Therapeutic plasma exchange and intravenous immunoglobulin as primary therapy for D alloimmunization in pregnancy precludes the need for intrauterine transfusion.

BACKGROUND: Maternal D alloimmunization detected in early gestation requires aggressive intervention to prevent severe fetal anemia. An intrauterine transfusion (IUT) is indicated to prevent fetal death once severe fetal anemia has been detected, but is not without risk. Protocols combining therapeutic plasma exchange (TPE) and intravenous immunoglobulin (IVIG) have been described, but they usually bridge to IUT. STUDY DESIGN AND METHODS: We describe a 27-year-old G4, P0-1-2-0 Caucasian female with a history of ruptured ectopic pregnancy presented at 12 weeks' gestation with a very high anti-D titer (2048). TPE was performed on that week and twice more in the following week, with a fourth final exchange during Week 14. A loading dose of IVIG (2 g/kg) was administered over 2 days after the third TPE and then 1 g/kg per week until Week 28 (total, 14 doses). RESULTS: The antibody titer decreased to 256 by the beginning of 15 weeks' gestation and remained stable at that level for the remainder of the pregnancy. Doppler ultrasonographic measurements of the fetal middle cerebral artery peak flow velocity performed throughout gestation showed no evidence of fetal anemia. A healthy male infant was delivered at 37 weeks' gestation with mild immune-mediated hemolysis. The infant underwent successful treatment with an IVIG dose of 750 mg/kg and a red blood cell exchange. CONCLUSION: Our unique TPE-IVIG protocol was successful at preventing the onset of severe fetal anemia in a patient with high titer anti-D. Since IUT may be fatal, our approach offers a safer and less-invasive treatment regime that can adequately sustain a fetus until term.

Genetic diagnosis of band 3 deficiency and sexing in bovine preimplantation embryos.

Band 3 deficiency with hereditary spherocytosis and hemolytic anemia in Japanese black cattle, band 3(Bov.Yamagata), is caused by a total lack of band 3 protein with an autosomal dominant inheritance. Genotyping for band 3 deficiency and sexing were successfully achieved in biopsied embryo cells with efficiencies of 98.4% and 97.4%, respectively. Transfer of the embryo that was determined as homozygous for the mutant allele into a recipient cow resulted in the production of a fetus exhibiting the genotype and red cell phenotypes characteristic of band 3(Bov.Yamagata). These results demonstrate that our procedure is reliable and applicable to produce animals free from or homozygous for the mutant allele by breeding carrier animals.

Clinical application of a rapid method using agarose gel electrophoresis and Western blotting to evaluate von Willebrand factor protease activity.

A method for evaluating the activity of the von Willebrand factor (vWF) protease is described, and a clinical application is illustrated. The procedure utilizes gel electrophoresis, Western blotting, and luminographic detection methods to evaluate the distribution of vWF multimers before and after incubation of clinical samples under conditions that favor proteolysis by this enzyme. Physiologically, the high-molecular-weight multimers of vWF are cleaved by the vWF protease under conditions of high shear stress in parts of the arterial circulation; cleavage of vWF multimers is also observed after exposure of vWF to denaturing agents in vitro and thus can serve as a laboratory test for the activity of the protease. vWF protease activity is decreased or absent in patients with thrombotic thrombocytopenic purpura due to an inhibiting autoantibody, and this leads to high levels of noncleaved vWF and to life-threatening thrombosis, thrombocytopenia and anemia. The assay evaluates the activity of the protease by assessing the cleavage of vWF multimers after patient plasmas are incubated in vitro under denaturing conditions. With the use of these electrophoresis and Western blotting techniques, patient plasmas can be rapidly assessed for the activity of the vWF protease which may aid in the treatment strategy for these patients.

Decreased fetal erythropoiesis and hemolysis in Kell hemolytic anemia.

OBJECTIVE: Lower changes in optical density (450 nm) measurements have been reported in fetuses with anti-Kell anemia compared with those anti-D anemia. The purpose of this investigation was to determine if hemolysis and erythropoiesis differ between anti-Kell and anti-D hemolytic disease. STUDY DESIGN: Ninety-three pregnancies complicated by either anti-D or anti-Kell alloimmunization were evaluated. Fetal blood samples obtained at the first cordocentesis were tested for the red blood cell antigen type, hemoglobin, hematocrit, reticulocyte count, nucleated red blood cells, total serum bilirubin concentration, umbilical venous respiratory blood gases, serum erythropoietin level, and strength of the direct Coombs test. To determine the evolution of hemolytic anemia in the two antigen groups, these laboratory parameters were repeated on the fetal blood samples triggering the decision to perform a fetal intravascular transfusion (hematocrit <30%). RESULTS: A total of 65 of 93 fetuses were antigen positive (11 for Kell and 54 for RhD). The mean gestational age and laboratory measurements of antigen- positive, nonanemic fetuses at first blood sampling did not differ significantly between groups. There was a strong inverse relationship observed between the hemoglobin concentration and reticulocyte count independent of gestational age in the anti-D group but not in the anti-Kell group. Eight (73%) fetuses with anti-Kell antibodies and 37 (69%) with anti-D antibodies underwent intravascular transfusion. At the cordocentesis when the decision for transfusion was made, anti-Kell anemic fetuses had lower reticulocyte counts and total bilirubin concentrations. The strong inverse relationship between the hemoglobin and reticulocyte count was again seen only in the anti-D group. In both groups, fetal erythropoietin increased significantly between the first and last blood samplings and in each group were negatively correlated with hemoglobin independent of gestational age. CONCLUSION: Anti-Kell anemic fetuses have lower reticulocyte counts and total serum bilirubin levels than do comparable anti-D anemic fetuses. This finding argues in favor of fetal blood sampling rather than amniotic fluid analyses for the management of fetal hemolytic disease resulting from Kell antibodies. Unlike RhD alloimmunized fetuses, these fetuses do not manifest an inverse relationship between hemoglobin concentration and reticulocyte count. We speculate that compared to anti-D fetal anemia, anti-Kell anemia is associated with increased hemolysis of nonhemoglobinized or incompletely hemoglobinized erythroid precursors.

Fetal serum ferritin and cobalamin in red blood cell isoimmunisation.

Fetal and maternal serum ferritin and cobalamin concentrations were examined in 40 red blood cell-isoimmunised pregnancies undergoing cordocentesis at 18-38 weeks of gestation and the values were compared to those of normal pregnancies. In the red blood cell-isoimmunised pregnancies, the fetal serum ferritin concentration was increased and the cobalamin concentration was decreased, whereas maternal serum ferritin was decreased and cobalamin was not significantly different from normals. There was a significant association between the degree of fetal anaemia and the increase in fetal serum ferritin concentration, but not with fetal serum cobalamin. This study suggests that fetal haemolytic anaemia is associated with iron overload and cobalamin deficiency.

Fetal compensation of the hemolytic anemia in mice homozygous for the normoblastosis (nb) mutation.

The mouse autosomal recessive mutation nb causes a deficiency of erythroid ankyrin and generates a life-threatening hemolytic anemia in adult mice; however, at birth, nb/nb mice appear to be robust and show no pallor. In our study, the time of disease onset was sought by comparison of nb/nb and +/? mice both in utero and postnatally. Erythroid ankyrin messenger RNA (mRNA) is expressed in fetal erythroid progenitors from normal mice, but is reduced to 10% of normal levels in mutant fetuses. Despite the deficiency of erythroid ankyrin mRNA, 16 and 18 day nb/nb fetuses have normal levels of red blood cells (RBCs) and the RBCs are morphologically normal by scanning electron microscopy. The earliest signs of any clinical anomaly are an increase in the number of circulating reticulocytes and the deposition of minor amounts of iron just before birth in the 18 day fetal nb/nb liver, suggesting that RBCs are being destroyed. Within 24 hours after birth, nb/nb neonates have a slight but significant decrease of their RBC counts. During the next 5 days, the nb/nb RBC counts decrease markedly, the reticulocyte counts assume the mutant adult levels of 60%, the erythrocytes become microcytic and fragmented, and iron deposits accumulate in the liver. The rapid onset of clinical disease postnatally, coupled with our findings that the erythroid ankyrin gene is transcribed in fetal erythroid cell precursors from normal mice, suggest that mechanisms exist in the nb/nb fetus to compensate for the erythroid ankyrin deficiency.