Mesothelial cells the notorious mimickers in pleural fluid
"Mesothelial cells are considered 'mimickers' because, when they become reactive due to inflammation or other conditions, they can sometimes appear very similar to cancerous cells under a microscope, particularly mimicking adenocarcinoma or mesothelioma, leading to potential misdiagnosis in cytology or biopsy samples".
MESOTHELIAL CELLS: HISTOLOGY AND CYTOLOGY On histology, the pleural surface lining the thoracic cavity consists of a single layer of flat, elongated mesothelial cells. Similar to their appearance in other body sites like the liver and spleen, these cells can sometimes adopt a more cuboidal shape. In these cuboidal mesothelial cells, the eosinophilic and granular nature of the cytoplasm becomes more pronounced. The nucleus is typically round to ovoid, with evenly distributed granular chromatin and a variably distinct nucleolus.
Irritation of the mesothelial surface can lead to reactive and hyperplastic changes. Mesothelial hyperplasia may present as an increased proliferation of mesothelial cells arranged in solid sheets, nests, papillary and tubulopapillary structures, gland-like formations, cord-like arrangements, or as single cells with variable nuclear alterations. Reactive changes, which can arise from various causes, may lead to nuclear enlargement (with an increased nuclear-to-cytoplasmic [N:C] ratio), irregular nuclear contours, coarse chromatin distribution, prominent nucleoli, and multinucleation. Additionally, reactive mesothelial cells may exhibit cytologic features that raise concerns for malignancy, such as mitotic figures, nuclear atypia, and background necrosis, as seen in rheumatoid effusions. While some reactive mesothelial changes can result in papillary structures embedded in fibrotic stroma, hyperplastic mesothelial cells do not invade underlying tissues and typically maintain a uniform appearance.
On cytologic examination of benign pleural effusion specimens, mesothelial cells are usually present in low numbers. They are characterized by low N:C ratios, smooth to slightly irregular nuclear contours, granular chromatin, and a moderate amount of densely granular cytoplasm. A distinctive "two-tone" cytoplasmic pattern may be observed, where the central cytoplasm (endoplasm) appears denser due to perinuclear filament accumulation, while the peripheral cytoplasm (ectoplasm) is paler. Multinucleation can be seen in both benign and reactive mesothelial cells. In pleural effusions, mesothelial cells typically appear singly or in small clusters. Their long, slender surface microvilli create well-defined cell borders, often forming intercellular "windows."
Reactive mesothelial cells, while retaining many features of benign mesothelial cells, can also exhibit morphological changes that raise concern for malignancy, including increased N:C ratios, coarse chromatin distribution, and prominent nucleoli. Additionally, reactive mesothelial cells may show cytoplasmic blebbing and vacuolization. These vacuoles can vary in appearance depending on their underlying cause. Some vacuoles result from cellular degeneration associated with hydropic change, appearing as multiple small vacuoles or a single large vacuole that displaces the nucleus peripherally. Other vacuoles may form due to the accumulation of glycogen, lipid, or hyaluronan.
Beyond these cytologic changes, reactive mesothelial proliferations can exhibit diverse architectural patterns, including small three-dimensional cell clusters, papillary-like aggregates, and pseudoacini. When present in crowded cell groups, reactive mesothelial cells often display a scalloped or "knobby" border. Background inflammatory cells may also be seen in some pleural effusions containing reactive mesothelial cells.
Figure 1 Reactive mesothelial cells can display significantly enlarged nuclear to cytoplasmic (N:C) ratios, coarse chromatin distribution, and prominent nucleoli in the background of variable amounts of inflammatory cells (A). Degenerative changes and intracytoplasmic accumulation of substances can induce cytoplasmic vacuolization within mesothelial cells. When arranged in clusters, mesothelial cells are shown to have scalloped or “knobby” borders (B) (pleural effusion ThinPrep, Papanicolaou, original magnification ×60 [A]; air-dried smear preparation, Diff-Quik, original magnification ×40 [B]).
Figure 2. Adenocarcinoma showing a pleomorphic population of crowded cells with varying degrees of nuclear enlargement and irregularity (A). In a subset of adenocarcinomas, a continuous, smooth “community” border may be appreciated, creating a distinct cell cluster outline (B) (pleural effusion, air-dried smear preparation, Diff-Quik, original magnifications ×40 [A] and ×10 [B]).
Overtly malignant pleural effusions are most often associated with metastatic disease, frequently originating from adenocarcinomas, which have been reported to account for the majority of malignant pleural effusions. In many cases, a malignant pleural effusion is expected as part of disease progression or widespread metastasis from a known primary tumor. However, in some instances, malignant effusions may serve as the first indication of an undiagnosed malignancy.
Adenocarcinomas in pleural effusion samples typically exhibit a biphasic cell population, consisting of a distinct group of malignant cells intermixed with background benign elements. The malignant cells often show pronounced pleomorphism, characterized by enlarged nuclei (resulting in an increased nuclear-to-cytoplasmic [N:C] ratio), irregular nuclear contours, coarse chromatin distribution, and prominent nucleoli. While some tumor types, such as lobular carcinoma of the breast and signet ring carcinoma of the gastrointestinal tract, may present with singly dispersed malignant cells, many adenocarcinomas form three-dimensional cell clusters. These clusters demonstrate varying degrees of nuclear overlap and possess smooth, "community" borders. These smooth borders contrast with the characteristically scalloped borders seen in mesothelial cell clusters.
The significant morphologic overlap between mesothelial cell proliferations and adenocarcinoma can make it difficult to accurately determine the cell origin based on morphology alone. As previously mentioned, pleomorphism can be observed in both malignant processes and reactive mesothelial cells. Conversely, some adenocarcinoma subtypes may lack prominent pleomorphism and instead display a more uniform population of malignant cells, such as certain subsets of breast and endometrioid adenocarcinomas.
Although "community" borders are often associated with adenocarcinoma cell clusters, they can occasionally be seen in mesothelial proliferations as well. Additional mesothelial cell characteristics can further complicate the distinction between mesothelial proliferations and adenocarcinoma. While cytoplasmic vacuolization is more commonly linked to adenocarcinoma, both reactive and malignant mesothelial cells may also exhibit varying degrees of vacuolization, differing in size and location (either peripheral or central). Large vacuoles can indent mesothelial cell nuclei, mimicking the appearance of signet ring adenocarcinoma.
Moreover, in Diff-Quik–stained smear preparations, peripheral cytoplasmic blebs are more readily apparent in mesothelial cells. Another potential diagnostic challenge is the presence of “collagen balls,” which are more frequently seen in pelvic washing cytology specimens. These structures consist of attenuated benign mesothelial cells surrounding a dense, acellular, rounded collagen fragment, which can resemble the three-dimensional clusters typically observed in adenocarcinoma.
When morphology fails to provide a clear distinction between mesothelioma and reactive mesothelial proliferations, ancillary studies can be useful. Traditionally, general epithelial markers such as MOC-31, BerEP4, B72.3, Leu-M1, and carcinoembryonic antigen (CEA) have been used to identify carcinomas in effusion specimens. However, similar to mesothelial markers, these markers also come with interpretative challenges. For instance, Ber-EP4, B72.3, and MOC-31 have been reported to show positivity in small subsets of mesotheliomas, which can lead to potential diagnostic pitfalls.
A newer marker, claudin-4, has demonstrated higher sensitivity and specificity than MOC-31 and Ber-EP4 in distinguishing metastatic adenocarcinoma from mesothelial proliferations. Additionally, organ-specific immunohistochemical markers are valuable in identifying metastatic adenocarcinomas in patients with known or suspected extrapleural primary tumors. Nuclear site-specific markers can be particularly useful in these cases, such as TTF-1 for lung adenocarcinomas, p63 or p40 for squamous cell carcinomas, and CDX-2 for adenocarcinomas with intestinal differentiation.
As pathologists fluid cytology is a routine investigation but it can shed wonderful insights to conclude the diagnosis, but this can be quite difficult once we come across a reactive mesothelial cell population, recently reported 2 such cases of pleural effusion, with minimal atypia which were later confirmed as adenocarcinoma on biopsy.
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