Heme-induced kallikrein-kinin system activation in hemolytic uremic syndrome.

Abstract

Heme is released from red blood cells during hemolytic uremic syndrome (HUS). Here, we investigated hemolysis-induced kallikrein-kinin system activation and the interaction between heme and clotting factor XII (FXII). Pediatric patients (19 male and 15 female, median age three years, range three months to 14 years) with HUS induced by enterohemorrhagic Escherichia coli (EHEC) and mice infected with EHEC were assessed for activation of the kallikrein-kinin system by degradation of high-molecular-weight kininogen ((HK) by immunoblotting) and levels of residual pre-kallikrein (by chromogenic assay). Heme induction of kallikrein-kinin system activation was evaluated by plasma kallikrein or FXII activity, bradykinin release (by ELISA) and heme binding to FXII (by surface plasmon resonance). Hemolysis was induced in vitro and in mice by phenylhydrazine in the presence or absence of C1-inhibition. Ten of the 34 patients with EHEC-HUS had HK degradation associated with the severity of hemolysis and 17 of 32 had reduced residual pre-kallikrein associated with kidney dysfunction. Similarly, EHEC-infected mice had HK degradation and pre-kallikrein consumption, the latter inversely correlated with hemolysis biomarkers lactic dehydrogenase and arginase 1 and with creatinine levels. Heme bound to and activated FXII with increased kallikrein activity, causing HK breakdown and releasing bradykinin in normal, but not FXII-deficient plasma; effects inhibited by hemopexin, C1-inhibitor, and corn trypsin inhibitor. Phenylhydrazine-induced hemolysis led to HK degradation in human and murine blood; not detected when red blood cells were added to FXII-deficient plasma, or in the presence of C1-inhibitor. Mice challenged with phenylhydrazine exhibited hemolysis and significant pre-kallikrein consumption as well as HK degradation in five of 10 mice. HK degradation was abrogated by pretreatment with C1-inhibitor. Our study demonstrates that heme binds to and activates FXII leading to HK degradation and bradykinin release. This could be a potent inflammatory mechanism in hemolytic disease, such as HUS, and amenable to pharmacological blockades.