Cytology for Primary Osteolytic Neoplasia

Jul 11, 2017 / Practitioner Updates

Osteoblasts with eccentrically placed nuclei and perinuclear clearing.

Aspiration of lytic bone lesions is becoming a more common technique in the diagnosis of primary bone tumors. This article describes what’s new in the diagnosis of primary osteolytic neoplasia.

Tumors associated with bone lysis or proliferation are categorized as primary bone tumors, i.e., tumors that arise from cells associated with bone. Just as there are various cells types responsible for the formation of healthy bone, there are various tumors types classified as primary bone tumors, including: fibrosarcoma, osteosarcoma, chondrosarcoma, synovial cell sarcoma, histiocytic sarcoma, and hemangiosarcoma.

Many of these tumors share similar cytologic features, making them difficult to distinguish with a standard stain used for cytology. It is critically important todifferentiate neoplastic bone from “reactive” proliferative bone in response to injury or lysis. Reactiveosteoblasts can be large and have prominent nucleoli, but they often have small nuclei relative to the amount of cytoplasm (Figure 1, top of page).

neoplastic cells

Figure 2: Neoplastic cells from a chondrosarcoma; the cells are embedded in thick mats of eosinophilic matrix. 500x magnification.

Generally, aspirates from neoplastic bone are much more cellular than aspirates of healthy or even reactive bone, because of the lysis of the matrix and the disorganized proliferation of neoplastic cells. Prominent criteria of malignancy are easily recognized. Often, standard cytology can give a diagnosis of sarcoma, while a more specific diagnosis requires special staining. In addition, each tumor has distinguishing features that may allow for ranking of a differential list, including: location of the lesion, radiographic appearance, and breed in addition to specific cytologic features.

The most distinguishing feature of chondrosarcomas is the presence of abundant eosinophilic matrix within the background (Figure 2). Fibrosarcomas are more likely to consist of spindle-shaped cells when compared with osteosarcomas and chondrosarcomas, which are more likely to have round cells.


Figure 3: Osteosarcoma from a dog. The cells exhibit marked criteria of malignancy including prominent nucleoli and marked anisocytosis and anisokaryosis. 1000x magnification.

Osteosarcoma cells are often described as plasmacytoid because they can be round and have an eccentrically placed nucleus and sometimes a perinuclear clearing (Figure 3). Osteosarcomas often exhibit remarkable criteria of malignancy. Aspirates of suspected osteosarcomas can be stained for alkaline phosphatase (ALP) activity to increase the sensitivity and specificity of diagnosis. This stain is easy to perform on unstained slides.

In studies of canine osteosarcoma performed at the University of Illinois, ALP staining resulted in 100% sensitivity and 89% specificity in differentiating osteosarcoma from other sarcomas. Positive staining is indicated by a grayish-black staining of the cytoplasm (Figure 4). It is critical to evaluate the sample cytologically to confirm a diagnosis of sarcoma before staining for ALP activity, because reactive bone can stain positive for ALP just as easily as osteosarcoma.


Figure 4: Osteosarcoma stained for ALP activity. The cytoplasm of the cells stain dark grey to black. 500x magnification.

Currently we are evaluating bone tumors in cats for ALP activity. So far, only osteosarcomas are positive for ALP activity in cats.

Synovial cell sarcomas share many of the features of other bone sarcomas and are challenging to diagnose with histopathology as well. Radiographic appearance of the lesion can be the most distinguishing feature, because synovial cellsarcomas are more likely to cross the joint. These tumors are known to have what is described as a biphasic appearance, exhibiting features of both epithelial and mesenchymal cells. Immunostaining of the tumors can reveal positive staining for both vimentin, a marker for mesenchymal cells, and cytokeratin, a marker for epithelial cells. Features of epithelial and mesenchymal cells can be identified with cytology as well as histopathology.

Histiocytic sarcomas commonly result in lytic bone lesions but can also cause disease in the spleen, lung, and other soft tissue structures. Breeds of dogs known to be predisposed to this tumor include Bernese mountain dogs, Rottweilers, golden retrievers, and flat-coated retrievers. Cytologically, these tumors are highly cellular and exhibit marked cellular atypia. These tumors can be distinguished from osteosarcoma with ALP staining because histiocytic sarcoma is negative for ALP activity.

Hemangiosarcoma of bone is fairly uncommon. Cytologically, the sample is often of very low cellularity and the cells can exhibit erythrophagia. Similar to histiocytic sarcoma, the cells are negative for ALP activity and, therefore, can be easily distinguished from osteosarcoma.

Anne Barger, DVM, MS, DACVP

Feature photo above shows Figure 1: Several reactive osteoblasts with eccentrically placed nuclei and perinuclear clearing. 500x magnification.