Hematology and Immunology A concise update Adlette Inati MD Pediatric Hematology Oncology Lebanese American University and Rafik Hariri University Hospital

Lebanese University Oct 25, 2013 Learning Objectives Concisely Review Anemia Hematopoietic and Stromal Cell Differentiation Erythropeisis Main players

Pluripotent stem cells

Chemical regulators  Cytokines  Erythroid specific growth factor  Erythropoietin (EPO)  Thyroid hormones, estrogens and androgens

Nutritional requirements  Proteins and amino acids

 Vitamin B12 and folic acid  Vitamin B6 (pyridoxine)   Other nutrients (Vitamin C, cupper, cobalt) Anemia Diagnosis

 History  Physical Examination  Blood count  Blood smear  Reticulocyte count Medical History

 Sudden onset of pallor, fatigue or exercise intolerance?

 Blood loss, easy bruisability?

 Cold intolerance, constipation, lethargy, poor growth

 Recent drug or fava bean intake or viral ?

 Neonatal jaundice or episodes of jaundice or dark urine in the past?

 The child’s diet? Any iron supplements? Any history of pica?

 Any history of recurrent bone or joint pain or diarrhea? Family History

 Anemia or low blood counts?

 Excessive bleeding, transfusion requirements ?

 Splenectomy ,cholecystectomy, hystereectomy at an early age?

 Autoimmune ?

 Consanguinity?

 Ethnic / geographical origin? Physical Examination

 Is the child active and playful or fatigued?

 Look for pallor, signs of heart failure, icterus, jaundice, glossitis

 Look for petechiae, , splenomegaly, pathologic lymphadenopathy, abdominal masses, skeletal abnormalities and abnormal facies Complete blood count (CBC)

Automated counter  Provides a panel of tests:

 Red Blood Cells (RBCs)  Number of RBCs per volume of blood  (Hb) – amount of Hb in the blood  Red cell Indices

 White Blood Count (WBCs)  Number of white blood cells per volume of blood  WBC differential - types of WBCs present

 Platelets (Plts)  Number of platelets per volume of blood  MPV Normal values are age and sex dependent Blood Smear

 Shows specific morphologic abnormalities  Reveals polychromatophilia which is a reflection of reticulocytosis  Recognizes abnormal cell morphology Reticulocyte Count An Underutilized Parameter

Immature mononucleated RBCs indicating active erythropoiesis

Elevated in blood loss or

Low to NL in bone marrow failure or ineffective erythropoiesis

Oski FA, Brugnara C, Nathan DG: A diagnostic approach to the anemic patient. In: Nathan and Oski’s Hematology of Infancy and Childhood. 5th ed. Harcourt Health Sciences; 1998:375-384 Polycythemia Anemia Lymhocytes(Increased Hb) (Decreased Hb) Decreased blood volume (dehydration, burns, diuretics) Decreased production

Increased RBC mass (polycythemia) Increased destruction - Chronic hypoxia (hemolysis) - Erythropoetin secreting Tumors - Primary (polycythemia vera)

Blood loss -

Sequestration Anemia Diagnostic Algorithm

History, Physical exam Hb, WBC, Platelets Microcytic Red cell indices Macrocytic Reticulocyte count Blood film Iron deficiency Folic acid deficiency Lead poisoning Vit. B12 deficiency Sideroblastic anemia Normocytic Hypothyroidism Unstable Liver Anemia of chronic Drugs disease

Low Reticulocyte count Nl reticulocyte count High reticulocyte count R/O Bone Marrow Chronic R/o hemorrhage Failure/Red Cell Aplasia Adlette Inati. Iron curriculum R/O hemolysis ESH, Webcast 2010 Iron-deficiency anemia (IDA) • • • • • • • • • •

• absent globinchainsynthesis Inherited anemias characterizedby Severe transfusion dependent Severe dependent anemia transfusion If Significant Significant Mild anemia, diagnosed usually in late childhood inlate usually diagnosed Mild anemia, Occasional blood transfusions may be required be may transfusions Occasional blood impaired moderately production chain Globin Various interactions genetic May require genetic counseling counseling May genetic require Asymptomatic Heterozygous condition Homozygous disorder Homozygous untreated, leads to death usually in usually death tountreated, leads imbalance of imbalance b a globin globin chains first decade decade first presenting early life early presenting in b b b d - - - intermedia thalassemia thalassemia thalassemia ecreased minor major or

Severity of disease Thalassemia Diagnosis Beta thalassemia blood smear Normal blood smear

Lab diagnosis

Low Hb / Hct Low MCV / MCH High HbF / HbA2 β -Thalassemia Major Management

Several ongoing thalassemia trials in Lebanon and the ME Macrocytic Anemias

 Large RBC s with high MCV > 100  Common in adults  Hypersegmented neutrophils

Megaloblastic: Impaired DNA synthesis and assembly resulting in delayed nuclear maturation with normal cytoplasmic development Vit B12 deficiency Folate deficiency Drug induced Myelodysplasia Non-megaloblastic: abnormal RBC maturation Alcohol Liver disease Hypothryroidism chemotherapy/drugs Megaloblastic anemia Symptoms

Folic Acid Vitamin B12 deficiency deficiency No neurologic symptoms Neurologic symptoms Pallor, lethargy, weakness Anorexia, diarrhea Light headedness Papitations, angina Macrocytic anemias Hemolytic anemias

 A group of inherited or acquired disorders characterized by premature RBC destruction and inability of the marrow to have sufficient erythrocyte production to keep up with accelerated RBC destruction, thus producing anemia

 Presentation ranges from an asymptomatic to a severe life threatening state with angina and cardiopulmonary decompensation in severe cases

 Usual symptoms and signs: pallor, lethargy, jaundice, icterus and dark urine, fever and sometimes back and abdominal pain Classification of Hemolytic Anemias Intra versus extra vascular hemolysis Intravascular Hemolysis Extravascular Hemolysis

Severe clinical course Mild to moderate course

↑ Free plasma hemoglobin No free plasma hemoglobin

↑ Free urinary Hb / hemosiderin Absent urinary Hb / hemosiderin

Schistocytes Spherocytes

Decreased serum haptoglobin Normal serum heptaglobin

↑ Indirect bilirubin reticulocytosis Urobilinogen in stool and urine Intra versus extra corpuscular hemolysis

INTRACORPUSCULAR EXTRACORPUSCULAR

•Membrane abnormalities •Immune Hereditary spherocytosis •Allo immune •Autoimmune •Enzyme abnormalities •Non Immune G6PD deficiency •Microangiopathic (TTP) •Infection(malaria) •Chemical agents (spider venom •Hb abnormalities / Thalassemia Membrane Abnormalities Hereditary Spherocytosis Enzyme abnormalities Favism

Adlette Inati, et. J Med Screen. 2012;19 (2):103-4 Hemoglobinopathies Pathophysiology of SCD

β-Globin Gene

Glutamic acid HbS Valine (ɑ2βˢ2) Deoxygenated Hb polymers Low oxygen tension

Sickle erythrocyte

Haemolysis Vaso occlusion

Pain, acute chest syndrome, joint , Anaemia, gallstones, priapism, stroke acute splenic sequestration, increased leg ulcers, pulmonary hypertension , multi-organ damage

Adapted from Inati A, et al. Pediatr Ann. 2008;37:311-25. SCD diagnosis SCD blood smear Normal blood smear

HPLC. Sickle Cell Disease Preventive strategies

Neonatal Screening, Penicillin prophylaxis and Vaccination

Disease Modifying Comprehensive Therapies Care and family (hydoxyurea, transfusions education and stem cell transplant)

Transcranial Screening for doppler pulmonary hypertension screening for

stroke risk Inati A etal. Pediatr Ann. 2008;37:311-25 Vichinsky et al. Pediatrics 1988; 81: 749-755 Sickle cell disease in Lebanon

Evelyne Khoriaty etal. & Adlette Inati. EHA 2012, Abstract no. 2021 Hemoglobinopathies Treatment Options in Development Treatment Option Clinical Effect Arginine es production of nitric oxide a potent vasodilator Decitabine es HbF

Butyrates short-chain FAs that inhibit histone deacetylase (HDAC), HbF production Clotrimazole Inhibits cation transport channels in RBC membranes thereby reducing cellular dehydration Gene Therapy Clinical evidence not yet available. Early promise shown in a transgenic mouse model

Several ongoing trials in Lebanon on HbF modulation Bone marrow failure syndromes (BMFSs) Bone marrow failure syndromes Hereditary Acquired

Diamond Blackfan anemia Idiopathic

Fanconi’s anemia Drugs: chemotherapy, chloramphenicol, benzene, sulfas…. Dyskeratosis congenita Radiation, chemicals Schwackman – Diamond anemia Viruses: EBV, HIV, parvovirus, hepatitis

Amegakaryocytic thrombocytopenia Malnutrition

Thrombocytopenia with absent radius Paroxysmal nocturnal hemoglobinuria syndrome (PNH)

Severe congenital neutropenia Disorders of immune system

Myelodysplastic syndromes (MDS) Bone marrow failure syndromes Symptoms Bone marrow failure Treatment Options

Hematopoetic growth Bone Marrow factors Transplant

Immune Suppressive Supportive Care Therapy White blood cells disorders

Benign disorders Proliferative disorders

 Quantitative  Leukemia  Leukopenia  Leukocytosis  Lymphoma

 Qualitative abnormalities  Myeloma Leucocytosis Leucopenias Increased WBCs Decreased WBCs

 Infections  Bone marrow failure  and or suppression  Cancers  Increased (leukemias….) destruction  Shift of neutrophils  Sequestration from marginating  Shift of neutrophils pool to circulating from CP to MP pool (margination) Neutropenia

Classification Neutrophil Count

Mild 1.0- 2.0 x 10⁹/L

Moderate 0.5-1.0x 10⁹/L

Severe <0.5 x 10⁹/L

Very severe <0.1 x 10⁹/L (agranulocytosis) Fever and Neutropenia