Free heme and hemopexin in acute kidney injury after cardiopulmonary bypass and transient renal ischemia.

Free heme is released from hemoproteins during hemolysis or ischemia reperfusion injury and can be pro-inflammatory. Most studies on nephrotoxicity of hemolysis-derived proteins focus on free hemoglobin (fHb) with heme as a prosthetic group. Measurement of heme in its free, non-protein bound, form is challenging and not commonly used in clinical routine diagnostics. In contrast to fHb, the role of free heme in acute kidney injury (AKI) after cardiopulmonary bypass (CPB) surgery is unknown. Using an apo-horseradish peroxidase-based assay, we identified free heme during CPB surgery as predictor of AKI in patients undergoing cardiac valve replacement (n = 37). Free heme levels during CPB surgery correlated with depletion of hemopexin (Hx), a heme scavenger-protein. In mice, the impact of high levels of circulating free heme on the development of AKI following transient renal ischemia and the therapeutic potential of Hx were investigated. C57BL/6 mice were subjected to bilateral renal ischemia/reperfusion injury for 15 min which did not cause AKI. However, additional administration of free heme in this model promoted overt AKI with reduced renal function, increased renal inflammation, and reduced renal perfusion on functional magnetic resonance imaging. Hx treatment attenuated AKI. Free heme administration to sham operated control mice did not cause AKI. In conclusion, free heme is a predictor of AKI in CPB surgery patients and promotes AKI in transient renal ischemia. Depletion of Hx in CPB surgery patients and attenuation of AKI by Hx in the in vivo model encourage further research on Hx therapy in patients with increased free heme levels during CPB surgery.

Renal medullary osmolytes NaCl and urea differentially modulate human tubular cell cytokine expression and monocyte recruitment.

Renal immune cells serve as sentinels against ascending bacteria but also promote detrimental inflammation. The kidney medulla is characterized by extreme electrolyte concentrations. We here address how its main osmolytes, NaCl and urea, regulate tubular cell cytokine expression and monocyte chemotaxis. In the healthy human kidney, more monocytes were detected in medulla than cortex. The monocyte gradient was attenuated in patients with medullary NaCl depletion by loop diuretic therapy and in the nephrotic syndrome. Renal tubular epithelial cell gene expression responded similarly to NaCl and tonicity control mannitol, but not urea. NaCl significantly upregulated chemotactic cytokines, most markedly CCL26, CCL2, and CSF1. This induction was inhibited by the ROS scavenger n-acetylcysteine. In contrast, urea, the main medullary osmolyte in catabolism, dampened tubular epithelial CCL26 and CSF1 expression. Renal medullary chemokine and monocyte marker expression decreased in catabolic mice. NaCl-, but not urea-stimulated tubular epithelium or CCL2 and CCL26, promoted human classical monocyte migration. CCL26 improved bactericidal function. In the human kidney medulla, monocyte densities correlated with tubular CCL26 protein abundance. In summary, medullary-range NaCl, but not urea, promotes tubular cytokine expression and monocyte recruitment. This may contribute to the pyelonephritis vulnerability in catabolism but can possibly be harnessed against pathologic inflammation.

Peritoneal dialysate-range hypertonic glucose promotes T-cell IL-17 production that induces mesothelial inflammation.

Peritoneal dialysis (PD) employs hypertonic glucose to remove excess water and uremic waste. Peritoneal membrane failure limits its long-term use. T-cell cytokines promote this decline. T-cell differentiation is critically determined by the microenvironment. We here study how PD-range hypertonic glucose regulates T-cell polarization and IL-17 production. In the human peritoneal cavity, CD3+ cell numbers increased in PD. Single cell RNA sequencing detected expression of T helper (Th) 17 signature genes RORC and IL23R. In vitro, PD-range glucose stimulated spontaneous and amplified cytokine-induced Th17 polarization. Osmotic controls l-glucose and d-mannose demonstrate that induction of IL-17A is a substance-independent, tonicity dose-dependent process. PD-range glucose upregulated glycolysis and increased the proportion of dysfunctional mitochondria. Blockade of reactive-oxygen species (ROS) prevented IL-17A induction in response to PD-range glucose. Peritoneal mesothelium cultured with IL-17A or IL17F produced pro-inflammatory cytokines IL-6, CCL2, and CX3CL1. In PD patients, peritoneal IL-17A positively correlated with CX3CL1 concentrations. PD-range glucose-stimulated, but neither identically treated Il17a-/- Il17f-/- nor T cells cultured with the ROS scavenger N-acetylcysteine enhanced mesothelial CX3CL1 expression. Our data delineate PD-range hypertonic glucose as a novel inducer of Th17 polarization in a mitochondrial-ROS-dependent manner. Modulation of tonicity-mediated effects of PD solutions may improve membrane survival.