Erythrocytosis and Lymph Node Cytology PDF

ERYTHROCYTOSIS AND POLYCYTHEMIA LEARNING OBJECTIVES  Appreciate the breed specific differences in RBC mass  Know the primary hormonal regulator of RBC production, where it is produced and what stimulates its production  Understand the causes of erythrocytosis and their mechanisms THE TERMIN...

ERYTHROCYTOSIS AND POLYCYTHEMIA LEARNING OBJECTIVES  Appreciate the breed specific differences in RBC mass  Know the primary hormonal regulator of RBC production, where it is produced and what stimulates its production  Understand the causes of erythrocytosis and their mechanisms THE TERMINOLOGY  Erythrocytosis = increase in RBC mass (HCT/PCV, RBC count, and/or Hgb)  Polycythemia = means “many cells in the blood”  Most pathologists restrict the use of “polycythemia” strictly for a neoplastic condition (polycythemia vera = neoplastic proliferation of RBCs)  Others use the terms interchangeably…. Confusing! WHY ISN’T MORE BLOOD BETTER?  Increases blood viscosity → sludging of RBCs in vessels  Leads to impaired blood flow  Decreased tissue oxygenation  Congested mucous membranes  Dilated retinal vessels  Seizures BREED SPECIFIC DIFFERENCES  Some breeds have a higher RBC mass in health  Sighthounds, greyhounds, some dachshunds  Racing horses, warm-blooded breeds RBC PRODUCTION – ABOUT EPO  Erythropoietin (EPO) → primary hormonal regulator of RBC production  Produced by the fetal liver and the adult kidney  Upregulated by renal hypoxia (not RBC mass)  Anemia  Poor renal perfusion  Poor oxygenation of blood  ERYTHROCYTOSIS RELATIVE ABSOLUTE NOT a true increase in RBC mass True increase in RBC mass Dehydration ↓ plasma volume (hemoconcentration) History, PE findings, ↑ protein, USG, azotemia Primary Secondary Low/normal Epo Increased Epo Splenic contraction Polycythemia vera Excitable animal (epinephrine response) Redistribution of RBCs from spleen Cats, horses Appropriate Inappropriate Hypoxia induced No systemic hypoxia High altitude Renal diseases Lung diseases Epo-secreting tumors Right-to-left cardiac shunts Exogenous RELATIVE ERYTHROCYTOSIS: DEHYDRATION  Most common cause of mild to moderate erythrocytosis  NOT an absolute increase in RBC mass → plasma volume decreased  PE findings  Dry or tacky mucous membranes  Tenting of skin  Laboratory findings  ↑ proteins (total protein, albumin)  Highly concentrated urine (high urine specific gravity/USG)  Possible pre-renal azotemia (↑ urea and creatinine)  Rehydrate the patient and recheck RELATIVE ERYTHROCYTOSIS: SPLENIC CONTRACTION  Dehydration absent  Not an increase in RBC mass → RBCs are redistributed into circulation from splenic contraction  Transient and secondary to fear, excitement, or exercise  Seen frequently in cats and horses > dogs  More excitable?  Larger splenic RBC reserves  May also see a lymphocytosis (epinephrine-induced) ABSOLUTE ERYTHROCYTOSIS  Due to a true increase in RBC mass  All causes of a relative erythrocytosis must be excluded  Primary (rare) → EPO-independent (EPO is N or ↓)  Known as polycythemia vera → neoplastic proliferation of mature RBCs independent of EPO  RBCs are morphologically unremarkable  Diagnosis of exclusion (all relative and secondary causes ABSOLUTE ERYTHROCYTOSIS  Secondary → EPO-dependent (EPO is ↑)  Appropriate  EPO production is ↑ secondary to hypoxia or hypoxemia  Causes → heart disease, lung disease, high altitudes  Inappropriate  EPO production is ↑ without systemic hypoxia/hypoxemia  Causes → renal lesions (e.g., cyst, tumor, infection) causing local hypoxia, non-renal tumors that produce EPO or EPO-like substances, exogenous (doping) IN SUMMARY….. Classification HCT Protein Hypoxia EPO Relative ↑ ↑ ---- ---- (Dehydration) Relative (Splenic) ↑ ---- ---- ---- Secondary Systemic ↑ ---- ↑ Appropriate Secondary Renal only, in ↑ ---- ↑ Inappropriate some cases Polycythemia vera ↑ ---- ---- N or ↓ CASE EXAMPLE: SALLY  7-year-old, female spayed mixed breed dog with a 5-day history of lethargy and anorexia Analyte Value Reference Interval Units WBC 10.2 4.39 – 11.6 x103 /mL RBC 13.2 H 4.98 – 7.92 x106 /mL HGB 27.1 H 12.6 – 19.9 g/dL HCT 82.3 H 36.6 – 55.6 % MCV 66 60 – 72 fL MCHC 34.5 32 – 36.4 g/dL Platelet 266 200 – 500 x103 /mL  ERYTHROCYTOSIS RELATIVE ABSOLUTE NOT a true increase in RBC mass True increase in RBC mass Dehydration ↓ plasma volume (hemoconcentration) History, PE findings, ↑ protein, USG, azotemia Primary Secondary Low/normal Epo Increased Epo Splenic contraction Polycythemia vera Excitable animal (epinephrine response) Redistribution of RBCs from spleen Cats, horses Appropriate Inappropriate Hypoxia induced No systemic hypoxia High altitude Renal diseases Lung diseases Epo-secreting tumors What else would you like to know? Right-to-left cardiac shunts Exogenous  CASE EXAMPLE: SALLY  Additional information:  Not clinically dehydrated  Total protein: 8.5 g/dL (RI: 5.7 – 9.4)  Unremarkable chemistry panel and urinalysis  ERYTHROCYTOSIS RELATIVE ABSOLUTE NOT a true increase in RBC mass True increase in RBC mass Dehydration ↓ plasma volume (hemoconcentration) History, PE findings, ↑ protein, USG, azotemia Primary Secondary Low/normal Epo Increased Epo Splenic contraction Polycythemia vera Excitable animal (epinephrine response) Redistribution of RBCs from spleen Cats, horses Appropriate Inappropriate Hypoxia induced No systemic hypoxia High altitude Renal diseases Lung diseases Epo-secreting tumors What else would you like to know? Right-to-left cardiac shunts Exogenous Additional diagnostics? CASE EXAMPLE: SALLY  Dog is from south Georgia  Thoracic radiographs:  No evidence of heart or lung disease  ERYTHROCYTOSIS RELATIVE ABSOLUTE NOT a true increase in RBC mass True increase in RBC mass Dehydration ↓ plasma volume (hemoconcentration) History, PE findings, ↑ protein, USG, azotemia Primary Secondary Low/normal Epo Increased Epo Splenic contraction Polycythemia vera Excitable animal (epinephrine response) Redistribution of RBCs from spleen Cats, horses Appropriate Inappropriate Hypoxia induced No systemic hypoxia High altitude Renal diseases Lung diseases Epo-secreting tumors What else would you like to know? Right-to-left cardiac shunts Exogenous Additional diagnostics? CASE EXAMPLE: SALLY  Abdominal ultrasound  A 6 cm mass identified on the cranial pole of the left kidney that expanded the renal capsule  Remainder unremarkable  Pre-operative erythropoietin level  32 mU/mL (RI 1.9-22.3 mU/mL)  ERYTHROCYTOSIS RELATIVE ABSOLUTE NOT a true increase in RBC mass True increase in RBC mass Dehydration ↓ plasma volume (hemoconcentration) History, PE findings, ↑ protein, USG, azotemia Primary Secondary Low/normal Epo Increased Epo Splenic contraction Polycythemia vera Excitable animal (epinephrine response) Redistribution of RBCs from spleen Cats, horses Appropriate Inappropriate Hypoxia induced No systemic hypoxia High altitude Renal diseases Lung diseases Epo-secreting tumors Right-to-left cardiac shunts Exogenous  CASE EXAMPLE: SALLY  FNA of renal mass → atypical mesenchymal cells diagnosed as sarcoma  Sally underwent surgery to remove the left kidney  Histologic examination revealed a grade II soft tissue sarcoma CASE EXAMPLE: SALLY  Two weeks post-op  Sally’s appetite and energy level returned to normal  PCV was 51%  Went on to live a long, happy life Lymph Node Cytology K a t ie M e t ca l f, D V M , M S , D A CV P LEARNING OBJECTIVES  Understand how lymphocytes are sized relative to other blood cells and be able to size a lymphocyte if given an image  Understand the normal cellular composition of a lymph node and be able to identify common cell types  Know the differentials for an enlarged lymph node and how they are diagnosed via cytology (i.e., differences in cellular composition) WHEN WOULD YOU ASPIRATE A LYMPH NODE?  Lymphadenomegaly or enlargement of one or multiple lymph nodes  Detected via palpation, radiography, or ultrasonography  Evaluating for the presence of metastatic disease  Also called staging COMMONLY SAMPLED LYMPH NODES Lymph Node Location Drainage Features Mandibular Ventral angle of the jaw Most of the head, including the rostral oral cavity Prescapular Cranial shoulder (in front of Caudal part of the head (pharynx, pinna), most (superficial cervical) supraspinatus m.) of the thoracic limb, and part of the thoracic wall Axillary Caudal and medial to the Most of the thoracic wall, deep structures of the shoulder joint thoracic limb and neck, cranial mammary glands Superficial inguinal Furrow between the Caudal mammary glands, ventral half of abdominal wall and medial abdominal wall, penis, prepuce, scrotum, tail, thigh ventral pelvis, medial thigh and stifle Popliteal Back of the stifle Area distal to the stifle Medial iliac Near the caudal vena cava and Skin of the pelvic area, pelvic limb, distal aorta (internal) intestinal and urogenital system MAKING A DIAGNOSTIC SMEAR  Goal: thin layer of intact cells, rapidly air-dried slides  Methods for sampling:  Impression Smears  Woodpecker or “needle off”  Fine-Needle Non-Aspiration Technique  Fine-Needle Aspiration Technique  Use “squash” prep technique  Allows for proper dispersion of cells and improves diagnostic ability MAKING A DIAGNOSTIC SMEAR  Using proper “squash” prep technique creates diagnostic smears ☺ MAKING A DIAGNOSTIC SMEAR  Avoid making “splat” or “shotgun blast” smears  Limits diagnostic ability as it creates dense smears and high numbers of lysed cells  A clinical pathologist’s worst nightmare! SAMPLE EVALUATION  Cellularity vs. degree of blood contamination  Evaluate the cell types present, their morphology, and relative proportions of each cell type  Is the sample lymph node? IS THE SAMPLE LYMPH NODE?  Other things you may see instead…  Salivary epithelium (samples from the mandibular lymph node)  Adipose tissue – aspirated perinodal fat  Muscle…..ouch! Salivary epithelial cells Mature adipocytes Skeletal muscle SIZING OF LYMPHOCYTES & CYTOMORPHOLOGY Small lymphocyte Intermediate lymphocyte Large lymphocyte Smaller in size than a neutrophil Similar in size to a neutrophil Larger in size than a neutrophil Scant basophilic cytoplasm Low to moderate amounts of Low to moderate amounts of basophilic cytoplasm basophilic cytoplasm Condensed (closed) chromatin with Lighter chromatin Finely stippled (open) chromatin indistinct nucleoli often with visible nucleoli NORMAL LYMPH NODE  Majority of small lymphocytes (>90%) with low numbers of intermediate and large lymphocytes (75%) but there are increased numbers of intermediate and large lymphocytes (>10% and up to 25%)  Large lymphocytes 25-50% of population → grey zone  Large lymphocytes >50% of the population → lymphoma REACTIVE LYMPH NODE  Plasma cells may be increased (variable numbers)  May see Mott cells → plasma cells filled with immunoglobulin-containing vacuoles (called Russell bodies)  REACTIVE LYMPH NODE Neutrophil  REACTIVE LYMPH NODE Large Lymphocytes  REACTIVE LYMPH NODE Plasma cells  REACTIVE LYMPH NODE Example from image: ~ 80% small lymphocytes ~ 10% intermediate and large lymphocytes ~ 10% plasma cells Interpretation: Reactive lymph node LYMPHADENITIS  Lymphadenitis = inflammation within the lymph node  Many causes → neutrophilic, eosinophilic, mixed  ***Inflammatory cells may also be blood-associated → watch out for blood contamination!  Predominant inflammatory cell type categorizes the inflammation present NEUTROPHILIC LYMPHADENITIS  May also be called purulent or suppurative lymphadenitis  Recognized by >5% neutrophils – remember…you need to exclude blood origin!  Fairly non-specific finding → may be associated with bacterial, neoplastic, or immune-mediated conditions  If mandibular LN → think dental disease! (PYO)GRANULOMATOUS LYMPHADENITIS  Called macrophagic or histiocytic lymphadenitis  Increased numbers of macrophages (variable #)  Pyogranulomatous → ↑neutrophils and macs  Causes: Cat with FIP  Chronic inflammation  Higher-order bacteria (e.g., filamentous, acid-fast)  Fungal infections (yeast, hyphae) Histoplasmosis EOSINOPHILIC LYMPHADENITIS  Greater than 3% eosinophils  Several causes:  Allergy/hypersensitivity  Local skin disease (e.g., atopic dermatitis)  Parasites (e.g., fleas)  Some fungal infections (e.g., Pythium)  Paraneoplastic (e.g., lymphoma, MCT) MIXED CELL LYMPHADENITIS  Most common  Non-specific, can be due to a combination of etiologies LYMPHOMA  Characterized by a predominance of a monomorphic lymphocyte population that is >50% of the lymphocytes  Most lymphomas are composed of large lymphocytes that exhibit atypia (e.g., open chromatin and prominent nucleoli)  Small cell, well-differentiated variants exist and will require additional testing to diagnose (e.g., PARR, flow cytometry)  Classified based on their immunophenotype (B- vs. T-cell) which yields relevant treatment and/or prognostic information  LYMPHOMA Example from image: >90% large lymphocytes