Figure 6.

Heme and nucleosomes participate of the immune-mediated thrombotic thrombocytopenic purpura patient’s plasma induced Ca²⁺-mediated endothelial activation. (A) Free heme concentration (mM) measured in control (n=13), immune-mediated thrombotic thrombocytopenic purpura (iTTP) patients in acute phase (iTTP-AP, n=62) or iTTP patients in remission (iTTP-R, n=14) plasma samples (*P<0.05, ***P<0.001). (B) Von Willebrand Factor (VWF) expression detected by fluorescence microscopy after permeabilized human microvascular endothelial cells from derm (HMVEC-d) incubation with heme or iTTP plasma samples in the absence (A, B) or in the presence (C, D) of hemopexin (+Hx) (original magnification X40, scale bar=30 mm). (C) Endothelin-1 concentrations in the supernatants of HMVEC-d incubated with iTTP plasma samples in the absence (representing 100%) or in the presence of hemopexin (n=14, *P<0.05). (D) Fluorescence intensity (AU) of the Ca²⁺ flux induction in HMVEC-d after a 20-second incubation with iTTP plasma samples alone or after preincubation with hemopexin (n=18, **P<0.01). (E) Total nucleosomes in control (n=13), iTTP patients in acute phase (iTTP-AP, n=62) or iTTP patients in remission (iTTP-R, n=14) plasma samples (***P<0.001). (F) VWF secretion detected by fluorescence microscopy after incubation of HMVEC-d with iTTP plasma samples in the absence (A) or in the presence of Annexin V (B) (original magnification X40, scale bar=30 mm). (G) Endothelin-1 concentrations in the supernatants of HMVEC-d incubated with iTTP plasma samples in the absence (representing 100%) or in the presence of Annexin V (n=16, **P<0.01). (H) Fluorescence intensity (AU) of the Ca²⁺ flux induction in HMVEC-d after a 20-second incubation with either iTTP plasma samples alone, or in the presence of Annexin V (n=17, **P<0.01).