Hereditary stomatocytosis, hereditary cryohydrocytosis, and hereditary xerocytosis

Hereditary stomatocytosis, hereditary cryohydrocytosis, and hereditary xerocytosis

What every physician needs to know:

Hereditary stomatocytosis and hereditary xerocytosis are rare, genetically distinct, autosomal dominant diseases of red blood cell sodium and potassium permeability. Hereditary cryohydrocytosis is a subtype of hereditary stomatocytosis. All three present with hemolysis and anemia, which can vary from mild to severe. The key signs and symptoms are those of all hemolytic anemias: jaundice, pallor, fatigue, splenomegaly, gallbladder disease, and susceptibility to an aplastic crisis following infection with parvovirus B19. The diseases begin at birth, but especially in the case of hereditary xerocytosis and hereditary cryohydrocytosis, may be mild enough to go undiscovered for years.

Hereditary stomatocytosis is characterized by excessively water-laden erythrocytes. The extremely high influx of Na⁺(sodium) and water from the plasma exceeds the loss of cellular K⁺ (potassium ion) and the resulting swollen red cells show a unique combination of large size (high mean corpuscular volume [MCV]), low mean corpuscular hemoglobin concentration (MCHC), and osmotic sensitivity in the osmotic fragility test. No other condition shows this unique combination. Blood smears feature stomatocytes, or a mixture of stomatocytes and spherocytes.

Hereditary cryohydrocytosis is a mild variant of hereditary stomatocytosis in which the abnormal Na⁺permeability increases below 20ºC, exceeding the ability of the red cell to pump the leaked Na⁺back out. As a consequence, red cells from patients with cryohydrocytosis swell and partially hemolyze when stored in the refrigerator overnight. This is a unique feature and is pathognomonic for the condition.

Hereditary xerocytosis (which is also called dehydrated hereditary stomatocytosis) is the most common of the inherited red cell cation permeability disorders. It is characterized by a mild increase in potassium permeability that is sufficient to lead to the gradual loss of red cell K⁺ and water, and to red cell dehydration, stiffness, and hemolysis. Red cells are macrocytic despite being dehydrated (high MCHC, resistant osmotic fragility), which is a unique and nearly diagnostic combination. Hemoglobin and hematocrit values are often normal (compensated hemolysis) and patient blood smears are surprisingly normal, featuring mostly a few target cells.

There are multiple recent reviews, see references below.

Are you sure your patient has hereditary stomatocytosis, hereditary cryohydrocytosis, or hereditary xerocytosis? What should you expect to find?

Common clinical features

These diseases show the typical features of red cell destruction and compensatory red cell production characteristic of all hemolytic anemias: indirect hyperbilirubinemia, low levels of haptoglobin, normal or mildly elevated lactate dehydrogenase (LDH) and reticulocytosis. Red cell destruction appears to occur primarily in the reticuloendothelial system, so signs of intravascular hemolysis (hemoglobinemia, hemoglobinuria, extremely high LDH levels) are not usually observed.

Anemia varies from none to severe, or even transfusion-dependent, depending on the underlying mutation. Splenomegaly and cholelithiasis with bilirubin gallstones are common. Signs and symptoms date from birth, though they may be overlooked for years, or even decades in the milder variants.

Signs of hemolysis are often more evident in the neonatal period, due to immaturity in the conjugation of bilirubin by the liver, so a history of neonatal jaundice or transfusions, or of treatment with phototherapy should be sought. All patients with hemolysis are susceptible to aplastic crises if infected with parvovirus B19, and patients with the more severe variants may suffer from intermittent hemolytic crises and increased jaundice with infections that activate macrophages or exacerbate splenomegaly.

Positive family history

A careful family history should be taken, looking for anemia, splenectomy, jaundice, early gallbladder disease, transfusion, neonatal jaundice or edema, hyperkalemia, iron overload, iron chelation therapy, or thromboembolic disease. The red cell membrane permeability diseases are transmitted from generation to generation with no gender preference (autosomal dominant pattern). As with most dominant diseases, symptoms may vary among affected family members. A careful family history should always be taken; however, the lack of a positive family history is not critical, as many patients have new mutations.

Unique features

The characteristic features of hereditary stomatocytosis (HSt), hereditary cryohydrocytosis (HC) and hereditary xerocytosis (HX) and related diseases are shown in
Table I. (Adapted from Lux SE. Hematology of Infancy and Childhood. 2015.)

Red blood cell morphology on peripheral blood smears is shown in Figure 1.

Figure 1.

Severe overhydrated, stomatin-deficient hereditary stomatocytosis

This rare disease is variously called hereditary stomatocytosis, overhydrated hereditary stomatocytosis, and hereditary hydrocytosis. The excess monovalent cations elevate cell water, producing large, osmotically fragile cells with a low MCHC. The diagnostic features observed in commonly available laboratory tests include, unique red cell morphology (5 to 40% stomatocytes, with or without spherocytes) (see Figure 1), moderate to severe hemolysis and anemia, macrocytosis (high mean corpuscular volume [MCV], that is, 95 to 150fL), low MCHC (24 to 30%), and a positive osmotic fragility test (osmotically fragile cells, curve shifted toward high ionic strengths). The combination of macrocytosis and a low MCHC is virtually diagnostic of hereditary stomatocytosis, especially when stomatocytes are present on the peripheral blood smear and the osmotic fragility test is positive.

Additional studies in specialty laboratories will show:

• An elevated erythrocyte Na⁺concentration (60 to 100mEq/L, normal 5 to 12mEq/L), a reduced erythrocyte K⁺concentration (20 to 55mEq/L, normal 90 to 103mEq/L), and an increased total Na⁺plus K+ concentration (110 to 140mEq/L, normal 95 to 110mEq/L)

• An extremely high passive membrane permeability of Na⁺and K⁺ (20 to 40 times normal)

• Abnormal red cell ektacytometry

• Nearly absent stomatin (31-kD protein) on SDS (sodium dodecyl sulfate) gel electrophoresis of red cell membranes or by immunostaining of erythrocytes

• In many, and perhaps all cases, mutations in the RHAG gene (see later section, “Pathophysiology”).

Patients with this severe variant of stomatocytosis usually present with signs of hemolysis and anemia in infancy. They may also present with pseudohyperkalemia due to loss of K⁺ from the excessively leaky red cells between the time blood is drawn and electrolytes are analyzed.

Milder variants of hereditary stomatocytosis

These patients are similar to the classic disease described above, except that hemolysis and anemia are milder, the cells are less leaky, and stomatocytosis may be less pronounced. The osmotic fragility test may be normal or show abnormally fragile cells. Patients often present later in life for a work-up of mild anemia, macrocytosis, reticulocytosis, stomatocytosis, pseudohyperkalemia, gallstones, or a palpable spleen tip. Stomatin concentrations are normal. Where tested, these patients have had mutations in band 3 (see later section on “Pathophysiology”).

Hereditary cryohydrocytosis, type 1

This rare form of HSt is milder than the classical overhydrated form and has unique monovalent cation (that is, Na⁺ and K⁺) ion permeability properties. Patients have mild to moderate hemolysis, with or without anemia. Their red cells are mildly macrocytic and slightly osmotically fragile. They tend to be stomatocytic (see Figure 1). We and others have observed a tendency for the slit of the “stoma” to be curved and eccentrically placed, which we have not observed in other conditions. Several such cells are seen at the top of Figure 1 (cryohydrocytosis). However, this characteristic is not always present. The MCHC is variable and is not consistently low as in severe overhydrated stomatocytosis. The primary defect is in the anion transporter, band 3 (see section on “Pathophysiology”).

Na⁺ and K⁺concentrations and permeabilities are less abnormal at physiological temperatures in HC1 than in overhydrated HSt, but instead of declining with temperature, as in classical HSt, Na⁺permeability increases below 20ºC, exceeding the ability of the red cell to pump the leaked Na⁺back out. As a consequence, red cells from patients with HC1, collected in EDTA (ethylenediamine tetraacetic acid) or heparin, swell and partially hemolyze when stored in the refrigerator overnight. This is a requirement for the diagnosis of hereditary cryohydrocytosis and is its signature feature.

Patients with HC1 may present later in life with macrocytosis, anemia, reticulocytosis, intermittent jaundice, gallstones, a palpable spleen tip, stomatocytosis, or because cold hemolysis was noted by an astute laboratory technician.

Hereditary cryohydrocytosis, type 2

This disorder has only been observed in two families so far. The hematological picture is similar to HC1, or maybe a bit more severe, but the patients also have cataracts and a variety of neurological features including, seizures, mental retardation, and spastic rigidity or ataxia. The reported patients also had massive hepatosplenomegaly. Unlike the other forms of cryohydrocytosis, this type is also stomatin deficient. The primary defect is in a glucose/ascorbic acid transporter, Glut1 (see section on “Pathophysiology”).

Southeast Asian ovalocytosis

Southeast Asian ovalocytosis (SAO) is usually classified as a subtype of hereditary elliptocytosis (HE), but the elliptical red cells are more rounded than in typical HE and some have a bar across the middle of the elliptical-shaped central clearing, giving them a characteristic theta (θ) shape (Figure 1). Recent studies show that SAO red cells have the cation permeability properties of cryohydrocytes, including the cold sensitivity. The homozygous state is lethal. Heterozygotes have little or no hemolysis or anemia except in the neonatal period when jaundice and mild hemolysis and anemia are common. However, the mutation enhances resistance to cerebral malaria and, as a consequence, is very common in regions of Southeast Asia where malaria is rife, especially in the lowland aboriginal tribes of New Guinea and other areas of Melanesia. It is rare in other parts of the world. The primary defect is in band 3 (see section on “Pathophysiology”).

Hereditary xerocytosis (also called dehydrated hereditary stomatocytosis)

This is the most common of the inherited red cell cation permeability disorders. It is characterized by a mild increase in potassium permeability that is sufficient to lead to the gradual loss of red cell K⁺and water, and to red cell dehydration, stiffness, and hemolysis. HX also goes by the name “dehydrated hereditary stomatocytosis”. This is a very unfortunate name since most patient’s blood smears show few, if any, stomatocytes and the term leads to confusion with classical HSt, which is an entirely different disease. In fact, most patient blood smears are surprisingly normal (Figure 1), with a few target cells, some irregular spiculated cells, and rare stomatocytes.

Patients with HX may present at any age for evaluation of reticulocytosis, stomatocytosis, pseudohyperkalemia, jaundice, gallstones, or a palpable spleen tip. Hemoglobin and hematocrit values are often normal, despite evidence for mild to moderate hemolysis, suggesting that patients are physiologically more anemic than the numbers suggest. There is some evidence that viscous, dehydrated red cells do not traverse the kidney well as more supple, better hydrated red cells, leading to less oxygen reaching the juxtaglomerular apparatus, which would stimulate more erythropoietin production and a higher hemoglobin level than would be expected for the degree of hemolysis.

HX red cells have a reduced K⁺concentration and reduced total monovalent cations (Na⁺+ K⁺) (Table I). Red cell morphology is very unimpressive. There are usually a few target cells, occasionally a modest number, and sometimes a few echinocytes, a few stomatocytes, or some dense, contracted, irregular cells (Figure 1). Stomatocytosis is more dramatic in the rare patients with homozygous HX. The MCV ranges from high normal to macrocytic and the MCHC is typically elevated. Because the red cells are dehydrated, the osmotic fragility (OF) curve is always shifted toward osmotic resistance, though the effect can sometimes be subtle. The combination of macrocytic cells that are dehydrated (resistant OF and high MCHC) should strongly suggest the diagnosis of hereditary xerocytosis.

Curiously, some patients with HX develop non-immune hydrops in utero, or perinatal edema and ascites. The edema regresses spontaneously, either in utero or soon after birth. The cause of this complication and how it relates to the red blood cell permeability defect is a complete mystery. A constellation of clinical findings including anemia, jaundice, pseudohyperkalemia, fetal or perinatal edema or ascites, and even hydrops fetalis has been described in the fetal and newborn period. Different combinations of these features occur in different patients.

The mechanism of the edema and ascites is unclear. It doesn’t seem to be related to anemia or hypoalbuminemia. Although treatments such as intrauterine transfusions and albumin infusions have been employed, the efficacy of these interventions is difficult to assess, due to the spontaneous resolution of symptoms in late gestation and the early neonatal period.

The clinical course is also remarkable for iron overload in the absence of transfusions, even more so in patients who are heterozygous for a hemochromatosis gene. The serum iron, serum ferritin, and iron saturation are usually high in affected adults. This often requires treatment.

Pseudohyperkalemia

Patients with HX (and HSt and HC) may present as pseudohyperkalemia due to leakage of red cell K⁺into the serum after blood is drawn but before it is analyzed. This is a clinically benign condition but it must be distinguished from true hyperkalemia to prevent inadvertent treatment of patients and potential harm.

Beware of other conditions that can mimic hereditary stomatocytosis, hereditary cryohydrocytosis, or hereditary xerocytosis:

Hemolytic anemias that can be confused with hereditary stomatocytosis and xerocytosis include those that are inherited in an autosomal dominant fashion, macrocytic hemolytic anemias and hemolytic anemias with similar red cell morphology.

Autosomal dominant hemolytic anemias

• Hereditary elliptocytosis (HE)

• Hereditary spherocytosis (HS)

• Hereditary stomatocytosis (HSt)

• Hereditary xerocytosis (HX)

• Unstable hemoglobinopathies

The biggest risk is that HSt or HX will be confused with HS. This happens fairly frequently and is dangerous because splenectomy is commonly performed in patients with HS and is very dangerous in HSt and HX (see section “If you decide the patient has hereditary stomatocytosis, hereditary cryohydrocytosis, or hereditary xerocytosis, what therapies should you initiate immediately?”). HS and HSt both show osmotically sensitive red cells on an osmotic fragility (OF) test and patients with HSt can have some spherocytes on peripheral blood smears, but HS red cells are dehydrated (high MCHC) and have a low normal to slightly low MCV, while HSt red cells are macrocytic (or at least have a high normal MCV) and are waterlogged with a low MCHC. The combination of macrocytosis and a low MCHC is unique to hereditary stomatocytosis.

Similarly, both HS and HX have an abnormal osmotic fragility test but the red cells are osmotically sensitive in HS and osmotically resistant in HX. This is a critical distinguishing feature; however, in today’s computerized medical environment, the OF test is sometimes just reported as abnormal without an actual picture of the OF curve, which can easily lead to mistakes.

Macrocytic hemolytic anemias

• Hemolytic anemias with reticulocytosis and no red cell dehydration

• Hereditary stomatocytosis

• Immunohemolytic anemias

• Paroxysmal nocturnal hemoglobinuria

• Red cell enzyme disorders

• Unstable hemoglobinopathies

• Hereditary xerocytosis

Reticulocytes are much larger than mature red blood cells and hemolytic anemias with significant reticulocytosis are macrocytic, unless the pathophysiologic process also causes red cell dehydration, as is the case with hemoglobins S and C, HS and HX. Patients with all of the latter diseases, except HX have a low normal or slightly low MCV despite reticulocytosis. For unknown reasons, HX red cells have a high normal or high MCV, even though they are quite dehydrated (high MCHC and resistant osmotic fragility). This is a key to recognizing the disease.

HSt and HX are not easily confused with the other nonhemolytic macrocytic anemias (for example, bone marrow failure, marrow dysplasia, megaloblastic anemias, thyroid or liver failure, chronic blood loss), which are either acquired or present with relatively low reticulocyte counts and often with multiple cytopenias.

One confusing situation is when patients with HX present during an aplastic crisis, most often due to infection with parvovirus B19. The combination of anemia and a low reticulocyte count at that point, combined with unimpressive red cell morphology, can mask the diagnosis temporarily.

Stomatocytosis

• Acute alcoholism

• Drugs that partition selectively into the inner half of the lipid bilayer

• Hereditary cryohydrocytosis

• Hereditary stomatocytosis

• Mediterranean stomatocytosis and macrothrombocytopenia

• Rh_null and Rh_mod diseases

• Tangier disease

• Vinca alkaloids

All of these conditions have distinctive features that make them relatively easy to distinguish from HSt. Mediterranean stomatocytosis is probably the most easily confused. It is a fascinating disorder that is due to phytosterolemia, a metabolic defect in which absorption of all sterols (cholesterol and phytosterols from plants) is massively increased. The patients have mild to moderate hemolytic anemia, very prominent stomatocytosis, splenomegaly, normal red cell cations, a normal osmotic fragility test, and macrothrombocytopenia (40,000 to 150,000 platelets that are 3μm³ to 120μm³ in size).

Hemolytic anemias that often have relatively nonspecific red cell morphology

• Cold agglutinin disease

• Congenital dyserythropoietic anemias

• Embden-Meyerhof pathway (glycolytic) defects

• Erythropoietic or hepatoerythropoietic porphyria

• Hemolysis with infections (sometimes)

• Hereditary xerocytosis

• Hexose monophosphate shunt defects

• Paroxysmal nocturnal hemoglobinuria

• Rh hemolytic disease in newborns

• Thalassemia (heterozygous)

• Unstable hemoglobinopathies

• Vitamin E deficiency

• Wilson disease

Fortunately, other than unstable hemoglobinopathies, none of these conditions mimics the combination of high MCHC, macrocytosis, and osmotic resistance observed in HX. However, unstable hemoglobins and HX can easily be confused, since both diseases are dominantly inherited and both feature macrocytic red cells with relatively nonspecific and unimpressive morphology. In addition, patients with unstable hemoglobinopathies can have a small subpopulation of osmotically resistant red cells on osmotic fragility tests, though not to the degree usually seen in HX. In most unstable hemoglobinopathies, red cells show prominent basophilic stippling, which is not seen in HX, but this is not always the case.

Which individuals are most at risk for developing hereditary stomatocytosis, hereditary cryohydrocytosis, or hereditary xerocytosis:

These inherited disorders are all rare and none show any gender predisposition or any sensitivity to environmental or drug exposures.

Hereditary xerocytosis is the most common and is estimated to have a prevalence of about 1 in 50,000, which would make it roughly 50 times less frequent than hereditary spherocytosis. However, the disease is almost certainly underdiagnosed, probably to a considerable degree.

Hereditary stomatocytosis and hereditary cryohydrocytosis are both very rare.

All three diseases are inherited in an autosomal dominant pattern, but new mutations are fairly frequent.

What laboratory studies should you order to help make the diagnosis and how should you interpret the results?

Once the diagnosis of hemolytic anemia is established, the key readily available diagnostic tests to distinguish HSt, HC, and HX are:

• MCV and MCHC

• Blood smear for erythrocyte morphology

• Osmotic fragility test (unincubated)

The results of these tests are shown in Table I and are described in the section entitled “Beware of other conditions that can mimic hereditary stomatocytosis, hereditary cryohydrocytosis, and hereditary xerocytosis.”

More definitive tests include:

• Analysis of red blood cell Na⁺ and K⁺ content and passive permeabilities

• Analysis of red cell membrane stomatin content (markedly deficient in HSt and type 2 HC)

• Cold hemolysis (abnormal in HC)

Cold hemolysis means significant hemolysis (about 50%) of blood anticoagulated in EDTA or heparin and kept overnight at 4ºC. Cold hemolysis is ameliorated if the blood is stored in solutions containing impermeant solutes such as citrate-phosphate-adenine-dextrose, the anticoagulant used when blood is collected for transfusion.

• Osmotic gradient ektacytometry (OGE)

OGE is a test of the deformation of red cells under varying degrees of shear stress and osmolality. The test measures red cell deformability and surface-to-volume ratio (as in the osmotic fragility test), and gives indirect information about red cell hydration. It is a valuable test; however, only a few laboratories in the world own an ektacytometer and can do the test.

• Specific genetic testing for known causal protein defects (see Table I and the section entitled “Pathophysiology”)

What imaging studies (if any) will be helpful in making or excluding the diagnosis of hereditary stomatocytosis, hereditary cryohydrocytosis, or hereditary xerocytosis?

N/A

If you decide the patient has hereditary stomatocytosis, hereditary cryohydrocytosis, or hereditary xerocytosis, what therapies should you initiate immediately?

If you decide the patient has hereditary stomatocytosis or hereditary xerocytosis, what therapies should you initiate and which should you avoid?

Hereditary stomatocytosis

Patients with HSt vary from mild hemolysis and anemia to severe, life-threatening, transfusion dependent disease. In most cases, the disease is constant throughout life, though we have seen patients who had severe anemia in infancy that became milder with time during the first year of life. There are no reports of how often this occurs.

The swollen, water-laden stomatocytes have trouble negotiating the spleen, where the acidotic and hypoglycemic environment impairs their ability to synthesize adenosine-5′-triphosphate (ATP) and expel the inward torrent of Na⁺and water via the Na⁺, K⁺-ATPase pump. As a consequence, the spleen is a graveyard for stomatocytes and splenectomy is an effective, though incomplete therapy (that is, the anemia is only partially relieved). However, the risks of splenectomy are very high in this disease, as will be discussed, so much so, that the operation is contraindicated.

In most cases, patients with HSt have a tolerable degree of hemolysis and anemia and do not require any dramatic interventions, or at most, occasional transfusions for intermittent hemolytic or aplastic crises. In occasional patients, the anemia is severe enough (usually a hemoglobin level of 5 to 7g/dL or less) to slow growth, greatly decrease normal energy, or cause congestive heart failure, in which case the physician is faced with the unpalatable choice of chronic transfusions coupled with iron chelation therapy, splenectomy followed by life-long anticoagulation, or bone marrow transplantation. The propensity for iron overload in this disease further complicates the options for chronic transfusion therapy and bone marrow transplantation. There is insufficient information about the relative risks of these unappealing therapeutic options in HSt patients to offer rational advice on how to choose among them.

Hereditary cryohydrocytosis

Patients with HC type 1 generally have sufficiently mild hemolysis and anemia that they do not require transfusions. In the few instances where splenectomy has been performed, it has not been clearly beneficial. No long term complications of splenectomy have been observed, but the numbers are very small.

In patients with HC type 2, seizures and other neurological complications are an important part of the disorder. The primary defect is in the glucose transport channel, Glut1 (see section on “Pathophysiology”) and the neurological symptoms are believed to be due to inadequate glucose transport into the brain. In other patients with defects in Glut1 who lack the hemolytic component, there is evidence that the seizures improve on a ketogenic diet, suggesting that such a diet would be useful in HC2 patients as well.

Hereditary xerocytosis

Most patients with HX have only mild anemia and mild to moderate hemolysis and do not require any specific therapy for their hemolytic anemia. Many have no anemia at all, by conventional laboratory definitions, but this may be both artifactual and physiologically deceptive. The stiff xerocytes do not deform normally. This leads to an artifactually high MCV in Coulter-type counters, and a corresponding increase in the hematocrit, which is calculated using the MCV. However, HX patients have high reticulocyte counts and other evidence of hemolysis even when their hemoglobin levels appear normal, which suggests they are physiologically more anemic than these parameters indicate. In other words, the erythropoietic drive is still abnormally high, even when the hemoglobin level suggests it shouldn’t be. Possibly, the rigid xerocytes have difficulty traversing the small blood vessels that service the renal juxtaglomerular apparatus, leading to diminished oxygen delivery and elevated erythropoietin production, even at apparently “normal” hemoglobin levels.

Also, when measured, 2,3-diphosphoglycerate (2,3-DPG) concentrations have been low in HX (and HSt) red cells. This would increase hemoglobin oxygen affinity and further stimulate erythropoiesis. The bottom line is, physicians should be aware that patients with hereditary xerocytosis are probably more anemic, physiologically, than the hemoglobin and hematocrit numbers suggest.

Avoid splenectomy

Physicians who care for patients with either HSt or HX agree that there is a high risk of thromboembolic complications following splenectomy. So much so, that the operation is contraindicated in these diseases in all but extreme situations. Unfortunately, the data backing up this prohibition are not very extensive. The recommendation flows primarily from a single study, a retrospective compilation of case reports of nine splenectomized adults collected by Stewart and his colleagues in 1996.

All the patients were labelled as having “hereditary stomatocytosis” but they actually had a mixture of severe, overhydrated HSt and HX. Both groups suffered from serious thrombotic complications, sometimes recurrent over years, including deep venous thrombosis, pulmonary embolism, superficial thrombophlebitis, portal vein thrombosis, intracardiac mural thrombosis, arterial thrombosis, and pulmonary hypertension. Four of the nine patients died.

In addition, the author is aware of another splenectomized adult patient with HSt, who died of thromboembolic complications years after the operation. No such complications were observed in six additional unsplenectomizedadults with HSt or HX in the Stewart study.

There have been six additional reports of individuals with HSt or HX who developed severe thrombotic complications in the 18 years since this initial paper. Given the retrospective, incomplete, and anecdotal nature of all these reports, the lack of paired unsplenectomized controls, and other inherent flaws, it is impossible to estimate the precise risk of splenectomy in these two diseases, but it is apparent that it is high, and that splenectomy, which is only partially effective in HSt and ineffective in HX (and probably ineffective in HC), should be avoided. The reason(s) why thrombotic complications are so much higher after splenectomy in these diseases than in similar conditions like hereditary spherocytosis, is unknown.

We recommend that patients who have already been splenectomized, often following a mistaken diagnosis of hereditary spherocytosis, be placed on chronic Coumadin therapy and that they avoid contraceptives and, if possible, pregnancy.

More definitive therapies?

N/A

What other therapies are helpful for reducing complications?

Folic acid supplementation

All patients with moderate to severe hemolytic anemias should probably receive supplemental folic acid (1mg/d PO) to be sure that deficiency of this cheap and harmless water-soluble vitamin does not limit erythropoiesis.

Assess iron overload and treat if necessary

For unknown reasons, patients with hereditary stomatocytosis and xerocytosis are susceptible to iron overload, even if they have not been transfused. This seems to be true in most patients, suggesting it is not due to co-inheritance of a gene for hemochromatosis, though if that occurs, it increases the risk even more.

In some patients. the problem is severe and tissue iron deposition is great enough to damage the liver and other organs. For this reason, all patients with hereditary stomatocytosis or xerocytosis should have laboratory studies to assess iron status, with follow-up studies as indicated. If necessary, they should be treated with Exjade or other iron chelators. The diagnosis and treatment of iron overload is discussed in more detail elsewhere.

What should you tell the patient and the family about prognosis?

The natural histories of hereditary stomatocytosis and hereditary xerocytosis are not well understood. There are no systematic studies of the lifespan of patients who have not been splenectomized or about late complications of the diseases. As noted earlier, it is clear that splenectomy poses a high risk of thromboembolic complications and there seems to be a higher than expected risk of iron accumulation, which may require intervention. Also, these patients, like all patients with hemolytic anemias, are at relatively high risk for developing bilirubin gallstones and cholecystitis or biliary obstruction.

Overall, however, the red cell membrane permeability diseases are often relatively well tolerated, and most patients probably live reasonably normal lives.

What if scenarios.

N/A

Pathophysiology

Red cell water content is largely regulated by the changes in the intracellular concentration of monovalent cations (Na⁺and K⁺), since water molecules surround the ions in a hydration shell. A net increase in Na⁺ and K⁺ causes water to enter, forming “stomatocytes” or “hydrocytes,” whereas a net loss of these ions produces dehydrated red cells or “xerocytes.” So far, the primary defects in hereditary stomatocytosis, xerocytosis, and cryohydrocytosis are mutations the permeability properties of normal red cell membrane ion channels, rather than mutations that create new channels from nonchannel proteins. Not all the mutations are known yet, but this is probably a general rule.

Hereditary stomatocytosis

All of the patients with classical overhydrated hereditary stomatocytosis studied so far have mutations in the RhAG protein (rhesus associated glycoprotein, gene name RhAG). RhAG associates with the RhD and RhCE proteins that carry the Rh (rhesus) antigens to form the Rh complex. This complex resides in a larger membrane protein complex that includes the red cell adhesion proteins CD47 and LW, band 3 (the Cl^–-HCO3^– exchange channel), and glycophorin B, which serves as a chaperone to bring RhAG to the membrane.

RhAG has the typical structure of a protein channel, with 12 transmembrane helices. It is believed to function as an ammonium or CO₂(carbon dioxide) channel. So far, all hereditary stomatocytosis patients have mutations in one of just two conserved amino acids in the second transmembrane helix: Ile61→Arg or Ser65→Phe. The limited spectrum of causative mutations may explain the rarity of this disease. The mutations are thought to disrupt the channel structure and, in fact, the mutant RhAG proteins induce large cation leaks when introduced into oocytes.

HSt red cell membranes are remarkably permeable to Na⁺and K⁺ions, particularly Na⁺. Permeabilities as great as 15 to 40 times normal are observed. Because the influx of Na⁺exceeds the loss of K⁺, HSt red cells progressively gain cations and water and swell. In solution, the “edematous” stomatocytes are bowl-shaped, but when they deposit on slides the extra volume encroaches on the normally circular central clearing, yielding the characteristic slit shape (Figure 1). More swollen red cells appear as spherocytes (Figure 1). The swollen, water-laden stomatocytes are large (high MCV), osmotically fragile, and have a low density (low MCHC).

Monovalent cation transporters are greatly stimulated by the tremendous influx of Na⁺ions, particularly the Na⁺, K⁺ pump, and attempt to extrude the Na⁺torrent, consuming much ATP in the process. If they fail, stomatocytes would be expected to succumb to osmotic lysis, but the lack of evidence for intravascular hemolysis suggests they are somehow recognized and removed by the reticuloendothelial system before bursting. It is known that overhydrated stomatocytes are vulnerable to splenic sequestration since red cell metabolism and ATP production are compromised in the stagnant, acidotic, hypoglycemic splenic cords.

HSt erythrocytes are also moderately deficient in 2,3-diphosphoglycerate (2,3-DPG). The reason is unknown. Perhaps a portion of the 1,3-DPG normally used for 2,3-DPG synthesis is diverted through phosphoglycerate kinase to provide extra ATP for cation transport. The 2,3-DPG deficiency mildly enhances oxygen affinity and causes additional water entry and cell swelling.

Curiously, the red cell membranes of all patients with the classic overhydrated form of HSt nearly lack a 31-kD protein called stomatin or band 7.2b. This small protein resides in membrane lipid rafts and appears to regulate ion channels. For a time, defective stomatin was believed to cause the severe form of HSt. However, the stomatin gene is entirely normal in the disease and a stomatin gene knockout in mice does not cause any Na⁺leak or hemolysis in red blood cells.

Some HSt red cells contain reduced amounts of stomatin; however, in most HSt red cells it is lost early in erythropoiesis. A clever recent study showed that stomatin normally binds to the major glucose transporter, Glut1 (glucose transporter 1), and switches Glut1 to an L-dehydroascorbic acid transporter. This only occurs in species like humans that lack the ability to synthesize ascorbic acid. Because stomatocytic red cells consume so much ATP trying to pump out Na⁺, it may be that the HSt red cells that survive and circulate are those that are able to lose stomatin and retain Glut1 as a glucose channel, to provide extra metabolic substrate.

Patients with milder forms of HSt are known, and in the few investigated cases mutations in band 3 (gene name SLC4A1) have been discovered. Band 3 is the major erythrocyte membrane protein and is the anion channel responsible for shuttling chloride and bicarbonate ions across the membrane, which aids in the transport of CO₂ from tissues to the lungs. Quantitative deficiency of band 3 causes hereditary spherocytosis not stomatocytosis. In these stomatocytosis patients, normal amounts of band 3 are made, but the protein contains missense mutations within a portion of the transmembrane domain that is believed to form the anion channel. Presumably the mutant band 3’s reach the membrane, and allow some Na⁺ions to slip into the red cell during anion exchange. Normal anion exchange through band 3 is vastly greater than Na⁺or K⁺permeability across the entire red cell membrane (millions of times greater), so even a tiny inward Na⁺leak would be serious.

Hereditary cryohydrocytosis

Type 1 cryohydrocytosis (HC1) is phenotypically similar to the mild forms of HSt and is also caused by missense mutations in band 3 in the region of the protein thought to form the anion channel. At physiological temperatures the Na⁺ leak and hemolysis are modest, but below 20ºC the leak increases and exceeds the ability of the Na⁺, K⁺ pump to compensate, leading to the characteristic cold-induced swelling and hemolysis that give the disease its name. Why some band 3 mutations are cold sensitive and others are not, is a mystery. Presumably the conformational changes produced by the temperature-sensitive mutations worsen at low temperatures, but this has not been studied.

Interestingly, there are also patients with the phenotype of hereditary spherocytosis (HS) who have band 3 missense mutations and a low temperature cation leak. Perhaps these mutations destabilize band 3, leading to band 3 deficiency and HS, yet allow enough of the leaky band 3 to reach the surface to alter Na⁺permeability.

Southeast Asian ovalocytosis (SAO) is another disease that is related to cryohydrocytosis. The disease is caused by deletion of nine amino acids (numbers 400 to 408) in band 3 at the beginning of the transmembrane domain. The misfolded protein does not function in anion transport but does get inserted in the membrane and, for uncertain reasons, makes the membrane rigid. SAO red cells are abnormally permeable to Na⁺and K⁺and, like cryohydrocytes, the leak is more pronounced at low temperatures.

Type 2 cryohydrocytosis (CH2) or stomatin-deficient cryohydrocytosis is caused by a mutation in glucose transporter 1 or Glut1 (gene name: SLC2A1), which, as noted earlier, binds stomatin and is converted to a L-dehydroascorbate transporter in human red cells and other tissues. Like stomatocytosis due to RhAG mutations, the red cells are stomatin deficient. The mutations block glucose transport. This causes the characteristic neurological symptoms (seizures, mental retardation, and movement disorders). The hemolysis and cataracts appear to be due to the altered cation permeability of the mutant channel.

Many patients have been described with a similar neurological phenotype who are heterozygous for defects in Glut1 (so-called “Glut1 deficiency disorder”), but these patients do not have hemolysis or cataracts, and their Glut1 mutations do not increase Na⁺or K⁺permeability. They just block glucose transport. A single family is known with a Glut1 mutation that increases the permeability of Ca²⁺ as well as Na⁺and K⁺. The affected members also have hemolysis but their red blood cells are dehydrated due to Ca²⁺ (calcium) stimulation of K⁺egress (the so-called “Gardos channel”), and are echinocytic instead of stomatocytic.

Hereditary xerocytosis

HX is the most common of the membrane cation permeability defects. Because blood smears are unimpressive, the anemia is often fully compensated (that is, normal hemoglobin levels), and because red cell electrolyte measurements are hard to obtain, it is likely that many (probably most) patients go undiagnosed.

Physiologically, the major red cell abnormality is a change in the relative permeability of the membrane to potassium. Efflux of K⁺ is increased two to four fold and approximates or slightly exceeds Na⁺influx. Na⁺, K⁺ pump activity is increased appropriately for the slightly elevated Na⁺content, but the pump cannot compensate for K⁺losses in excess of Na⁺gain, because the pump expels three Na⁺ions for every two K⁺ions it returns, which is the normal ratio of the Na⁺_in and K⁺_out leaks. As a consequence, the K⁺permeability defect, though modest, causes HX red cells to gradually become cation depleted and dehydrated.

The defective gene has been traced to a relatively small segment on the long arm of chromosome 16 (q24.2-qter), and was recently pinpointed to a protein called Piezo1 (gene name FAM38A) in multiple HX kindreds. Missense mutations near the center or tail end of this very large protein have been found in all families studied to date.

There are two Piezo proteins (Piezo1 and Piezo2). They are widely distributed and form multimeric subunit cation channels that are triggered by mechanical distortion of the membrane. Piezo proteins are involved in pain sensing and perhaps in other mechanical sensations like hearing and touch. The presence of Piezo1 in the red blood cell membrane suggests they may also be involved in homeostatic adjustments to changes in volume. The Piezo1 mutations in HX lengthen the time it takes to activate the Piezo1 channel after membrane deformation and slows the time it takes to deactivate it. How these changes in channel function lead to K+ loss and cell dehydration in hereditary xerocytes is not yet clear. Perhaps the lost K+ ions simply flow out through the mutant Piezo1 channel. Alternatively, K+ loss may be a secondary consequence of excess Ca2+ entry through the mutant channel. It is known that Ca2+ enters normal red cells when they are deformed. It is likely that this occurs through Piezo1 and triggers Ca2+-stimulated K+-loss via the Gardos channel. Why defects in this critical channel in HX don’t have more widespread pathological effects, especially in homozygotes, is a mystery. Perhaps red cells only have Piezo1 while other cells have both Piezo1 and Piezo2, providing redundancy.

Hereditary xerocytes have an increased proportion of the phospholipid phosphatidylcholine (PC) in their membranes (12 to 20 femtomole/cell, normal range 10 to 12 femtomole/cell). Indeed, hereditary xerocytosis was given the name high phosphatidylcholine hemolytic anemia. Early studies suggested the excess PC was due to diminished transfer of PC fatty acids to phosphatidylethanolamine, a pathway that is normally stimulated by cellular dehydration. It is not clear why this pathway is inhibited in hereditary xerocytosis, or how it relates to the underlying membrane leakiness and hemolysis.

Xerocytes are also shear sensitive and are exceptionally prone to membrane fragmentation in response to metabolic stress. Similar shear sensitivity can be induced in normal red cells by dehydrating them. It is also observed in other conditions, like sickle cell disease, where red cell dehydration occurs. One consequence is that strenuous exercise can exacerbate hemolysis in hereditary xerocytosis.

Familial pseudohyperkalemia

This autosomal dominant condition maps to the same locus (16q23-q24) as hereditary xerocytosis and appears to be just a mild form of that disease. Patients have no significant hemolysis or anemia, though they may have macrocytosis. Because the red cells have a passive K⁺ leak that is exaggerated in the cold, electrolyte analyses of plasma obtained from blood samples stored for a few hours at room temperature or below may falsely suggest that the patient is hyperkalemic. The key clinical point is to recognize that the “hyperkalemia” is artifactual and check that the patient has a normal electrocardiogram, so that he or she does not mistakenly receive hyperkalemic therapy, which could cause dangerous hypokalemia.

Familial pseudohyperkalemia has also been mapped to chromosome 2q35-q36 in some families and traced to mutations in ABCB6, a protein that normally functions as a mitochondrial porphyrin transporter.

What other clinical manifestations may help me to diagnose hereditary stomatocytosis, hereditary cryohydrocytosis, or hereditary xerocytosis?

N/A

What other additional laboratory studies may be ordered?

N/A

What’s the evidence?

Gallagher, PG. “Disorders of red cell volume regulation”. Curr Opin Hematol. vol. 20. 2013. pp. 201-207. (The best recent review of the molecular defects and pathophysiology of the stomatocytosis and xerocytosis syndromes by one of the leading investigators in the field.)

Bruce, LJ, Guizouarn, H, Burton, NM. “The monovalent cation leak in overhydrated stomatocytic red blood cells results from amino acid substitutions in the Rh-associated glycoprotein”. Blood. vol. 113. 2009. pp. 1350-1357. (The original description of molecular defects in the RhAG protein in the severe form of overhydrated hereditary spherocytosis.)

Bruce, LJ, Robinson, HC, Guizouarn, H. “Monovalent cation leaks in human red cells caused by single amino-acid substitutions in the transport domain of the band 3 chloride-bicarbonate exchanger, AE1”. Nat Genet. vol. 37. 2005. pp. 1258-1263. (The original description of missense mutations in the transport domain of band 3 in patients with cryohydrocytosis, the mild form of hereditary stomatocytosis, and a rare form of hereditary spherocytosis with a low temperature cation leak.)

Flatt, JF, Guizouarn, H, Burton, NM. “Stomatin-deficient cryohydrocytosis results from mutations in SLC2A1: a novel form of GLUT1 deficiency syndrome”. Blood. vol. 118. 2011. pp. 5267-5277. (Identification of the molecular defect in Glut1 in stomatin-deficient cryohydrocytosis.)

Glader, BE, Fortier, N, Albala, NM, Nathan, DG. “Congenital hemolytic anemia associated with dehydrated erythrocytes and increased potassium loss”. N Engl J Med. vol. 291. 1974. pp. 491-496. (The first thorough description of hereditary xerocytosis (which the authors call hereditary desiccytosis) and its permeability defect.)

Lux, SE, Orkin, SH, Ginsburg, D, Nathan, DG, Look, AT, Fisher, DE, Lux, SE. “Disorders of the red cell membrane”. Nathan and Oski’s Hematology of Infancy and Childhood. 2015. (A very detailed and thorough recent review of the clinical and pathophysiological features of red cell membrane disorders.)

Miller, G, Townes, PL, MacWhinney, JB. “A new congenital hemolytic anemia with deformed erythrocytes (“stomatocytes”) and remarkable susceptibility or erythrocytes to cold hemolysis in vitro. I. Clinical and hematologic studies”. Pediatrics. vol. 35. 1965. pp. 9-6-915(First clinical description of cryohydrocytosis and the characteristic cold-sensitive hemolysis.)

McLean, D, Gray, E. “K-characteristic photon absorption from intensifying screens and other materials: theoretical calculations and measurements”. Med Phys. vol. 23. 1996. pp. 1253-1261. (The initial warning that splenectomy carries a high long-term risk of thromboembolic complication in patients with hereditary stomatocytosis or hereditary xerocytosis. The paper is flawed by its anecdotal and retrospective analysis, by the relatively small number of patients, and by lumping patients with hereditary stomatocytosis and hereditary xerocytosis (which the authors call overhydrated and dehydrated stomatocytosis, respectively) together as if they were the same condition. Nevertheless, the data are persuasive and this is still the best evidence that splenectomy is contraindicated in these diseases. The authors also note the high incidence of iron overload in these patients.)

Zarkowsky, HS, Oski, FA, Sha’afi, R. “Congenital hemolytic anemia with high sodium, low potassium red cells. I. Studies of membrane permeability”. N Engl J Med. vol. 278. 1968. pp. 573-581. (The first detailed study of the clinical and pathophysiological features of hereditary stomatocytosis and its permeability defect.)

Zarychanski, R, Schulz, VP, Houston, BL. “Mutations in the mechanotransduction protein PIEZO1 are associated with hereditary xerocytosis”. Blood. vol. 120. 2012. pp. 1908-1915. (Evidence in two kindreds with hereditary xerocytosis linked to the known locus on chromosome 16q, that the disease is caused by mutations in the PIEZO1 channel, encoded by the FAM38A gene. This channel mediates mechanotransduction in mammalian cells and is probably involved in erythrocyte volume homeostasis.])

Andolfo, I, Alper, SL, De Franceschi. “Multiple clinical forms of dehydrated hereditary stomatocytosis arise from mutations in PIEZO1”. Blood. vol. 121. 2013. pp. 3925-3935. (Important additional evidence that PIEZO1 mutations are responsible for hereditary xerocytosis.)

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