Ultrasound-guided fine-needle aspiration cytology of solid pseudopapillary tumor of the pancreas: a tertiary care centre experience

Background: Solid pseudopapillary tumours (SPT) of pancreas is an unusual neoplasm with uncertain malignant potential. It is seen predominantly in adolescent girls and young women. We present the clinical, ultrasound-guided fine needle aspiration (US-FNA) cytologic features, differential diagnosis and immunohistochemical findings in SPT. Methods: A retrospective 4-year (2007–2010) fine needle aspiration samples of 8 solid pseudopapillary tumours of pancreas were reviewed. Results: The entire cases were cured with exploratory laparotomy. This study includes 8 cases (1 male, 7 female) of solid pseudopapillary tumours of pancreas. All these cases showed characteristic cytomorphological features displaying hypercellular smears with presence of several papillary fragments lined by multilayered anisomorphic atypical cells having fine chromatin with delicate nuclear groove, myxoid stroma and foamy macrophages in a haemorrhagic background. Conclusions: Correct preoperative cytological diagnosis of solid pseudopapillary tumours of pancreas may be accurately possible by US-FNA technique and also aids in treatment of the surgically curable cancer in the patients.


Introduction
Solid-pseudopapillary tumour of the pancreas (SPT) is a rare uncertain malignant potential tumour which has distinctive cytopathologic features. It is also known as "Frantz tumor" which was first described by Dr. Frantz in 1959 [1]. SPT occurs more frequently in the body and tail of the pancreas of adolescent girls and young women. This tumour can be large and can undergo degenerative cystic changes that are evidenced on clinical and radiographic studies [2]. SPT typically is limited to the pancreas at the time of preliminary diagnosis, and a complete surgical excision is treatment of choice. Metastases are rare after excision, and even patients with metastasis at initial diagnosis often survive for years or decades [3,4]. On the contrary pancreatic adenocarcinoma is an aggressive neoplasm. So targeted therapy and chemotherapy have been found to be beneficial in the patients which were highlighted in some studies [5]. Interestingly, IFN-β was significantly more effective than IFN-α in inducing cell growth inhibition in pancreatic cancer because it induces a more potent and early cell cycle arrest and apoptosis activation compared with IFN-α [6].
SPT from other pancreatic cystic neoplasms of similar radiologic and cytologic appearance Endoscopic FNA as well as US-FNA can have an important role and provide an accurate preoperative cytodiagnosis, particularly when the USG findings of the mass are inconclusive. US-FNA can differentiate but with different biologic behavior and treatment, such as pancreatic endocrine tumours, acinar cell carcinoma, and papillary mucinous carcinoma [7][8][9][10][11][12][13]. SPT behaves like a low-grade malignancy and has an excellent prognosis and hence its differentiation from its other counterparts is important [14]. This study highlights doi: 10.7243/2052-6199-2-2 mainly the cytomorphologic features, differential diagnosis and immunohistochemistry of the cases of SPT diagnosed by US-FNA.

Materials and methods
This retrospective study in which US guided FNA of pancreatic cases between years 2007-2010 were reviewed. Total number of 150 US guided FNA of pancreas were performed at our institute. Eight out of them were diagnosed as SPT on USguided FNA smears and surgical resection. The clinical details, radiology, cytopathology and histopathology data was retrieved for re-evaluation from the records. US-FNA was performed using 22 gauge needles via a transhepatic (6 cases) or a transgastric (2 cases) approach. The aspirated material was quickly smeared into glass slides, air dried, wet fixed in 95% ethyl alcohol for subsequent Papanicolaou staining and remaining for cellblock (if any). Adequacy of aspirated material was checked under light microscope during the procedures. Immunohistochemical stains for CD10, vimentin, cytokeratins, chromogranin, synaptophysin, neuron-specific enolase and progesterone were performed on 4 cases consisting of surgical resection (two cases) and cellblock samples (two cases). All smears, cell block and tissue sections were reviewed with emphasis on the evaluation of cytomorphologic features and immunohistochemical results. Histopathological tissue sections of all resected specimens were reviewed and findings correlated with those obtained by US-FNA.

Results
A total of 8 patients with cytological diagnosis of solid pseudopapillary tumours of pancreas were identified. Patients' age ranged from 15 to 47 years. Average tumour size was 5.5x4.5 to 16.0x15.0 cm and majorities were located in the pancreatic body and head. Sonologically the tumours were solid in 3 patients (37.5%), mixed solid and cystic in 4 patients (50.0%), and cystic in 1 patients (12.5%). A preoperative diagnosis of solid pseudopapillary tumour was made in 8 patients on the basis of US-FNA cytology. Surgical resection was performed in all cases of SPT.
Patients clinical, USG findings and Cytohistologic findings are summarized in Table 1. The smears showed variable degree of cellularity and characteristically branching papillary fragments in 7 out of 8 cases which composed of vascular stalks with a perivascular amorphous myxoid substance festooned by bland-appearing tumor cells (Figure 1). These tumor cells have variable amounts of scant to moderate cytoplasm with focally projections in some; uniform oval to round nuclei with nuclear grooves, fine chromatin, 1-2 small nucleoli and variable amount of cytoplasm in all cases (Figure 2). The cytology smears also showed areas of necrosis (1 case), mitosis (1 case), cytoplasmic hyaline globules, nuclear groove (7 cases) and cluster of atypical cells with pseudorosette forming structure in 2 cases (Figure 3). In case 5, smear showed prominent large nucleoli in some cells. Some atypical cells have binucleation  and multinucleation with abundant plasmacytoid cytoplasm (2 cases). Background showed areas of haemorrhage (all cases), foam cells (5 cases) and cholesterol crystals in 2 cases (Figure 4). 1 of 8 cases US-FNA in cystic mass of the pancreas yielded haemorrhagic fluid and smear shows predominantly myxoid material and occasional singly scattered small to round atypical cells with nuclear groove, fine chromatin and foamy histiocytes. Cytomorphologic findings with histopathologic diagnosis were described in Table 2 a large heterogeneous circumscribed solid and solid-cystic mass in head, body and tail regions of the pancreas (Figure 5). Histopathological examination were done in all eight cases and showed similar microscopic findings. Outer surface of gross specimen showed well circumscribed mass having variegated solid to cystic cut surface with areas of haemorrhage and necrosis (Figure 6). Tumour cells arranged in papillary fronds, sheets and cords around delicate fibrovascular septae (Figure 7). These cells were small to medium size displays uniform round to oval nuclei with groove, fine chromatin and mild to moderate cytoplasm. Marked degenerative changes were also seen in all the cases. Immunohistochemistry was performed only in four cases and results showed strong positivity for CD10 (Figure 8     neuron-specific enolase (NSE) and progesterone receptors. These immunohistochemical stain results were consistent with previously described immunohistochemistry of the solid peudopapillary tumour of the pancreas.

Discussion
Solid pseudopapillary tumour of the pancreas (SPT) is very rare tumour with low malignant potential having a distinctive clinicopathological profile [15,16]. SPT is most commonly seen in young women with a mean age of 27 years (range, 2-81 years). It is rare in men pediatric population [17][18][19][20]. Similarly our study showed female dominance with average age of 27.25 years. Clinical symptoms of the patients usually present on and off abdominal pain in epigastrium and palpable abdominal mass with compression of adjacent organs when the mass is large. In one case of the patient also showed symptoms of vomiting, jaundice and anorexia. Thus, a cystic mass in the pancreas of a young woman raises a clinical suspicion for SPT. Out of eight cases, five of them showed cystic component on ultrasound. In most patients, the tumour follows an indolent clinical course, and complete resection often is curative. However, approximately 10% to 15% of SPT are malignant and show local infiltration, recurrence, or distant metastasis [21,22]. SPT with malignant behaviour seems more prevalent in the pediatric population, it has been doi: 10.7243/2052-6199-2-2 found in older men and also has indolent behavior [23,24]. The benign-appearing SPT might contrast with the cellular anaplasia present in the metastatic deposits [25]. Thus, there are no histologic features that can predict aggressive clinical behavior. Proposed pathologic features related to aggressive behavior or metastatic potential include diffuse growth pattern, venous invasion, nuclear pleomorphism, mitotic rate, necrosis, and areas of dedifferentiation [26].
Imaging studies of SPT show a well-circumscribed, hetero-  geneous, solid and cystic mass with sometimes calcification and have often central cystic degeneration. Similarly, tumoral calcification can be seen in the often cystic and nonfunctional large pancreatic endocrine tumor, serous cystadenoma, and mucinous cystadenocarcinoma and less commonly in ductal adenocarcinoma [27]. Two of our cases showed calcification. Thus, image diagnosis is not specific [28]. EUS permits a better evaluation of SPT, but the findings also are not specific. The majority of preoperative diagnoses of SPT have been made by percutaneous approach via EUS or US-FNA techniques. These US-FNA cytomorphologic features are highly characteristic and distinct from those of other cystic or solid tumors of the pancreas [29]. US-guided FNA diagnosis of SPT is simple, accurate and cost effective technique. In most of our SPT cases, US-FNA cytology smears showed numerous isolated branching papillaroid fronds with capillaries having multilayered mildly anisomorphic round to oval atypical cells displaying nuclear groove, fine chromatin, scant to moderate cytoplasm alongwith myxoid stroma, foamy histiocytes, occasional bi/multinucleated cells, cellular debris, psammoma bodies and eosinophilic cytoplasmic hyaline globules on a hemorrhagic background. These hyaline globule are periodic acid-Schiff-positive with diastase-resistant inclusions and similar findings are also described in some previous cases [30][31][32][33][34][35][36]. Thus we recommend that when these cytological features are present in the smears,cell block and in combination with immunohistochemical profile, the diagnosis of SPT can be made straight forward. However, pancreatic endocrine tumors, papillary mucinous carcinoma, acinar cell carcinoma and, occasionally, intraductal papillary mucinous tumour and serous cystadenoma must be excluded. In one of our case the US-FNA in cystic mass of the pancreas yielded haemorrhagic fluid and smear shows myxoid material and few singly scattered small to large atypical cells with round to oval eccentric nuclei with nuclear groove in some, fine chromatin and moderate to abundant cytoplasm along withfew foamy histiocytes on the background.
Immunohistochemically, most SPT are immunoreactive for vimentin (vim), a1-antitrypsin, a1-antichymotrypsin [37], occasionally positive for neuron specific enolase and synaptophysin [38] and nonreactive for S-100, CA 19.9 and chromogranin A. In our cases, SPT showed strong expression of vimentin, CD10 alongwith focal for neuron-specific enolase, synaptophysin, progesterone receptors and variable cytokeratin. Lack of expression of epithelial membrane antigen and chromogranin is inconsistent with any of the normal pancreatic cell types and reinforces the epithelial, mesenchymal, and endocrine cell phenotype that suggests a pluripotential embryonic stem cell origin for SPT [39,40]. Immunohistochemical detection of progesterone receptors has been found in the neoplastic epithelium of SPT in women but not in men [41].
The main differential diagnoses of SPT includes pancreatic neuroendocrine tumour, acinar cell carcinoma, papillary doi: 10.7243/2052-6199-2-2 mucinous carcinoma and intraductal papillary mucinous tumour (IPMT) ( Table 3). Pancreatic endocrine tumours mainly occur in older patients and may be associated with a variety of clinical syndromes. Pancreatic neuroendocrine tumours might show similar cytomorphologic features, and, in the absence of papillary structures with perivascular myxoid substance, the distinction between the two is difficult. The presence of rosettes, without papillary structures, occasional multinucleated giant cells, and spackled nuclear chromatin or "salt and pepper" chromatin indicates in favor of a pancreatic endocrine tumour [42,43]. Cystic neuroendocrine tumours of the pancreas usually are functional but occasionally might be nonfunctional and are more likely to be confused with SPT on imaging studies [44]. These tumors express neuroendocrine markers such as chromogranin, neuron-specific enolase, and synaptophysin which are usually nonreactive in cases of SPT.
Pancreatoblastoma is a tumour of childhood that lacks the pseudopapillary pattern and fibrovascular stalks with myxoid stroma and consistently is negative for vimentin and positive for pancreatic enzymes, features that allow its distinction from SPT. Acinar cell carcinomas with an endocrine component having immunostains and ultrastructural features of both tumours have been described [42]. Serous cystadenoma showed a well-demarcated solid or cystic mass by EUS and might exhibit prominent papillary differentiation on FNA.
Papillary mucinous carcinoma presents a single, large, often unilocular cystic mass in patients [45]. In compare with SPT, the cells in papillary mucinous carcinoma are columnar with variable nuclear anaplasia, irregular chromatin, and prominent nucleoli [46]. The abundant mucinous background of papillary mucinous carcinoma must not be confused with the myxoid stroma and globoid structures seen in SPT. Thick glistening and viscid mucus material almost always present in intraductal papillary mucinous tumour is an important feature that distinguishes this neoplasm from SPT [47].
Ricardo H et al., focused on the cytomorphologic features and differential diagnosis of 6 new cases of SPT diagnosed EUS-guided FNA and briefly addressed the recent advances in his understanding of the histogenesis, immunohistochemical analysis, and genetic events of this tumor [48]. Salla et al., reported the clinical, imaging, cytomorphologic features and differential diagnosis of a 17-year-old woman having SPT diagnosed by EUS-guided FNA with a review of the literature in which patient has complaints of an unexplained episodic pain for 2 months and a short history of hypertension. The cytomorphology of the tumor is highly characteristic, with features that are distinctive from those of other cystic and solid tumors of the pancreas [49]. Mehta et al., also described the cytomorphology of solid pseudopapillary tumor of the pancreas and its differential diagnosis in six cases diagnosed preoperatively on US-FNA [50].
An accurate FNA diagnosis will alter the eagerness of patient and surgeon to a complete resection of the tumour, since recurrence has not been reported after complete local resection. In addition, while surgery might not be different, the diagnosis is important if considering preoperative (neoadjuvant) chemotherapy, radiotherapy, or both for acknowledged carcinoma.

Conclusions
We consider as that US-FNA can present as best cellular yield with an overall diagnostic accuracy and sensitivity for the correct and early cytological diagnosis of SPT