Cytology of Malignant Mesothelioma

31 Cytology of Malignant Mesothelioma Richard M. DeMay Because patients with mesotheliomas frequently present with effusions, cytologic examination of the effusion fluid may be the first diagnostic study. The fluid is often yellowish, but many are bloody (1 3). It is characteristically thick and mucoid owing to hyaluronic acid content. The cell count is high, and typically remains high after repeated taps, which is unusual in benign effusions. In theory, the diagnosis of mesothelioma is easy, based on malignant cells that look like mesothelial cells (Figs. 31.1 and 31.2). There is no foreign, alien, extra, or discrete population of tumor cells, in contrast with metastatic malignancy (4 7). Instead, there is a morphologic kinship of the malignant cells with native mesothelial cells, forming a continuous spectrum from apparently benign to atypical to malignantappearing mesothelial cells (4 13). In practice, the diagnosis of mesothelioma can be difficult. For example, the kinship that is so characteristic of mesothelioma is a two-edged sword (4). This same morphologic feature that makes the diagnosis possible can also make the diagnosis impossible in some cases. Some mesotheliomas show only subtle cytologic abnormalities (5,9,14). The malignant cells can be indistinguishable from benign, reactive mesothelial cells or even macrophages (12,15,16). At the other end of the spectrum, in poorly differentiated tumors, the diagnosis of malignancy may be obvious, but the mesothelial origin may not be (4). A key observation in the cytologic diagnosis of mesothelioma one that can be made at low microscopic power is the presence of more and bigger cells, in more and bigger clusters (4). Extreme mesothelial cellularity suggests the diagnosis of mesothelioma (9 11,17,18), although abundant mesothelial cells can also occur in some benign conditions and not all mesotheliomas yield highly cellular specimens (4). Furthermore, too many large clusters of cells suggest a diagnosis of malignancy, particularly in pleural effusions, although again, not every case has this feature (19). Cell groups can range from small to large; large groups are sometimes composed of hundreds of cells (12,13,20). Single cells are usually present in mesothelioma, and predominate in some cases (6,8,9,11,12,14,21). 481

482 Chapter 31 Cytology of Malignant Mesothelioma Figure 31.1. Malignant mesothelioma: malignant mesothelial like cells. Pap stain (400 ). When single cells predominate, the diagnosis can be more difficult; a possible clue may be the sheer number of mesothelial cells (14). Cell aggregates in mesothelioma characteristically are large, and have knobby, flower-like, berry-like, or highly complex contours (Fig. 31.3) (4,11,17). In contrast, cell aggregates in benign effusions are fewer, smaller, and less complex (13,18). Cell aggregates in adenocarcinoma can be large, but tend to have smooth, community borders (5). The clus- Figure 31.2. Malignant mesothelioma: malignant mesothelial like cells. Romanovsky stain (400 ).

R.M. DeMay 483 Figure 31.3. Malignant mesothelioma: complex aggregate of malignant mesothelial cells. Pap stain (200 ). ters are usually solid in mesothelioma, but can be hollow, mimicking glandular acini (5,11,18). True acinar formation is characteristic of adenocarcinoma. Cell aggregates with central cores of collagen usually indicate mesothelial cells (benign or malignant) (22). Collagen cores are rare in adenocarcinoma. The collagenous material is homogeneous and translucent. It stains cyanophilic in the Papanicolaou (Pap) stain and is slightly metachromatic in Romanovsky stains (Fig. 31.4). Papillary aggregates, though nonspecific, are more common in mesothelioma than in either adenocarcinoma or benign effusions (16,21,23). Cell-in-cell patterns (also known as cell-embracing, pincer-like grip, cell engulfment, and cannibalism ) (Fig. 31.5) are more Figure 31.4. Malignant mesothelioma: metachromatic collagen core. Romanovsky stain (400 ).

484 Chapter 31 Cytology of Malignant Mesothelioma Figure 31.5. Malignant mesothelioma: cell-in-cell pattern, pincer-like grip, characteristic of mesothelial cells. Pap stain (400 ). common and more complex in mesothelioma than benign effusions (4,5,7,14,17,23). Windows (openings between adjacent cells) are a characteristic feature of (benign or malignant) mesothelial cells, and are more common in mesothelioma than in adenocarcinoma (24) (Fig. 31.1). Unfortunately, there are exceptions to these general rules presented above, such as community borders in mesothelioma and knobby contours in adenocarcinoma (5). Malignant mesothelial cells are typically more variable in size than benign mesothelial cells and frequently larger and sometimes even giant (Fig. 31.6) (5,25). However, frankly bizarre-appearing cells favor Figure 31.6. Malignant mesothelioma: giant mesothelial cell. Pap stain (400 ).

a diagnosis of carcinoma. Classically, the nucleus (N) and cytoplasm (C) tend to change in size proportionately, so that the N/C ratio remains relatively constant. This imparts a certain degree of uniformity to mesothelioma, not usually seen in adenocarcinoma (4,26). The cytoplasm of mesothelial cells is characteristically dense in the center (endoplasm) and delicate toward the edges (ectoplasm) (Figs. 31.1 and 31.2) (9). A characteristic two-tone staining pattern is sometimes seen, in which the endoplasm stains pink to orange and the ectoplasm blue to green (4,9,19). Sometimes, dense, coagulated mummified cells (similar to Councilman bodies in the liver) are seen in mesothelioma. These cells look a bit like dyskeratocytes, with pink-to-orange cytoplasm and dark, pyknotic nuclei (Fig. 31.7) (4). Though rare, they are a marker for mesothelioma (exclude squamous cell carcinoma) (14,19). Microvilli can sometimes be appreciated in the Pap stain as fine cytoplasmic projections that are metachromatic (pink) in Romanovsky stains (4). Microvilli are rarely seen on adenocarcinoma cells in effusions, and even when they are, the microvilli are stubby and only present on the luminal surface (in cell blocks). Similarly, cytoplasmic blebs (prominent cytoplasmic outpouchings, probably degenerated microvilli that coalesce) are often accentuated in malignant mesothelial cells, compared with benign ones, and are uncommonly seen in adenocarcinomas (23). Several kinds of vacuoles can be seen in mesothelioma, including tiny lipid vacuoles, larger glycogen vacuoles, hard-edged hyaluronic acid vacuoles, and clear, degenerative vacuoles. Both the number of vacuoles and the number of vacuolated cells are highly variable. Adenocarcinoma can also have lipid, glycogen, mucin, or degenerative vacuoles. Cytoplasmic vacuoles are often best appreciated with R.M. DeMay 485 Figure 31.7. Malignant mesothelioma: mummified mesothelial cell. Pap stain (400 ).

486 Chapter 31 Cytology of Malignant Mesothelioma Romanovsky stains (4,12,23,27). Special stains can be helpful in differential diagnosis of vacuoles. Mesothelial nuclei are usually located near the center of the cell, while eccentrically located nuclei are more characteristic of adenocarcinoma (4,8,12). Binucleation and trinucleation are fairly common, and multinucleation occurs, although these are nonspecific findings. Although the usual nuclear criteria of malignancy (pleomorphism, enlargement, abnormal chromatin, nucleoli, etc.) apply in the diagnosis of mesothelioma (19), nuclear atypia can be subtle in some cases (6,18). Conversely, marked nuclear atypia can be seen in benign conditions such as hepatitis, uremia, pancreatitis, and postradiation, as well as adenocarcinoma. Degeneration can cause changes that mimic malignancy. Marked nuclear membrane irregularity is associated with malignancy, but may not be a prominent feature. Irregular nuclear membranes can also be seen in benign mesothelial cells, particularly in washing specimens ( daisy cells ). Intranuclear cytoplasmic invaginations, rare in benign mesothelial cells, can be seen in either mesothelioma or adenocarcinoma (4,12,28,29). Chromatin abnormalities range from subtle to obvious. However, marked hyperchromasia is usually absent, unless the cells are degenerated (which usually gives the chromatin a smudgy, homogeneous appearance) (14). Nucleoli are usually seen in mesothelioma and may be enlarged, multiple, or irregular in outline (30). Macronucleoli, if present, suggest malignancy (19). Mitoses are uncommon, and not helpful in diagnosis unless they are clearly abnormal (9,12,14). The hyaluronic acid that is characteristic of mesothelioma can sometimes be seen in cytologic specimens as a flocculent material in the background of the slide (4,9). In Romanovsky stains it resembles synovial fluid: coarsely granular, pink (metachromatic) precipitate (16). In the Pap stain, it ranges from granular to fluffy to bubbly in appearance (Fig. 31.8) (31). Psammoma bodies or marked lymphocytic infiltration can occur in mesothelioma (12,32); both are nonspecific (23). Necrosis and debris are not common in mesothelioma, but favor malignancy when seen (with exceptions, particularly infections) (12,33). False-negative diagnoses are well known in mesothelioma. Most false negatives are due to inadequate sampling (i.e., unsatisfactory specimens with few or no mesothelial cells) (19,34). Sarcomatous mesothelioma is rarely diagnosed in exfoliative cytology, since few or no diagnostic cells are exfoliated (35,36). Benign mesothelial proliferations with reactive ( atypical ) mesothelial cells can be difficult to distinguish from mesotheliomas (see below) (30). Conversely, cytologically bland mesotheliomas, composed of cells resembling benign mesothelial cells or histiocytes, are difficult to recognize as malignant (15,16,37). At the other extreme, sometimes the malignant cells are highly abnormal appearing, with clearly malignant features. In such cases, the diagnosis of malignancy may be obvious, but the cell of origin may not be evident (13,38).

R.M. DeMay 487 Figure 31.8. Malignant mesothelioma: flocculent background, hyaluronic acid. Pap stain (400 ). False-positive diagnoses of mesothelioma are rare, but can occur (14,19). Although large clusters of benign mesothelial cells can occasionally be seen in benign effusions, particularly in pericardial effusions or ascites, they are unusual (19,39 42). Diagnosis of mesothelioma based on complex aggregates of atypical mesothelial cells is reliable, but diagnosis based on single atypical mesothelial cells is more difficult (19). References 1. Antman KH. Clinical presentation and natural history of benign and malignant mesothelioma. Semin Oncol 1981;8:313 319. 2. Pisani RJ, Colby TV, Williams DE. Malignant mesothelioma of the pleura. Mayo Clin Proc 1988;63:1234 1244. 3. Ribak J, Lilis R, Suzuki Y, et al. Malignant mesothelioma in a cohort of asbestos insulation workers: clinical presentation, diagnosis, and causes of death. Br J Ind Med 1988;45:182 187. 4. DeMay RM. The Art and Science of Cytopathology. Vol I: Exfoliative Cytology, Vol II: Aspiration Cytology. Chicago: American Society of Clinical Pathologists, 1966. 5. Ehya H. The cytologic diagnosis of mesothelioma. Semin Diagn Pathol 1986;3:196 203. 6. Triol JH, Conston AS, Chandler SV. Malignant mesothelioma: cytopathology of 75 cases seen in a New Jersey community hospital. Acta Cytol 1984; 28:37 45. 7. Whitaker D, Shilkin KB. The cytology of malignant mesothelioma in Western Australia. Acta Cytol 1978;22: 67 70. 8. Klempman S. The exfoliative cytology of diffuse pleural mesothelioma. Cancer 1962;15:691 704.

488 Chapter 31 Cytology of Malignant Mesothelioma 9. Naylor B. The exfoliative cytology of diffuse malignant mesothelioma. J Pathol Bact 1963;86:293 298. 10. Nguyen G-K. Cytopathology of pleural mesotheliomas. Am J Clin Pathol 2000;114(suppl 1, Pathol Patterns Rev):S68 S81. 11. Roberts GH, Campbell GM. Exfoliative cytology of diffuse mesothelioma. J Clin Pathol 1972;25:577 582. 12. Sherman ME, Mark EJ. Effusion cytology in the diagnosis of malignant epithelioid and biphasic pleural mesothelioma. Arch Pathol Lab Med 1990; 114:845 851. 13. Whitaker D, Shilkin KB. Diagnosis of pleural malignant mesothelioma in life a practical approach. J Pathol 1984;143:147 175. 14. Whitaker D. The cytology of malignant mesothelioma. Cytopathology 2000;11:139 151. 15. Guffanti MC, Faleri ML. Benign-appearing mesothelioma cells in a serous effusion. Acta Cytol 1985;29:90 92. 16. Spriggs AI, Grunze H. An unusual cytologic presentation of mesothelioma in serous effusions. Acta Cytol 1983;27:288 292. 17. DiBonito L, Falconieri G, Colautti I, et al. Cytopathology of malignant mesothelioma: a study of its patterns and histological bases. Diagn Cytopathol 1993;9:25 31. 18. Leong AS-Y, Stevens MW, Mukherjee TM. Malignant mesothelioma: cytologic diagnosis with histologic, immunohistochemical, and ultrastructural correlation. Semin Diagn Pathol 1992;9:141 150. 19. Whitaker D, Shilkin KB, Sterrett GF. Cytological appearances of malignant mesothelioma. In: Henderson DW, Shilkin KB, Langlois SLP, et al, eds. Malignant Mesothelioma. New York: Hemisphere, 1991:167 182. 20. Berge T, Gröntoft O. Cytologic diagnosis of malignant pleural mesothelioma. Acta Cytol 1965;9:207 212. 21. Kho-Duffin J, Tao L-C, Cramer H, et al. Cytologic diagnosis of malignant mesothelioma, with particular emphasis on the epithelial noncohesive cell type. Diagn Cytopathol 1999;20:57 62. 22. Takagi F. Studies on tumor cells in serous effusion. Am J Clin Pathol 1954; 24:663 675. 23. Stevens MW, Leong AS-Y, Fazzalari NL, et al. Cytopathology of malignant mesothelioma: a stepwise logistic regression analysis. Diagn Cytopathol 1992;8:333 341. 24. Yu GH, Baloch ZW, Gupta PK. Cytomorphology of metastatic mesothelioma in fine-needle aspiration specimens. Diagn Cytopathol 1999;20:328 332. 25. Kwee W-S, Veldhuizen RW, Alons CA, et al. Quantitative and qualitative differences between benign and malignant mesothelial cells in pleural fluid. Acta Cytol 1982;26:401 406. 26. Adams VI, Unni KK. Diffuse malignant mesothelioma of pleura: diagnostic criteria based on an autopsy study. Am J Clin Pathol 1984;82:15 23. 27. Boon ME, Veldhuizen RW, Ruinaard C, et al. Qualitative distinctive differences between the vacuoles of mesothelioma cells and of cells from metastatic carcinoma exfoliated in pleural fluid. Acta Cytol 1984;28:443 449. 28. Baddoura FK, Varma VA. Cytologic findings in multicystic peritoneal mesothelioma. Acta Cytol 1990;34:524 528. 29. Herrera GA, Wilkerson JA. Ultrastructural studies of malignant cells in fluids. Diagn Cytopathol 1985;1:272 275. 30. Gupta PK, Frost JK. Cytologic changes associated with asbestos exposure. Semin Oncol 1981;8:283 289.

31. Whitaker D. Hyaluronic acid in serous effusions smears. Acta Cytol 1986; 30:90 91. 32. Japko L, Horta AA, Schreiber K, et al. Malignant mesothelioma of the tunica vaginalis testis: report of first case with preoperative diagnosis. Cancer 1982;49:119 127. 33. U.S. Canadian Mesothelioma Reference Panel. The separation of benign and malignant mesothelial proliferations. Am J Surg Pathol 2000;24:1183 1200. 34. Renshaw AA, Dean BR, Antman KH, et al. The role of cytologic evaluation of pleural fluid in the diagnosis of malignant mesothelioma. Chest 1997;111: 106 109. 35. Kobayashi TK, Teraoka S, Tsujioka T, et al. Ciliated ovarian adenocarcinoma cells in ascitic fluid cytology: report of a case with immunocytochemical features. Diagn Cytopathol 1988;4:234 238. 36. Tao L-C. The cytopathology of mesothelioma. Acta Cytol 1979;23:209 213. 37. Boon ME, Posthuma HS, Ruiter DJ, et al. Secreting peritoneal mesothelioma: report of a case with cytological, ultrastructural, morphometric and histologic studies. Virchows Arch Pathol Anat 1981;392:33 44. 38. Whitaker D. The validity of a cytological diagnosis of mesothelioma. Aust NZ J Med 1987;17(S2):519 (abstract). 39. Becker SN, Pepin DW, Rosenthal DL. Mesothelial papilloma: a case of mistaken identity in a pericardial effusion. Acta Cytol 1976;20:266 268. 40. Rosai J, Dehner LP. Nodular mesothelial hyperplasia in hernia sacs: a benign reactive condition simulating a neoplastic process. Cancer 1975;35: 165 175. 41. Selvaggi SM, Migdal S. Cytologic features of atypical mesothelial cells in peritoneal dialysis fluid. Diagn Cytopathol 1900;6:22 26. 42. Spriggs AI, Jerrome DW. Benign mesothelial proliferation with collagen formation in pericardial fluid. Acta Cytol 1979;23:428 430. R.M. DeMay 489