Summary.A family with an unusual form of hereditary stomatocytosis is described. The affected members showed a mild, dominantly-inherited, haemolytic anaemia with intracellular Na and K levels of 41-48 and 44-53 mmol/(l cells) respectively. This anaemia was associated with marked 'pseudohyperkalaemia': that is, loss of K from red cells on storage at room temperature. At 37ЊC, 'leak' tracer flux rates (assessed as the ouabain þ bumetanide-resistant K fluxes) showed a roughly 5-fold acceleration compared to normal, and an abnormal temperature dependence with a shallow slope between 37 and 20ЊC (mean Q 10 (ratio of reaction rates at temperature T and T ¹ 10) over this interval, 1·6; normal 2·2). The pseudohyperkalaemia could be attributed to the disparity between pump and leak at 20ЊC. This is an identical mechanism to that previously shown for the haematologically trivial condition, 'familial pseudohyperkalaemia'. No protein or lipid abnormality was found in the membrane of these cells.

In overhydrated hereditary stomatocytosis (OHSt), Coomassie-and silver-stained polyacrylamide gels show an apparently complete deficit of the 32-kDa membrane protein, stomatin. We have used an antistomatin antibody to examine peripheral blood films, bone marrow, splenic tissue, and hepatic tissue from these patients by immunocytochemistry. This technique revealed that, in fact, some red cells did show positive stomatin immunoreactivity; and consistent with this result, Western blot analysis of the red cell membranes confirmed that about one twentieth to one fiftieth of the normal amount of stomatin was in fact present. Flow cytometry, combining immunoreactive quantitation of stomatin expression with thiazole orange staining for reticulocytes, showed that in OHSt, it was the young cells that had more stomatin. Magnetic-activated cell separation studies, using beads to which an anti-transferrin receptor antibody was conjugated, confirmed that in OHSt there was a correspondence between expression of stomatin and the transferrin receptor. Immunocytochemistry and Western blotting revealed that in OHSt patients, the protein was present in spleen, liver, neutrophils, platelets, monocytes, and about 50% of the peripheral lymphocytes, with the same distribution as in healthy controls. Neither Southern blots, nor direct sequencing of multiple subclones of the cDNA, nor sequencing of amplicons from genomic DNA revealed any significant abnormality in stomatin gene sequence in these patients. The deficiency of stomatin from red cells appears to be due to a loss of stomatin from these red cells on maturation in the bone marrow and in the circulation. IntroductionThe name "stomatocytosis" was coined to describe the morphology in a dominantly inherited hemolytic anemia distinguished from hereditary spherocytosis by the cell shape. 1 This original condition is now known as overhydrated hereditary stomatocytosis (OHSt). Aside from the morphology, the anemia shows 2 other features: a catastrophic "leak" across the plasma membrane to the univalent cations Na ϩ and K ϩ (Zarkowsky et al 2 ), and the deficiency of the 32-kDa integral membrane protein, stomatin, or erythrocyte membrane protein 7.2b. [3][4][5] The condition represents an unusual genetic situation, in which a protein is apparently missing in a dominant, and therefore presumably heterozygous, condition. The gene coding for the missing protein has been cloned and sequenced. 5,6 Preliminary reports have indicated that no mutation has been found in American OHSt families, in the protein-coding region at least. 7,8 The pedigrees of OHSt are too small for effective genome-wide searches, but other variants of this leaky-cell class of red cell disease have been mapped. An American pedigree with nonstomatin-deficient dehydrated hereditary stomatocytosis (HSt) did not map to the stomatin locus on chromosome 9, 9,10 and European pedigrees of this non-stomatin-deficient dehydrated condition map to chromosome 16. 11 Other HSt pedigrees with different variants, none of which showed s...

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Summary.We describe two British families with similar, dominantly-inherited, temperature-related variants of hereditary stomatocytosis, consistent with the original description of 'cryohydrocytosis'. The cells show a 5-6-fold increase in passive permeability at 37ЊC with abnormal intracellular Na and K levels at 15-20 and 60-65 mmol/(l cells) respectively. Marked temperature effects were evident: lysis of red cells on storage in the cold was blatant and when whole heparinized blood was stored at room temperature, K accumulated in the plasma, producing 'pseudohyperkalaemia'. Studies of the temperature dependence of passive permeability showed that the minimum in the passive permeability, which is seen in normal cells at 8-10ЊC, was shifted up to 23ЊC in these abnormal cells, such that the permeability at 0ЊC exceeded that at 37ЊC. The abnormal temperature dependence in these genetically abnormal red cells strongly resembles that seen in normal cells when suspended in media in which either Na or Cl has been replaced by an organic cation or anion: it could be said these cells had a genetic mutation that somehow rendered the cell resistant to the stabilizing action of NaCl at low temperatures.

Summary This report concerns congenitally Na+–K+ leaky red cells of the ‘hereditary stomatocytosis’ class. Three new isolated cases and one new pedigree are described, and one previously reported case is expanded. In all cases, Western blotting of red cell membranes revealed a deficiency in the 32 kDa membrane protein, stomatin. All showed pronounced cation leaks at 37°C with markedly abnormal intracellular Na+ and K+ concentrations, like all other such stomatin‐deficient cases. Consistent with recent findings in two previously described British pedigrees, immunocytochemistry demonstrated that the deficiency of stomatin was not complete. On typical blood films, some red cells showed positive stomatin immunoreactivity, while most were negative, although in one case only a minority were negative. All platelets and neutrophils were stomatin positive. The cases differed markedly between themselves with regard to the temperature dependence of the passive leak to K+. Three showed a simple monotonic temperature dependence, while two showed a minimum at around 20–25°C, such that the cells were extremely leaky at 0°C, giving the phenotype known as ‘cryohydrocytosis’. These patients are the only two known cases of stomatin‐deficient cryohydrocytosis. Both showed a congenital syndrome of mental retardation, seizures, cataracts and massive hepatosplenomegaly, probably defining a new haemato‐neurological syndrome.