Life‐threatening hemolytic anemia due to an autoanti‐Pr cold agglutinin: evidence that glycophorin A antibodies may induce lipid bilayer exposure and cation permeability independent of agglutination
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BACKGROUND: The hemoglobin of a 29‐year‐old man fell below 35 g/L over 5 days, despite 14 units of red blood cells (RBCs), due to an anti‐Pr cold agglutinin (CA). His hemolytic anemia necessitated respiratory support in intensive care for 4 weeks.STUDY DESIGN AND METHODS: The hemolysis was investigated by the effects on blood group–compatible RBCs of this anti‐Pr and an anti‐I CA and of a rabbit anti‐human glycophorin A (GPA) immunoglobulin G (IgG) antibody on Ca2+ permeability and of phosphatidylethanolamine (PE) exposure. 1) The anti‐Pr CA (in a plasmapheresis product from the patient) was absorbed and eluted from RBC ghosts and its immunophenotype was determined by agarose electrophoresis and immunofixation. 2) Ca2+ permeability was measured by the response of Fluo‐3–labeled RBCs to addition of external Ca2+. 3) Exposed PE was measured with streptavidin‐labeled biotinylated peptide Ro 09‐0198 (cinnamycin).RESULTS: 1) The patient's anti‐Pr CA was a polyclonal IgG. 2) The anti‐Pr and the rabbit anti‐human glycophorin IgG, but not an anti‐I CA, rapidly increased Ca2+‐dependent fluorescence upon addition of external Ca2+ in a fraction (15%‐25%) of RBCs that also became positive for cinnamycin. 3) Trypsin treatment of RBCs reduced the Ca2+ influx due to the anti‐Pr IgG, but neither trypsin nor neuraminidase changed the responses to the rabbit anti‐human GPA IgG.CONCLUSIONS: The anti‐Pr CA and rabbit anti‐human GPA increased exposure of PE and increased membrane Ca2+ permeability that may have caused hemolysis. The difference in the responses to these antibodies to enzyme treatment of RBCs suggests that they react with different epitopes on GPA.
