Investigation of the Erythrocyte Series PDF

PAGE 1 https://www.umfst.ro PA 5- Investigation of the erythrocyte https://edu.umch.de series. 2024 May Assist. Prof. Irina-Bianca K...

PAGE 1 https://www.umfst.ro PA 5- Investigation of the erythrocyte https://edu.umch.de series. 2024 May Assist. Prof. Irina-Bianca Kosovski, MD, Ph Assist. Prof. Andreea-Catalina Tinca, MD, Ph Assist. Prof. Andreea Cozac Szoke, MD, Ph Structure PAGE 2 1. Introduction 2. Methods of RBC analyses CBC The reticulocyte count The peripheral blood smear The bone marrow examination 3. Specific tests modifications for: Anemia Polycythemia Hemolytic anemia Introduction PAGE 3 The blood is a mixture of cellular elements, fluid, proteins and metabolites. Blood has four major components: 1. Red blood cells (erythrocytes): transport oxygen from the lungs to the peripheral tissues. 2. White blood cells (leukocytes): have a defensive role, destroying infecting organisms, such as bacteria and viruses, as well as assisting in the removal of dead or damaged tissues. Netter's Essential Histology, Ovalle, William K., PhD; Nahirney, Patrick C., PhD. Pages 169-185. © 2021. Introduction PAGE 4 3. Platelets (thrombocytes): are the first line of defense against damage to blood vessels, adhering to defects and participating in the blood clotting system. 4. Plasma: is the proteinaceous solution in which the above-mentioned cells circulate, and carries nutrients, metabolites, antibodies, hormones, proteins of the blood clotting system and other molecules throughout the body. Netter's Essential Histology, Ovalle, William K., PhD; Nahirney, Patrick C., PhD. Pages 169-185. © 2021. The complete blood count (CBC) PAGE 6 The usual complete blood count provided by automated cell counters includes a variety of directly measured and calculated parameters: 1. the red blood cell count 2. the hemoglobin concentration 3. the hematocrit 4. the red cell indices https://ww w.selectscie nce.net/pro 5. the white blood cell count ducts/xe- 5000- hematology 6. the white blood cells formulas - analyzer/?p rodID=1132 7. the platelet count 86 8. the platelet indices Indications: if signs and symptoms of anemia, inflammation or blood coagulation impairment are present in case of routine follow up of patients The complete blood count PAGE 7 1. Hemoglobin Normal general values (in clinic, REFERENCE RANGES-> can be slightly different for each analyzer/laboratory!!!) Age Female (g/dl) Male (g/dl) Adult (>18 y) 12.0-16.0 13.5-17.5 6-18 y 10.0-15.5 1-6 y 9.5-14.0 6 month-1 y 9.5-14.0 2-6 month 10.0-17.0 0-2 month 12-20 NN 14-24 The complete blood count PAGE 8 1. Hemoglobin Interpretation of hemoglobin: 1. Low hemoglobin Anemic syndrome The anemic syndrome is defined as a polyetiological syndrome characterized by decrease of the circulating hemoglobin (Hb) below the normal values. Although the number of red blood cells is often decreased, only the hemoglobin (Hb) concentration really reflects the capacity to transport oxygen. Blood dilution (artifact) The complete blood count PAGE 9 1. Hemoglobin Interpretation of hemoglobin: 2. High hemoglobin Hemoconcentration – Severe burns – Dehydration Primary polycythemia: Polycythemia Vera (PV) - is a myeloproliferative disorder associated with a low serum level of erythropoietin (EPO). Secondary polycythemia – Obstructive lung disease – EPO mediated. – High altitudes – EPO mediated. – Congenital cardiac diseases – EPO mediated. The complete blood count PAGE 11 2. RBC Typical appearance of a mature red cell in a stained smear of the Normal values= millions peripheral blood (a). 1. Newborns: 4.8 - 7.2 million/µL Scanning electron micrograph 2. Children: 3.8 – 5.5 million/µL showing the typical biconcave Stevens & Lowe's Human Histology, Lowe, James S., BMedSci, BMBS, DM, FRCPath; Anderson, Peter G., DVM, PhD; Anderson, Susan I., BSc, 3. Adults: structure of a mature red cell (b). MMedSci, PhD. © 2020. o Males: 4.6 - 6.0 million/µL o Females: 4.2-5.0 million/µL o Pregnancy: slightly lower than normal adult values The complete blood count PAGE 12 3. Hematocrit (HCT) This is the ratio of the volume of red cells to The HCT will be low in case of: the volume of whole blood (RBC x MCV/10). Low number or small volume of RBC Increased plasma volume Increased HCT: Normal range for hematocrit is different Increased number or volume of RBC between the sexes: Decreased plasma volume Men: 45 - 52% Women: 37 - 48% The complete blood count PAGE 14 4. Erythroid parameters A. Mean corpuscular volume (MCV) is the average volume of a red blood cell. This indirect parameter is calculated from the hematocrit and red cell count. Normal range: 80 - 90 femtoliters (fl- a fraction of one millionth of a liter; 10−15 L ) Interpretation MCV: Normocyte = normal size of the RBCs -> normocytic anemia Microcyte = small RBCs-> microcytic anemia Macrocyte = big RBCs-> macrocytic anemia The complete blood count PAGE 15 4. Erythroid parameters B. Mean corpuscular hemoglobin (MCH) is the average amount of hemoglobin in the average red cell. This is an indirect parameter calculated from hemoglobin concentration and the red cell count. Normal range: 27 - 32 pg/cell The complete blood count PAGE 16 4. Erythroid parameters C. Mean Corpuscular Hemoglobin Concentration (MCHC) is the average concentration of hemoglobin in a given volume of red cells. This is also an indirect parameter calculated from the hemoglobin concentration and the hematocrit. Normal range: 32 – 36 g/dl Interpretation MCH+MCHC: Normochromic RBC-> normochromic anemia Hypochromic RBC-> hypochromic anemia The complete blood count PAGE 17 4. Erythroid parameters D. Red Cell Distribution Width (RDW) is a measurement of the variability of red cell size and shape. Normal range: RDW-CV: 12-15% (the coefficient of variation of the RDW) RDW-SD: 37-47 fl (the standard deviation of the RDW) Interpretation RDW: ↑-> marker of red cell anisocytosis and poikilocytosis (variation in size and shape) The complete blood count PAGE 18 5. The Price Jones curve Is usually displayed by the hematology analyzer and it shows the distribution of red blood cells according to their diameters. If the curve is to: the right of the normal curve it indicates larger diameters the left of the normal curve it indicates smaller diameters. The reticulocyte count PAGE 20 Reticulocytes are immature red blood cells and the normal values are 0.5% to 1.5%. Can be identified base on the network of ribosomal RNA (=reticulum). Can be expressed by: the reticulocyte percentage the absolute reticulocyte count (the reticulocyte percentage multiplied by the red blood cell count) the reticulocyte production index (RPI) (corrects the reticulocyte count for the degree of anemia and then adjusts by a factor that compensates for the release of less mature reticulocytes into the circulation under circumstances of more severe anemia) The reticulocyte count PAGE 21 It takes ~ 1 week for a stem cell to differentiate fully into a mature erythrocyte. Crash Course Haematology and Immunology, Redhouse White, Gus, BSc (Hons); Vanbergen, Olivia, MA Oxon, MSc, MBBS (distinction). © 2019. The reticulocyte count PAGE 23 Polychromasia = increased number of reticulocyte in Romanowsky stain/the Wright-Giemsa stain (the classic stain for the peripheral smear). Polychromatic cells are erythrocytes with a blue tinge; because polychromatic cells are larger than normal mature erythrocytes, they are known as polychromatic macrocytes. Goldman-Cecil Medicine, Bain, Barbara J. Published January 1, 2020. Pages 1020- 1027.e2. © 2020. The reticulocyte count PAGE 24 The importance of the reticulocyte count 1. DIAGNOSIS: Reticulocyte counts - a good indicator of the bone marrow's ability to produce red blood cells; they are helpful in the initial approach to anemia in distinguishing anemias due to: underproduction of red cells adequate or overproduction but shortened survival in the circulation. Examples: Decreased : iron deficiency anemia, aplastic anemia, megaloblastic anemia Increased: hemolytic anemia, posthemorrhagic anemia, marrow infiltrative processes The reticulocyte count PAGE 25 The importance of the reticulocyte count 2. TREATMENT: During treatment of anemia, increases in the reticulocyte count can predict response to therapy prior to increments in the hemoglobin concentration or hematocrit (ex: iron treatment for iron deficiency anemia-> after 5-10 days: the reticulocyte crisis). Alternately, failure of the reticulocyte count to increase at the expected time can predict failure of current therapy and dictate a change of approach. The peripheral blood smear PAGE 26 Microscopic examination of peripheral blood smear is useful for diagnosis of some types of anemia: infiltrative processes of the bone marrow (leukemia) iron deficiency anemia megaloblastic anemia hemolytic anemia others. The peripheral blood smear PAGE 27 The blood sample is spread across a glass slide, forming a film of blood on the glass. This is then dried and stained, usually with a May-Grunwald stain. The blood film allows assessment of the morphology of blood cells and can show intracellular inclusions. Haematology: An Illustrated Colour Text, Howard, Martin R, MBChB MD FRCP FRCPath; Hamilton, Peter J, MA BM BCh FRCP FRCPath (retired). © 2013. The peripheral blood smear PAGE 30 Normal peripheral blood film Crash Course Haematology and Immunology, Redhouse White, Gus, BSc (Hons); Vanbergen, Olivia, MA Oxon, MSc, MBBS (distinction). © 2019. The peripheral blood smear PAGE 31 Normal description: round red blood cells with central pallor about one-third of the cell’s diameter-> normochromic RBC the size is compared with a nucleus of the small lymphocyte-> normocytic RBC Haematology: An Illustrated Colour Text, Howard, Martin R, MBChB MD FRCP FRCPath; Hamilton, Peter J, MA BM BCh FRCP FRCPath (retired). © 2013. The peripheral blood smear PAGE 32 Variation in: size of red cells is called anisocytosis shape of red cells is called poikilocytosis. Underwood's Pathology, Culligan, Dominic; Watson, Henry G.. Pages 550-607. © 2019. The peripheral blood smear PAGE 33 Acanthocytes (severe liver disease ) Macrocytes (Megaloblastic anemia) Target cells (liver disease) Hematology, basic principles and practice , ed 7, Philadelphia, 2018, Elsevier. The peripheral blood smear PAGE 34 Iron deficiency anaemia Sickle cell disease Hematology, basic principles and practice , ed 7, Philadelphia, 2018, Elsevier. The peripheral blood smear PAGE 35 Polycythemia Iron-deficiency anaemia Talley & O'Connor's Clinical Examination, Talley, Nicholas J, MBBS (Hons)(NSW), MD (NSW), PhD (Syd), MMedSci (Clin Epi)(Newc.), FRACP, FAFPHM, FAHMS, FRCP (Lond. & Edin.), FACP, FACG, AGAF, FAMS, FRCPI (Hon); O'Connor, Simon, FRACP, DDU, FCSANZ. © 2018. Davidson's 100 Clinical Cases, Das, T. © 2012. The peripheral blood smear PAGE 36 RBCs can also have a number of intracellular inclusions. Some of the major inclusions are: Howell-Jolly bodies. These are singular remnants of DNA that would ordinarily be removed by splenic macrophages. Their presence may be seen in splenectomized patients. Basophilic stippling. This is characterized by the presence of numerous basophilic granules in the cytoplasm. These granules contain RNA. They can be seen in many conditions and are classically described in sideroblastic anemia and lead poisoning. Clinical Hematology Atlas, Bernadette F. Rodak Jacqueline H. Carr, Elsevier, 2016 The peripheral blood smear PAGE 37 Cabot ring. Loop, ring, or figure eight dark blue to purple; may look like beads on a string. 1 to 2/cell. Thought to be remnants of mitotic spindle and are associated with myelodysplastic syndrome, megaloblastic anemia. Clinical Hematology Atlas, Bernadette F. Rodak Jacqueline H. Carr, Elsevier, 2016 Microscopic examination of the bone marrow PAGE 38 The bone marrow is the main site of hematopoiesis. Bone marrow occupies the spaces between the trabeculae of medullary bone and consists of highly branched vascular sinuses and a reticulin scaffolding, with the interstices packed with hematopoietic cells. The microscopic examination of bone marrow (BM) sometimes shows: the underlying cause of anemia due to cancers of the bone marrow (leukemia, lymphoma, multiple myeloma, or other cancers) the type of anemia (e.g. megaloblastic anemia). Microscopic examination of the bone marrow PAGE 39 The BM aspiration technique. Haematology: An Illustrated Colour Text, Howard, Martin. R, MBChB MD FRCP FRCPath; Hamilton, Peter J, MA BM BCh Kumar and Clark's Clinical Medicine, Murphy, Michael F.; Pasi, K. John; Roy, Noemi. 2021. Pages 319-378. FRCP FRCPath. © 2013. Microscopic examination of the bone marrow PAGE 40 Haematology: An Illustrated Colour Text, Howard, Martin. R, MBChB MD FRCP FRCPath; Hamilton, Peter J, MA BM BCh FRCP FRCPath. © 2013. Microscopic examination of the bone marrow PAGE 41 Normal bone marrow: megakaryocytes (∗) erythroid precursors ( ) granulocytic precursors (▲) Robbins and Cotran Atlas of Pathology, Klatt, Edward C., MD. © 2021. Microscopic examination of the bone marrow PAGE 42 Marrow aspirate smear from a patient with hemolytic anemia. There is an increased number of maturing erythroid progenitors (normoblasts). Robbins & Cotran Pathologic Basis of Disease, Kumar, Vinay, MBBS, MD, FRCPath; Abbas, Abul K., MBBS; Aster, Jon C., MD, PhD. © 2021. Types of anemia PAGE 43 According to the parameters in the CBC we can distinguish the following types of anemia: 1. Normocytic normochromic anemia (MCV N, MCH N): Iron deficiency (initial) Recent bleeding Hemolytic anemia Aplastic anemia Chronic infections (initial) Types of anemia PAGE 44 2. Microcytic hypochromic anemia (MCV↓, MCH↓, MCHC↓): Iron deficiency Chronic infections Thalassemia Cancer 3. Normocytic hypochromic anemia (MCV N, MCH↓) Renal diseases (erythropoietin deficiency) 4. Macrocytic normochromic anemia (MCV ↑, MCH ↑, MCHC N): B12, folic acid deficiency – megaloblastic anemia Chemotherapy Reactive reticulocytosis Liver diseases (including alcohol intake) Investigation of the hemolytic syndrome PAGE 45 1. CBC: ↓ RBC, ↓ HCT, ↓/N Hg 2. The reticulocyte count is usually high as an expression of the bone marrow compensation for the destruction. 3. Peripheral blood smear: polychromasia, anisocytosis, presence of nucleated RBCs (immature). Medical Sciences, Newland, Adrian C.; MacCallum, Peter; Davies, Jeff. © 2019. Investigation of the hemolytic syndrome PAGE 46 4. Biochemical tests: the breakdown of RBC is reflected by: High serum unconjugated bilirubin (indirect) Urinary hemosiderin present Urinary hemoglobin present High activity of serum lactate dehydrogenase (LDH, the erythrocyte enzyme) 5. Coombs Test This test detects antibodies that bind to red blood cells surface, causing premature red blood cell destruction (hemolysis) - useful for diagnosis of auto-immune hemolytic anemia. Medical Sciences, Newland, Adrian C.; MacCallum, Peter; Davies, Jeff. © 2019. Polycythemia PAGE 47 Polycythemia is an increase in hemoglobin concentration, HCT and red cell count. Depending on whether there is: a real increase in the RBC a reduction in plasma volume. Haematology: An Illustrated Colour Text, Howard, Martin R, MBChB MD FRCP FRCPath; Hamilton, Peter J, MA BM BCh FRCP FRCPath (retired). © 2013 Polycythemia PAGE 48 Causes and classification 2. Apparent polycythemia 1. True polycythemia (increased red cell mass) (reduced plasma volume, normal red cell mass) Primary polycythemia Secondary polycythemia Polycythemia vera Appropriate EPO production (hypoxia) Low plasma volume polycythemia Smoking (due to carbon monoxide Dehydration exposure) Burns COPD Enteropathy Long-term exposure to high altitude Cyanotic heart disease High-affinity haemoglobinopathy Inappropriate EPO production Renal pathology (carcinoma or cysts, artery stenosis, amyloidosis) Hepatocellular carcinoma Identify the following elements PAGE 49 Normal smear: round red blood cells with central pallor about one-third of the cell’s diameter, scattered platelets, occasional white blood cells. Goldman-Cecil Medicine, Bain, Barbara J. © 2020. Identify the following elements PAGE 50 Poikilocytosis Hypochromic/microcytic RBC Ovalocytes Target cells Goldman-Cecil Medicine, Bain, Barbara J. © 2020. Identify the following elements PAGE 51 Anisocytosis Poikilocytosis Howell-Jolly bodies Goldman-Cecil Medicine, Bain, Barbara J. © 2020. Identify the following elements PAGE 52 Spherocytosis Goldman-Cecil Medicine, Bain, Barbara J. © 2020. Identify the following elements PAGE 53 Target cells Goldman-Cecil Medicine, Bain, Barbara J. © 2020. Identify the following elements PAGE 54 Megaloblastic anaemia: macrocytic with many oval forms the neutrophil is hypersegmented Talley & O'Connor's Clinical Examination, Talley, Nicholas J, MBBS (Hons)(NSW), MD (NSW), PhD (Syd), MMedSci (Clin Epi)(Newc.), FRACP, FAFPHM, FAHMS, FRCP (Lond. & Edin.), FACP, FACG, AGAF, FAMS, FRCPI (Hon); O'Connor, Simon, FRACP, DDU, FCSANZ. © 2018. References PAGE 55 Netter's Essential Histology, Ovalle, William K., PhD; Nahirney, Patrick C., PhD. Pages 169-185. © 2021. Netter's Integrated Review of Medicine, Lim, Hana I.; Lee, Alfred. © 2021. https://www.selectscience.net/products/xe-5000-hematology-analyzer/?prodID=113286 Clinical Hematology Atlas, Bernadette F. Rodak Jacqueline H. Carr, Elsevier, 2016 Kumar and Clark's Clinical Medicine, Murphy, Michael F.; Pasi, K. John; Roy, Noemi. © 2021. Netter Collection of Medical Illustrations: The Respiratory System, Kaminsky, David A., MD. © 2011. Stevens & Lowe's Human Histology, Lowe, James S., BMedSci, BMBS, DM, FRCPath; Anderson, Peter G., DVM, PhD; Anderson, Susan I., BSc, MMedSci, PhD. © 2020. Crash Course Haematology and Immunology, Redhouse White, Gus, BSc (Hons); Vanbergen, Olivia, MA Oxon, MSc, MBBS (distinction). © 2019. Hematology, basic principles and practice , ed 7, Philadelphia, 2018, Elsevier. Underwood's Pathology, Culligan, Dominic; Watson, Henry G.. Pages 550-607. © 2019. Rodak's Hematology Clinical Principles and Application 5th Edition, Elsevier Immunology, Male, David, BA, MA, PhD; Peebles, R. Stokes, MD; Male, Victoria, BA, MA, PhD. Published January 1, 2021. Pages 352-355. © 2021. Goldman-Cecil Medicine, Bain, Barbara J. © 2020.