High-fat diet activates splenic NOD1 and enhances neutrophil recruitment and neutrophil extracellular traps release in the spleen of ApoE-deficient mice

In the course of atherogenesis, the spleen plays an important role in the regulation of extramedullary hematopoiesis, and in the control of circulating immune cells, which contributes to plaque progression. Here, we have investigated the role of splenic nucleotide-binding oligomerization domain 1 (NOD1) in the recruitment of circulating immune cells, as well as the involvement of this immune organ in extramedullary hematopoiesis in mice fed on a high-fat high-cholesterol diet (HFD). Under HFD conditions, the absence of NOD1 enhances the mobilization of immune cells, mainly neutrophils, from the bone marrow to the blood. To determine the effect of NOD1-dependent mobilization of immune cells under pro-atherogenic conditions, Apoe(-/-) and Apoe(-/-)Nod1(-/-) mice fed on HFD for 4 weeks were used. Splenic NOD1 from Apoe(-/-) mice was activated after feeding HFD as inferred by the phosphorylation of the NOD1 downstream targets RIPK2 and TAK1. Moreover, this activation was accompanied by the release of neutrophil extracellular traps (NETs), as determined by the increase in the expression of peptidyl arginine deiminase 4, and the identification of citrullinated histone H3 in this organ. This formation of NETs was significantly reduced in Apoe(-/-)Nod1(-/-) mice. Indeed, the presence of Ly6G(+) cells and the lipidic content in the spleen of mice deficient in Apoe and Nod1 was reduced when compared to the Apoe(-/-) counterparts, which suggests that the mobilization and activation of circulating immune cells are altered in the absence of NOD1. Furthermore, confirming previous studies, Apoe(-/-)Nod1(-/-) mice showed a reduced atherogenic disease, and diminished recruitment of neutrophils in the spleen, compared to Apoe(-/-) mice. However, splenic artery ligation reduced the atherogenic burden in Apoe(-/-) mice an effect that, unexpectedly was lost in Apoe(-/-)Nod1(-/-) mice. Together, these results suggest that neutrophil accumulation and activity in the spleen are driven in part by NOD1 activation in mice fed on HFD, contributing in this way to regulating atherogenic progression.