana Alemany

doi:10.1074/jbc.m110.169409

Abstract

Cot/tpl2 (also known as MAP3K8) has emerged as a new and potentially interesting therapeutic anti-inflammatory target. Here, we report the first study of Cot/tpl2 involvement in acute peripheral inflammation in vivo. Six hours after an intraplantar injection of zymosan, Cot/tpl2(-/-) mice showed a 47% reduction in myeloperoxidase activity, concomitant with a 46% lower neutrophil recruitment and a 40% decreased luminol-mediated bioluminescence imaging in vivo. Accordingly, Cot/tpl2 deficiency provoked a 25-30% reduction in luminol-mediated bioluminescence and neutrophil recruitment together with a 65% lower macrophage recruitment 4 h following zymosan-induced peritonitis. Significantly impaired levels of G-CSF and GM-CSF and of other cytokines such as TNFα, IL-1β, and IL-6, as well as some chemokines such as MCP-1, MIP-1β, and keratinocyte-derived chemokine, were detected during the acute zymosan-induced intraplantar inflammatory response in Cot/tpl2(-/-) mice. Moreover, Cot/tpl2 deficiency dramatically decreased the production of the hypernociceptive ligand NGF at the inflammatory site during the course of inflammation. Most importantly, Cot/tpl2 deficiency significantly reduced zymosan-induced inflammatory hypernociception in mice, with a most pronounced effect of a 50% decrease compared with wild type (WT) at 24 h following intraplantar injection of zymosan. At this time, Cot/tpl2(-/-) mice showed significantly reduced NGF, TNFα, and prostaglandin E(2) levels compared with WT littermates. In conclusion, our study demonstrates an important role of Cot/tpl2 in the NGF, G-CSF, and GM-CSF production and myeloperoxidase activity in the acute inflammatory response process and its implication in inflammatory hypernociception.

Highlights

Like receptors, and C-type lectin receptors, by pathogen-associated molecular patterns starts a program that culminates in the development of an inflammatory process
To ensure a role of Cot/tpl2 in the zymosan intracellular signaling through TLR2/6, we stimulated WT and Cot/tpl2Ϫ/Ϫ Bone marrow derived macrophages (BMDM), which express low levels of dectin-1 but high levels of TLR2/6 [45], with zymosan and found no increase in the phosphorylation state of ERK1/2 in Cot/tpl2Ϫ/Ϫ BMDM
The bioluminescence observed 24 h later in both WT and Cot/tpl2Ϫ/Ϫ hindpaws had returned to almost basal levels (Fig. 1C). In concordance with these data, Cot/tpl2Ϫ/Ϫ hindpaw extracts 6 h following zymosan injection showed a 47% decreased MPO specific activity compared with zymosan-injected WT hindpaws extracts (Fig. 1F). Because this decrease in MPO activity in Cot/tpl2Ϫ/Ϫ hindpaws could indicate a decrease in the number of infiltrated neutrophils, the number of inflammatory cells was determined in zymosan-injected WT and Cot/tpl2Ϫ/Ϫ hindpaws

Summary

Introduction

Like receptors, and C-type lectin receptors, by pathogen-associated molecular patterns starts a program that culminates in the development of an inflammatory process (for review, see Ref. 1). We demonstrate that Cot/tpl2 modulates the number of neutrophils recruited to inflammatory foci induced by zymosan injection in vivo as well as the level of cytokines/chemokines, NGF, PGE2, and leukotriene (LT) B4 production at those sites.

Results

Conclusion