Hemolysis and plasma heme liberated from Hb induce stasis in transgenic sickle mice. (A) Percent stasis was measured in the subcutaneous venules of NY1DD, HbSS, HbAS, HbAA, and C57BL/6 mice with DSFCs. Flowing venules were selected and mapped at baseline (20-35 venules per mouse). Mice were given a bolus infusion (0.012 mL/g) of the following treatments at the indicated Hb doses: saline (control), water (to induce hemolysis in vivo), HbA, metHbA, cyanometHbA, methylene blue (2 mg/kg, IV) + HbA, haptoglobin (3.2 µmol/kg, IV) + HbA, hemopexin (1.6 µmol/kg, IV) + HbA, or HbS. Percent stasis was measured using intravital microscopy 1 hour after infusion. The numbers of mice (n) in each treatment group are indicated. Bars are mean % stasis + standard deviation (SD) with mean stasis values written above the bars. (B) Percent stasis was measured in the subcutaneous venules of NY1DD and C57BL/6 mice with DSFCs as described in panel A. Mice were given a bolus infusion (0.012 ml/g) of the following treatments at the indicated heme dosages: heme, hemopexin (0.4 µmol/kg, IV) + heme, and PPIX (40 µmol/kg, IP 60 minutes preheme) + heme. (C) Correlation between percent stasis and total plasma heme concentrations. NY1DD sickle mice (n = 3/treatment) were infused with saline, heme, Hb, metHb or cyanometHb, or exposed to 1 hour of hypoxia (7%O₂) followed by 1 hour of hypoxia-reoxygenation (H/R). Percent stasis and total plasma heme were measured 1 hour after treatments. Values are mean % stasis and mean total plasma heme. (D) Percent stasis was measured in HbSS and HbAA mice in steady state. DSFCs were implanted on day −3. Flowing venules were selected at baseline on day 0. The same venules were reexamined for stasis on days 1 and 2. HbAA mice were untreated. HbSS mice were untreated or infused with human haptoglobin (900 µg/g) or hemopexin (34 µg/g) on days 0, 1, and 2. n = 3 mice per group; *P < .05 for HbSS vs HbSS + haptoglobin, HbSS + hemopexin or HbAA mice.