WBC Disorders - Week 14 PDF

WBC Disorders Cicero Kent Nasser, RMT Mirabele Camoro, RMT, MLS (ASCPi) Dianne Linas, RMT, MLS (ASCPi) Ian Polestico, RMT, MLS (ASCPi) At the end of the session, the students with 75% accuracy will be able to: 1. Describe correctly the basic genetic defect and the morphologic consequences in Pel...

WBC Disorders Cicero Kent Nasser, RMT Mirabele Camoro, RMT, MLS (ASCPi) Dianne Linas, RMT, MLS (ASCPi) Ian Polestico, RMT, MLS (ASCPi) At the end of the session, the students with 75% accuracy will be able to: 1. Describe correctly the basic genetic defect and the morphologic consequences in Pelger-Huet anomaly, Alder-Reilly anomaly, Chediak-Higashi syndrome, and May–Hegglin anomaly. 2. Describe accurately the cellular deficiencies and functional consequences of leukocyte adhesion disorders. 3. Describe correctly the characteristic macrophage morphology associated with the mucopolysaccharies, Gaucher disease, and Niemann-Pick disease At the end of the session, the students with 75% accuracy will be able to: 4. Discuss thoroughly the causes and development of acute leukemia, and interpret the results of diagnostic tests for acute leukemias. 5. Characterize accurately the diagnostic criteria used for acute myeloid and acute lymphoblastic leukemias. 6. Compare and contrast correctly acute lymphoblastic and myeloid leukemias by morphology, presenting signs and symptoms, laboratory findings, and prognosis. At the end of the session, the students with 75% accuracy will be able to: 8. Define correctly chronic myelogenous leukemia, and describe the cell lines involved, the clinical phases, and the expected clinical manifestations, key peripheral blood and bone marrow findings, and diagnostic criteria applicable to each stage. 9. Describe accurately the peripheral blood findings in chronic lymphocytic leukemia and hairy cell leukemia. Congenital Defects of Leukocyte Number and Function Severe Combined Immune Deficiency A group of genetic immunodeficiencies affecting both cellular and humoral immunity. Marked decrease in circulating T cells, poorly functioning B cells, hypogammaglobulinemia, and profound clinical manifestations. Gamma Chain Deficiency X-linked SCID, (Most common form of SCID) Caused by mutations in the IL2RG gene. ○ IL2RG normally codes for the common g chain in leukocyte receptors that bind with interleukins 2, 4, 7, 9, 15 and 21. Circulating T and natural killer (NK) lymphocytes are nearly absent. B cells are adequate in number but are dysfunctional. ADA Deficiency Autosomal recessive adenosine deaminase deficiency. ○ Key component of the metabolic breakdown of adenosine triphosphate and RNA. Profound decreases in T, B, and NK cells. Skeletal abnormalities, neurologic deficits, and skin rashes. Wiskott-Aldrich Syndrome Rare X-linked disease caused by one of more than 400 mutations in the WAS gene. WASp is important in cytoskeletal remodeling and nuclear transcription in hematopoietic cells. T cells are decreased; B cells, T cells and NK cells, neutrophils and monocytes are dysfunctional. 22q11 Syndromes A microdeletion in chromosome band 22q11.2, most likely involving TBX1. ○ DiGeorge syndrome, autosomal dominant Opitz GBBB, Sedlackova syndrome, Caylor cardiofacial syndrome, Shprintzen syndrome, and conotruncal anomaly face syndrome. Variable degrees of immunodeficiency because of the absence or decreased size of the thymus and low numbers of T lymphocytes. 22q11 Syndromes DiGeorge syndrome Sedlackova syndrome Caylor cardiofacial syndrome Sphrintzen syndrome Conotruncal anomaly face syndrome Bruton Tyrosine Kinase Deficiency Classified as an antibody deficiency. Reductions in all serum immunoglobulin isotypes and profoundly decreased or absent B cells. Chédiak-Higashi Syndrome A rare autosomal recessive disease of immune dysregulation. Mutation in the CHS1 LYST gene on chromosome 1q42.1-2 that encodes for a protein that regulates the morphology and function of lysosome-related organelles. Many types of cells in the body are affected and exhibit abnormally large lysosomes, which contain fused dysfunctional granules. Congenital Defects of Phagocytes The congenital neutropenias (CNs) are a rare group of genetic diseases characterized by low neutrophil count, increased risk of infection, organ dysfunction, and a high rate of leukemic transformation. Leukocyte Adhesion Disorders (Defects of Motility) Rare autosomal recessive inherited conditions resulting in the inability of neutrophils and monocytes to move from circulation to the site of extravasation. ○ LAD I - B2 Integrin ○ LAD II - Selectin synthesis ○ LAD III - Kindlin 3 Shwachman-Diamond syndrome Is a defect in leukocyte motility. SDS is a rare autosomal recessive disease caused by mutations in the SBDS gene. Defects of Respiratory Burst Chronic granulomatous disease (CGD) is a rare condition caused by the decreased ability of neutrophils to undergo a respiratory burst after phagocytosis of foreign organisms. Patients experience life-threatening catalase-positive bacterial and fungal infections WHIM Syndrome WHIM (warts, hypogammaglobulinemia, infections, and myelokathexis syndrome) is classified as a defect in intrinsic and innate immunity. Mutations in the CXCR4 gene. ○ Regulates movement of white blood cells between the bone marrow and peripheral blood. Neutropenia, lymphopenia, monocytopenia, and hypogammaglobulinemia are present. Morphologic Abnormalities of Leukocytes Without Associated Immunodeficiency Pelger-Huët Anomaly Also known as true or congenital PHA Decreased nuclear segmentation and distinctive coarse chromatin clumping pattern. Affects all leukocytes Mutation in the lamin beta-receptor gene Pelger-Huët Anomaly Nuclei may appear round, ovoid, or peanut shaped. ○ Bilobed forms—the characteristic spectacle-like (“pince-nez”) morphology with the nuclei attached by a thin filament can also be seen. Pseudo- or Acquired Pelger-Huët Anomaly Neutrophils with similar morphology to PHA can be seen in patients with MDS, acute myeloid leukemia, and myeloproliferative Neoplasm. Acquired or pseudo-PHA neutrophils are also associated with severe bacterial infections, HIV, tuberculosis, and mycoplasma pneumonia.  TRUE PHA PSEUDO PHA Number of cells affected > 68% < 68% Types of WBC affected All WBC lineages Neutrophil only Neutrophil Hypersegmentation Hypersegmented neutrophils have more than five lobes and are most often associated with megaloblastic anemia, in which hypersegmented neutrophils are usually larger than normal. Alder-Reilly Anomaly A rare inherited disorder characterized by granulocytes (monocytes and lymphocytes less often) with large, darkly staining metachromatic cytoplasmic granules. Initially reported in patients with gargoylism. May-Hegglin Anomaly A rare, autosomal dominant disorder characterized by variable thrombocytopenia, giant platelets, and large Döhle body-like inclusions in neutrophils, eosinophils, basophils, and monocytes. Mutation in the MYH9 gene on chromosome 22q12-13.3 Lysosomal Storage Disorders Lysosomal Storage Disorders Are a group of more than 50 inherited enzyme deficiencies resulting from mutations in genes that code for the production of lysosomal enzymes Are classified according to the under degraded macromolecule that accumulates in the cell Mucopolysaccharidoses A family of inherited disorders of mucopolysaccharide or glycoaminoglycan (GAG) degradation Deficient activity of an enzyme necessary for the degradation of dermatan sulfate, heparan sulfate, keratan sulfate, and/or chondroitin sulfate Gaucher Disease Most common of the lysosomal lipid storage diseases. Caused by a defect or deficiency in the catabolic enzyme b-glucocerebrosidase. Bone marrow replacement by Gaucher cells contribute to anemia and thrombocytopenia. ○ Gaucher cells are distinctive macrophages, single or in clusters, exhibiting abundant fibrillar blue-gray cytoplasm with a striated or wrinkled appearance. Niemann-Pick Disease Characterized by an accumulation of fat in cellular lysosomes of vital organs. Foam cells and sea-blue histiocytes can be seen in the bone marrow. ○ Foam cells are macrophages with cytoplasm packed with lipid-filled lysosomes that appear as small vacuoles (foam) after staining. ○ Sea blue histiocytes are macrophages with lipofuscin-, glycophospholipid-, and sphingomyelin contained in -cytoplasmic granules, 1 to 3 m in diameter, that appear blue with Wright stain. Leukocytes: The Quantitative Abnormalities Neutrophils Neutrophilia ○ Neutrophils greater than 7.0X10^9/L in adults or 85.x10^9/L in children ○ Can occur as a result of catecholamine-induced shift in neutrophils Neutrophils Neutropenia ○ A decrease in the ANC to less than 2.0x10^9/L in white adults and 1.3x10^9/L in black adults ○ Causes of neutropenia: Increased rate of removal or destruction of peripheral blood neutrophils Fewer neutrophils released from the bone marrow Decreased ratio of circulating versus marginal pool of neutrophils A combination of the three Eosinophils Eosinophilia ○ Absolute eosinophil count greater than 0.4x10^9/L ○ Associated with parasitic infections and allergic reactions Eosinopenia ○ Absolute eosinophil count of less than 0.9x10^9/L Basophils Basophilia ○ Absolute basophil count greater than 0.15x10^9/L ○ Chronic myeloid leukemia, allergic rhinitis, hypersensitivity to drugs or food, chronic infections, hypothyroidism, chronic inflammatory conditions, radiation therapy, and bee stings Monocytes Monocytosis ○ Absolute monocyte count greater than 1.0x10^9/L in adults and greater than 3.5x10^9/L in neonates ○ First sign of recovery after myelosuppression Monocytopenia ○ Absolute monocyte count of less than 0.2x10^9/L Lymphocytes Lymphocytosis ○ In children, it is defined as an absolute lymphocyte count greater than 5.0x10^9/L Lymphocytopenia ○ In children, it is defined as an absolute lymphocyte count less than 1.0x10^9/L Myelogenous Leukemias: Non-Lymphocytic Leukemias General Characteristics Myeloperoxidase: positive (+) Sudan Black B: positive (+) Divided into two types: Acute versus Chronic Duration of symptoms Number of immature or mature cell forms WBC count Acute versus Chronic Acute Chronic Duration of Short Long Symptoms Cells in BM and PB Numerous immature Mostly mature cells cells WBC count Increased Total WBC count range from extremely elevated to lower than normal Acute Myelogenous Leukemias (AML) There is a breach in the differentiation in the myeloid stem cell line Increased immature cells Predominating cells (PBF): myeloblast and promyelocyte Presence of Auer rods Most common type of leukemia in adults Laboratory Findings (AML): Anemia is usually present Elevated WBC count Thrombocytopenia Hyperuricemia Hyperphosphatemia Hypocalcemia High M:E ratio Clinical Presentations (AML): Pallor, fatigue, and fever with infections Bruising and bleeding Disseminated intravascular coagulopathy (DIC) Bone and joint pain Splenomegaly Tumor lysis syndrome Investigation and Diagnosis (AML): Investigation: ○ CBC ○ PBF examination ○ BM aspirate ○ BM biopsy Diagnosis: ○ BM - hypercellular (greater than 20% of cells are blasts) Classification of AML World Health Organization French-American-British (FAB) (WHO) Based on cytogenetics and Based on morphology and molecular studies cytochemistry At least 20% blasts in bone Based on marrow to diagnose AML Romanowsky-Stained smears Pseudo-pelger-huet cells in the granulocytic cell line Auer rods M0 (AML, Minimally Differentiated) Blasts have CD13, CD33, CD34, and CD117 No evidence of cellular maturation of blasts Auer rods (-), MPO (-), SBB (-) Less than 5% of all AML Patients are usually infants or older adults M1 (AML, Without Maturation) Blasts having CD13, CD33, CD34, and CD117 similar with those of M0 90% of cells in BM are blasts Found in all age groups with highest incidence in adults Has no male or female predominance M1 (AML, without maturation) Nuclear:Cytoplasmic Maturational Asynchrony ○ Common finding in M1 leukemia ○ Morphologic abnormality: nucleus appears more immature than cytoplasm ○ Functional abnormality: leukemic blasts exhibits phagocytosis which is a property only of a mature WBC Auer rods (+), MPO (+), SBB (+) Chloroacetate esterase (+), acetate esterase (-) M2 (AML, with maturation) Greater than 20% Type I and II blasts in BM ○ At least 10% granulocyte at various stages of maturation Distinguished from M1 by presence of granulocytic cells at or beyond the promyelocytic stage of maturation Pseudo-Pelger-Huet (+): rod-shaped or dumbbell-shaped or nonsegmented nuclei M2 (AML, with maturation) Hypogranular Neutrophils (+): leads to deficient phagocytosis, deficient microbial killing, and deficient chemotaxis Auer rods (+), MPO (+), and SBB (+) Aspects of dysplasia are present M3 (Acute Promyelocytic Leukemia) aka Hypergranular Promyelocytic Leukemia Found in all age groups similar to M1 and M2 Greater predilection for males Frequently more associated with DIC Abnormal promyelocytes with heavy granulation M3 (Acute Promyelocytic Leukemia) Presents with leukopenia Auer rods (+) and intensely positive for MPO and SBB Faggot cells (+) Reniform or bilobed nuclei M3m (Microgranular Promyelocytic Leukemia) Numerous granules present but can only be detected by electron microscopy hence the term “Microgranular” Has worse prognosis than M3 due to initial high blast counts Caused by a chromosomal translocation t(15:17) M4 (Acute Myelomonocytic Leukemia) aka Naegili Monocytic Leukemia Positive for Myeloid antigens: CD13 and CD33 Positive for Monocytic antigens: CD4, 11c, 14, 36, and 64 Auer rods (+), MPO (+), SBB (+), specific and non-specific esterases (+) M4 (Acute Myelomonocytic Leukemia) Lysozyme-muramidase ○ Contained in larger amounts in monocytes and excreted in large amounts in urine when there is M4 leukemia ○ Serum or Urine lysozyme: diagnostically important for M4 leukemia ○ 3x the upper limit is significant Caused by a problem in Chromosome 16 M4eo (Acute Myelomonocytic Leukemia with Eosinophilia) Increased marrow eosinophils Cells exhibit large basophilic granules mixed with smaller eosinophilic granules Uniquely exhibits distinct chloroacetate esterase and PAS (+) which differentiates it from normal eosinophils M5 (Acute Monocytic Leukemia) aka Schilling Leukemia Presents with highest incidence of organomegaly and organ involvement of all AMLs Greater than 80% of marrow cells are monoblasts, promonocytes, or monocytes M5 (Acute Monocytic Leukemia) Auer rods (+), MPO (-), SBB (-), Specific esterase (-) Associated with problems in Chromosome 11, t(9,11) Divided into M5a (poorly differentiated) and M5b (well differentiated) M5a (Acute Monocytic Leukemia, Poorly Differentiated) Characterized by large blast cells with delicate, lacy chromatin in both blood and bone marrow ○ 1-3 large, prominent vesicular nucleoli are present ○ Voluminous cytoplasm with 1 or more pseudopods More than 80% of monocytic compartment predominance are blasts M5b (Acute Monocytic Leukemia, Well Differentiated) Characterized by presence of all stages of monocyte development (monoblasts, promonocytes, and monocytes) Predominant cell in BM: Promonocyte Associated with diffuse erythematous skin rash (differentiates it from M5a clinically) M6 (Acute Erythroleukemia) DiGuglielmo’s syndrome Variable WBC count and pancytopenia occurs Presence of numerous nucleated RBCs Mixed populations of hypochromic and normochromic RBCs M6 (Acute Erythroleukemia) M6 leukemia frequently progresses to M1, M2, or M4 leukemia M6 erythroblasts: alpha-naphthly acetate esterase (+) Auer rods (+) Defect in Chromosome 5 and 7 M7 (Acute Megakaryocytic Leukemia) Myelosclerosis: distinct feature of M7 Previously classified as Undifferentiated Leukemia since MPO (-), SBB (-), and esterase (-) Unique cytochemistry for megakaryoblasts: alpha-naphthyl acetate esterase (+), alpha-naphthyl butyrate esterase (-) PAS (+) CD41, CD42b, and CD61 Defect in Chromosome 21 Chronic Myelogenous Leukemia Chronic granulocytic leukemia An MPN that originates in the abnormal pluripotent bone marrow stem cell and is consistently associated with the BCR-ABL (1) fusion gene located in the Philadelphia chromosome Results in a disordered proliferation of cells (granulocytic cell line) Cells still have the ability to differentiate Chronic Myelogenous Leukemia Increased WBC, mature WBC, PMNs Occurs at any age but most commonly at 60–65 years old Men have slightly greater rate of disease Pathophysiology (CML) Chromosome 9 and 22 translocation Translocation of c-abl from chromosome 9 to location of bcr on chromosome 22 to form fusion gene bcr-abl (active tyrosine kinase) Signs and symptoms (CML) Splenomegaly Constitutional symptoms (fatigue, malaise, LG fever, weight loss, excess sweating) Anemia Bleeding Pruritus Investigation and Diagnosis (CML) Investigation ○ Leukopenia ○ Basophilia ○ Low RBC count Diagnosis ○ Evidence of BCR-ABL fusion product or presence of Philadelphia chromosome CML versus Leukemoid Reaction Leukemoid Reaction ○ Excessive leukocytic response in the peripheral blood ○ A result of severe infection, inflammation, metabolic disease, or malignancy ○ Leukocyte (neutrophil) alkaline phosphatase (LAP) test: Used to distinguish LR from CML Uses Kaplow’s method Kaplow’s (LAP) Score Score Description 0 No reddish brown to black ppt 1+ Slightly diffused reddish brown to black ppt 2+ Moderately diffused reddish brown to black ppt 3+ Heavily diffused reddish brown to black ppt 4+ Very heavily diffused reddish brown to black ppt Kaplow’s (LAP) Score Normal Kaplow’s score: 20-100 Generally: ○ Increased LAP score: leukemoid reaction ○ Decreased LAP score: CML Disorders with increased LAP Score: ○ Infections ○ Polycythemia vera ○ Intoxications Disorders with decreased LAP Score: ○ MDS ○ Sideroblastic anemia ○ PNH ○ CML CML versus Leukemoid Reaction CML Leukemoid Reaction Leukocyte in PB Blasts/promyelocyte Usually myelocytes Toxic granulation Absent Present Eosinophils/Basophils Increased Decreased LAP Decreased Increased Philadelphia Chromosome Usually present Absent Splenomegaly Usually prominent Mild (if present) Platelet count >600 or 30% bone marrow blasts Clinical syndrome Easier to use and is still widely taught Defines acute leukemia as: >20% bone marrow blasts Cytochemical Stains in Hematology Cytochemical Stains in Hematology Cytochemistry is the use of special stains for the microscopic detection of cellular constituents such as lipids, carbohydrates, and enzymes Prussian Blue Fe+ Reagent: Potassium ferricyanide – HCl Blue or green color Leukocyte Alkaline Phosphatase (LAP) Neutrophils, Band forms of granulocytes Increased in leukemoid reaction Decreased in CML Myeloperoxidase (MPO) Granules of neutrophils and eosinophils Differentiates acute myelogenous or monocytic leukemia from ALL Similar result to Sudan Black B Auer rods (lyso structures): Peroxidase + (seen in AMLs) Sudan Black B (SBB) Lipids Stain granulocytes ONLY Differentiates acute myelogenous and myelomonocytic leukemias from ALLs Periodic Acid Schiff (PAS) Helpful in: ○ AMLs - FAB M6 ○ ALL - FAB L1, L2 All cells stained EXCEPT for erythroblasts Chloroacetate Esterase Stain Specific for granulocytic cells ONLY Blue colored precipitate aka specific esterase Nonspecific Esterases Differentiates granulocytic from monocytic leukemias Substrate: alpha-naphthyl Specific for monocytes Positive: monocytes, platelets, macrophages, megakaryocytes Tartrate Resistant ACP (TRAP) aka ACP For the diagnosis of hairy cell leukemia (leukemic reticuloendotheliosis) Nitroblue Tetrazolium Test (NBT Test) Screening for CGD (Chronic Granulomatous Disease) Terminal Deoxynucleotidyl Transferase Primitive lymphoid cell (lymphoblasts) marker Stain for ALLs ALL vs. AML ALL AML PBS and BM: Blast and Genetically heterogeneous immature leukocyte clonal disorder of myeloid (lymphoid lineage) lineage Disease of childhood and Increase in the number of adolescent immature cells due to Total WBC is usually hematopoietic insufficiency elevated with or without leukocytosis Most common in children Most common in adult FAB CLASSIFICATION (Subgroup ALLs) FAB ALL Characteristic Small lymphoblast Homogenous L1 Regular nucleus Most T-cell ALLs are FAB L1 Most common childhood ALL Large lymphoblast Heterogenous L2 Irregular nucleus (cleft) Most common ALL in adults Leukemic phase of Burkitt’s Lymphoma Large lymphoblast Homogenous and vacuolated L3 Round or oval nucleus All FAB L3 are B cell Chromosomal abnormality: T(8,14) the MYC proto-oncogene on chromosome 8 is fused with the immunoglobulin heavy chain locus on Chromosome 14  WHO (World Health Organization) B-cell lineage Precursor B lymphoblastic leukemia/lymphoblastic ALL Classification lymphoma (B-ALL) T-cell lineage Precursor T lymphoblastic leukemia/lymphoblastic lymphoma (T-ALL) CLL vs CML CLL CML Older adults in Western Absence of Philadelphia countries Chromosome Disease of mainly B-cells, T(9,22) BCR/ABL gene some T cells mutation CLL smudge cells: JAK2 Decreased LAP score V617K gene positive JAK2 V617F gene negative Middle-aged adults Hairy Cells TRAP positive B-cell malignancy CD markers: CD19, CD20, CD22 Most specific markers: CD123, ANNEXIN A1 Mycosis Fungoides Most common Cutaneous Lymphoma Presence of Sezary cells T-cell malignancy -END-