Peritoneal washing cytology in an adult granulosa cell tumor: A case report and review of literature.

Adult-type granulosa cell tumors (AGCT) account for 1-2% of all ovarian tumors and 95% of granulosa cell tumors. In AGCT, at the time of peritoneal washing, tumor cells are rarely seen to exfoliate and FIGO stage is raised from IA/IB or IIA/IIB to stage IC or IIC despite the absence of a gross intraepithelial tumor. Patients with positive peritoneal washing cytology must be followed up for pelvic recurrence and metastasis. A more sensitive cytologic evaluation, histopathologic correlation and immunohistochemical staining can advance our practice. Here, we describe a case of AGCT with the emphasis on cytologic features observed in specimens obtained from peritoneal washing fluid.

Introduction

Peritoneal washing is routinely performed at the time of abdominal surgery and used to detect subclinical intraperitoneal metastases.[1] However, while it is important for tumor staging, it is very difficult to identify the presence of rare tumor cells and to diagnose AGCT using peritoneal washing. Cytological examination of tumor cells has mostly focused on samples obtained using fine-needle aspiration and ascitic fluids. As no study of cytology samples obtained from peritoneal washing is known, we describe a case of AGCT with emphasis on the cytologic features observed in specimens obtained from peritoneal washing fluid.

Case Report

A 30-year-old woman presented with menometrorrhagia and abdominal pain 6 years ago. She subsequently underwent left salpingo-oophorectomy (USO) for an adnexal mass, which was diagnosed as AGCT. Then, chemotherapy (Paclitaxel + Carboplatin) was prescribed on the basis of clinical records. In the follow-up, a hetero-echoic mass lesion measuring 6 × 4 cm with multicystic areas in the right adnexal region and multiple peritoneal and omental implants were seen on computed tomography (CT) scans. Then total abdominal hysterectomy with right USO, omentectomy, appendectomy and peritoneal washing were done.

Peritoneal washing fluid was prepared using conventional methods. Cytology was hypercellular with both large and small overlapping cell clusters [Figure 1]. Tumor cells were round to oval, with hyperchromatic nuclei containing granular chromatin. Micronucleoli and cytoplasmic vacuoles were not observed. Characteristic Call-Exner bodies showing microfollicular structures with amorphous material and rare, coffee-bean-like nuclear grooves were seen [Figure 2]. Theca cells were not observed.

Figure 1

Photomicrograph of cytology specimen which have overlapping cell clusters. (H and E, ×400)

Figure 2

Photomicrograph of cytology specimen. Tumor cells surrounding eosinophilic hyaline globules resembling call-Exner bodies. (H and E, ×100)

Gross examination of the right ovarian mass showed a tan and solid tumor with small cystic structures. Microscopically, the tumor was composed of granulosa cells arranged in the trabecular and microfollicular pattern [Figure 3]. Mild cytological atypia and 6 mitosis per 10 high power field (×400) were observed in the tumor, while there was no necrosis. Immunohistochemical stains exhibited that the tumor cells were strongly and diffusely cytoplasmic positive for inhibin and negative for epithelial membrane antigen (EMA) [Figure 4]. Both the cytological and histological findings were consistent with the diagnosis of AGCT. The histological examination of the implants in the peritoneal cavity and omentum also confirmed the diagnosis of AGCT.

Figure 3

Photomicrograph of adult granulosa cell tumor histopathology. Tumor cells with round to oval nuclei. Characteristic Call-Exner bodies showing microfollicular structures with amorphous material are seen. (H and E, ×20)

Figure 4

Photomicrograph of positive immunohistochemical stain for inhibin. (IHC, ×40)

Discussion

AGCT is the ovarian sex cord stromal tumor derived from the granulosa cells and luteinised cells[2] and accounts for 1% of all ovarian tumors.[3] Although the histology of ovarian AGCTs is well documented, it is rarely encountered in cytological specimens and identification of tumor cells is very difficult. Care is necessary when evaluating ovarian tumors using specimens obtained by fine-needle aspiration, peritoneal washing, ascitic fluid and pleural fluid, since tumor cells can be easily overlooked. Cytological preparations of AGCT demonstrate three-dimensional clusters of monotonous cells comprising a loose monolayer and individual cells showing uniform, pale, round to oval nuclei, rare small nucleoli with or without coffee-bean-like nuclear grooves and granular chromatin. Characteristic microfollicular structures with amorphous material, called Call-Exner bodies, might also be seen.

When we reviewed the literature, we found that AGCT was rarely diagnosed by peritoneal washing cytology. Gupta et al.[4] described a case in which tumor cells exfoliated in the pleural and ascitic fluids with exudative effusions. The largest series of AGCT in cytological preparations were reported by Ali et al.[5] who described 10 AGCT cases that were diagnosed by fine-needle aspiration. In nine of these cases, large and small overlapping cell clusters, individual cells and nuclear grooves were observed, whereas Call-Exner bodies were seen in only seven cases. Tumor cell necrosis, small-punctuate cytoplasmic vacuoles and naked nuclei were also noted. Atypical mitotic activity was not identified in any of 10 cases. They also noted that prominent nucleoli, cytoplasmic vacuoles, single cell necrosis and increased mitotic figures were correlated with aggressive behaviour and increased risk of recurrence, and clinicians should be informed of these findings in pathology reports.

Kavuri[2] described the cytologic features of seven ovarian GCTs (six AGCT and one JGCT). The peritoneal washing and ascitic fluids of these patients were examined. Imprints prepared from tumors, intraoperatively during frozen section, were also evaluated. They found that fluids were less cellular than imprints that had fewer grooves, single cells and naked nuclei. Call–Exner bodies were not identified in fluids but were prominent in imprints.

Lal[6] reported the cytomorphologic features of seven histologically confirmed GCTs in fine-needle aspiration and peritoneal washing specimens. In their report, they emphasized the importance of some cytologic features, such as theca cells and the presence of prominent arborisation of vessels, which were not previously mentioned. They also suggested that immunostaining for inhibin antibody on cell block specimens might be useful for a definitive diagnosis of GCT in cytologic samples.

In our case, peritoneal washing fluid was hypercellular with large and small overlapping cell clusters. We observed loose monolayer cells showing microfollicular structures with amorphous material resembling Call-Exner bodies and rare, coffee bean like nuclear grooves were seen.

The differential diagnosis of AGCT in fluids includes follicle cysts, carcinoid tumor, small cell carcinoma of ovary pulmonary-type, undifferentiated carcinoma, Brenner tumor, other sex cord stromal tumor like Sertoli-Leydig cell tumor and sex-cord tumor with annular tubules (SCTAT).

As the cytomorphology of a follicle cyst mimics that of an AGCT, correlation with sonographic and laparoscopic findings is essential.[7] The nuclei of the cells in carcinoids, which have coarse chromatin, contrast with the pale nuclei of the AGCT. The presence of characteristic nuclei with ‘salt and pepper’ chromatin and indistinct nucleoli, eosinophilic granular cytoplasm with distinct outlines and chromogranin or synaptophsin positivity is helpful in the identification of a carcinoid tumor. Additionally, the acini of carcinoids often contain a dense eosinophilic secretion, which is sometimes calcified.[37] In undifferentiated carcinoma, the nuclei are usually unequal in size and shape and are rarely grooved.[37] In pulmonary-type small cell carcinoma of the ovary, dark, uniform, small, ungrooved nuclei, inconspicuous nucleoli and scanty cytoplasm are mostly seen.[37] Large germ cells with clear cytoplasm along with smaller sex-cord-type cells and OCT-4 immunopositivity in gonadoblastoma are helpful in the identification of these tumors.[37] The presence of two distinctive cell types, one containing abundant eosinophilic cytoplasm, and the other with scanty cytoplasm and ring-shaped simple tubules with central hyaline bodies, help to distinguish SCTAT. Hyaline deposits in these tumors are typically larger than Call–Exner bodies, which are often calcified.[37] Like AGCT, a Brenner tumor has pale cytoplasm and often has grooved nuclei. It could be difficult to distinguish these two entities but immunohistochemistry is useful in resolving this difficulty.[37]

Similarly, it could be difficult to differentiate sertoli cell tumors (SCT) from AGCT, since SCTs are also formed by small cells with uniform oval nuclei and prominent nucleoli, and irregularly distributed chromatin.[37]

Peritoneal washing is routinely performed at the time of surgery and used to detect subclinical intraperitoneal metastases. Exfoliation of tumor cells in peritoneal washing samples raises the FIGO stage from IA/IB or IIA/IIB to stage IC or IIC despite the absence of a grossly visible tumor.[1] However, stage is the most important prognostic factor for AGCT.[5] Sun et al.'s[8] follow-up study of 176 cases with adult-type ovarian granulose cell tumors found that 77.8% of the patients were diagnosed at stage I and nearly 95% survived for 5-10 years. However, AGCT also has the potential for aggressive behaviour, which is characterized by local invasion, recurrence and metastasis many years after initial treatment,[269] necessitating long-term clinical follow-up of AGCT patients.[59]

Conclusion

Patients with positive peritoneal washing cytology must be followed up for pelvic recurrence and metastasis. This requirement calls for care in the evaluation of peritoneal washing cytology. Due to their rarity, adult-type granulosa cell tumors present a diagnostic challenge for cytologic preparations. We believe that sensitive cytologic evaluation, histopathologic correlation, taking clinical history and a positive immunohistochemical reaction with inhibin could help us in practice.