In order to make the diagnosis, the clinician must consider the ethnicity, family history, clinical presentation, physical exam as well as laboratory parameters. The laboratory diagnosis of the thalassemia syndromes has traditionally been based on careful interpretation of the complete blood count, including the red cell indices, the review of the peripheral blood smear, and characterization of the type of Hb produced. Hb levels vary based on the severity of the underlying genotype, but the anemia is always microcytic with low red cell indices. The smear shows variable hypochromia and the presence of abnormal shapes and sizes of cells, all more so in individuals with increasing severity of anemia. There is a reduction or absence of production of HbA, and thus an increase in HbF and HbA2 on Hb fractionation. Special staining, α -, β -, and γ -globin assessments are performed in some sophisticated laboratories, along with some degree of genotyping, especially for known mutations. In the developed world, it is now possible to characterize the disease more completely, including all of the possible genetic modifiers with the easy availability of sophisticated DNA testing, including whole exome or genome sequencing. While this is not routinely done for all patients, especially those with mild clinical manifestations, in more complex cases, it may be used to fully explain the clinical picture. There may also be some predictive value in genotyping for some of the newer therapies, as will be described later. Family members should also have testing if they have not been tested previously.

The diagnosis is usually fairly easy to make, though the following may be considered if it is not immediately clear:

 • congenital sideroblastic anemia, one of the rarer instances when a BM exam or specific genetic testing may be needed to make the diagnosis

• juvenile chronic myelogenous leukemia may be superficially considered, because of the elevated HbF levels, but other testing is quite straightforward to make this diagnosis

Homozygous β0 Thalassemia

Infants are born with relative anemia and microcytosis. The Hb level decreases progressively during the first months of life. When the child becomes symptomatic, the Hb level may be as low as 3 to 4 g/dL. RBC morphology is strikingly abnormal, with many microcytes, bizarre poikilocytes, teardrop cells, and target cells (Fig. 1). A characteristic finding is the presence of extraordinarily hypochromic, often wrinkled and folded cells (leptocytes) containing irregular inclusion bodies of precipitated α -globin chains. Nucleated RBCs are frequently present in peripheral circulation. The reticulocyte count is 2% to 8%, lower than would be expected in view of the extreme erythroid hyperplasia and hemolysis. The low count reflects the severity of intramedullary erythroblast destruction. The white blood cell count is elevated. A moderate polymorphonuclear leukocytosis and normal platelet count are typical unless hypersplenism has developed. The BM (not necessary for diagnosis) exhibits marked hypercellularity caused by erythroid hyperplasia. The RBC precursors show a maturational arrest with defective hemoglobinization and reduced amounts of cytoplasm.

Fig1. MORPHOLOGIC APPEARANCE OF THE PERIPHERAL BLOOD FILM IN A CASE OF SEVERE β-THALASSEMIA. Note the bizarre cells, the hypochromia, nucleated red blood cells, target cells, and polychromasia.

The osmotic fragility is strikingly abnormal. The RBCs are so markedly resistant to hemolysis in hypotonic sodium chloride solution that some are not entirely hemolyzed even in distilled water. Even before transfusion therapy is initiated, the serum iron and transferrin saturation and serum ferritin may already be increased as a result of increased iron absorption, thus ruling out iron deficiency as an alternate diagnosis.

The Hb profile reveals predominantly HbF. In patients with homozygous β 0-thalassemia, HbA is absent throughout life, unless transfused. In β +-thalassemia HbA may be very low or undetectable in the newborn but present in reduced amounts in later life. The levels of HbA 2 in thalassemia major are variable, probably because of increased numbers of F cells that have a decreased HbA 2 content.  Other biochemical abnormalities of the RBC in cases of thalassemia major include a postnatal persistence of the RBC in antigen and a decrease of RBC carbonic anhydrase; these findings are probably also caused by the elevated levels of circulating F cells.

The intraerythrocytic inclusions in the peripheral blood cells of patients with thalassemia, termed Fessas bodies, are especially prominent after splenectomy. These inclusions, best seen by staining with supravital staining (Brilliant Cresyl Blue) or by phase microscopy, are aggregates of precipitated, denatured α -chains. They are also found in large numbers within erythroid precursors in the BM.

RBC survival in cases of thalassemia major is variable but usually markedly decreased. The 53Cr half-life ranges between 6.5 and 19.5 days compared with the normal half-life of 25 to 35 days,  but there is some variability with cells containing more fetal Hb surviving longer. Increased plasma iron turnover and poor use of radiolabeled iron provide further evidence of IE.

Unconjugated bilirubin levels are elevated, in the range of 2.0 to 4.0 mg/dL at the time of diagnosis but may rise substantially as the anemia worsens in the absence of transfusion. Serum aspartate aminotransferase levels are frequently increased at diagnosis because of hemolysis. Alanine aminotransferase levels are usually normal before transfusion therapy but may rise subsequently because of iron induced hepatic damage or viral hepatitis. Lactate dehydrogenase levels are markedly elevated as a consequence of IE. Haptoglobin and hemopexin are reduced or absent.

Hypersplenic patients may have low white cell and platelet counts, and after splenectomy, leukocytosis and thrombocytosis are common. Asplenic individuals usually have large, flat “pseudo-macrocytes,” or leptocytes that appear larger in two dimensions because they are so devoid of Hb that they collapse flat on the slide like a deflated balloon, and small, deformed microcytes, as well as markedly increased nucleated red cell counts, and the white cell count frequently has to be corrected for their presence. Staining of the blood with methyl violet, particularly in splenectomized subjects, reveals stippling or ragged inclusion bodies in the red cells.

When transfusions are initiated, the Hb level rises to the mild to moderate anemia range, being higher in patients on regular transfusions and generally lower in those receiving on-demand transfusions. The mean corpuscular volume (MCV) and Hb fractionation reflects transfused blood as well. Iron parameters are altered to reflect iron overload as transfusions continue. Serum iron and transferrin saturation, as well as ferritin levels are elevated, and remain so, with the ferritin level declining in response to chelation therapy.

Transfusion complications include the development of auto- or alloantibodies with a positive DAT and antibody screen, iron-related organ dysfunction, hepatic, and endocrine. All of these parameters must be regularly monitored in transfusion-dependent patients.

Homozygous or Compound Heterozygous β+ Thalassemia

 In these individuals, the Hb levels are generally higher, but otherwise the hematologic changes are similar. The more severe the anemia, with greater IE, the more the peripheral smear resembles that of a thalassemia major patient prior to starting transfusions, as noted above. Examination of the BM, if performed, would confirm IE to a variable degree, with more severe patients having the picture of untreated thalassemia major. Hb fractionation shows a variable amount of HbA depending on the specific genotype, and the rest is HbF and A2, or in cases of HbE-thalassemia, a proportion of HbE. Even in the absence of transfusions, iron studies will show a steady rise in serum iron, transferrin saturation, and ferritin as a result of increased intestinal absorption, the rate of rise determined by the degree of IE. With intermittent or regular transfusions, the iron levels will rise more rapidly, and follow the same course as individuals with thalassemia major.

Heterozygous β-Thalassemia

Inheritance of a single β -thalassemia allele usually results in a mild anemia. The classic picture of β -thalassemia minor is a mild microcytic anemia (Hb averages 1 or 2 g/dL lower than that seen in normal persons of the same age and gender, typically 9 to 11 g/dL, MCV 50 to 70 fL). The red cell count is usually normal or elevated. HbF levels decline more slowly than usual in the first year of life, and the diagnostic elevated HbA 2 levels are established by approximately 6 months of age. 59–61 Strong intrafamilial correlations of both HbA 2 and MCV are noted. 62,63 The HbA2 level is increased to 3.5% to 7%. The level of fetal Hb is elevated in approximately 50% of cases, usually to 1% to 3% and rarely to greater than 5%. The RBC count is increased or normal. The RBCs are characteristically hypochromic (mean corpuscular hemoglobin [MCH] < 26 pg). Osmotic fragility is decreased; indeed, a one-tube osmotic fragility test has been used in the past for mass screening. 60 The smear shows varying numbers of target cells, poikilocytes, ovalocytes, and basophilic stippling (Fig. 2). The reticulocyte count is normal or slightly elevated. RBC survival is normal, iron utilization is decreased, and slight IE is present.  Iron studies are normal, unless concomitant iron deficiency is also present, in which case the transferrin saturation and ferritin are low. In this situation, the Hb is lower, as are the red cell indices, and the red cell count. The BM (not usually done) in heterozygous β -thalassemia shows slight erythroid hyperplasia with rare red cell inclusions. Megaloblastic transformation as a result of folic acid deficiency occurs occasionally, particularly during pregnancy. A mild degree of IE is noted, but red cell survival is normal or nearly normal.

Fig2. MORPHOLOGY OF THE PERIPHERAL BLOOD FILM IN A PATIENT WITH HETEROZYGOUS β-THALASSEMIA (A) AND A PATIENT WITH HETEROZYGOUS α-THALASSEMIA (B). Note the profound hypochromia and microcytosis and the many target cells. (From Pearson HA, Benz EJ Jr. Thalassemia syndromes. In: Miller DR, Baehner RL, McMillan CW, eds. Smith’s Blood Diseases of Infancy and Childhood. 5th ed. St. Louis: CV Mosby; 1984:439.)

β-Thalassemia With Normal A2 Level

Rare forms of β -thalassemia are seen in which heterozygotes have a completely normal hemogram, or with mild microcytic anemia but with a completely normal Hb pattern on electrophoresis or high-pressure liquid chromatography (HPLC). This can be confused with the more common heterozygous α -thalassemia and may cause difficulties in genetic counseling and prenatal diagnosis. Based on hematologic studies, two main classes of “normal HbA2 β -thalassemia”—some times called types 1 and 2—are seen.

Type 1, the “silent” form of β -thalassemia, 2 is characterized in heterozygotes by a normal hemogram—there is no anemia, and the MCV is normal. Although this condition can be partly identified by demonstrating a mild degree of globin-chain imbalance, with α -to- β synthesis ratios of approximately 1.5:1, it can only be diagnosed with certainty by DNA analysis. Compound heterozygotes for this condition and β 0-thalassemia have a mild form of β -thalassemia intermedia.

Normal HbA2 β -thalassemia type 2 in heterozygotes has a hematologic profile of mild microcytic anemia which is indistinguishable from typical β -thalassemia with elevated HbA2 levels.  The homozygous state has not been described. The compound heterozygous state for this gene and for β -thalassemia with raised HbA2 levels is characterized by a clinical picture of severe transfusion-dependent β -thalassemia.

Dominant β-Thalassemia

 The clinical features of the dominant β -thalassemias resemble the features of thalassemia intermedia. The blood picture shows the usual thalassemia changes, and there is moderate anemia and splenomegaly. The marrow shows erythroid hyperplasia with well-marked inclusion bodies in the red cell precursors, which may be seen in the peripheral blood after splenectomy. Hb analysis shows HbA and elevated HbA2, but the HbF level is not usually elevated much higher than that seen in β -thalassemia heterozygotes.

اخر الاخبار

اخبار العتبة العباسية المقدسة

عبر ممثلية مواكب بابل.. قسم الشعائر يشارك في حملة إغاثة الشعبين الإيراني واللبناني

المجمع العلمي يواصل تنفيذ برنامج قراءة صفحة من القرآن الكريم في صلاح الدين

قسم شؤون المعارف: موسوعة مؤلفات الإمامية تُعدّ أحد أبرز المشاريع الموسوعية التي أنجزناها

قسم الشؤون الفكرية يباشر بتوزيع عدد من إصدارات العتبة العباسية المقدّسة في محافظة واسط

المزيد

الآخبار الصحية

عدد مرات التبرز يكشف أسرارا عن صحة الأمعاء

علامات جينية طويلة الأمد تتركها الرضاعة الطبيعية في دم الطفل

هل يمكن إعادة برمجة الجهاز المناعي لإنتاج أجسام مضادة نادرة؟

دراسة: أدوية الزهايمر "لا تحدث فرقا يذكر لدى المرضى"

المزيد

الآخبار التكنلوجية

روسيا تطور تقنية جديدة لتعديل خصائص البلاديوم

الصين: طاقم مهمة شينتشو-21 سيبقى في الفضاء شهرا آخر

"أبل" تهدد بحذف تطبيق "غروك" من متجرها وتكشف السبب

"أبل" تهدد بحذف تطبيق "غروك" من متجرها وتكشف السبب

المزيد

مواضيع ذات صلة

Complement Fixation Assays

Flocculation Tests

Particle Agglutination Tests

Direct Whole Pathogen Agglutination Assays

Separating IgM From IgG For Serologic Testing

Serodiagnosis of Infectious Diseases

TSH- Receptor Antibodies

Thyroglobulin and Thyroid Peroxidase Antibodies

Interpretation of Serologic Tests

Features of the Humoral Immune Response Useful in Diagnostic Testing

Enzyme Immunoassays

Immunofluorescent assays