Sustained induction of IP-10 by MRP8/14 via the IFNβ-IRF7 axis in macrophages exaggerates lung injury in endotoxemic mice.

BACKGROUND

As a damage-associated molecular pattern, the myeloid-related protein 8/14 (MRP8/14) heterodimer mediates various inflammatory diseases, such as sepsis. However, how MRP8/14 promotes lung injury by regulating the inflammatory response during endotoxemia remains largely unknown. This study aims at illuminating the pathological functions of MRP8/14 in endotoxemia.

METHODS

An endotoxemic model was prepared with wild-type and myeloid cell-specific deletion () mice for evaluating plasma cytokine levels. Lung injury was evaluated by hematoxylin and eosin (H&E) staining, injury scoring and wet-to-dry weight (W/D) ratio. The dynamic profile of interferon γ (IFNγ)-inducible protein 10 (IP-10) mRNA expression induced by macrophage MRP8/14 was determined by quantitative real-time polymerase chain reaction (qPCR). Immunoblotting was used to evaluate the increase in IP-10 level induced by activation of the JAK-STAT signaling pathway. Luciferase reporter assay was performed to detect the involvement of IRF7 in gene transcription. air pouch experiments were performed to determine the biological function of IP-10 induced by MRP8/14.

RESULTS

Experiments with mice showed that MRP8/14 promoted the production of cytokines, including IP-10, in the bronchoalveolar lavage fluid (BALF) and lung injury in endotoxic mice. The result of qPCR showed sustained expression of mRNA in macrophages after treatment with MRP8/14 for 12 h. Neutralization experiments showed that the MRP8/14-induced expression in RAW264.7 cells was mediated by extracellular IFNβ. Western blotting with phosphorylation-specific antibodies showed that the JAK1/TYK2-STAT1 signaling pathway was activated in MRP8/14-treated RAW264.7 cells, leading to the upregulation of gene expression. IRF7 was further identified as a downstream regulator of the JAK-STAT pathway that mediated gene expression in macrophages treated with MRP8/14. air pouch experiments confirmed that the IFNβ-JAK1/TYK2-STAT1-IRF7 pathway was required for chemokine (C-X-C motif) receptor 3 (CXCR3) T lymphocyte migration, which promoted lung injury in the context of endotoxemia.

CONCLUSIONS

In summary, our study demonstrates that MRP8/14 induces sustained production of IP-10 via the IFNβ-JAK1/TYK2-STAT1-IRF7 pathway to attract CXCR3 T lymphocytes into lung tissues and ultimately results in lung injury by an excessive inflammatory response in the context of endotoxemia.