Lymph Node Cytology. Jed Overmann, DVM, DACVP. Assistant Clinical Professor, Clinical Pathology College of Veterinary Medicine University of Minnesota

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1 Lymph Node Cytology Jed Overmann, DVM, DACVP Assistant Clinical Professor, Clinical Pathology College of Veterinary Medicine University of Minnesota

2 Objectives Become familiar with the cytologic appearance of normal lymph nodes Become familiar with criteria for diagnosis and the cytologic appearance of common pathologic processes of lymph nodes Inflammation Metastatic neoplasia Reactivity/hyperplasia Lymphoma Understand the limitations of lymph node cytology

3 Overview Indications Sample acquisition/preparation Cytology of normal lymph nodes Cytology of abnormal lymph nodes

4 Indications for Lymph Node Aspiration Lymphadenomegaly** Detection of metastatic disease** Classification of lymphoma

5 Indications for Lymph Node Aspiration Lymphadenomegaly Enlargement of one or more lymph nodes If multiple LNs affected, sample > 1 Submandibular LNs tend to be reactive

6 Indications for Lymph Node Aspiration Detection of metastatic disease Even normal sized LNs can contain evidence of metastatic disease

7 Indications for Lymph Node Aspiration Classification of Lymphoma Immunocytochemistry Flow cytometry PCR for antigen receptor rearrangement (PARR) Often done via HISTOPATHOLOGY and IHC

8 Sample Acquisition 22-gauge needle, 6 or 12 ml syringe Aspiration vs Non-aspiration technique

9 Sample preparation Slide preparation **Slide-over-slide smears ( squash prep )** Blood smear technique Be gentle, lymphocytes are fragile cells! Quickly dry Stain Generally Romanowsky type stains e.g. Wright s stain, Diff-Quik Leave some unstained if sending to pathologist

10 Ruptured Cells

11 Cytology of Normal Lymph Nodes

12 Normal Lymph Node 85-90% small lymphocytes Smaller than a neutrophil or 1-1.5x diameter of RBC Scant amount of basophilic cytoplasm Round nucleus with clumped chromatin pattern No nucleoli

15 Normal Lymph Node 10-15% medium lymphocytes and lymphoblasts Size of neutrophil or larger or > 2-3x diameter of RBC Lighter, smoother chromatin pattern Slightly more cytoplasm +/- nucleoli

19 Normal Lymph Node Low numbers of plasma cells Moderate amount of deeply basophilic cytoplasm Eccentric nucleus Clumped chromatin pattern Golgi zone (perinuclear clearing)

23 Normal Lymph Node Small numbers of macrophages and rare mast cells

25 Normal Lymph Node Occasional neutrophils and eosinophils Often primarily blood associated

26 Abnormal Lymph Nodes Reactive/hyperplastic Lymphoma Lymphadenitis (inflammation) Metastatic neoplasia

27 Reactive/hyperplastic Small lymphocytes still predominate **Increased numbers of: Plasma cells Can see occasional Mott cells Medium lymphoctyes and lymphoblasts

30 Mott Cells Highly activated plasma cells Cytoplasm filled with pale vacuoles Often round to oval Glassy appearance Represent packets of immunoglobulin

33 Reactive/hyperplastic May also see small increases in: Macrophages Neutrophils Eosinophils Mast cells

34 Atypical reactive/hyperplastic processes Expanded population of intermediately sized lymphocytes Minimal increase in plasma cells Can be difficult to distinguish from lymphoma e.g. ehrlichiosis, FeLV

36 Lymphoma 50% lymphoblasts Large cells Smooth chromatin pattern Often have visible nucleoli +/- increased # s of mitotic figures +/- cytophagic macrophages

42 Lymphoma of Granular Lymphocytes (LGL) Seen most often in cats Affected sites GI tract Mesenteric LNs Liver Spleen

43 Lymphoma Diagnostic difficulties Small cell lymphoma GI lymphoma vs IBD Atypical hyperplasia vs lymphoma Histopathology (+/- IHC), PARR

44 Lymphadenitis Increased numbers of neutrophils (>5%), eosinophils (>3%), or macrophages Must take into consideration degree of blood contamination Named for the predominant inflammatory cell type(s) Reactivity/hyperplasia often present as well

45 Lymphadenitis Suppurative or (pyo)granulomatous Must look for an etiologic agent Metastatic neoplasia can cause this type of inflammation

46 Bacteria Blastomyces sp.

47 Histoplasma sp.

48 Lymphadenitis Eosinophilic Hypersensitivity reactions Parasitic Paraneoplastic eg Mast cell tumor

49 Metastatic Neoplasia Defined by the presence of an atypical/malignant cell population eg metastatic carcinoma eg metastatic mast cell tumor eg metastatic melanoma

53 A few other things

54 Perinodal Fat

55 Salivary gland Streaming pink background, windrowing of RBCs Glandular epithelial cells Large amount of foamy cytoplasm Small, round, dark nucleus

57 Questions/Comments

Cytology of Lymph Nodes

Indications Cytology of Lymph Nodes Lymph node enlargement That was easy Mary Anna Thrall Don Meuten Indications Lymph node enlargement Suspect metastasis Normal sized lymph nodes are Normal Do NOT aspirate