This document provides an introduction to microscopic interpretation in dermatopathology. It outlines the steps to examine a histopathology slide which include: examining the slide with the naked eye to gain initial clues, scanning at low magnification to identify the type of specimen and site of involvement, and examining at higher magnifications to confirm features and identify specific cell types. It then describes the normal histology of skin, identification of common cell types found in skin such as keratinocytes, melanocytes and fibroblasts. Finally, it discusses methods of diagnosis including pattern recognition and algorithms to categorize inflammatory dermatopathology patterns.

1. Introduction to microscopic interpretation Dr. Santosh Rathod

2. Have your eyes and mind open Processing the acquired visual information Arriving at a tentative diagnosis (ie, model building) Testing the preliminary diagnosis with further examination Confirming the diagnosis Correlating available clinical information Finalizing the diagnosis

3. From where to start?

4. Examination of the Slide with the Naked Eye To gain some appreciation of the size, number, and nature of the histologic sections on the slide The tinctorial properties (histochemical staining) also may provide clues to diagnosis; For example,bluish cellular aggregates or nodules suggest high nuclear-to-cytoplasmic ratios because of basophilic staining of nuclei and, as a result of processes such as basal cell carcinoma, small cell carcinoma, and infil- trates of small lymphocytes or calcium deposition.

5. Examination of the MicroslideatScanning(2 X or 4X) Magnification Firstly, one should attempt to identify the type of specimen submitted Then, inspect the specimen with the idea of determining in general terms from what anatomic site the tissue was taken. Entire specimen (ie, epidermis, dermis, or subcutis) should be scanned - for the principal site of involvement by a disease process, if any, and - the nature of the process, whether inflammatory proliferative, inflammatory and proliferative or non-inflammatory.

6. Examination at IntermediateMagnification The tendency to go to higher magnification too soon should be resisted because one often will overlook a crucial feature, and thus, in effect, one “cannot see the forest for the trees.” The reasons for closer inspection of the specimen (with 10Xand 40X objectives) are to confirm particular features of pathologic processes For identification of specific cell types, such as lymphocytes or granulocytes

7. Normal histology of skin St. Corneum St. Granulosum St. Spinosum St. Basale Pappilary dermis Reticular Dermis

8. Identification of cells Type of cells normally present in epidermis : Majority are - Keratinocytes (90%) Minority population of – Langerhans cells Melanocytes Neuroendocrine(Merkel Cells) Unmyelinated axons Occasional Cells – Toker cells found in nipple epidermis in approximately 10% individuals

9. s Granular keratinocyte Spinouskeratinocyte Basal keratinocyte

10. Langerhans cells Marrow derived Dendritic Antigen presenting cells In H&E sections, appear as clear cells Special stains are generally required for their detection and enumeration

11. melanocyte Melanin synthesizing dendritic cells Located within basal layer of epidermis, hair bulb, ORS In H&E stained section, dendritis are not visible, cell bodies can be seen dispersed in basal layer Contain round to oval, dark stained nuclei that are generally smaller than basal keratinocyte

12. Dermis Cellular content : Fibroblasts Dermal dendritic cells Macrophages Mast cells Extra-cellular content : Collagen Elastic fibres Ground substance

13. Dermal fibroblast Appear as inconspicuous bipolar spindle cells with elongated ovoid nuclei Can’t be reliable distinguished from other dermal spindle shaped and dendritic cells ( dermal dendrocytes) Synthesizes collagen IH stain – Vimentin

14. Phagocytic macrophages Also, called Histiocytes Are of bone marrow origin, circulate in blood as precursors and enter tissue as monocytes Activated monocytes – macrophages Aggregation of activated macrophages – granulomas Macrophages that have ingested melanin- melanophages Macrophages that have ingested hemosiderin - siderophages

15. Monocytes are indistinguishable by routine histology from lymphocytes as both have a small, dark, rounded nuclei with very scanty cytoplasm

16. Macrophages are larger cells than monocyte and possess a vesicular, light staining, elongated nuclei with a clearly visible nuclear membrane

17. Emigrant inflammatory cells Neutrophilic granulocytes Eosinophilic granulocytes Basophilic granulocytes Lymphocytes Plasma cell

18. Neutrophilic granulocyte Polymorphonuclearleucocyte Lobated “pop-corn” shaped nuclei within pale pink fairly granular cytoplasm Nuclear breakdown due to local necrosis or by autodigestion of nuclear lobes as in vasculitis results in “nuclear dust” of vasculitis

19. Eosinophilic granulocyte Characterized by strongly eosinophilic granules in cytoplasm and a characteristically bilobed nuclei Although visible with routine stains, these granules stand out more clearly in brilliant red when stained with giemsa

20. Plasma cell Have abundant cytoplasm that is deeply basophilic, homogenous and sharply defined Round eccentrically placed nuclei along its membrane it shows course, deeply basophilic, regularly distributed chromatin particles which gives “cart-wheel” appearance

21. Methods of diagnosis in dermatopathogy Initial stage of pattern - by the process of hypothesis recognition by a “gestalt” generation and differ- based or instant recognition tial diagnosis Pattern recognition method by Ackerman

22. Inflammatory Dermatopathology by Pattern and Algorithm Steps Categorize the pattern Assess the inflammatory cell population Look for specific findings that direct the algorithm as far as it can be taken Correlate the histologic assessment with known clinical information

23. Superficial PerivascularDermatitis Superficial and Deep Perivascular Dermatitis Nodular and Diffuse Dermatitis Panniculitis Vasculitis Folliculitisand Perifolliculitis IntraepidermalVesicular and Pustular Dermatitis Subepidermal Vesicular Dermatitis

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