blood17 Hematology: Anemias MUDr. Stanislav Matoušek, PhD images HEMATOLOGY l. hemolyticanemia40x01small ¡ Anemias and -penias (not having enough elements) thrombocytopenia Leukemia etc. (oncology – lympho- and myeloproliferative disorders) E.g. chronic myeloid leukemia ¡ Disorders of blood clotting Primary hemostasis Secondary hemostasis Bleeding disorders/ thrombo-embolism Clinical symptoms of hematologic disease l Anemia ¡ → signs of hypoxia – tiredness, weakness, dyspnea ¡ → signs of low levels of hemoglobin - paleness ¡ → cardiovascular symptoms – palpitation l l Polycytemia → hyperviscose blood → risk of thrombosis l l Bleeding, spontaneous bleeding, unceasing bleeding l l Thrombosis → embolism – local symptoms of swelling or ischemia – DVT -, pulmonary embolism lFrequent infections l Anemia •Hb •M: < 135 g/L •F: < 120 g/L •Hct •M: < 40 % •F: < 37 % •Ery •M: < 4,3 * 1012 /L •F: < 3,9 * 1012 /L • images2 haematocrit HEME002 Principal criterion: Hb< 120 g/L in w or < 135 in m ¡ •low hemoglobin concentration paleness • •Deficient oxygen delivery into tissues • •Tissue hypoxia sympaticus •activated •weakness,dyspnea • palpitations hyperkinetic circulation ¡ ¡ ¡ Pathophysiology of anemia symptoms ¡ Causes of hypoxia lAltitude hypoxia – lack of O2 in the inspired air = low pO2 lRespiratory insufficiency– hypoxic hypoxia lLack of hemoglobin – transport hypoxia = anemia lCirculatory disturbance – circulatory hypoxia lImpaired oxidation in mitochondria – histotoxic hypoxia l l l l Laboratory tests: •Principal: –Complete (full) blood count (CBC or FBC) •Complementary: –Tests of iron metabolism –Erythropoietin levels –Detecting antibodies against RBC – Coombs test = antiglobuline test AGT –Osmotic fragility test –Historically: Ham’s test (resistance in acidic environment) –Blood film/smear microscopic examination –Bone marrow cytology/ aspiration (Sternal puncture) – – Anemias classified by RBC morfology (CBC) •by MCV – microcytic - e.g. iron deficiency –normocytic - e.g. acute bleeding –macrocytic (megaloblastic) - pernicious •by MCHC (color) –hypochromic – lack of iron –normochromic – – ¡ ¡ ¡ ¡ ¡ ¡ ¡ Stem cells, growth factors Cellular division, maturation: DNA synthesis: vitamin B12, folic acid erythropoietin: Hemoglobin synthesis: globin, porphyrine, Fe Other factors BONE MARROW PERIPHERAL BLOOD ¡ ¡ bleeding PRODUCTION hemolysis LOSSES Erythrocytes ¡ ¡ hemoglobin, RBC count, hematocrit MCV, MCH, MCHC shape etc. Anemias by their etiology/patho • decreased production –Stem cell failure or failure to differentiate –disorder in DNA synthesis –Disorder in hemoglobin synthesis –Lack of erytropoetin / renal failure •Complete loss of erythropoiesis – decrease of RBC count 10% / wk •increased destruction - hemolysis –Defect of erythrocytes –Causes outside of RBC • increased loss - bleeding • misdistribution and loss (hyperslenism, pooling in spleen) • Reticulocyte count •Daily replenishment rate –0.5 – 1.5% of total RBC count –Mature during the 1st day in peripheral blood •Criterion of bone marrow activity – •Key test in distinguishing anemias –Reticulocytosis •Reaction of the BM to a blood loss (hemolytic anemias, severe bleeding) •Response to a correct anemia therapy (e.g. defic. B12 or Fe) –Reticulocytopenia •Defective erythropoiesis • • Blood loss anemia •Acute blood loss –shortly after massive blood loss Hb normal due to vasoconstriction –normochromic - normocytic •Chronic blood loss –results in iron deficiency •Excessive hemolysis (RBC destruction) • Excessive hemolysis (RBC destruction) •Extrinsic RBC defect (normocytic-normochromic RBC ) •Immunologic abnormalities (AIHA, PNH) •Mechanical injury (trauma, infection) •Intrinsic RBC defect •Membrane alterations –congenital (spherocytosis, elliptocytosis) –Aquired (hypophosphatemia) •Metabolic disorders (G6PD deficiency) •Hemoglobinopaties (Sicle cell disease, Thalassemia) reticulocytosis, LDH is increased, unconjugated bilirubin accumulates G6PD deficienc over 100 mutant forms identified. X-linked, drug senzitive variety most common (drugs which produce peroxide) AIHA - autoimmune hemolytic anemia ¡ HEMOLYSIS ¡ INTRAVASCULAR Hb EXTRAVASCULAR ¡ ¡ ¡ spleen spleen, bone marrow, liver (macrophages) haptoglobin kidneys SYMPTOMS OF HEMOLYSIS loss of red blood cells anemia BM activation reticulocytosis loose Hb hemoglobinemia, hemoglobinuria hemosiderinuria intravascular increased prodution of bilirubin jaundice (icterus) extravascular damage to the kidneys kidney splenomegaly spleen Reticulocyte-(counter-stain TESTS FOR HEMOLYSIS immune mechanisms – direct Coombs (antiglobulin) test coombstest2 Search for antibodies against proper RBC Antibodies other than AB0 These Abs are responsible for hemolysis Paroxysmal nocturnal hemoglobinuria Molecular and cellular differences between PIGA- and PIGT-PNH. (A) In he... •Paroxismal Nocturnal Hemoglobin- uria • • • •From: Fattizzo B, Serpenti F, Giannotta JA, Barcellini W. Difficult Cases of Paroxysmal Nocturnal Hemoglobinuria: Diagnosis and Therapeutic Novelties. J Clin Med. 2021;10(5):948. Published 2021 Mar 1. doi:10.3390/jcm10050948 Jcm 10 00948 g001 Direct antiglobulin (Coombs’) test (DAT) •Detection of antibodies to erythrocyte surface antigens •AIHA •Antiglobulin serum is added to washed RBC from the patient ------ agglutination indicates presence of immunoglobulins or complement components bound to RBC TESTS FOR HEMOLYSIS Test of osmotic resistence RBC survive only in isotonic surrounding but have some toleration to its changes RBC in some hemolytic states have decreased tolerance Special tests membrane properties (electrophoresis of proteins) properties of hemoglobin genetic tests rbc skeleton 2 Deficient erythropoiesis •Iron deficiency –microcytic-anisocytosis, ↓ reticulocytes •Vitamin B12 or Folate deficiency –macrocytes-anisocytosis •BM failure - chronic diseases, aplastic anemia, myelodysplasia, leukemia –normochromatosis-normocytosis –BM hypoplasia Iron deficiency: Include = anemia of chronic blood loss; Hypochromic-microcytic anemia; Hypochromic anemia of pregnancy, infancy, and childhood) BLOOD LOSS leadind cause; DEFECT IN FE UTILIZATION – hemoglobinopaties, sideroblstic and myelodysplastic anemia HIGH RDW --- ringed sideroblasts in BM SIDEROBLASTIC ANEMIA associated with myelodysplastic sy. Vitamin B12: RISK OF GASTRIC CANCER --- GI X-ray advised REQUIRES INTRINSIC FACTOR from gastric mucosa to absorb in the intestine Folate PROLONGD COOKING DESTROYS FOLATE, Peaple who eat marginal diet = TEA-and –TOASTERS, CHRONIC ALCOHOLICS Defective DNA synthesis ---- RNA synthesis continue ---- increased cytoplasmatic volume and maturation ---- increased intramedullar cell death = inefective erythropoiesis Chronic disease: JZ6]Kidney, endocrine – thyroid, pituitary, or protein depletion Tests for iron •iron concentration in serum (age , sex) •TIBC (total iron binding capacity for Fe) •transferrin saturation (N 20-55 %) •serum ferritin •serum (solubile) transferrin receptor (sTfR) • Tests of iron metabolism •Serum iron ( SI) •F: 600-1400 mg/L, 11-25mmol/L; M: 750-1500 mg/L, 13-27mmol/L •Low in Fe deficiency and chronic disease •High in hemolytic syndromes and iron overload •Total iron binding capacity (TIBC) •2500 – 4500 mg/L , 45-82 mmol/L •High in Fe deficiency •Low in chronic disease •Serum ferritin (30-300 ng/mL) •Fe storage glycoprotein •Closely correlates with total body Fe stores •<12 ng/mL Fe deficiency •Elevated in Fe overload, liver injury, tumors (Acute phase protein) • Tests for iron metabolism •Serum transferin receptor •Increase in increased erythropoiesis and early Fe deficiency •RBC ferritin •storage status over the previous 3 month (Fe deficiency/overload) •unaffected by liver function or acute illness •Free RBC porphyrin •increased when heme synthesis altered ¡ anemia Manifest Latent iron deficiency erythropoiesis Prelatent no stores serum iron transferin satur. transferin serum ferritin TIBC ~ transferin iron in BM ¡ ¡ ¡ ¡ ¡ ¡ ¡ Regulation of iron Lakomec ¡Hepcidin – signal molecule – keep your iron inside the cell! - anemia of chronic disease Microcytic Hypochromic Anemia (MCV<83; MCHC<31) Lead poisoning Basophilic stippling of RBCs Sideroblastic Ring sideroblasts in marrow Hemoglobi-nopaties Hemoglobin electrophoresis Iron deficiency Responsive to iron therapy Chronic inflammation Unresponsive to iron therapy Thalassemia major Reticulocytosis and indirect bilirubinemia Thalassemia minor Elevation of fetal hemoglobin, target cells, and poikilocytosis normal hypochrom iron deficiency Microcytic Hypochromic Anemia (MCV<83; MCHC<31) Serum Iron Total Iron-Binding Capacity (TIBC) Bone Marrow Iron Comment Lead poisoning Basophilic stippling of RBCs Sideroblastic Ring sideroblasts in marrow Hemoglobinopaties Hemoglobin electrophoresis N N ++ N N ++ N ¡ ++++ Microcytic Hypochromic Anemia (MCV<83; MCHC<31) Serum Iron Total Iron-Binding Capacity (TIBC) Bone Marrow Iron Comment Iron deficiency Responsive to iron therapy Chronic inflammation Unresponsive to iron therapy Thalassemia major Reticulocytosis and indirect bilirubinemia Thalassemia minor Elevation of A of fetal hemoglobin, target cells, and poikilocytosis ¡ N ++++ N N ++ ¡ ¡ ++ ¡ ¡ 0 Macrocytic Anemia (MCV, >95) Megaloblastic bone marrow Deficiency of vitamin B-12 Deficiency of folic acid Drugs affecting DNA synthesis Inherited disorders of DNA synthesis Nonmegaloblastic bone marrow Liver disease Hypothyroidism and hypopituitarism Accelerated erythropoiesis (reticulocytes) Hypoplastic and aplastic anemia Infiltrated bone marrow megalo normal Absorption of vit. B12 Vitamin B12 Digestion, Absorption and Metabolism - YouTube Lack of vit. B12 - causes •Not enough ingestion – strict vegans if they do not take care •Autoimmune inflammation of gastric mucosa (atrophic gastritis) leading to deficiency in intrinsic factor •Diseases of terminal ileum (celiac disease, Crohn disease) Blood smear •Morphology of blood elements –Anisocytosis = variation in size –Poikilocytosis = variation in shape (schistocytes=RBC fragments; ovalocytes; spherocytes) Various Forms of RBCs Spherocyte Loss of central pallor, stains more densely, often microcytic. Hereditary spherocytosis and certain acquired hemolytic anemias. Target cell Hypochromic with central "target" of hemoglobin. Liver disease, thalassemia, hemoglobin D, postsplenectomy. Elliptocyte Oval to cigar shaped. Hereditary elliptocytosis, certain anemias (particularly vitamin B-12 and folate deficiency). Schistocyte Fragmented helmet- or triangular-shaped RBCs. Microangiopathic anemia, artificial heart values, uremia, malignant hypertension. Stomatocyte Slitlike area of central pallor in erythrocyte. Liver disease, acute alcoholism, malignancies, hereditary stomatocytosis, and artifact. Sickle cell Elongated cell with pointed ends. Hemoglobin S and certain types of hemoglobin C and l. Sicle Cell Disease SCD_1000x Hemoglobin (Hb) S arises from a mutation substituting thymine for adenine in the sixth codon of the beta-chain gene GAG to GTG. This causes coding of valine instead of glutamine in position 6 of the hemoglobin beta chain. The resulting hemoglobin has the physical properties of forming polymers under deoxy conditions. It also exhibits changes in solubility and molecular stability. These properties are responsible for the profound clinical expressions of the sickling syndromes. Under deoxy conditions, Hb S undergoes marked decrease in solubility, increased viscosity, and polymer formation at concentrations exceeding 30 g/dL. It forms a gel-like substance containing hemoglobin crystals called tactoids. The gel-like form of hemoglobin is in equilibrium with its liquid soluble form. A number of factors influence this equilibrium, including the following: Oxygen tension Polymer formation occurs only in the deoxy state. If oxygen is present, the liquid state prevails. Concentration of Hb S The normal cellular hemoglobin concentration is 30 g/dL. Gelation of Hb S occurs at concentrations greater than 20.8 g/dL. The presence of other hemoglobins Hemoglobin A and hemoglobin F have an inhibitory effect on gelation. These and other hemoglobin interactions affect the severity of clinical syndromes. SS hemoglobin produces a more severe disease than SC, SD, SO Arab, and SA hemoglobin. When to do CBC? •suspected hematologic, inflammatory, neoplastic, or infectious disease •screening of infants (<1 yr.), pregnant women, elderly patients, and patients with nutritional abnormalities •routine patient evaluation, admission to hospital