American Society of Hematology

Figure 7

$Laboratory indicators of hemolytic anemia. Mean ± SEM are shown for 6 to 10 animals of each genotype. (A) Hemoglobin levels of sickle mice were lower than hemizygotes, but both were below wild-type, consistent with previous reports.23,27 The hemizygous mouse is anemic but does not have erythrocyte sickling. (B) Elevated reticulocyte fraction indicated markedly increased erythropoiesis in sickle mice. (C) Spleen as a fraction of body weight was elevated in sickle mice. The spleen is a normal site of erythropoiesis in adult mice and greatly increases in conditions of increased erythropoiesis. (D) Indirect bilirubin was elevated in sickle mouse plasma. (E) Lactate dehydrogenase in plasma was elevated in sickle mice as a nonspecific indicator of hemolysis. (F) Plasma NO consumption in blood processed under low shear to avoid artifactual hemolysis was measured using the gas-phase chemiluminescent NO analyzer (Sievers, Boulder, CO) as previously described.6 Nitric oxide consumption measurement was elevated in sickle mice (n = 21) compared with hemizygotes (n = 11) or wild-type controls (n = 6) by Kruskal-Wallis test. Absorbance spectrophotometry in the Soret band determined that plasma hemoglobin concentration correlated with NO consumption in mice (r2 = 0.96, n = 49, data not shown). The wide variability in NO consumption in the sickle mice was similar to the variability of plasma NO consumption in humans with sickle cell disease.6 (G) Whole blood nitrite levels were obtained by mixing fresh blood immediately with nitrite preservation solution63 and then assaying by chemiluminescence (Sievers, Boulder, CO) as an indirect measure of NO synthase activity. Whole blood nitrite levels were lower in sickle mice than hemizygous and wild-type controls by the Mann-Whitney test (n = 4 to 7 per group). Nitrite in the BM-S mice (n = 9) was lower than the nitrite in the controls receiving transplants with wild-type marrow (BM-C, n = 4). (H) Lung homogenate assays demonstrated significantly higher arginase activity in sickle mice and in BM-S mice (n = 4 to 7 mice per group). Statistically significant results are indicated as follows: *P < .05 versus hemizygous controls; #P < .05 versus BM-C controls.$

Laboratory indicators of hemolytic anemia. Mean ± SEM are shown for 6 to 10 animals of each genotype. (A) Hemoglobin levels of sickle mice were lower than hemizygotes, but both were below wild-type, consistent with previous reports.^23,27 The hemizygous mouse is anemic but does not have erythrocyte sickling. (B) Elevated reticulocyte fraction indicated markedly increased erythropoiesis in sickle mice. (C) Spleen as a fraction of body weight was elevated in sickle mice. The spleen is a normal site of erythropoiesis in adult mice and greatly increases in conditions of increased erythropoiesis. (D) Indirect bilirubin was elevated in sickle mouse plasma. (E) Lactate dehydrogenase in plasma was elevated in sickle mice as a nonspecific indicator of hemolysis. (F) Plasma NO consumption in blood processed under low shear to avoid artifactual hemolysis was measured using the gas-phase chemiluminescent NO analyzer (Sievers, Boulder, CO) as previously described.⁶ Nitric oxide consumption measurement was elevated in sickle mice (n = 21) compared with hemizygotes (n = 11) or wild-type controls (n = 6) by Kruskal-Wallis test. Absorbance spectrophotometry in the Soret band determined that plasma hemoglobin concentration correlated with NO consumption in mice (r² = 0.96, n = 49, data not shown). The wide variability in NO consumption in the sickle mice was similar to the variability of plasma NO consumption in humans with sickle cell disease.⁶ (G) Whole blood nitrite levels were obtained by mixing fresh blood immediately with nitrite preservation solution⁶³ and then assaying by chemiluminescence (Sievers, Boulder, CO) as an indirect measure of NO synthase activity. Whole blood nitrite levels were lower in sickle mice than hemizygous and wild-type controls by the Mann-Whitney test (n = 4 to 7 per group). Nitrite in the BM-S mice (n = 9) was lower than the nitrite in the controls receiving transplants with wild-type marrow (BM-C, n = 4). (H) Lung homogenate assays demonstrated significantly higher arginase activity in sickle mice and in BM-S mice (n = 4 to 7 mice per group). Statistically significant results are indicated as follows: *P < .05 versus hemizygous controls; #P < .05 versus BM-C controls.

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