PATHOLOGY

A 28 yo male presents with complaints of weight loss, easy fatigability, and night sweats. Physical eamination shows cervical, axillary and inguinal lymphadenopathy. His blood tests shows HB-9.5 g/dl (13.5-17.5), TLC - 13,000/cu mm (4.5-11*10^3), Plt - 100,000/cumm (150-400* 10^3). Abdominal CT shows enlargement of celiac group of lymph nodes. Lymph node biopsy shows feature suggestive of mixed cellularity type of Hodgkin lymphoma. Which of the following infectious agent is responsible for the development of this disease? A. CMV B. HIV C. EBV D. HHV-8 E. HTLV-1

C. EBV

Neutropenia and Agranulocytosis -With this, pt will have signs of infection

Clinical manifestations -Infection- fever, chills, fatigue. -Gingival pain and swelling, skin abscess, recurrent sinusitis, recurrent otitis, symptoms of pneumonia (cough with green/yellow sputum, difficulty in breathing/dyspnea). -With agranulocytosis the infections are overwhelming and often fatal.

Leukopenia

-Abnormally low white cell count (leukopenia) usually results from reduced numbers of neutrophils (neutropenia, granulocytopenia) and Lymphocytes. -Neutropenia / granulocytopenia is more common -Lymphopenia seen in HIV infection, following glucocorticoid therapy, cytotoxic drugs, autoimmune disorders, malnutrition, congenital immunodeficiency diseases. -Incidence of Lymphopenia is less common compared to neutropenia.

ALL/Lymphoma (B cell and T cell)

-Acute Lymphoblastic Leukemia/Lymphoma: -Cell of origin : immature B cells (Pre-B) and T cells (Pre-T). -Clinical presentation: B-ALL- childhood (3-11 yr), peak incidence at the age of 3. *T cell ALL: Adolescence age, majority < 15 yr (10-19yr) (thymus is of max size at this age) *CNS involvement: meningeal spread and involvement of dura. *Involvement of testis *Mediastinal involvement: thymus is common in T-ALL -Pathogenesis: *B-ALL: t(12;21) involving ETV6 and RUNX1. * T-ALL: gain-of-function mutation in NOTCH1 *t(9;22) - bad prognosis -Factors associated with favorable prognosis: *Age 2-10 yrs *Low white cell count *Presence of t(12;21) -When blast cell involve lymph node/ solid organ , it is known as acute lymphoblastic lymphoma.

Anemias of Blood Loss

-Acute blood loss: alterations mainly reflect the loss of intravascular volume *Soon after acute blood loss, the RBCs (red blood cells) appear normocytic & normochromic. *As the marrow regenerates, an increase in the reticulocyte count in the peripheral blood is most striking -Chronic blood loss: leads to anemia, when the rate of loss exceeds the regenerative capacity, or when iron reserves are depleted

A 50 yo male presents with chronic back pain for the past 8 months. He also complaints of weakness, inability to concentrate in work, easy fatigability and constipation which started ten months ago. His blood workup shows Hb-8.5g/dl (13.5-17.5 g/dl), TLC-9,000/cumm (4,500-11,000), Plt - 200,000/cu mm (150,000-400,000), ESR 120 mm/1st hr (0-22 mm/hr), serum calcium - 13.5 mg/dl (8.5-10.2mg/dL), serum creatinine-3.2 mg/dl (0.5-1.2 mg/dL), BUN 52 mg/dl (7-20 mg/dL). What is the most likely serum electrophoresis finding seen in this patient? A. Increased IgG with kappa chain restriction B. Inscreased IgM with kappa chain restriction C. Increased IgG with increase in both kappa chain and lambda chain D. Increased IgM with increase in both kappa chain and lambda chain

A. Increased IgG with kappa chain restriction

Hairy cell Leukemia (cont) -Often when a bone aspiration is done in these individuals to find the cause of pancytopenia, often they have a dry tap on the aspiration because the fibrosis which is occupying the bone marrow. However if you take a bone biopsy you can clearly see these FRIED EGG APPEARANCE eggs, central blue staining nucleus with empty cytoplasm

-Hairy cell leukemia: Immunophenotype- Pan B cell markers CD 19 and CD 20 positive along with distinctive markers like CD103, and Annexin A1 (DO NOT NEED TO KNOW THIS). -Prognosis: indolent tumor, responds well to mild chemotherapy. -"Fried egg" appearance

A 13 yo boy is brought by his parents with complaints of difficulty in swallowing which is worsening, difficulty in breathing, intermittent fever and weight loss for the past few weeks. Physical examination reveals cervical lymphadenopathy, and petechial hemorrhages over the palate and limbs. A chest X-ray reveals enlarged thymus. His blood work up shows hemoglobin - 7.3/dl (12.5-14.5), total leukocyte counts of 100,000/cu mm (4500-11,000), platelets - 25,000/cumm (150,000-450,000/cu mm) and blasts of 80%. Image of the peripheral smear is given below. What is the most likely diagnosis? A. AML B. ALL C. CML D. CLL

B. ALL

Plasma cell neoplasms The pathogenesis with the kidney here is that intact immunoglobulins are not filtered through the kidney, but the light chains filter through and get deposited in the tubules and the glomeruli

-secrete a monoclonal Ig or Ig fragment and have pathologic consequences. -synthesize excess light chains along with complete Igs. -Ig kappa light chains are synthesized more than lambda -Light chains are small in size, they are also excreted in the urine, where they are referred to as Bence-Jones proteins. -Plasma cell disorders are: *Multiple myeloma *Waldenstrom macroglobulinemia: commonly associated with lymphoplasmacytic lymphoma -Monoclonal gammopathy of undetermined significance (MGUS)

A 65 yo male presents with complaints of weakness, easy fatigability, fullness in the sotmach and dragging sensation in the abdomen for the past 8 months. Physical examination shows pallor and splenomegaly. His blood work up is given Hb - 10.5 g/dl, TLC - 102,000/cu mm Platelets 500,000/cu mm Differential count shows Neutrophils 30% Band forms 10% Metamyelocytes 06% Myelocytes 25% Promyelocytes 04% Lymphocytes 07% Monocytes 05% Basophils 08% Eosinophils 05% What is the most likely diagnosis? A. Chronic Myelogenous Leukemia B. Polycythemia Vera C. Myelodysplastic syndrome D. Chronic lymphocytic Leukemia E. Leukemoid reaction

A. Chronic Myelogenous Leukemia

A 56 yo female presents with slowly enlarging neck mass for the past many months. She also states that some times the neck mass reduces in size, but it does not completely go away. Physical examination reveals enalrged spleen, but there is no liver enlargement. The biopsy of the lymph node reveals nodular proliferation of centrocytes and centroblasts with pale centsrs and these cells are positive for CD19, CD20, and BCL2 but negative for CD5. What is the most likley diagnosis? A. Small lymphocytic lymphoma B. Follicular lymphoma C. Diffuse large B cell lymphoma D. Burkittlymphoma E. Hair cell leukemia

B. Follicular lymphoma

A 62 yo male presents with fever, productive cough associated with green colored sputum. Patient also noted weight loss from few months and sensation of fullness after a light meal (early satiety). Physical examination reveals: splenomegaly and cervical lymphadenopathy. Blood tests: Hb-10.5g/dl (13.5-17.5 mg/dl), total leukocyte counts 35,000/cu mm (4,500-11,000), and platelets 175,000/cu mm (150,000-400,000). Hematologists reports spherocytes, abnormal lymphocytes with large number of smudge cells in the peripheral blood. Flow cytometry studies (blood) reveal that the abnormal lymphocytes are of B cell origin. What is the most likely mechanism for the reduction of hemoglobin levels and development of the spherocytes in this patient? A. Immune hemolytic anemia B. Suppression of the bone marrow C. Ineffective erythropoiesis D. Iron deficiency E. Megaloblastic anemia

A. Immune hemolytic anemia

A 56 yo female presents with slowly enlarging neck mass for the past many months. She also states that some times the neck mass reduces in size, but it does not completely go away. Physical examination reveals enalrged spleen, but there is no liver enlargement. The biopsy of the lymph node reveals nodular proliferation of centrocytes and centroblasts with pale centsrs and these cells are positive for CD19, CD20, and BCL2 but negative for CD5. What is the genetic abnormality associated with this disorder? A. t (8;14) B. t (14;18) C. t (11;14) D. t (15;17) E. t (9;22)

B. t (14;18)

A 62 yo male presents with fever, productive cough associated with green colored sputum. Patient also noted weight loss from few months and sensation of fullness after a light meal (early satiety). Physical examination reveals: splenomegaly and cervical lymphadenopathy. Blood tests: Hb-10.5g/dl (13.5-17.5 mg/dl), total leukocyte counts 35,000/cu mm (4,500-11,000), and platelets 175,000/cu mm (150,000-400,000). Hematologists reports spherocytes, abnormal lymphocytes with large number of smudge cells in the peripheral blood. Flow cytometry studies (blood) reveal that the abnormal lymphocytes are of B cell origin. What is the most likely diagnosis in this patient? A. Diffuse large B cell lymphoma B. Follicular lymphoma C. Chronic lymphocytic leukemia D. Acute lymphoblastic leukemia E. Viral infection

C. Chronic lymphocytic leukemia

Leukemia Vs Lymphoma Lymphoma - arises in the tissue Leukemia - arises in the blood -Nonhodgkins lymphoma is unpredictable - seen in elderly (except Burkitt's lymphoma) -Hodgkins lymphonma is predictable -bimodal age distribution (young and old)

Leukemia -Neoplasm of blood forming cells, primarily affecting bone marrow, can progress to involve soft tissue and lymph node -Age group: children (ALL), adults (AML), elderly (CML, CLL) -Presentation: Abrupt onset in acute leukemia, slow onset in chronic leukemia Lymphoma -Neoplasm of cells of immune system (B, T and NK cell) and affect LN, it can progress to involve BM. -Age group: can occur at any age but, NHL commonly affect adults and elderly (except Burkitt lymphoma, it is seen in children and young adults) and HL has bimodal age distribution (young and old). -Presentation; indolent onset for most of lymphomas (except high grade NHL- DLBCL and Burkitt lymphoma). -Leukemia can become lymphoma and lymphoma can become leukemia *T-Cell ALL - can involve your thymus where it is called a lymphoblastic lymphoma *CLL - when involving a tissue its called small cell lymphoma

A 30 yo male presents with complaints of fever, night sweats, chills, and pruritis from the past 6 weeks. Physical examination shows enlarged supraclavicular lymph node. Biopsy of the lymph node is given below. What is the most likely diagnosis in this patient? A. Small cell Lymphoma B. Burkitt lymphoma C. Lymphoblastic lymphoma D. Hodgkin lymphoma E. Follicular lymphoma F. Langerhans cell histiocytosis

D. Hodgkin lymphoma

Eosinophilia

Eosinophils >450 cells (0.45x109/ L) causes- -allergic disorders -parasitic infestations -skin disorders -Hodgkin lymphoma -collagen vascular disorders -hyper eosinophilic syndrome

Myeloproliferative neoplasms / MPNs

Few common features of MPN: -Mutated, constitutively activated tyrosine kinases. -Growth factor independent proliferation and survival of marrow progenitors. -Most of the myeloproliferative disorders originate in multipotent myeloid progenitors -Homing of the neoplastic stem cells to secondary hematopoietic organs Extramedullary hematopoiesis -Transformation to a spent phase characterized by marrow fibrosis and peripheral blood cytopenias. -Variable transformation to acute leukemia (AML/ALL)

Lymphocytosis

Lymphocytes > 4000 cells (4x109/ L) causes- -Viral infections -Toxoplasmosis -Chronic infections-Tuberculosis

Small Lymphocytic lymphoma

Microscopy: diffuse effacement of nodal architecture, replaced by predominantly small lymphocytes with round nuclei, condensed chromatin, and scant cytoplasm (picture A). Loose aggregates of larger lymphocytes (Picture B, black arrow) known as proliferation centers - pathognomonic of CLL/SLL

Polycythemia

Primary polycythemia -Polycythemia Vera Secondary polycythemia -Chronic lung disease: chronic obstructive lung disease in smokers -Cyanotic heart disease -Increased erythropoietin production by tumors: renal cell carcinoma, uterine tumors and cerebellar hemangioblastoma -Relative or apparent polycythaemia: where plasma is reduced, seen in dehydration (diarrhoea, vomiting, burns) and diuretic use.

Lymphoplasmacytic lymphoma Aka WALDENSTROM MACROGLOBULINEMIA Tumor arising from mature B cells which further mature into plasma cells. Hence the neoplastic component is both mature B cells and plasma cells -The clinical manifestations are due to excessive secretion of abnormal IgM, so classified under Plasma Neoplasms -Clinically these pt's do not have hypercalcemia, lytic bone lesions or renal failure but they do have the anemia

-A B-cell neoplasm of older adults: neoplastic cells are both B lymphocytes and Plasma cells -Age- 60-70 yr -The plasma cell component secretes monoclonal IgM, sufficient to cause hyper viscosity syndrome known as known as Waldenstrom macroglobulinemia -No bone destruction -Renal involvement is rare -Clinical feature: -Lymphadenopathy, hepatomegaly and splenomegaly. *Anemia: due to marrow infiltration or autoimmune hemolysis caused by cold agglutinins (IgM). Hyperviscosity syndrome: -Visual impairment, -Neurologic problems (headache, dizziness, and stupor) -Bleeding (formation of IgM + clotting factor complexes), -Cryoglobulinemia (precipitation of Ig at low temp -> Raynaud phenomenon and cold utricaria)

Multiple myeloma Plasma Cell Neoplasms CRAB symptoms -increased CALCIUM -RENAL failure -ANEMIA -Lytic BONE lesions

-A plasma cell neoplasm commonly associated with hyperCalcemia, Renal failure, Anemia, lytic Bone lesions (CRAB symptoms) and acquired immune abnormalities. -Incidence is higher in men and of African decent -Peak age of incidence is 60-70 yr -Pathogenesis: rearrangement involving IgH locus and various proto-oncogenes. -T(11;14) (cyclin D1; IgH), t(6;14) (cyclin D3;IgH) *Because of this, there is proliferation of abnormal immunoglobulins -Acquisition of MYC mutations -> plasma cell leukemia

Lymph node enlargement: lymphadenitis/ lymphadenopathy *Refer to immunology slides: Organs and Tissues of the Immune System -When a pt has skin infection, which part will show the changes? PARACORTEX (because the skin has numerous T lymphocytes) -Whenever there is an autoimmune disease, the cortex is expected to enlarge because B cells are expected to proliferate -Whenever there is an infection involving the mucosa (pharyngitis, tonsilitis), the cortex enlarges, because B cells are proliferating

-Acute or Chronic. -Increased large irregular lymphoid follicles with normal architecture. (mantle zone present). -Lymphadenitis: Enlarged lymph nodes due to inflammation or infections. Painful. *Can be acute: Acute bacterial infection like staph *Can be chronic: Like RA, autoimmune diseases *In chronic infections there is chronic enlargement of the lymph node which is usually painless. *If the infection is acute, the lymph nodes will be painful, in chronic infection the lymph nodes will be painless -Lymphadenopathy - any enlargement- may be Painless. *Ie metastasis from the breast *Or any lymphoma -Etiology: *Infections: bacterial, viral or fungal *Immune: rheumatoid arthritis, *Cancers: reactive/metastasis. Germinal Center: light zone WITHIN the Mantle zone: dark zone WITHIN the Marginal zone

Adult T-cell Leukemia/lymphoma A Mature T Cell Lymphoma -Most T cells are present in the skin -Most B cells are present in the mucosa

-Adults infected with human T-cell leukemia retrovirus type-1 (HTLV-1) -Endemic areas: Southern Japan, West Africa, and the Caribbean basin. -Clinical presentation: skin lesions (erythmatous plaques/ nodules), generalized lymphadenopathy, hepatosplenomegaly, peripheral blood lymphocytosis, and hypercalcemia -Pathogenesis: HTLV-1 encodes a protein, Tax a potent activator of NF-κB (a transcription factor) -> enhances lymphocyte growth and survival. -Morphology: cells with multilobated nuclei ("cloverleaf" or "flower" cells) -Malignant lymphocytes spill into the blood now called as leukemia -This leukemia/lymphoma is very aggressive -Prognosis: rapidly progressive aggressive diseases.

Clinical presentation of acute leukemia

-Age: young adults or children - ALL , adults - AML -Abrupt onset (days to few weeks) -Decreased Hematopoiesis: *Erythropoiesis - decreased RBC- Anemia and increase in fatigue. *Leukopoiesis - decreased WBC - fever and Infections *Thrombopoiesis - decreased PLT - excess Bleeding or bleeding from/in unusual sites -Bone marrow expansion/destruction: *Bone pains. -Extramedullary Hematopoiesis / spread: *Splenomegaly- early satiety *Hepatomegaly- dragging sensation or pain in right part of the abdomen *Lymphadenopathy (more in lymphoblastic malignancy) -Headache, vision changes, nonfocal neurologic abnormalities (CNS spread or bleeding due to low platelet count)

Differentiation of WBCs

-All lymphocytes derive from a common progenator cells - haemopoetic cells -These haemopoetic cells are present in bone marrow (specifically in the medulla/spongy bone) -These cells require factors for their proliferation and to maintain their stem cell pool (ie. stem cell factor and Ligand for FTL3) -These substances are secreted from the cells also present in the bone marrow which help in maintain the stem cell pool and help with differentiation -When the cells commit, they commit either to a lymphoid series or a myeloid series -Lymphoid: They mature to T cell, B - cells, or NK cells -Myeloid: They mature into granulocytes, erythrocytes or megakaryotype series -When it matures into granulocyte series it can become neutrophils, basophils or eosinophils -When it commits, it can also produce monocytes -Similarly when you see a neoplasm arising from these cells, it has certain characteristics *Ie. if there is leukemia arising from a hemopoetic cell, it will not show any differentiation or markers. However if it is arising from an early progenitor cell with lymphoid potential, it will show markers specific to lymphoid series and hence called ALL or ACUTE LYMPHOBLASTIC LEUKEMIA. When it is arising from the progenitor with myeoloid potential it is called AML, ACUTE MYEOLID LEUKEMIA. There are also leukemias arising from the mature cells, like chronic leukemias arising from the granulocyte series called CHRONIC MYELOGENOUS LEUKEMIA, POLYCYTHEMIA VERA (arising from erythrocyte series), ESSENTIAL THROMBOCYTOSIS (arising from megakaryocyte series), CML(arising from granulocyte series), etc.

ANEMIAS

-Anemia : reduction below normal limits of the total circulating red cell mass -Classification of Anemia according to underlying mechanism: *Blood Loss: acute or chronic *Increased Rate of Destruction (Hemolytic anemia) *Decreased / Impaired Red Cell Production

Essential thrombocytosis (ET)

-Associated with activating point mutations in JAK2 (50% of cases) or MPL (5% to 10% of cases). -No polycythemia and no marrow fibrosis. -Causes of reactive Thrombocytosis has to be ruled out. -Causes of reactive thrombocytosis, bleeding, inflammatory disorders and iron deficiency, must be excluded before the diagnosis of ET -Clinical presentation: Age after 60yr. -Due to thrombocytosis: thrombosis of blood vessels (MI, Hepatic vein, DVT) and gangrene. -Eryhtromelalgia; a throbbing and burning pain in hands and feet caused by occlusion of small arterioles by plts. -Mortality is due to thrombotic complication -Increased platelets in peripheral blood With abnormal large platelets (black arrow)

Precursor cell neoplasm: ALL/L (B cell and T cell)

-Blood tests: Anemia, Thrombocytopenia, leukocytosis (total WBC count > 1,00,000) -Blasts are present in peripheral blood, Neutropenia (+). -Blasts: nuclear size is large. Nuclear chromatin is delicate and finely dispersed, less conspicuous nucleoli, scant cytoplasm, no cytoplasmic granules. -Blasts are MPO negative and PAS positive -Bone marrow: Hypercellular, replaced by blasts. -Mitotic rate is high = Aggressive tumor

Multiple myeloma: Investigations Xray shows multiple lytic lesions involving the bones of the skull

-CBC; anemia, elevated ESR, normal or decreased WBC count, and platelet count. -Serum calcium increased -S. Creatinine and BUN: elevated -Urine: proteinuria and increased excretion of immunoglobulin light chain > 6 gm/dl *Usually not picked up with dipstick test -Serum electrophoresis; M protein >3 gm/dl *M protein is immunoglobulin G which is more than 3 gm/dl

G6PD deficiency

-G6PD reduces NADP to NADPH, while oxidising Glucose-6 -phosphate -NADPH then provides the reducing power and converts oxidized glutathione to reduced glutathione *The reduced glutathione protects against oxidant injury by catalyzing the breakdown of oxidant compounds, e.g. H2O2 *Synthesis of NADPH is dependent on activity of G6PD. -Removal of H2O2, a potential oxidant, is dependent on the adequacy of reduced glutathione (GSH),which is generated by the action of NADPH

Differences between HL and NHL

-HL arises in a single node or chain of nodes, and spreads first to anatomically contiguous lymphoid tissues. -Characteristic histological pattern: RS cells -NHLs frequently occur at extranodal sites and spread in an unpredictable fashion

Leukemoid reaction -You have to identify a leukemoid reaction from leukemia -When you see a blood count, it could be very similar and look like leukemia so you won't know whether to call it a reactive condition or a malignant condition -In the Leukemoid reaction is the reactive response of the bone marrow to reactive stimuli -Here you will see immature cells in the peripheral blood You'll know if it is reactive if you see DOHL BODIES, TOXIC GRANULES, OR CYTOPLASMIC VACULATION! In acute myeloid leukemia - you'll see basophilia In polycythemia vera - you'll see incrased platelets, leukocytes and RBCs

-Cause: infection , severe hemorrhage or acute hemolysis, stress-post op/trauma, post splenectomy. -WBC count > 15,000 cells/cumm - 50,000 cells/ cumm , can reach up to 100,000 cells/cumm. -Mature neutrophils, Left shift (+). *You will see more mature cells than immature cells -Neutrophils show toxic granules, cytoplasmic vacuolations and Dohle bodies -Blasts (myeloid blasts) are absent. -LAP (leukocyte alkaline phosphatase) score is increased. -In viral fever (infectious mononucleosis); atypical lymphocytes seen in peripheral blood, which can be confused for leukemic cells. -Differential diagnosis: *Chronic myeloid leukemia (LAP score decreased & Basophilia +), *Polycythaemia Vera (LAP score is increased & hemoglobin increased), *Leukemia (blasts are present & LAP score decreased)

Acute Myeloid Leukemia

-Cell of origin: hematopoietic progenitor cells. -It primarily involve the bone marrow . -Usually present with symptoms related to altered hematopoiesis. -Children with trisomy 21 (Down's syndrome) have an increased risk for childhood acute lymphoblastic leukemia as well as acute myeloid leukemia. -Recent WHO classification is based on presence of cytogenetic abnormalities, dysplasia and therapy associated AML. -The Important types to consider are: *AML with t(8;21), has full range of myelocytic maturation. *Acute Myelocytic and Monocytic leukemia with inv(16) *Acute promyelocytic leukemia with t(15;17) *AMLs following myelodysplastic syndromes or exposure to DNA-damaging agents (chemotherapy or radiation therapy) have deletions involving Ch 5 and 7.

Myelodysplastic Syndromes

-Characterized by maturation defects -Associated with ineffective hematopoiesis -Increased risk for transformation to AML. -Has to be differentiated from Megaloblastic anemia -Primary (idiopathic): mean age of onset is 70 years -Secondary to previous genotoxic drug or radiation therapy (t-MDS): appear 2-8 yr after the therapy -Clinical presentation: Asymptomatic or weakness, infections, and hemorrhages (all due to pancytopenia). -Multiple abnormalities seen including Pseudo-Pelger-Huet cells, neutrophils with 2 nuclear lobes (green arrow).

Chronic lymphocytic leukemia / small lymphocytic lymphoma: CLL/SLL *A type of PERIPHERAL LYMPHOMA

-Chronic lymphocytic leukemia (CLL): tumor cells in peripheral blood -Small lymphocytic lymphoma (SLL): tumor cells involving lymph node -Age: 60 yr -M:F= 2:1 -Clinical presentation: Asymptomatic, or infection or anemia. -Generalized lymphadenopathy and hepatosplenomegaly in 50-60% cases. -CLL/SLL associated with immune dysregulation: *Hemolytic anemia (immune), *thrombocytopenia (immune) and *Hypogammaglobulinemia which increases the susceptibility for infection. -Pathogenesis: deletions of 13q, and trisomy 12.

AML

-Clinical presentation: Average age of onset is 60 years. Uncommon before 45 years -presents within few weeks to moths after the onset of disease -Features of : anemia, neutropenia, and thrombocytopenia -Infection with: Pseudomonas, fungi and commensals. -Gum hypertrophy and bleeding in Monocytic leukemia -DIC in promyelocytic type.

Hodgkin lymphoma (continued)

-Clinical presentation: Painless lymphadenopathy (most common) . -Nodular sclerosis or lymphocyte predominance types: have stage I-II disease and are usually free of systemic manifestations. -Disseminated HL (stages III-IV) or mixed-cellularity or lymphocyte depletion subtypes are more likely to have constitutional symptoms. -Constitutional symptoms (B symptoms): fever, night sweats, weight loss. Spread of HL is stereotypical : nodal -> splenic -> hepatic -> BM -> extranodal -Investigation needed for staging: physical examination, radiologic imaging of the abdomen, pelvis, and chest, and biopsy of the bone marrow. -Staging: prognostic value, also important in choice of therapy -Ann Arbor clinical staging of Hodgkin and Non-Hodgkin lymphoma

Hodgkin lymphoma : Mixed-Cellularity

-Constitutes 20% to 25% of cases -Biphasic age distribution ; young adults and older age (>55) -Systemic symptoms such as night sweats and weight loss, and advanced stage at presentation -Morphology: LN is diffusely effaced by a heterogeneous cellular infiltrate, which includes T cells, eosinophils (arrow head), plasma cells, and benign macrophages admixed with mononuclear variant of RS cells. -Immunophenotype: same as nodular sclerosis type , infected with EBV.

Polycythemia Vera (PCV)

-Diagnosis of PCV: -Hemoglobin in F: > 16.5 g/dl and M:>18.5 g/dl Presence of JAK2 mutations -Marrow biopsy: Hypercellular (for age) with increase in all the three lineages. -Serum erythropoietin : low -Absence of other causes for (secondary) polycythemia. -Other lab findings: Total WBC count range from 12,000-50,000, Platelets > 500,000 with abnormal forms seen (giant forms) -Natural history and complications: 25% of patients first come to attention due to deep venous thrombosis. -~20% progress to terminal stage and develop myelofibrosis (marrow failure with EMH) , ~ 10 % can develop AML.

Mycosis Fungoides / Sézary Syndrome

-Different manifestations of a tumor of CD4+ helper -T cells that home to the skin. Mycosis Fungoides: -Clinical presentation: skin lesion progress through three distinct stages. -An inflammatory premycotic phase: erythmatous patches -A plaque phase -A tumor phase -Microscopy: epidermis and upper dermis are infiltrated by neoplastic T cells, with cerebriform nuclei -Late in the disease tumor cells spread to lymph node and BM

Chronic Myelogenous Leukemia (CML)

-Distinguished from other myeloproliferative disorders by the presence of a chimeric BCR-ABL gene. -Pathogenesis: In more than 90% of cases, BCR-ABL is created by a reciprocal (9;22)(q34;q11) translocation (the so-called Philadelphia chromosome [Ph]). -Clinical presentation; primarily a disease of adults, peak incidence is in the 50-60 yr of life. -Insidious onset: mild-to-moderate anemia and hyper-metabolism due to increased cell turnover -> fatigability, weakness, weight loss, and anorexia. -A dragging sensation in the abdomen caused by splenomegaly, or the acute onset of left upper quadrant pain due to splenic infarction. -Three phases of disease: chronic phase, accelerated phase and Blast crisis.

Hemolytic Anemias (HA)

-Examples of Intrinsic hemolytic anemia : -Hereditary spherocytosis -G6PD deficiency -Paroxysmal nocturnal hemoglobinuria -Sickle cell anemia -Examples of Extrinsic hemolytic anemia : -Autoimmune -Microangiopathic -Infections

Normal Bone marrow (BM)

-Hematopoietic cells; erythroid, mature megakaryocytes and mature myeloid cells in the center around the sinusoids. -Immature myeloid cells and megakaryocytes near trabeculae/ paratrabacular location. -White balls = adipocytes -Pink = bony trabeculae -Normal cellularity (for age) %= 100-Age *As the age progresses, cellularity decreases *Ie. A 25 yo individual undergoing bone marrow biopsy was told they have normal cellularity on biopsy. This means this individual has a proliferating population of hemopoetic cells occupying 75% of the bone marrow -Bone biopsy done to check irregularities of the bone marrow -Bone aspiration is to count the blast in the bone marrow evaluating for leukemia -Bone marrow biopsy plays a very important role in diagnosis of any malignancies -The mature megakaryocytes, their cytoplasm buds off to give platelets, they are situation around the sinosoids -Mature myeloid elements, granulocytes, are also situated near the sinosoids, usually situated in the center of the pockets -More immature cells are mostly arranged around the trabeculae -The erythroid series, usually associated with macrophage, called nursing cells, because they help with maturation of erythroid series by providing various growth factors and IL and iron

G6PD deficiency

-Hemolysis sequence: infection or exposure to oxidants → oxidation of globin chains, then denaturation of Hb, and formation of 'Heinz bodies'. These damage the membrane and cause an intravascular hemolysis -Less severe membrane damage may lead to → decreased RC deformability -'Bite cells' : macrophages pluck out the Heinz bodies in the spleen, & a bite of cytoplasm appears to be removed

Hemolytic Anemias (HA)

-Hemolytic anemias are further classified according to the : -Cause of the hemolysis (intrinsic vs extrinsic to the RBC ) & by the -Location of the hemolysis (intravascular vs extravascular)

Hemolytic Anemias

-Hereditary Spherocytosis -Red Cell Enzyme Defects: Glucose-6-Phosphate Dehydrogenase Deficiency -Sickle Cell Disease -Thalassemia Syndromes -Paroxysmal Nocturnal Hemoglobinuria -Immunohemolytic Anemia -Hemolytic Anemia Resulting from Trauma to Red Cells

Diffuse large B cell lymphoma -Many ways to acquire it but they all have one feature in common which is presence of large malignant cells If Richter syndrome -Transformation of CLL into DLBCL -There will be CD5 and CD23 Follicular Lymphoma to DLBCL -There will BCL2 expression Mutations of BCL6 -There will be BCL6 expression

-Histopathology: diffuse infiltrates of large B cells which are malignant *When we say large cell, it is 5x more of that of a small lymphocyte -Round to oval vesicular nucleus (black arrow), some are multilobated or cleaved. Moderate amount of pale cytoplasm. *Vesicular means a nucleus which is empty -Immunophenotype; CD19 and CD20 (+) and germinal center B-cell marker like CD10 and BCL6 (+) -DLBCL are aggressive and rapidly fatal require aggressive chemo.

Toxic Change/Toxic Granules If you see any of the 3 changes: 1. TOXIC CHANGE/ DARK or TOXIC GRANULES 2. CYTOPLASMIC VACULATION 3. DOHL BODIES They are associated with INFECTION!

-If these are seen in a patient with increased neutrophil count, this lets you know that the pt HAS AN INFECTION! -The dark, course, toxic granules can be seen in the cytoplasm of the neutrophils -CYTOPLASMIC VACULATION - this is also a change that is seen in the neutrophil during infection *They are situated near the nucleus -You will also see DOHL BODIES which are enlarged ER present in the cytoplasm of the neutrophil

Follicular lymphoma (cont) Normally in a lymphoid follicle there is no BCL2 expression in the germinal center as it is minimal or not present, but with follicular lymphoma you will see the center of the follicles with increased BCL2 expression

-Immunophenotype: Pan-B cell markers CD19 and CD20 positive. *This is a tumor arising from a mature B cell so they will be positive for the markers -BCL2 Positive (>90% cases) help to distinguish follicular hyperplasia from follicular lymphoma

In contrast to Myeloblasts

-In contrast to Lymphoblasts, -Myeloblasts: have *More voluminous cytoplasm *Prominent nucleoli, delicate chromatin *Fine azurophilic cytoplasmic granules *MPO positive, PAS negative

Extravascular hemolysis in HA

-In the majority of HAs, premature destruction of red cells occurs within phagocytes, known as extravascular hemolysis. If persistent, manifested by splenomegaly -Extravascular hemolysis: caused by alterations that render the red cell less deformable -The principal clinical features of extravascular hemolysis are: anemia, splenomegaly and jaundice. *Patients often benefit from splenectomy -(No hemoglobinuria and no hemosiderinuria present )

Polycythemia Vera (PCV)

-Increased marrow production of red cells, granulocytes, and platelets (panmyelosis). -Pathogenesis: a point mutation in JAK2. -Has to be distinguished from other causes of polycythemia (secondary). -Serum erythropoietin levels in PCV are low, whereas secondary forms of polycythemia have high erythropoietin levels. -Clinical presentation Insidious onset; in adults Most symptoms are related to the increased red cell mass and hematocrit (>60) -Increased viscosity of blood -> abnormal blood flow, Headache, dizziness, hypertension. -Patients are plethoric and cyanotic (due to deoxygenated blood in peripheral circulation) -Intense pruritus and peptic ulceration due to release of basophilic granules

Hereditary Spherocytosis (HS)

-Inherited AD disorder: intrinsic defect in the red cell membrane -Diverse mutations may be present → an insufficiency of membrane skeletal components -Red cells are spheroidal and less deformable → splenic sequestration and destruction -Clinical: anemia, splenomegaly and jaundice -Various proteins including spectrin, ankyrin and glycophorin A maintain the normal shape, strength and flexibility of the RC membrane -In HS, the interactions between proteins weakens -RBCs are exposed to shear stresses in the circulation, and this leads to a loss of membrane fragments resulting in a spheroidal red cell

AML with monocytic differentiation

-Lack Auer rods; usually myeloperoxidase negative -Nonspecific esterase positive (NSE): positive -Folded or lobulated nuclei -Often infiltrate skin (leukemia cutis) & gingiva, other organs, lymphadenopathy -Clinical presentation: gingival hypertrophy, extravasation of abnormal monocytes in to tissue.

Intravascular hemolysis in HA

-Less common is intravascular hemolysis : rupture or lysis of red cells within the circulation -Causes : *Mechanical hemolysis e.g. prosthetic valves, cardiac valves, thrombosis in microcirculation (MAHA) *Intracellular parasites (e.g. malaria ) *PNH (paroxysmal nocturnal hemoglobinuria) -Clinical : anemia, decreased haptoglobin, hemoglobinemia, hemoglobinuria, hemosiderinuria, jaundice -Large amounts of free Hb are released from lysed RBCs & are immediately bound by haptoglobin. -As serum haptoglobin is depleted, free Hb oxidizes to methemoglobin, which is brown in color and some passes out in the urine leading to a → red-brown color -Concomitantly, heme groups are catabolized to bilirubin leading to jaundice. -Unlike extravascular hemolysis, splenomegaly is not seen

Langerhans Cell Histiocytosis (LCH)

-Letterer -Siwe disease / multifocal multisystem LCH: -Age < 2yr (MC) occasionally affects adults. -Erythmatous, scaly lesions over trunk , abdomen and scalp. -Hepatomegaly, splenomegaly, lymph node enlargement and destructive osteolytic bone lesions -Recurrent infections: otitis media and mastoiditis -Fatal if untreated -Unifocal unisystem LCH : most commonly affect skeletal system of older children and adults . -Arises with-in the medullary cavities of bones: calvarium, ribs and femur -Multifocal unisystem LCH also known as Hand-Schuller-Christian disease : -Clinical triad of calvarial bone defects, diabetes insipidus (involvement of posterior pituitary stalk), and exophthalmos.

Leukemia: Cancer of blood cells

-Leukemia arising from common progenator cells - lymphoid series : Acute Lymphatic Leukemia -Leukemia arising from mature B or T cells: Chronic Lymphocytic Leukemia Acute: arising from immature cells Chronic: arising from mature cells -Same concept with the myeloid series

Leukocytosis: Neutrophilia

-Leukocytosis: increased number of neutrophils in the blood (absolute neutrophil count > 6,500 cells or >6.5 X 109 / L). Causes -Acute inflammation; stimulation o f BM by TNF, IL-6/1 in bacterial Infection - Chronic inflammation: growth factor dependent

Clinical presentation of lymphoma

-Lymphadenopathy: painless and slowly progressive *Most commonly seen in Hodgkins Lymphoma. Non-hodgkins can have this but is most commonly presented with extranodal involvement -Weight loss, fever, night sweats (B symptosm) *Both in HL and NHL Extra-nodal involvement: common sites are -The GI tract (including the waldayer ring) -Skin -Bone marrow -Sinuses -GU tract -Thyroid -CNS

Monoclonal gammopathy of undetermined significance (MGUS)

-MGUS is a most common plasma cell dyscrasias, 3% of individuals >50 yrs, 5% individuals >70 yrs. -Definition: Asymptomatic and the serum M protein levels < 3 gm/dl -1% progress in to Multiple myeloma /yr

Hodgkin lymphoma: Lymphocyte Predominance Type (non-classical HL)

-Majority of patients are males, usually younger than 35 years of age -Typically present with cervical or axillary lymphadenopathy. -Not associated with EBV -Morphology: *nodular infiltrate of small lymphocytes admixed with variable numbers of macrophages, *L-H variant of RS cells which have a multilobed nucleus resembling a popcorn kernel ("popcorn cell"). -Immunophenotype: CD15 and CD30 Negative. -CD 20 and BCL 6 positive

Adult reference ranges for Red Cells*

-Mean cell volume (MCV): the average volume of a red cell -Mean cell hemoglobin concentration (MCHC ) : the average concentration of hemoglobin in a given volume of packed red cells

Burkitt lymphoma (BL) continued -This tumor arises from mature B cells -Fastest growing tumor in the body, you will see A LOT of apoptosis -On histology: You will see a lot of monomorphic tumor cells with macrophages in between trying to clear away all macrophotic cells (the black arrow). The light stain around the highly proliferating tumor cells are called STARRY SKY PATTERN -These are intermediate sized tumor cells (1-2 times larger than a small lymphocyte), with very high mitotic activity -These lymphoma cells can spill over to the blood where its called Burkitts Leukemia

-Microscopy: diffuse infiltrate of intermediate sized cells. -With high mitotic activity (picture B, green arrow) and numerous apoptotic cells. -Apoptotic cells phagocytised by macrophages and cytoplasm contain nuclear debris (picture A, black arrow) creates "starry sky" pattern. -When tumour cells spill into blood called Burkitt leukemia (L3) and show characteristic morphology - cytoplasmic vacuolation

Multiple myeloma (continued) -Clinically the pt will have altered renal function test, increased calcium levels. -Bone marrow aspiration will show increased plasma cells -Some plasma cells remain their morphology (eccentric nucleus with cogwill type of nucleus, clumped chromatin, with perinuclear halo) -You also see abnormal, binculeate forms of plasma cells.

-Morphology: Bone marrow plasma cells increased in number which counts for >30% marrow cellularity (normal 2-6%). -Plasma cell morphology: round eccentrically placed clumped nuclei with basophilic cytoplasm and a perinuclear clearing (Golgi apparatus) -Abnormal plasma cells can spill in to peripheral circulation -> plasma cell Leukemia -Immunophenotype: Positive for CD138 (syndecan -1) and CD56.

Lymphoplasmacytic lymphoma (cont) -Has both mature B cell and plasma cell component -In the bone marrow there will be increase in both mature lymphocytes that are neoplastic in origin as well as plasma cells

-Morphology: the marrow contains an infiltrate of lymphocytes, plasma cells, and plasmacytoid lymphocytes in varying proportions. -Tumor cell involvement seen in lymph node, spleen, and liver. -Immunophenotype: tumor cells show B-cell maturation CD20, CD 19 and cytoplasmic IgM +ve

Hodgkin lymphoma : Nodular Sclerosis

-Most common form of HL -65-70% -Equal incidence among males and females, young adults are more commonly affected -Has a propensity to involve the lower cervical, supraclavicular, and mediastinal lymph nodes -Characterized by presence of Lacunar type of RS cells and deposition of dense collagenous bands (green arrow) dividing lymph node in to lobules -Immunophenotype: positive for PAX 5, CD15 and CD30. CD45 (common leukocyte antigen) negative -Excellent prognosis

Mantle cell lymphoma -Arise from Late Pre-B cells -You must be able to identify Mantle Cell Lymphoma from CLL because Mantle requires much more aggressive treatment -Mantle zone in the germinal center. Mantel cell zone is homogenous and small cells. CD5 + and CD23-

-Uncommon lymphoid neoplasm in USA (2.5%) -Age 50-60 yr , with male predominance -Clinical presentation: The most common presentation is painless lymphadenopathy or generalized lymphadenopathy. *Splenomegaly in 50 % cases. *GIT involvement also seen in the form of polyps (lymphomatoid polyposis). -Pathogenesis: virtually all have t(11;14) (Cyclin D1;IgH) -Morphology; proliferation of a homogeneous population of small lymphocytes.

Diffuse large B cell lymphoma (DLBCL) -Heterogenous group of neoplasms because there is no specific etiology -This is an aggressive, high grade tumor

-Most common form of high grade NHL in USA -Median age of presentation ; 60 yr. -Clinical presentation; Rapidly enlarging mass at nodal or extra nodal site. -Most common nodal site: waldeyer ring, the oropharyngeal lymphoid tissue (tonsils and adenoid). -Extra nodal sites include: GIT (most common), Liver, spleen, skin, bone, brain. *However it can involve the lymph node which is there in the oropharynx -Special subtypes: *Immunodeficiency associated large B cell lymphoma: advanced HIV, allogeneic BM transplantation. Associated with EBV infection. **Ie. People with HIV/AIDS will often come in with CNS or liver/spleen involvement *Primary effusion lymphoma: KSHV/HHV-8 infection. Advanced HIV infection or in older adults. **Primary effusion lymphoma is a lymphoma arising in the serous cavity, seen in advanced states of HIV or HHV-8/Kaposi Sarcoma *Pathogenesis: dysregulation of BCL6 (30%), other- t(14;18) with overexpression of BCL2, and MYC mutation. *Development of DLBCL from preexisting CLL - Richter Syndrome *A patient with Follicular Lymphoma and there is translocation involving BCL2 and it can develop into DLBCL

Follicular Lymphoma -2nd most common type of Non-hodgkins lymphoma -Low grade lymphoma

-Most common form of indolent (low grade) NHL in USA. -Cell of origin: Germinal center B cells -Age: middle age -M:F= equal incidence Clinical presentation: painless generalized lymphadenopathy, with waxing and waning course. *Not associated with infection or inflammation outside of the lymph node enlarged for a large period of time -Pathogenesis: strong association with ch translocations involving BCL2 *Here there is increased expression of BCL2 *BCL2 is an anti-apoptotic protein in the intrinsic pathway. It inhibits the escape of cytochrome C and ROS from the mitochondria -T(14;18) IGH;BCL2, and MLL2 mutation (90%) -Prognosis; median survival 7-9yr -Transformation to DLBCL occurs in 30-50% cases.

Hereditary Spherocytosis (HS)

-Most specific morphologic feature: spheroidal shaped & abnormally dark staining (hyperchromic ) small RBCs that lack a central zone of pallor -Reduced deformability, so RBCs get trapped in the splenic cords and lead to → Splenomegaly -Increased MCHC due to dehydration, caused by loss of H2O -Lab: RBCs show increased susceptibility to osmotic fragility (lysis) and rupture easily -Other features : reticulocytosis, erythroid hyperplasia of the BM & mild jaundice. Pigmented gallstones may be seen -Most patients benefit from Splenectomy

Mucosa-associated lymphoid tumors/ "MALToma" Extranodal arising from the MARGINAL ZONE -Treatment at the appropriate stage can cause regression of this tumor as these conditions only cause the maltoma because of continuous inflammation Additional notes: The disease begins as a polyclonal immune reaction -> acquisition of initiating mutations -> B cell clonal population emerges -> this clonal population is dependent on antigen stimulated T helper cells for growth and survival.

-Mucosa-associated lymphoid tumors is a Marginal Zone Lymphoma -Arise in the tissue involved by chronic inflammation -Either autoimmune or infectious: *Salivary gland in Sjogren disease, *The thyroid gland in Hashimoto Thyroiditis, and *The stomach in Helicobacter gastritis -They remain localized for prolonged periods. -They may regress if the inciting agent (e.g., Helicobacter pylori) is eradicated

Sezary Syndrome

-Mycosis Fungoides + leukemia (spilling of tumor cells in to blood) -Mycosis Fungoides / Sezary Syndrome immunophenotype : CD2, CD3, CD5, CD4 positive and CD8 negative

CML

-Natural history and progression: Slow progression; even without treatment, the median survival is about 3 years. -50% of patients enter an "accelerated phase": increased basophil count (>20% in PB), Myeloblasts (10-19%) and then blast crisis phase. -In other 50% of patients, blast crises occur abruptly without an accelerated phase: peripheral blood blasts equal/more than 20% with extramedullary hematopoiesis. -In Blast crisis: 70% develop AML, 30% develop B cell-ALL

Primary myelofibrosis (PMF)

-Obliterative marrow fibrosis. -Age > 60 yr *Early stage: clinical presentation and morphological features are similar to PCV and ET *Late stage: progressive anemia and splenomegaly (fullness in left upper quadrant and after food). *Non-specific symptoms: fatigue, weakness, weight loss, night sweats. -Pathogenesis: activating JAK2 mutations -Increased secretion of TGF beta from marrow cells with JAK2 mutation -> fibrosis As marrow fibrosis progresses, circulating hematopoietic stem cells take up residence in secondary hematopoietic organs -> extra-medullary hematopoiesis -> hepatomegaly, splenomegaly.

Anemia

-On the basis of MCV (Mean cell volume ) I : Microcytic anemia (MCV < 80 fL) -Iron deficiency -Anemia of chronic 8disease (late stages) -Thalassemia -Sideroblastic anemia II: Macrocytic anemia (MCV > 100 fL) -Vitamin B12 deficiency -Folate deficiency -Reticulocytosis -Myelodysplastic syndrome III. Normocytic anemia (MCV 80-100 fL) -Acute blood loss (< 7 days) -Aplastic anemia -Renal disease

Mature Lymphocytes - markers for Lymphomas

-Pan B Cell Markers expressed on a mature B cell: -CD19 -CD20 -CD21 -Cd22 -These are messed up for Burkitt Lymphoma and Follicular Lymphoma -Usually when B cells become Plasma cells they lose these markers and get CD38 and CD138 -Late B Cells: -CD19 -CD10 -CD2 -NO TdT -This is messed up in: *Mantle Cell Lymphoma *CLL - maturation arrest -You differentiate them from each other because CLL is + for CD23, and Mantle cell does not express that

Hodgkin lymphoma (continued)

-Pathogenesis: -Activation of the transcription factor NF-κB is a common event in classical HL: -Activated by EBV infection (seen in 70%) -EBV+ tumor cells express latent membrane protein-1 (LMP-1) it up-regulates NF-κB. -Loss-of-function mutations involving negative regulators of NF-κB. -Accumulation of reactive cells in HL, is in response to cytokines and chemokines secreted by RS cells : IL-5. IL-10, M-CSF, Eotaxin, and immunomodulatory factor galectin-1. -Reactive cells secrete factors that support the growth and survival of RS cells. -Eosinophils and T cells express ligands that activate CD30 and CD40 receptors on RS cells and activation of transcription factor NF-κB.

Glucose-6-phosphate Dehydrogenase (G6PD) deficiency

-X-linked -G6PD deficiency : reduced ability of the red cell to protect itself against oxidative injury→ hemolytic disease -Common feature : Hemolysis occurs after exposure to oxidant stress Many variants identified -G6PD 'B': normal type -G6PD 'A': in about 10% of American blacks -The enzyme activity is normal in reticulocytes, but markedly deficient in older red cells

Common Leukocyte Antigens How to differentiate lymphoid from myeloid series - done by TdT! -TdT is present in immature cells which are commited to the lymphoid series -It then matures to either Pre-B or Pre-T cells -When it goes further into maturation to the Pre-B or Pre-T cells, the acquired certain antigens will tell you whether they are B cell lineage (CD19, CD10) or T cell lineage (CD2, CD7). Here they are also still positive fore TdT -As they mature into B cells and T cells, they lose the TdT *So if you see a TdT positive cell, it is an immature cell *Ie. ALL is TdT +, CD19 -, CD2- *Ie. B-Cell ALL is TdT+, CD19+, CD2- *Ie. T-Cell ALL is TdT+, CD19-, CD2+

-Pathway of normal B-cell differentiation and relationship to B cell lymphomas. -Terminal deoxynucleotidyl transferase (TdT) -expressed in immature, pre-B, pre-T lymphoid cells, and acute lymphoblastic leukemia/lymphoma cells. -CD10, CD 19, Cd20, CD21, CD22 are cell markers used to distinguish B cells at various stages of development and the stage of differentiation associated with particular lymphoma

Chronic lymphocytic leukemia: Peripheral blood findings

-Peripheral blood: increased leukocyte count and increase in absolute lymphocyte count. -smudge cells (green arrow). -The Spherocytes (red arrow) indicating immune hemolytic anemia. The BM is almost always involved by tumor cells either in the form of aggregates or infiltrates. -Can transform in to DLBCL (~5-10% cases) k/as Richter syndrome, this is heralded by rapidly enlarging LN or spleen -Immunophenotype: *Positive for Pan-B cell markers: CD19 and CD 20 *Positive for CD23 and CD 5

Hemolytic Anemias (HA)

-Premature destruction of red cells (RBCs): shortened life span i.e. less than < 120 days Features : -Accumulation of products of Hb catabolism: bilirubin -Compensatory increase in erythropoiesis within the BM, & an increase in EPO levels -Reticulocytosis in the peripheral blood Image : Marrow smear from a patient with hemolytic anemia. Increased numbers of maturing erythroid progenitors (normoblasts)

Langerhans Cell Histiocytosis (LCH)

-Proliferation of immature dendritic cells. -Most common mutation is activating mutations involving BRAF. -Morphology of neoplastic Langerhans cells: abundant vacuolated cytoplasm, with linear nuclear folds or grooves, surrounded by reactive eosinophils, lymphocytes, plasma cells and neutrophils. -Immunophenotype: CD1a, S-100 positive -Clinical presentation: Three forms 1. Letter -Siwe disease / multifocal multisystem LCH 2. Unifocal unisystem LCH or Multifocal unisystem LCH also known as Hand-Schuller-Christian disease : eosinophilic granuloma 3. Pulmonary LCH: seen in adult smokers, regress spontaneously on cessation of smoking.

SICKLE CELL DISEASE (SCD)

-Prototype of hereditary hemoglobinopathies *Point mutation → valine for glutamic acid at the 6th beta-globin chain leads to a → structurally abnormal HbS, & SCD -About 8% to 10 % of African - Americans are heterozygous for HbS (SC trait) *In homozygotes, almost all the Hb is HbS (SCD) -HbS molecules undergo aggregation & polymerization when deoxygenated→ long needle-like fibers within red cells→ a distorted sickle or holly-leaf shape -Major pathologic manifestations: 1. Chronic hemolysis 2. Microvascular occlusion 3. Tissue damage

Neutropenia ABSOLUTE COUNT should be less than 1,400

-Reduction in number of neutrophils in blood (absolute neutrophil count < 1,400 cells/cumm) -Serious infections can occur when neutrophil counts reduce to <500 cells/cumm -When severe - it is called AGRANULOCYTOSIS (Ie levels less than 200) Causes -Decreased production from the marrow: *Marrow suppression/ failure: Drugs, aplastic anemia *Ineffective granulopoiesis: Vitamin B12 deficiency/ folate deficiency -Increased destruction: SLE (immune complex mediated), Sepsis/ severe infection, drugs, splenomegaly.

Hodgkin lymphoma

-Reed- Sternberg (RS) cells. -RS cells release factors that induce the accumulation of reactive lymphocytes, macrophages, and granulocytes. -Mostly the neoplastic RS cells are derived from germinal center or postgerminal center B cells. -It is one of the most common cancers of young adults and adolescents, but can also occur in older individuals -WHO classification: 1. Nodular sclerosis 2. Mixed cellularity 3. Lymphocyte-rich 4. Lymphocyte depletion 5. Lymphocyte predominance -Classical HL : The neoplastic B cells have similar immunophenotype

Additional investigations

-Reticulocyte count -Bone Marrow (BM) Examination -Erythrocyte sedimentation rate (ESR)

Normal Blood Film Giemsa stain is the most common stain used to stain the blood

-See block 3 images file -Any abnormality in the peripheral blood will be indicative of whats happening in the bone marrow

AML with t (15;17): Acute Promyelocytic leukemia

-T(15;17) PML-RARα fusion gene → inhibition of granulocytic maturation -Pro-coagulants and fibrinolytic factors released by leukemic cells -> DIC, bleeding tendency -Respond to therapy with all-trans retinoic acid (ATRA) -ATRA binds to the PML-RARα fusion protein & antagonizes its inhibitory effect -In Blood and BM the tumor cells show predominant promyelocytes with Auer rods (red arrow)

Diagnosis of AML

-The diagnosis of AML is based on the presence of at least 20% myeloid blasts in the bone marrow -Myeloblasts have delicate nuclear chromatin, two to four nucleoli, and more voluminous cytoplasm (Green arrow) -than lymphoblasts. -The cytoplasm often contains fine, myeloperoxidase positive azurophilic granules. -Auer rods (red arrow), distinctive needle-like azurophilic granules, numerous in AML with t(15;17) (acute promyelocytic leukemia)

Sickle Cell Disease

-The rate & degree of sickling is affected by the amount of HbS & its interaction with other Hb types. -In heterozygotes ( sickle cell trait ): very little tendency to sickle Hb F inhibits polymerization of Hb S (therefore, usually no symptoms before 5 months of age )

Hairy Cell Leukemia

-This rare but distinctive B-cell neoplasm. -Middle-aged white males, with a median age of 55. M:F= 5:1 -Clinical presentation: splenomegaly most common. *Tumor cells which are proliferating and occupying the bone marrow are pushing away all the progenitor cells from the bone marrow into the blood, and these progenitor cells take up their home in the spleen and liver *Pancytopenia resulting from marrow involvement and splenic sequestration **Marrow suppresion **All the symptoms of pancytopenia are seen here *Frequent infections: atypical mycobacterial infections, and unexplained monocytopenia. *Not sure why they have monocytopenia but it is just common with this disease -Pathogenesis: activating point mutations in BRAF. -Morphology: fine hair like projections that are best recognized under the phase-contrast microscope. -Tumor cells induce production of extracellular matrix in Bone Marrow -> "dry tap"

WHO classification of lymphoid neoplasms -Most common neoplasm arising from the lymphocytes: NON-HODGKINS Among the Non-Hodgkins lymphoma the common ones are: -Diffuse Large B Cell Lymphoma -Follicular Lymphoma -Mucosa Associated Lymphoma -Mature T Cell Lymphoma T Cell Lymphomas -Adult T - Cell Leukemia Lymphoma associated with Human T cell Leukemia Virus *Sezary Syndrome B Cell Lymphomas: -CLL bar SLL? -Follicular Lymphoma -Diffuse Large B Cell Lymphoma -Burkitt's Lymphoma

1. Precursor B cell neoplasm -B cell - Acute lymphoblastic leukemia/lymphoma 2. Peripheral B-cell neoplasm- Non-Hodgkin lymphoma -Chronic lymphocytic leukemia/ small lymphocytic lymphoma - CLL/SLL -Follicular lymphoma -Diffuse large B cell lymphoma -Burkitt lymphoma -Mantle cell lymphoma -Hairy cell leukemia -Multiple myeloma - plasma cell disorder -Lymphoplasmacytic lymphoma - plasma cell and B cell disorder 3. Precursor T cell neoplasm -T cell - Acute lymphoblastic leukemia/lymphoma 4. Peripheral T-cell neoplasm - Non-Hodgkin lymphoma -Adult T-cell Leukemia/lymphoma -Mycosis Fungoides/Sezary Syndrome 5. Hodgkin lymphoma -Nodular sclerosis -Mixed cellularity -Lymphocyte rich -Lymphocyte depletion -Lymphocyte predominant

Basophilia

>150 cells (0.15x109/ L) -causes- Allergic reactions (immediate hypersensitivity), CML

Burkitt lymphoma (BL) -High grade tumor

Account for 30% childhood NHL in USA *Can occur in adults too but more common in children -All 3 from MYC mutations, acquired in 3 different ways 1. African (endemic) Burkitt lymphoma *Acquired by infection with EBV and usually vector born 2. Sporadic (nonendemic) Burkitt lymphoma *Associated with mutations of MYC, EBV association is not that strong here 3. Occurring in HIV patients *Associated with mutations of MYC, EBV association is not that strong here -Both endemic and sporadic BL are mainly found in children and young adults -Most tumors manifest at extranodal sites -Prognosis: very aggressive tumor , responds well to aggressive chemotherapy. -Clinical presentation: Endemic BL present as mandibular mass (swelling in the jaw). In other forms abdominal viscera, (kidneys, ovaries, and adrenals). -Pathogenesis: All forms are associated with MYC translocations . *T(8;14) partners are MYC; IgH locus, t(2;8) with Ig kappa or t(8;22) with Ig lambda light chain *MYC is a transcription factor for various proteins, causes progression of cell cycle, increase apoptosis and causes differentiation. Because of these mutations, BL is a highly aggressive tumor *Strong association with EBV infection

Cytochemistry in leukemia -All the myeloid series express MPO

Acute leukemia -> MYELOPEROXIDASE (MPO) and PERIODIC ACID SCHIFF (PAS - stains glycogen in the cell) 1. MPO Positive and PAS Negative -Acute Myeloid Leukemia THEN, add nonspecific esterase (NSE) -NSE negative -> Granulocytic M1, M2, M3 -NSE positive -> Myelo-monocytic M4 -NSE positive -> Monocytic M5a, M5b 2. MPO Negative and PAS Positive -Acute Lymphocytic Leukemia -Acute undifferentiated leukemia THEN, add ACID PHOSPHATASE -Acid phosphatase Positive -> ALL -Acid phosphatase Negative -> AUL

Hypocellular Bone Marrow

Causes include: -Aplastic anemia (congenital or acquired) -Bone marrow suppression by drug or autoimmune causes -Paraxysmal nocturnal hemoglobinemia

Hypercellular Bone Marrow

Causes include: -Leukemias (AML, ALL, CML, CLL, polycythemia vara) -Granulomas -Essential thrombocytosis -Early stages of primary myelofibrosis -Metastatic regions -Adipocytes are not visible, completely occupied by cells

WBC Absolute counts in Disease PENIA - decrease in number PHILIA - increased in number CYTOSIS - increased in number

Causes of NEUTROPHILIA -When counts are above 6,500 cells -Commonly aused by any infection (pna, deep tissue infection, pharyngitis, laryngitis) -Due to bone marrow stimulation by TNF, IL-1, and IL-6 -In both acute and chronic infections -Also seen in conditions like trauma, hemorrhage, splenectomy, increased corticosteroids Causes for EOSINOPHILIA -parasitic infection -allergic reaction Causes for decreased MONOCYTE count -Hairy cell Leukemia Causes for increased MONOCYTE count -Chronic bacterial infection (Mycobacterial infections) -infection mononucleousis -autoimmune diseases Causes of decreased LYMPHOCYTE counts -AIDS (selective decrease in CD4 cell counts) -DiGeorge Syndrome (congenital immunodeficiencies) -Malnutrition Causes of increased LEUKOCYTE count -Here you see atypical lymphocytes *Usually a lymphocyte has a round condensed nucleus with small cytoplasm. However here the nucleus will open up with nuclear chromatin exposed and there will be a lot of basophilic cytoplasm -Viral infections (HIV, CML, Ebsten Barr) -Toxoplasmosis

Pathophysiology of clinical presentation in Multiple myeloma These plasma cells are proliferating in the bone marrow. They are producing both intact immunoglobulins and light chains, as well as cytokines, IL-1 and MIP1 ALPHA -Most common sx's is chronic low back pain, or pathalogical fracture

Clinical presentation: pathological fracture, chronic bone pain (lytic bone lesion), renal failure, and anemia. -Myeloma derived MIP1α -> upregulates expression of RANKL -> activates osteoclasts -> bone resorption -> hypercalcemia and pathological fracture *RANK-L ligand -Hypercalcemia: weakness, confusion, lethargy, constipation and polyuria -Renal insufficiency and renal failure: due to deposition of immunoglobulins with in the glomeruli -> renal amyloidosis and renal failure -Bence-Jones proteinuria: excretion of immunoglobulin light chains in urine -Anemia: marrow infiltration -Production of abnormal immunoglobulins -> recurrent bacterial infections.

G6PD deficiency

Clinical: Most common triggers include: -Infections: Oxygen - derived free radicals are produced by activated leukocytes -Drugs : antimalarials, sulfonamides, nitrofurantoins and certain foods (including fava beans in the Mediterranean variant) -Oxidants can cause both intra- and extra-vascular hemolysis *If acute intravascular hemolysis occurs : anemia, hemoglobinemia, hemoglobinuria occurs 2 to 3 days after exposure to oxidants -The recovery phase is heralded by reticulocytosis

Stages

Common progenitor -> Myeloblast -> Promyelocytes -> Melocyte -> Metamyelocyte -> Band cell ->-> -Myeloblast: commited cell which will mature into neutrophil -Neutrophils are about 60% of the leukocytes present -3-5% of bands are seen in a normal individual *If this percentage is increase, it is abnormal

Precursor cell neoplasm: ALL/L (B cell and T cell)

Immunophenotype: -Immature B and T cell ALL are TdT + -B-ALL: CD19, CD 10, CD20 and cytoplasmic IgM heavy chain. -T-ALL: CD1, CD2, CD5, and CD7. They are double negative i.e., CD4 and CD8 - ve and CD 3 - ve -85% of ALL is B cell type. -B- Gated population is TdT positive and CD22 positive. C- Majority of Gated population is positive for CD10 and CD19 B-ALL

Follicular Lymphoma (continued) -They proliferate in the form of nodules. It is very difficult to distinguish reactive follicular hyperplasia with nodular follicular lymphoma -We use the BCL2 marker to highlight the neoplastic cells

Microscopy: predominantly nodular growth pattern -Low power: nodular architecture -High power: 2 cell types present Two principle types of cells : -centrocytes (small cells with irregular or cleaved nucleus)- as shown in picture B with black arrow *Intermediate sized cells with cleaved nucleus and condesed chromatin -centroblasts ( larger cells with open chromatin, several nucleoli, and modest cytoplasm) - as shown in picture B with red arrow. -This is low grade but has the capacity to transform into high grade DIFFUSE LARGE B CELL LYMPHOMA

Primary myelofibrosis (PMF) (cont)

Morphology: Bone marrow shows progressive reticulin positive fibrosis -> fibrotic obliteration of the marrow -> osteosclerosis Peripheral blood shows anemia, normocytic normochromic, with characteristic 'tear drop cells (dacrocytes)'

CML

Morphology: Bone marrow: Hypercellular for age, increase in number of granulocytic precursors and Megakaryocytes. Peripheral blood (PB): Chronic phase total leukocyte counts > 100,000/cumm, increased precursors of neutrophils peak in myelocytes and segmented neutrophils. Eosinophilia (+), Basophilia (+) usually < 20 %, Blasts < 2%. Splenic enlargement : seen due to extramedullary hematopoiesis (EMH) Hepatomegaly and lymphadenopathy: due to EMH

Anemia : Clinical features/ Investigations

Signs & Symptoms : Fatigue, pallor, menstrual disturbances, etc. Peripheral Blood Smear (PBS) -Macrocytes : Megaloblastic anemia -Microcytes : Iron deficiency anemia, thalassemia, spherocytosis -Anisocytosis : variation in size -Poikilocytosis :variation in shape RBC indices : -Decreased MCV seen in : Iron deficiency anemia & Thalassemia -Increased MCV : in Megaloblastic anemia