Two cases of breast carcinoma with osteoclastic giant cells: Are the osteoclastic giant cells pro-tumoural differentiation of macrophages?

Case 1 grossly showed a well-circumscribed solid tumor, measuring 3.5 × 2.5 cm on the maximum cut plane. The tumor appeared gray to white in the center, but hemorrhagic dark brown at the periphery (Figure 1A). Microscopically, the tumor was surrounded by hyperemic blood vessels (Figure 1B). The ductal carcinoma cells, which had relatively small and round nuclei with mild atypia and infrequent mitosis, formed distinct glandular and cribriform structures, equivalent to invasive ductal carcinoma, grade 1 (well-differentiated) (Figure 1C, D). The hypervascular stroma contained numerous inflammatory cells and multinucleated giant cells (Figure 1E). Some giant cells showed a polarized cell body resembling activated osteoclasts (Figure 1F). Immunohistochemically, strong staining for cytokeratin (CK) AE1/AE3 was observed in all tumor cells, but not in OGCs (Figure 1G). About 40% of tumor cells were positive for estrogen receptor (ER) (Figure 1H) and progesterone receptor (PgR), but negative for HER2, phenotypically corresponding to Luminal A type. Following the WHO classification, the diagnosis was breast carcinoma with OGCs.

The tumor in Case 2 also showed a well-circumscribed gross appearance, measuring 2.2 × 1.5 cm on the maximum cut plane. The tumor was rather whitish, but contained a small (5 mm) hemorrhagic area (Figure 1I). Microscopically, the main part of tumor consisted of spindle-shaped sarcomatous cells with frequent mitotic figures, and there were also foci of ductal carcinoma cells with an intraductal component (Figure 1J-L). Inflammatory cells and multinucleated OGCs were seen in the hypervascular stroma (Figure 1M, N). Immunohistochemically, the ductal carcinoma cells were positive for CK AE1/AE3 (Figure 1O) but not ER, PgR, or HER2. The spindle-shaped tumor cells constituting most of the tumor were not reactive for CK AE1/AE3, but instead were strongly positive for vimentin (Figure 1P). Thus, the case was diagnosed as breast carcinoma with OGCs, and the tumor was equivalent to metaplastic carcinoma with mesenchymal component, corresponding to so-called carcinosarcoma.

Despite the different histological features of the breast carcinoma cells, the OGCs in both cases showed similar morphology, and large OGCs contained 20-30 nuclei. In both cases, OGCs preferentially appeared in hypervascular stroma, but the topography of OGCs and blood vessels differed between Case 1 and Case 2. In Case 1, CD31 immunostaining demonstrated that the smaller blood vessels were relatively evenly distributed within the tumor; in contrast, numerous enlarged blood vessels were seen in the periphery (Figure 2A). Large OGCs with over 20 nuclei showed concentric topography; they were mainly seen in the central invasive lesion, while mono-, bi-, tri-, or oligonucleate CD68-positive cells were usually observed in the periphery (Figure 2B). Typical OGCs containing more than 20 nuclei were scarcely detected in the intraductal components, although some CD68-positive cells, including mono-, bi- or trinucleate cells, were seen within the ducts (Figure 2C). In Case 2, unlike Case 1, the distribution of OGCs and blood vessels was irregular, not concentric. OGCs were usually clustered in irregularly distributed hypervascular areas (Figure 2D). Mono-, bi-, tri-, or oligonucleated cells were also observed around the OGCs (Figure 2E). However, CD68-positive cells, regardless of the number of the nuclei, were scarcely seen within the minor intraductal lesions (Figure 2F).

To assess the microenvironment of the tumors, we examined the expression of two chemoattractants for macrophage migration and angiogenesis, VEGF and MMP12. Prominent expression of VEGF and MMP12 was observed in most tumor cells, most inflammatory cells, and even neighboring normal mammary glands both in Case 1 and Case 2 (Figure 2G-R). The OGCs, in both cases, also displayed marked expression of VEGF and MMP12.

In hypervascular stroma of both cases, microvessel density was evaluated according to the Chalkley count on CD-31-stained sections, as described earlier. The mean values of the three most vascular areas (hot spot) were 9.6 for Case 1 and 10.7 for Case 2, respectively. These counts were much higher than the average Chalkley count of a total 330 invasive breast carcinoma cases, 5.75 (range 2.33 - 10.67, median 5.67, SD 1.54), reported in a previous study[11, 12].

Phenotypic characteristics of OGCs in the two cases were evaluated and compared with those of osteoclastic cells in fibrous dysplasia of the rib bone (from a 45-year-old woman) and foreign-body giant cells in re-operated breast tissue (a 42-year-old woman). CD68, a histiocytic marker, was detected on all macrophage lineage cells, including OGCs and oligonucleated giant cells in both Case 1 and Case 2, osteoclastic cells in the bone sample, and foreign-body giant cells (Figure 3A-D). MMP9 is a broad marker for macrophage-osteoclast lineage cells, including mononuclear precursors, fused polykaryons, and mature osteoclasts in bone. MMP9 expression was distinctly detected in the osteoclastic cells (Figure 3E) and the OGCs in Case 1 (Figure 3F), but markedly weaker in Case 2 (Figure 3G) and foreign-body giant cells (Figure 3H). TRAP and cathepsin K, lytic enzymes for bone resorption, are functional markers for osteoclast-lineage cells. Their expression was distinct in the osteoclastic cells and the OGCs in both Case 1 and Case 2 (Figure 3I-K, M-O). Unexpectedly, foreign body giant cells were also strongly positive for TRAP (Figure 3L), although they were negative for cathepsin K (Figure 3P). HLA-DR, an MHC class II antigen, is generally expressed in antigen-presenting cells including macrophages and peripheral blood mononuclear cells. Only foreign body giant cells were positive for HLA-DR; the bone osteoclasts and OGCs were negative (Figure 3Q-T).