Analysis and manipulation of cell-mediated immune responses in shiga toxin-induced kidney injury

Shiga toxin (Stx)-‐producing enterohemorrhagic Escherichia coli (STEC) are the primary cause of hemolytic uremic syndrome (HUS), a life-‐threatening disease characterized by hemolytic anemia, thrombocytopenia and acute kidney failure. Endothelial cells were shown to be the main target for the cytotoxic effects of Stx leading to endothelial damage, recruitment of leukocytes, and thrombus formation. However, the precise contribution of monocytes, macrophages and neutrophils in the pathogenesis of HUS remains unclear and a therapy for STEC-HUS is still elusive. In this study, we have investigated the contribution of specific monocyte/macrophage subsets and the role of neutrophil recruitment to the pathology of HUS. To this end, a murine HUS model was employed in which mice were intravenously injected with a combination of Stx and lipopolysaccharide (LPS) to mimic a STEC infection. We observed impaired renal function as well as elevated levels of monocyte/macrophage-‐ and neutrophil-‐recruiting chemokines in the kidney of Stx/LPS-‐injected mice. Accordingly, flow cytometric analysis revealed increased numbers of CCR2-‐dependent Gr1high monocytes and neutrophils within the kidney, demonstrating myeloid cell infiltration during HUS. Positron emission tomography-‐magnetic resonance (PET-‐MR) imaging revealed renal injury mainly in the kidney cortex coinciding with the detection of Gr1high monocytes and neutrophils in this renal compartment. Indeed, reduced recruitment of Gr1high monocytes in Ccr2-‐deficient (Ccr2-‐/-‐) mice significantly ameliorated kidney injury and improved survival of these mice. Moreover, Ccr2-‐/-‐ x Cx3cr1gfp/gfp mice, which showed reduced abundance of Gr1high monocytes and Gr1low macrophages in the kidney, were completely protected from renal damage, indicating an essential role of both cell subsets. Furthermore, targeting CXCR2-‐dependent recruitment of neutrophils in either Cxcr2-‐deficient mice or by pharmacological inhibition (SB 225002) significantly ameliorated HUS and reduced mortality. Conclusively, this study identifies PET-‐MR imaging as a potential non-‐invasive approach to evaluate kidney damage in STEC-‐HUS. Moreover, these data demonstrate the crucial role of Gr1high monocytes, Gr1low macrophages and CXCR2-‐mediated neutrophil recruitment in renal injury in the murine model of HUS. Targeting the recruitment of these myeloid cells may serve as a first therapy against this devastating and severe disease.