Haemoglobinopathies
Normal haemoglobin is composed of two alpha and two non-alpha globin chains. Alpha globin chains are produced throughout life, including in the fetus, so severe mutations may cause intrauterine death.Production of non-alpha chains varies with age; fetal haemoglobin (HbF-αα/γγ) has two gamma chains, while the predominant adult haemoglobin (HbA-αα/ββ) has two beta chains. Thus, disorders affecting the beta chains do not present until after 6 months of age.Defective haemoglobin(Qualitative)
Sickle cell anaemia Sickle-cell disease results from a single glutamic acid to valine substitution at position 6 of the beta globin polypeptide chain. It is inherited as an autosomal recessive trait Hemoglobin Beta Gene (HB-B) also known as Beta Globin is a protein that resides in the red blood cells. The HBB is 146 amino acids long and its molecular weight is 15,867 Daltons. The molecules of the hemoglobin are responsible to carry oxygen through the body. The HBB is found in part 15.5 of the chromosome 11. This leads to an amino acid theSickle cell anaemia
Hb S is insoluble and forms crystals when exposed to low oxygen tension.Deoxygenated sickle Hb polymerizes into long fibrils‘tactoids’. The abnormal haemoglobin C variant participates in polymerisation more readily than haemoglobin A, whereas haemoglobin F strongly inhibits polymerisationThe red cells sickle may block the different areas of the microcirculation or large vessels causing infarcts of various organs.CLINICAL MANIFESTATION
1-Painful vaso-occlusive crisis. Plugging of small vessels inthe bone produces acute severe bone pain.This affects areas of active marrow: the hands and feet in children (so-called dactylitis) or the femora, humeri, ribs, pelvis and vertebrae in adults2-Stroke. silent stroke occurs in 10–15% of children with sickle-cell disease. Children at risk of stroke can be identified by screening with transcranial Doppler ultrasound, with fast flow associated with increased stroke risk. 3-Sickle chest syndrome. This may follow a vaso-occlusivecrisis and is the most common cause of death in adult sickle-cell disease. Bone marrow infarction results in fat emboli to the lungs, which cause further sickling and infarction, leading to ventilatory failure if not treated..CLINICAL MANIFESTATION
4-Sequestration crisis. Thrombosis of the venous outflow from an organ causes loss of function and acute painful enlargement. In children, the spleen is the most common site. Massive splenic enlargement may result in severe anaemia, circulatory collapse and death. Recurrent sickling in the spleen in childhood results in infarction and adults may have no functional spleen. In adults, the liver may undergo sequestration with severe pain due to capsular stretching. Priapism is a complication seen in affected men. 5-Aplastic crisis. Infection of adult sicklers with human parvovirus B19 (erythrovirus) may result in a severe but self-limiting red cell aplasiaDIAGNOSIS
Peripheral blood smears : Patients with sickle-cell disease have a compensated anaemia,usually around 60–80 g/L. The blood film shows sickle cells,target cells and features of hyposplenism from a young age.A reticulocytosis is present.lls, target cells, Howell-Jolly bies,d opresence of HbS can be demonstrated by exposing red cells to a reducing agent such as sodium dithionite; HbA gives a clear solution,whereas HbS polymerises to produce a turbid solution.Haemoglobin ElectrophoresisDDD 9.7%, Hb-A2 = 3.3%)Howell Jolly Body
Erythroblast
Haemoglobin Electrophoresis (alkaline pH )
Hb CHb S
Moves in same position as Hb A2
HbA
Anode
Haemolysate applied
Cathode
δ δ α α α s s α α γ γ α Hb S
Hb A2Hb F
Genotype αααα βsβs δδ γγ Haemoglobins Produced :
Diagnosis: Hb SS Disease
Laboratory diagnosis of sickle cell anaemia made by presence of only Hb S, Hb A2, and Hb F on Hb electrophoresis with no Hb A, a positive sickling test and presence of sickle cells in blood film
Diagnosis of sickle cell anaemia Cont.
Haemoglobin Electrophoresis: Hb A 0 % Hb S 87.0 % Hb F 9.7 % Hb A2 3.3 % Both parents of the affected individual will have sickle-cell traitTREATMENT
Painful vaso-occlusive crisis 1.hydration 2.precepitating factors 3.oxygen therapy 4.analgesic 5.exchange transfusion *Antisickling agent (Hydroxyurea) increase Hb F reduce sickling. *Bone marrow transplantation(Allogeneic-BMT). * Gene therapy.Treatment –cont. A regular transfusion programme to suppress HbS production and maintain the HbS level below 30% may be indicated in patients with recurrent severe complications, such as cerebrovascular accidents in children and chest syndrome in adult. Exchange transfusion,in which a patient is simultaneously venesected and transfused to replace HbS with HbA, may be used in life-threatening crises or to prepare patients for surgery.
The thalassemia syndromes
Thalassemia refers to a spectrum of disorders characterized by reduced or absent production of one (or, rarely, two or more) of the globin chains, thus disrupting the delicate balance between the production of alpha and non-alpha (eg, gamma or beta) globin chains. The two most common forms are: Alpha thalassemia Beta thalassemia These imbalance in chain synthesis lead to formation of unstable Hb. or decrease Hb. lead to hypochromic microcytic anaemia .THALASSEMIA
Pathophysiology of thalassemiaAlpha thalassemia – Alpha thalassemia is due to impaired or absent production of alpha globin chains, which leads to a relative excess of gamma globin chains in the fetus and newborn, and excess beta globin chains in children and adults. While the excess beta globin chains are capable of forming soluble homotetramers (beta-4, HbH), they are unstable and some precipitate within the cell, leading to a variety of clinical manifestations. Homozygous alpha (0) thalassemia, in which no alpha globin chains can be produced, is incompatible with extrauterine life, leading to hydrops fetalis and death shortly after deliveryPathophysiology of thalassemia-Cont.
Beta thalassemia – Beta thalassemia is due to impaired production of beta globin chains, which leads to a variable excess of alpha globin chains. For reasons that are not entirely clear, excess alpha globin chains cannot form soluble homotetramers and begin aggregating as soon as they accumulate in erythroid precursors, producing insoluble aggregates even in very early marrow erythroid precursors, affecting membrane assembly and accelerating programmed cell deathIneffecive erythropoiesisPathophysiology of thalassemia-Cont
A critical factor underlying the difference in pathophysiology between alpha and beta thalassemia is that, in the alpha thalassemias, the excess beta or gamma globin chains can form partially soluble but ineffective hemoglobin homotetramers. These homotetramers do not precipitate extensively until they are exposed to damaging effects in the circulation, mostly oxidant in nature. The protection against severe malaria may be mediated by increased susceptibility to infection with the nonlethal P. vivax, particularly in young children, thereby inducing limited cross-species protection against subsequent P. falciparum infectionClinical manifestations of thalassemia
Clinical manifestations of thalassemia
δ δ α α α γ γ αHb A2
Hb F
Diagnosis: β Thalassemia major
Genotype αααα - - δδ γγ Haemoglobins Produced
Laboratory diagnosis of β thalassemia major made by CBC, absence of Hb A, with increased Hb F. Some patients have small amounts of Hb A if some β globin chain is produced.
α-THALASSAEMIAS DEFINITIONS each individual normally carries four functional alpha chain genes (ie,aa/aa).Alpha (0) thalassemia Heterozygous alpha (0) thalassemia that result in the deletion of both alpha chain loci on one chromosome 16 (ie, aa/--)Alpha (+) thalassemia; ●Inheritance of three normal alpha genes (aa/a-) has been termed alpha thalassemia minima, silent carrier of alpha thalassemia, alpha thalassemia-2 trait ●Inheritance of two normal alpha genes has been termed alpha thalassemia minor or alpha thalassemia-1 trait, and is due either to heterozygosity for alpha (0) thalassemia (aa/--) or homozygosity for alpha(+)thalassemia (a-/a-).●Inheritance of one normal alpha gene (a-/--) is termed hemoglobin H (HbH) disease, because of the formation of HbH, which is composed of tetramers of the resulting excess beta chains (beta 4). ●inheritance of no normal alpha genes (--/--) is incompatible with extrauterine life, since the affected fetus will be unable to make any of the hemoglobins normally produced after birth (eg, hemoglobins A, F, and A2), all of which require the presence of alpha globin chains('Hydrops fetalis )
Clinical features
Alpha (0) thalassemia Affected subjects are clinically normal and may also be hematologically normal; the diagnosis can be reliably made only via DNA analysis.●Inheritance of two normal alpha ; These subjects are clinically normal but may have minimal anemia along with reductions in mean corpuscular volume (MCV) and mean corpuscular hemoglobin (MCH). ●Hb-H:These patients have moderate to severe degrees of lifelong hemolytic anemia, very modest degrees of ineffective erythropoiesis, splenomegaly, variable bony changes, and iron overload ●No normal alpha genes (--/--) ;This condition is incompatible with extrauterine life ('Hydrops fetalis and hemoglobin Bart's' )DIAGNOSIS OF α-THALA Positive family history with lab finding non specific findings : Blood smear show microcytic, hypochromic red cells, target cells, aniso-poikilocytosis, and decrease MCV. Heinz bodies are evident. Specific findings : definitive diagnosis is finding of HbH by Hb electrophoresis.
Normal Control
Abnormal ControlAbnormal Control
Patient Hb H
HbA
HbA2
Hb –H Diagnosis Haemoglobin Electrophoresis: Hb A 91.5 % Fast moving band 8.5% Hb A2 and Hb F decreased
Hb H Preparation
Hb H inclusions in RBCsDiagnostic features of beta-thalassaemia
Beta-thalassaemia major (homozygotes) • Profound hypochromic anaemia • Evidence of severe red cell dysplasia • Erythroblastosis • Absence or gross reduction of the amount of haemoglobin A • Raised levels of haemoglobin F • Evidence that both parents have thalassaemia minor Beta-thalassaemia minor (heterozygotes) • Mild anaemia • Microcytic hypochromic erythrocytes (not iron-deficient) • Some target cells • Punctate basophilia • Raised haemoglobin A2 fractionTREATMENT OF α- AND Я- THALASSEMIA 1.Regular red cell transfusions : hypertransfusion program (keep the level of Hb>100g/L) 2.Neocytes transfusion (increase RBC survival, decrease frequency of transfusion, and decrease iron over load). 2-3 units every 4-6 weeks . 3.Leukocyte filter will lowers rate of transfusion reaction. 4.Folic acid supplementation (5 mg) to prevent aplastic crisis.
5.Iron chelation: *Desferioxamine (Desferal) either with each unit of transfused blood (2 g) or by slow subcutaneous daily infusions by pump ( (1-4g over 8-12 *(Exjade ) Deferasirox *Deferiprone 6.Splenectomy ;indication: mechanical difficulty,hypersplenism. 7.Bone Marrow Transplantation : prior to development of hepatomegaly, portal fibrosis & iron over load.
TREATMENT –cont.
Treatment of beta-thalassaemia major
Problem Management Erythropoietic failure Allogeneic HSCT from HLA- compatible siblingTransfusion to maintain Hb > 100 g/L Folic acid 5 mg daily Iron overload Iron therapy contraindicated Iron chelation therapy Splenomegaly causing mechanical problems, Splenectomy excessive transfusion needs
PRENATAL DIAGNOSIS OF THALASSAEMIA
Guide parents and physicians in deciding whether to complete pregnancy.Both parent carriers. Fetal diagnosis:fetoscope to sample fetal venous blood show α/Я chain synthesis ratio.Amniocentesis or trophoblast (chorionic villus) biopsy for DAN analysis using DNA probes.