The place of FNA in the investigative sequence

The utility and practice of tFNA of the prostate and the reasons for its success in our hospital have been reported previously.^8-10 Provided the pathologist is able to pinpoint nodules and indurations of the prostate by palpation with the index finger, and has dexterity in performing tFNA, the results of FNA match those obtained by TNCB.¹⁰

The three-tiered cytological classification for grading prostate cancer proposed by Esposti²⁷ has been shown to have prognostic value. An estimation of Gleason score can also be made on tFNA samples.¹⁰ As recently as 2007, Maksen et al.⁵ stated that gland size roughly determines Gleason pattern in liquid-fixed FNA samples. Prostatic intraepithelial neoplasia (PIN) should not be diagnosed by FNA alone. However, a highly cellular smear with pronounced atypia seems to rule out PIN.²⁸ Immunostaining has proven useful in FNA of the prostate.^5,29

tFNA is likely to continue to be used in view of economic considerations and recent health budget restraints because it is a simple and comparatively much cheaper technique than TCNB. We acknowledge that nowadays tFNA has been relegated in most Western countries, but it may still play a key role in developing countries.

Indications

Accuracy of diagnosis

Transrectal fine needle aspiration and US-guided TNCB using automated biopsy devices have a high and essentially equal accuracy in diagnosing prostate cancer.^4,5 A recent review of the literature¹⁰ produced a list of references commenting on the correlation between cytological grading and Gleason score and the accuracy of digitally guided tFNA compared with thick and US-guided TNCB. Prerequisites for acquisition of an acceptable level of competence in performing tFNA and interpreting the smears have been defined.³⁴

Tannenbaum et al.⁴ and Maksen et al.⁵ have described the features of benign prostatic cytology, the sources of atypical cells, the cytology of carcinoma, and the causes of false-negative and false-positive cytological diagnoses in liquid-fixed FNA collections, and Willems et al.³⁴ in conventional smears. The most common cause of a false-negative FNA is an inadequate sample. Dexterous pathologists rarely make a false-positive diagnosis. We have previously commented on the diagnostic challenges posed by atypical cells in smears of granulomatous prostatitis⁸ and on pitfalls and infrequent findings in FNA of the prostate.⁹ Basic rules to avoid pitfalls in FNA in different contexts have been put forward.³⁵ See also Chapter 5 in the seminal monograph by Zajicek³⁶ and Chapter 11 in the book by Linsk and Franzén.³⁷

Complications

The rare complications of tFNA were analyzed in a review of 14 000 patients with follow-up.³⁸ Transient hematuria is not uncommon. Fever may develop within 24 hours. Patients should be instructed to report promptly in both situations. In our experience of more than 10 000 tFNAs (using 23-gauge needles), no tumor seeding in the needle track has been observed after 5 to 10 years’ follow-up.

Contraindications

The only contraindication is symptomatic febrile prostatitis.³⁶ We have performed tFNAs in patients with inadvertent acute prostatitis without complications. tFNA can be performed in patients with hemorrhagic diathesis and in patients on anticoagulation therapy.

Technical considerations

Patients should be verbally informed of the whole procedure by the examining pathologist and a written informed consent requested. No special preparations for biopsy and no sedation are necessary. tFNA is carried out as an outpatient procedure. A urological lubricant helps to introduce the palpating finger. The most comfortable position for both patient and examining pathologist is the lithotomy position on a gynecological examination couch (Fig. 13.1A,B). A rapid feedback to urologists is essential to achieve success.

Fig. 13.1 t-FNA of the prostate

(A) The Franzén guide fitted to the left index finger covered with a finger cot; (B) Performing a tFNA of the prostate.

Appropriate training should ensure that the operator properly masters the technique of tFNA using the Franzén guide and a 20-cm long 23-gauge needle fitted to a disposable syringe and a Cameco holder (Cameco AB, Box 5519, Täby, S-183 05 Sweden, Pat no. 3819091). This is essential to minimise the proportion of unsatisfactory specimens and to increase the accuracy of the procedure. The examining pathologist must be proficient in digital examination of the prostate. Details about the biopsy technique can be found elsewhere.^34,37

We perform one or two tFNAs from each abnormal palpable zone; additional aspirates are taken from the opposite lobe even if apparently normal on palpation. This approach enables us to suggest at least a T2C stage if cancer cells are found in aspirates from both lobes. We use Diff-Quik (DQ)™ and Pap stain in parallel.

The gross aspect of the yield may predict the diagnosis even before microscopic examination: abundant, granular yield suggests carcinoma; a drop of blood-tinged fluid with particulate material suggests hyperplasia; hemorrhagic yield indicates that the needle is out of the prostate; a ‘non-representative’ smear obtained by a dexterous operator suggests stromal hyperplasia; abundant fluid suggests either urine or a cyst.

The importance of proficient training in smearing and distributing the yield onto several slides cannot be overemphasised (see Chapter 2). The three-step smearing technique is recommended for fluid yields in order to concentrate solid fragments into two bands. The yield is suitable for molecular studies.

Benign prostatic hyperplasia (BPH) (Fig. 13.2)^{4,5,34,36,37,39}

Fig. 13.2 Prostate, benign hyperplasia

Honeycomb-like sheets of uniform glandular epithelial cells; note visible cell borders and coarse cytoplasmic granules in A (A, DQ; B, Pap, HP).

Cohesive monolayered sheets of glandular epithelial cells can be quite large with distinct boundaries; most cells are seen on end and appear polygonal with centrally placed nuclei. The abundant pale cytoplasm and the distinct cell membranes give the sheet a honeycomb appearance (Fig. 13.2A,B). Only at the periphery are some cells seen in profile as columnar.

The main criteria of benignity are the uniform distribution of nuclei within monolayered sheets, distinct cell membranes, low N : C ratio and intracytoplasmic secretory granules (Fig. 13.2A). The granules stain dark magenta with DQ but are less conspicuous in alcohol-fixed Pap-stained smears. Although granules are not present in all benign epithelial cells, they are rarely present in carcinoma cells and absent in epithelial cells from rectal mucosa.

Other common findings in smears of BPH are: inflammatory cells, macrophages, metaplastic squamous epithelial cells, clumps of condensed secretion, fragments of calculi and corpora amylacea. Tiny fragments of stromal smooth muscle tissue are sometimes seen. Cytologic features of the central zone have been described in scrape smears of surgical specimens.⁴⁰

Prostatitis (Figs 13.3 and 13.4)^4,36,37,39

Fig. 13.3 Prostatitis (acute)

Irregular epithelial sheet with mildly enlarged nuclei; background of neutrophils (DQ, HP).

Fig. 13.4 Prostatitis (granulomatous)

Many epithelioid histiocytes, multinucleated giant cell with phagocytosed secretion; many neutrophils (DQ, IP).

Mild epithelial atypia is acceptable in the presence of significant inflammation (Fig. 13.3). The distribution of cells in epithelial sheets may be less regular than normal and the cell membranes less distinct. Nuclei may be mildly enlarged and varying in size. Cytoplasmic granules are often absent, while degenerative changes such as cytoplasmic vacuolation are often seen. However, prominent nuclear enlargement and pleomorphism, nucleolar enlargement and chromatin abnormalities do not occur. There is little tendency to dissociation of epithelial sheets, and microacini are not seen. As inflammation may coexist with carcinoma, epithelial atypia must be carefully evaluated. We require the presence of epithelial sheets encrusted with polymorphonuclears to arrive at the diagnosis of acute prostatitis.

Granulomatous prostatitis (Fig. 13.4)^36,37,39 remains a diagnostic dilemma since both clinical and cytological findings may mimic carcinoma. The cytological diagnosis of non-specific granulomatous prostatitis or tuberculous prostatitis, respectively, is based on the presence of epithelioid granulomas or obvious caseous necrosis. Nuclear overlapping, anisonucleosis, occasionally striking atypia, naked nuclei and some acinar formation may result in a false-positive diagnosis. Epithelial atypical cells in granulomatous prostatitis show a typical basophilia in DQ-stained smears; this basophilia is not seen in prostate carcinoma cells.

The diagnostic challenges posed by epithelioid aggregates and reactive changes in both duct/acinar and metaplastic cells have been discussed elsewhere.⁸

The cytological findings in malacoplakia of the prostate by FNA have been described.⁴¹

Adenocarcinoma of prostate (Figs 13.5-13.8)^{4,5,27,34,36,37,39}

Fig. 13.5 Well-differentiated adenocarcinoma

Cohesive sheets of atypical glandular epithelium; mild nuclear enlargement and anisokaryosis; crowding of nuclei; absence of cytoplasmic granules. Note microacinar pattern in B and prominent nucleoli and a mitotic figure in C (A and B, DQ; C, Pap, HP).

Fig. 13.6 Moderately-differentiated adenocarcinoma

Aggregate of malignant cells; marked nuclear enlargement and nuclear pleomorphism; nuclear crowding; microacinar pattern: indistinct cell borders. Note contrasting honeycomb sheet of benign epithelium (DQ, HP).

Fig. 13.7 Moderately-differentiated adenocarcinoma

Cellular smears of less cohesive malignant glandular epithelial cells; vesicular nuclei; prominent nucleoli. Note fragile vacuolated cytoplasm suggestive of a clear cell pattern in B (A, Pap; B, DQ, HP).

Fig. 13.8 Poorly-differentiated adenocarcinoma

Poorly cohesive malignant cells with large vesicular malignant nuclei and large nucleoli. Note small sheet of benign epithelium on right in A (A, DQ; B, Pap, HP).

In smears of prostatic carcinoma, sheets of benign glandular epithelial cells are commonly seen side-by-side with aggregates of malignant cells (Fig. 13.6), reflecting the diffusely infiltrative growth of the tumor. Benign and malignant cells can be directly compared (very helpful clue) and differences in cytoarchitectural features are easily appreciated. Nuclear enlargement is one of the most important criteria of malignancy. Nucleolar enlargement is better demonstrated in Pap-stained smears.

Absence of visible cell membranes, nuclear crowding and overlapping and dissociation of cells are other important criteria. The presence of coarse intracytoplasmic secretory granules makes malignancy unlikely, but they can occasionally be found in cells from well-differentiated adenocarcinoma. Cytoplasmic vacuolation may be seen in both benign and malignant cells. Nuclear pleomorphism and chromatin abnormalities are obvious in less well-differentiated cancers but may be subtle in well-differentiated carcinomas, rendering a definitive malignant diagnosis difficult (Fig. 13.5). Demonstration of basal epithelial cells by immunocytochemistry may be of help in the distinction between well-differentiated adenocarcinoma and adenosis or basal cell hyperplasia,⁵ but interpretation is more difficult than in histologic sections. Immunostaining is a useful tool for the diagnosis of prostate cancer at metastatic sites.⁴² Results may be conflicting in metastases of poorly differentiated carcinomas.

If only a small number of atypical cells are found in a predominantly benign cell population, great caution should be observed in making a definitive diagnosis of malignancy. Sources of atypical cells have been referred to elsewhere.^4,5,8,9 In case of doubt, the patient should be submitted to systematic TNCB taken according to a standardized protocol.

Regarding cytological grading of prostatic adenocarcinoma, see below and Figures 13.5 to 13.8.

Not infrequently, smears from BPH contain a few cohesive aggregates of cells with frank atypia that may represent PIN, basal cell or atypical hyperplasia, and can be mistaken for malignancy (Figs 13.9 and 13.10). However, they fall short of clear-cut criteria of malignancy; cell cohesion is maintained and microacini are rarely seen. The atypical cells usually constitute only a minor proportion of the cell population. The presence of nucleolar enlargement in some cells should not lead to the diagnosis of carcinoma since they may correspond to focal atypical hyperplasia.

Fig. 13.9 Basal cell (atypical) hyperplasia

Sheets of glandular epithelial cells showing mild nuclear enlargement and nuclear crowding contrasting with normal epithelium (A, DQ; B, Pap, HP).

Fig. 13.10 Basal cell hyperplasia

Corresponding tissue section (H&E, IP).

PIN should not be diagnosed by tFNA alone.²⁸ We use the term ‘atypical cells’ when the atypia and cellularity are below the requirements needed to reach a confident diagnosis of malignancy, and submit the patient for systematic TNCB. Histology of these cases almost always reveals either high-grade PIN or low-grade carcinoma. Pathologists should refrain from making a definitive diagnosis of malignancy when the smears contain only a small proportion of cells with malignant features.

Contamination of samples by epithelial cells from the rectal mucosa is common when tFNA is performed by an inexperienced operator.^36,37 Isolated tall cylindrical cells, palisaded rows, glandular structures and goblet cells intermingled with mucin and rectal content indicate rectal mucosal origin. Rectal cells lack intracytoplasmic granules (Fig. 13.11).

Fig. 13.11 Rectal contamination

Microglandular aggregates and palisading columnar epithelial cells; some mucin (DQ, IP).

Inadvertent aspiration of the seminal vesicle may yield large atypical cells that may mislead the unwary into an erroneous diagnosis of poorly differentiated carcinoma. Large hyperchromatic, often multilobated, pleomorphic, even bizarre, nuclei are seen.^9,36,37 Coarse intracytoplasmic granules of lipofuscin, which stain dark green–blue with DQ, brown with Pap or H&E, quite different from the secretory granules of prostate epithelium (Figs 13.12 and 13.13), dense aggregates of basophilic amorphous material and spermatozoa in the background indicate origin from seminal vesicle.

Fig. 13.12 Seminal vesicle epithelium

(A) Large bizarre nuclei; note occasional spermatozoa in the background (arrow) (DQ, HP); (B) Single epithelial cells with large pleomorphic nuclei; note coarse intracytoplasmic pigment (DQ, HP).

Fig. 13.13 Seminal vesicle epithelium

(A) Vaguely glandular aggregates of epithelial cells with pleomorphic and hyperchromatic nuclei; abundant cytoplasm with vacuoles and brown pigment granules (Pap, HP); (B) Corresponding tissue section (H&E, HP).

Ganglion cells may mimic malignant cells (Fig. 13.14).⁹

Fig. 13.14 Ganglion cells

Three large ganglion cells with prominent nucleoli next to a cluster of malignant cells. Notice a malignant acinus at the bottom-right corner. (DQ IP).

The differential diagnosis between prostatic adenocarcinoma and transitional cell carcinoma is discussed below. Pitfalls and infrequent findings in prostate aspirates, a list of benign conditions mimicking carcinoma and clues to diagnosis have been published elsewhere.⁹

Effects of hormonal treatment and radiotherapy (Fig. 13.15)^{9,27,31,36,37,39}

Fig. 13.15 Prostate, hormonal effect

Metaplastic squamous cells; large glycogenized cells and clusters of epithelial cells, some with enlarged nuclei. Hormonal response in confirmed prostatic adenocarcinoma (DQ, IP).

Criteria for diagnosis

♦ Squamous metaplasia,

♦ Large cells with abundant transparent cytoplasm resembling a fried egg (‘glycogen cells’),

♦ Nuclear pyknosis,

♦ Loss of nucleoli,

♦ Nuclear shrinkage,

♦ Well-preserved malignant cells may persist associated with squamous metaplasia or ‘glycogen cells’.

Transitional cell carcinoma (Figs 13.16 and 13.17)^9,37

Fig. 13.16 Transitional cell carcinoma

(A) Pleomorphic malignant cells; hyperchromatic eccentric nuclei; dense cytoplasm (tFNA of prostate; DQ, HP); (B) Corresponding tissue section of transitional cell carcinoma invading the prostate (H&E, IP).

Fig. 13.17 Combined transitional/adenocarcinoma

Aggregate of well-differentiated adenocarcinoma with nuclear crowding and microacinar pattern; adjacent group of larger, more pleomorphic malignant transitional cells (DQ, HP).

Transitional cell carcinoma may invade the prostate from the urinary bladder or it may arise from periurethral ducts within the prostate itself. Coexistence of transitional cell carcinoma and adenocarcinoma of the prostate is not rare (Fig. 13.17). It is important to distinguish transitional cell carcinoma from adenocarcinoma, since the former does not respond to hormonal treatment.

Transitional cell carcinomas involving the prostate are usually deeply invasive, high-grade tumors (see also Figs 12.29–12.31). Squamous differentiation is seen in some tumors, whereas papillary structures are rare.

Rare tumors of the prostate⁹

Cytologic descriptions and differential diagnoses of ‘foamy cell carcinoma’, prostatic ductal adenocarcinoma (papillary or endometrioid) (Fig. 13.18), mucinous (colloid), small cell carcinoma, squamous cell carcinoma (Fig. 13.19), metastatic solid tumors within the prostate, and mesenchymal tumors have been described elsewhere.⁹ The differential diagnosis between ductal cell carcinoma, prostatic adenocarcinoma and transitional cell carcinoma has been discussed. Nuclear grooves are a helpful clue for diagnosis of ductal cell carcinoma.⁴³

Fig. 13.18 Ductal cell adenocarcinoma

A papillary cluster. Notice nuclear palisading and monomorphism ( DQ, HP).

Fig. 13.19 Squamous cell carcinoma of prostate

Clusters of small malignant cells, some showing squamous differentiation including a few orangeophilic cells (center and upper left) (Pap, IP).

A morphologic and immunohistochemical study of small cell carcinomas of the prostate has been published.⁴⁴ The cytological pattern is similar to small cell tumors in other sites (Fig. 13.20). A review of adenoid cystic/basal cell carcinoma of the prostate has been reported.⁴⁵ We are not aware of any report of cytological findings of this tumor.

Fig. 13.20 Small cell (neuroendocrine) carcinoma of prostate

(A) Clusters of malignant cells with a high N : C ratio and some nuclear molding (DQ, HP); (B) Tissue section, same case (H&E, IP). Immunostaining for chromogranin was positive.

Rarely, lymphomas may involve the prostate and cause enlargement and clinical symptoms. This could be mistaken for small-cell or anaplastic carcinoma in smears. The diagnosis can be confirmed by immunostaining.

Sarcomas of the prostate are rare. These are mainly rhabdomyosarcomas in children, leiomyosarcomas (Fig. 13.21) or fibrosarcomas in adults (see Chapter 15).^9,46

Fig. 13.21 Leiomyosarcoma of prostate

Tissue fragment of moderately pleomorphic spindle cells with eosinophilic cytoplasm; some loss of cell cohesion (H&E, HP).

The place of FNA in the investigative sequence

Pathologists attempting cytological diagnosis of testicular masses must be fully conversant with the WHO classification of tumors of the testis¹² and with the bewildering patterns shown by these tumors.^12,47-49 This is also essential in the proficient identification of extragonadal germ cell tumors, primary and metastatic, in FNA samples.^50-56 FNA sampling and subsequent microscopy should ideally be performed by the same pathologist to enable cytological interpretation in the light of clinical findings.^14,18

In our hospital, after clinical examination by the urologist, the patient is referred to the clinical cytology unit. The pathologist explains the procedure and obtains a written consent from the patient. A provisional diagnosis of benign or malignant, and if malignant of seminoma versus non-seminoma, is provided to the urologist on site. The whole procedure takes less than 30 minutes. In case of malignancy, orchidectomy follows 3–7 days after diagnosis. Rapid feedback to the urologist is essential.

FNA does not replace histological diagnosis. The aim is mainly to provide a triage of cases of testicular swelling into those who do not require surgery as the first-choice treatment and those who do.¹⁴ Seminoma represents about 50% of TGCT and 40–45% of all testicular neoplasms.⁴⁸ This means that familiarity with the cytologic patterns of seminoma enables the pathologist to identify about 50% of TGCT in FNA samples.

Accuracy of diagnosis

The experience of cytologic features of germ cell tumors (GCT), primary and metastatic, accrued over two decades, permits not only a confident diagnosis of malignancy but also correct tumor typing in most cases.^14,50-66 The proper use of FNA of testicular masses reduces the need for surgery considerably.^21,58-61 Gupta et al.²⁰ underlined the role of FNA in the differential diagnosis of epididymal nodules, avoiding surgical biopsy and other investigations. See also Chapter 12 in the monograph by Linsk and Franzén,⁶⁵ and Chapter 4 in that by Zajicek.⁶⁶

Limitations of FNA of testicular tumors are:¹⁴

A few additional specific problems should be emphasized:

Complications

Fine needle aspiration may be painful in benign conditions and in fertility studies. The pain usually irradiates to the homolateral inguinal zone. It subsides in an hour and can be relieved by painkillers. FNA of malignant conditions and acute scrotum syndrome is almost painless. We have seen microscopic hemorrhage and necrosis in the subsequent surgical specimen, possibly caused by the needle, but this has never prevented histological diagnosis.

The only contraindication to testicular FNA mentioned in the literature is acute orchitis accompanied by cellulitis of the scrotum.⁶⁵

Tumor seeding in the needle track has not been recorded in a number of studies.^58,65-67 Tumor stage (TNM) was not modified following FNA in a series of malignant testicular tumors.⁶¹ In our experience of more than 120 TGCT with 5–10 years’ follow-up, there has been no evidence of dissemination caused by FNA. The benefit of a rapid and reliable diagnosis of malignancy outweighs the unproven risk of dissemination.

Technical considerations

We do not use local anesthesia for testicular FNA. Infiltration of the spermatic cord has been advocated in sequential aspirations for the evaluation of male infertility and for sperm retrieval.⁶⁸ We use 25-gauge needles, only exceptionally 23 gauge, and only one or two biopsies with one or two passes each time. US guidance may be helpful in partly cystic tumors, in non palpable US-detected lesions, in cases of retroperitoneal GCT (burnt-out TCGT), and in the follow-up of patients with lymphoma or leukemia.

Adequate training and experience in performing FNA and handling of samples is of paramount importance (see Chapter 2). Some points are of particular importance in testicular tumors. It is essential that the needle remains within the target during aspiration and that the negative pressure is released before withdrawing the needle. Smear pressure must be carefully balanced to avoid crush artifacts, particularly in seminoma. Bloody samples must be smeared quickly and are most often suboptimal.

Gross examination of the samples at smearing often gives valuable hints to the nature of the target lesion (author’s unpublished results). We strongly recommend the parallel use of DQ and Pap staining, but DQ takes preference over Pap stain. We do not use immunocytochemistry routinely in FNA of suspected testicular tumors. We believe that its use exceeds the basic aim of FNA in this field: a triage separating benign from malignant lesions. However, immunostaining for OCT 3/4, EMA, AE1/AE3, S-100, and appropriate lymphoid markers is useful to establish germ cell origin for a metastatic poorly differentiated neoplasm in a young man because of the potential for specific curative chemotherapy.⁶⁹ Immunostaining for PLAP has been used in an attempt to detect noninvasive testicular cancer in cryptorchid men.⁷⁰

The non-neoplastic testis (Figs 13.22 and 13.23)

Smears from normal testicular tissue contain cells which represent all stages of spermiogenesis, from spermatogonia to spermatozoa, in varying proportions. Spermatozoa and late spermatids are easily recognized by the nuclear size, shape and hyperchromasia, whereas the less mature forms resemble lymphoid cells of blastic type, particularly in air-dried DQ-stained smears (Fig. 13.22). Dark and pale spermatogonia can be recognized by their chromatin density, primary spermatocytes in pachytene; the latter are larger and have easily discernible thick, long chromatin threads frequently parallel to each other. Normal spermiogenesis must not be mistaken for lymphoma or TCGT. The presence of normal spermiogenesis excludes neoplasia, unless the target has been missed.

Fig. 13.22 Non-neoplastic testis

Range of spermatogenetic cells including a few mature sperms; note resemblance to lymphoid cells (DQ HP).

Fig. 13.23 Sertoli cells

Clustered cells with abundant cytoplasm, some vacuolation, and indistinct cell borders (DQ, HP).

Sertoli cells, single or in groups, are easily identified, particularly in smears from atrophic testicular tissue. Sertoli cells have pale, round nuclei, prominent nucleoli and abundant, relatively dense, vacuolated cytoplasm with indistinct borders. They often appear as naked nuclei (Fig. 13.23). Sertoli cells predominate in infancy, cryptorchidism, cirrhosis, infections and following hormonal blockage for prostate cancer. In ‘Sertoli cell only syndrome’ there is a complete absence of the germinal line; this condition cannot be distinguished from diffuse severe atrophy.

Leydig cells are difficult to recognize in FNA smears of normal testis regardless of the staining method. Nuclei are perfectly round and characteristically have one or two eccentric nucleoli. The chromatin pattern is denser than that of Sertoli cells, and the cytoplasm is better defined, dense eosinophilic and granular. Reinke’s crystals are difficult to spot except in Leydig cell tumors.⁷¹ Leydig cells predominate in aspirates from patients with Klinefelter’s syndrome. In the appropriate clinical setting, this diagnosis is suggested if smears show abundant Leydig cells and connective tissue strands and absence of cells of germ lineage.

Detailed accounts of the cytology of spermatogenesis as seen in FNA smears were published more than two decades ago.^72,73 FNA has achieved great popularity in India and Middle Eastern countries for the study of male infertility.^21-23,74-77 It has been used to locate areas of sperm production to guide sperm extraction procedures in men with non-obstructive azoospermia.^68,78 An interesting scheme for reporting FNA testicular biopsies has been proposed by Dajani.⁷⁵ See also Linsk & Franzen⁶⁵ and Zajicek.⁶⁶

In acute orchitis.^60,65,66 the testicle is swollen and hard, and FNA is painful. Aspirates usually have abundant entangled fibrin threads, leukocytes, histiocytes, debris and fragments of necrotic seminiferous tubules. Chromatin threads may be found (Fig. 13.24).

Fig. 13.24 Acute orchitis

(A) Two intersected seminiferous tubules filled with necrotic and inflammatory cells; (B) Dirty background, naked nuclei of Sertoli cells, degenerate germinal cells (upper-right corner), tailless spermatozoa and debris (A Pap, LP; B DQ, HP).

Embryonal carcinoma, mixed TGCT and seminoma must be ruled out before making a diagnosis of acute orchitis.

Chronic orchitis⁶⁶ is a rare condition. Aspirates are scanty due to fibrosis. Lymphocytes, plasma cells, histiocytes and sparse polymorphs may be present.

Granulomatous orchitis^60,65,66 (Fig. 13.25) is usually cryptogenic but may be tuberculous or due to other infections. Aspirates resemble chronic orchitis but also include aggregates of epithelioid histiocytes and multinucleated giant cells. Caseous necrosis is frequently seen in tuberculous orchitis.

Fig. 13.25 Granulomatous orchitis

An epithelioid granuloma (left). Notice a primary spermatocyte in the upper-right corner and reactive atypical nuclei at the lower-right corner. (DQ, HP)

Tuberculous orchitis is almost always associated with tuberculous epididymitis, but isolated tuberculous epididymitis does occur. Increased consistency of a mobile epididymis does not suggest tumor, but an enlarged epididymis fixed to the testis as occurs in tuberculous epididymitis can mimic a tumor. Tuberculous orchitis can also mimic testicular malignancy.⁷⁹ The correct diagnosis of tuberculous epididymitis and epididymo-orchitis may prevent unnecessary orchidectomy.^17,25,80

Chronic and granulomatous orchitis must be distinguished from seminoma. Eighty per cent of seminomas contain lymphocytic infiltrates and 50% contain epithelioid granulomas.⁶⁶ Seminoma may present with a clinical history suggestive of chronic orchitis.⁶⁶ Seminoma must be ruled out whenever smears contain abundant lymphocytes or epithelioid cells. In our experience, the differential diagnosis between granulomatous orchitis, seminoma and non-Hodgkin lymphoma is easier cytologically than in tissue sections. Spermatic granuloma also enters the differential diagnosis in granulomatous lesions.⁸¹

Epididymitis^60,66 FNA smears from acute and chronic epididymitis are dominated by inflammatory cells intermingled with sparse epididymal cells, single or forming monolayered sheets. Quite atypical-looking cells may be found, probably representing regenerative atypia (Fig. 13.26). Surgical exploration may be inevitable if the lesion cannot be clearly separated from the testis clinically or radiologically. Tuberculosis should always be considered in epididymal lesions with or without testicular enlargement (see above). FNA provides adequate material for both cytologic and microbiologic examination and avoids orchidectomy.²⁵ We amplify by PCR the highly conserved IS6110 genomic sequence for the identification of Mycobacterium tuberculosis species.

Fig. 13.26 Chronic epididymitis

(A) Cluster of atypical, somewhat degenerate epithelial cells and inflammatory cells (MGG, HP); (B) Corresponding tissue section (H&E, IP).

The role of FNA in the diagnosis of epididymal nodules has been discussed by Gupta et al.²⁰

Miscellaneous

Hydrocele^60,65 usually presents as a concentric swelling. The aspirate is pale or dark amber. A pale aspirate is sparsely cellular of just a few mesothelial cells and lymphocytes. A dark amber sample contains numerous cells including polymorphs, typical and reactive mesothelial cells singly or forming three-dimensional balls, histiocytes and debris (Fig. 13.27B) representing a hydrocele of long evolution. If numerous three-dimensional balls with spherical or scalloped borders are seen, a mesothelioma of the tunica vaginalis should be considered. Japko et al. gave the first description of the cytology of malignant mesothelioma of the tunica vaginalis in hydrocele fluid.⁸²

Fig. 13.27 Testicular/epididymal cysts

(A) Spermatocele. Numerous sperms, some nucleated cells, probably sperm precursor cells (Pap, IP). (B) Hydrocele. Clusters of vacuolated and pigmented macrophages (MGG, IP);

Hematocele^60,65 aspirates are hemorrhagic and bright or dark red in color. A bright aspirate contains preserved erythrocytes, a dark aspirate lysed erythrocytes and hemosiderophages. Hematoceles are generally secondary to problems of venous drainage, coagulation disorders, or to trauma. The possibility of underlying neoplasia must always be considered and the testis reexamined after evacuation of the fluid with repeat FNA in case of doubt.⁶⁶

Spermatocele is usually peripheral and may mimic a tumor. The aspirate is typically milky and the presence of numerous immobile spermatozoids, balls of spermatozoids, histiocytes, spermiophages and some monolayered sheets of epididymal cells allows a confident diagnosis (Fig. 13.27A). A spermatocele can often be completely evacuated by FNA, leaving no residual lesion. FNA is both diagnostic and therapeutic in hydroceles, hematoceles and spermatoceles.⁶⁰

Scrotal hernia presents as a scrotal tumor with negative transillumination. Aspirates contain clusters of typical or reactive mesothelial cells and fibroadipose tissue. Mucosecretory epithelium is usually not seen since the needle generally only penetrates as far as the pre-hernial lipoma. The possibility of teratoma must be considered if mucous and intestinal epithelium is aspirated.⁶⁰

Spontaneous spermatic cord torsion and testicular trauma⁶⁰ account for an increasing number of consultations. Cytological findings are very similar. FNA is completely painless and aspirates consist of blood in variable quantity and state of preservation, cell debris and ghost germinal cells. Fragments of necrotic seminiferous tubules may be seen (Fig. 13.28A,B).

Fig. 13.28 Acute torsion

(A) A twisted seminiferous tubule filled with debris. (Pap, LP); (B) A cast of a seminiferous tubule with ghost cells (DQ, IP).

Spermatic granuloma smears show inflammatory cells and tailless spermatozoids, epithelioid granulomas and macrophages engulfing spermatozoids. Caseous necrosis is not seen. Neoplasia or benign tumor may be suspected clinically. The differential diagnosis has been discussed above (Fig. 13.29).⁸¹

Fig. 13.29 Spermatic granuloma

A cluster of histiocytes with phagocytosed spermatozoa (Pap, HP).

Scrotal epidermal cysts are common, easily recognized on clinical examination. Smears show keratinized or non-keratinized squames, multinucleated foreign body giant cells, polymorphs and histiocytes.

Neoplasms^{14,58-61,63,65,66}

The great majority of malignant testicular tumors are GCT. The cytologic features of TGCT described here are based on our personal experience on FNA of primary testicular tumors.

Since TGCT can include more than one component not all necessarily sampled by FNA, a definitive type-specific diagnosis is generally deferred to histological examination. Familiarity with the cytology of TGCT permits the recognition of extragonadal GCT, mainly in the mediastinum and retroperitoneum. The diagnosis is of great clinical importance since treatment and prognosis are different from that of other malignancies occurring in these sites. The diagnosis may be supported by immunostaining.^51,69 Note that spermiogenesis is not seen in satisfactory FNA samples of TGCT.

Detailed accounts of the utility and limitations of FNA at extragonadal sites and of cytologic features of extragonadal GCT have been published.^51,52

Seminoma (Figs 13.30 and 13.31)^{14,50,57-63,65,66}

Fig. 13.30 Seminoma

(A) Dispersed cells with large pale nuclei and poorly defined cytoplasm; note ‘tigroid’ background, smudged nuclei and small lymphocytes (MGG, HP); (B) Dispersed cells; moderately pleomorphic vesicular nuclei; single or multiple prominent nucleoli; many scattered lymphocytes (Pap, HP).

Fig. 13.31 Mediastinal seminoma

Smear from large mediastinal mass clinically thought to be thymic carcinoma. Cluster of poorly cohesive, obviously malignant cells and some necrotic debris. Cytoplasmic vacuolation and nuclear fragility suggest malignant germ cell tumor; some lymphocytes present but specific typing not possible. Histology confirmed seminoma (MGG, HP).

Seminoma is a highly cellular neoplasm of poorly cohesive cells and little stroma. The presence of a lace-like tigroid background (TB), and the high cellularity including lymphocytes account for the intense navy-blue color of the smears. This may suggest the diagnosis even before microscopic study. Comments about the origin of TB and the distinction from ‘lymphoglandular bodies’ have been made elsewhere.⁶⁰ TB may go unnoticed in Pap-stained smears, but is more conspicuous in DQ smears (Fig. 13.30). A diagnosis of seminoma should not be based solely on the presence of TB or TB-like material. We have observed TB-like material also in aspirates of embryonal carcinoma. The contrary is also true: seminoma should not be ruled out simply because the pathologist is unable to identify TB.¹⁴ Large seminomas may show prominent necrosis. Necrosis is a major cause of false negatives in testicular FNA.⁵⁹ Tumor giant cells or syncytiotrophoblastic cells may be seen in aspirates of seminoma, but can also be found in embryonal carcinoma, mixed TGCT and in trophoblastic tumors.

The plasmatic membrane is well-defined in seminoma, but it is difficult to see in embryonal carcinoma. When lymphocytes, plasma cells, histiocytes and epithelioid granulomas are abundant and seminoma cells scarce, a diagnosis of granulomatous orchitis may be entertained: the absence of spermatozoids and of the germinal line suggest a seminoma.⁶⁰

Chromatin trailing may be prominent, particularly if the sample was smeared vigorously. This artifact has no diagnostic value in itself, since it may occur also in embryonal carcinoma, yolk sac tumors, lymphomas and acute orchitis. When the smear is dominated by this artifact, and there are no acute inflammatory cells and no spermatogenesis, the pathologist must search carefully for preserved diagnostic cells. In this situation, a tentative diagnosis of seminoma may be made.¹⁴

Lymphocytes, plasma cells and even ‘lymphoglandular bodies’ may be prominent in smears of seminoma and may lead to an erroneous diagnosis of lymphoma. Lymphoblasts may also be mistaken for seminoma cells.

Familiarity with the cytology of testicular seminoma helps in the recognition of extragonadal seminoma and metastatic seminomas (Fig. 13.31). Immunostaining may be of help in doubtful cases.⁶⁹

Seminomas with a greater degree of cellular pleomorphism, higher mitotic rate and scarce stromal lymphocytes have been called anaplastic seminoma or atypical seminoma.¹² These seminomas may be mistaken for embryonal carcinoma.⁶⁵

Spermatocytic seminoma (Fig. 13.32)^14,64

Fig. 13.32 Spermatocytic seminoma

Note evident nuclear size variation, spireme chromatin (half left side) and absence of tigroid substance (DQ, HP).

Saran et al.⁶⁴ have listed the cytomorphological features that distinguish spermatocytic seminoma from classical seminoma. The markers useful in other TGCT are generally negative.¹²

Embryonal carcinoma (Figs 13.33 and 13.34)^{14,58-61,63,65,66}

Fig. 13.33 Embryonal carcinoma

(A) Adenocarcinoma-like tissue fragment of large malignant cells with large vesicular nuclei, coarse chromatin, prominent nucleoli; prominent cytoplasmic vacuolation (MGG, HP); (B) Fragment of undifferentiated mesenchymal tissue right; cluster of malignant epithelial cells with large vesicular nuclei and prominent large nucleoli (H&E, HP) left.

Fig.13.34 Embryonal carcinoma

Tissue section corresponding to Fig. 13.33B (H&E, IP).

The vesicular nuclei of embryonal carcinoma are larger and more pleomorphic than those of seminoma, the chromatin is coarse and irregular and nucleoli are large, occasionally huge, and eosinophilic. The cytoplasm is pale and distinctly vacuolated, but not ‘bubbly’. It is not highly fragile and a TB as in seminoma is not seen. Cytoplasmic boundaries are poorly defined (Fig. 13.33). Hemorrhage and tumor necrosis may be prominent and may hamper the identification of diagnostic cells. It is usually not seen in smears of seminoma. Chromatin threads and reticulated material resembling TB can occur. Large syncytiotrophoblastic cells may be found. Prominent lymphoid/plasma cell infiltrates or mucus are not seen. The presence of cellular mesenchyme on its own does not warrant a designation of teratoma (Figs. 13.33B and 13.34).⁴⁹

Tumors of more than one histologic type (mixed forms) (Figs 13.35 and 13.36)^14,60,63,65

While the basic GCT types are infrequent in pure forms they are very frequent in mixed forms. Embryonal carcinoma and teratoma are each present in 47% of cases, and yolk sac tumors in 41%; 40% of TGCT contain varying numbers of syncytiotrophoblastic cells.¹²

Fig. 13.35 Tumor of more than one histological type (mixed form)

Large bisected tumor occupying most of the enlarged testicle in a male 19 years of age. Note hyalinized, degenerate and cystic areas.

Fig. 13.36 Tumor of more than one histological type (mixed form)

(A) Well-differentiated glandular epithelium (right), large malignant cells similar to embryonal carcinoma (left) (MGG, HP); (B) Corresponding tissue section (A, H&E, IP; B, PAP, IP).

The diagnosis of mixed forms is simple when malignant elements of an epithelial nature coexist with clear-cut teratomatous structures. Necrosis may obscure the neoplastic cells and the teratomatous component may be only minor or even absent. This can be explained by its greater cohesion causing under-representation in FNA samples. As a result, the differential diagnosis between mixed TGCT and embryonal carcinoma is sometimes difficult or impossible.^{14,59-61,65,66} The presence of multinucleated syncytial cells is not diagnostic of choriocarcinoma.

Yolk sac tumor (Figs 13.37 and 13.38)^{14,53-56,59-61,63}

Most cytologic reports are based on FNA performed on metastatic sites. A detailed account of the morphologic spectrum of endodermal sinus tumor has been reported by Akhtar et al.⁵³

Fig. 13.37 Endodermal sinus tumor

(A) Polypoid epithelial fragment of large malignant cells with pale, almost clear, vacuolated cytoplasm (MGG, HP); (B) Corresponding tissue section (H&E, IP).

Fig. 13.38 Endodermal sinus tumor

(A) A glomeruloid cell cluster with vacuolated cytoplasm on a mucoid background (DQ, IP); (B) Cell cluster with a vascular stalk and several hyaline globules.

Criteria for diagnosis

♦ Mucoid background,

♦ Glomeruloid structures,

♦ Schiller-Duval bodies (characteristic finding when present),

♦ Very immature cells, prominent nucleolus, basophilic cytoplasm,

♦ Small naked nuclei,

♦ Intra- or extracellular PAS-positive hyaline globules (AFP positive),

♦ Prominent cytoplasmic vacuoles and distinct cell boundaries confer a clear-cell appearance,

♦ Immunocytochemistry: focal staining for AFP, AE1/AE3 and PLAP; negative for OCT .

Pure yolk sac tumors are almost always encountered in infants and young children but rare in the first 6 months of life.⁴⁹ In adults, this pattern is seen in approximately 40% of nonseminomatous GCT;¹² however, it can easily be mistaken for embryonal carcinoma in FNA smears. This is why most reported cases are of metastatic tumors. Hyaline globules and eosinophilic membrane-like material are hints to the diagnosis of this tumor type (Fig. 13.38B). In infants and young children, the aforementioned cytological criteria permit a confident diagnosis by FNA.

Choriocarcinoma (Fig. 13.39)^59,63,66

These tumors most commonly present with symptoms referable to metastases¹² and most FNA reports are of extratesticular metastases. Immunostaining for βHCG is helpful to identify syncytiotrophoblastic cells. Large syncytiotrophoblastic cells may also be aspirated from classical seminomas, embryonal carcinomas and mixed TGCT.

Fig. 13.39 Choriocarcinoma

(A) Cluster of cytotrophoblastic and syncytiotrophoblastic cells (MGG, HP); (B) Corresponding tissue section (H&E, IP).

Teratoma (Fig. 13.40)^14,59,63,66

Fig. 13.40 Testicular teratoma

(A) Mature components (squamous and mucus secretory) (Pap, IP); (B) Primitive neuroectoderm; (C) Immature mesenchyma (fetal-like tissue) (B,C; DQ, IP).

It is difficult to specify diagnostic criteria for this entity because of the variability of histologic patterns.^14,58 In our experience, the main problem is that small foci of embryonal carcinoma may be missed. Organising testicular infarction may lead the unwary to an erroneous diagnosis of teratoma.¹⁴ If the needle samples a cystic component of a teratoma, acellular fluid may be obtained, resulting in a false-negative diagnosis. In such cases, the pathologist must reexamine the testis, and any persistent testicular mass must be rebiopsied.

The finding of palisading cylindrical cells and mucus should raise the possibility of a scrotal hernia and this diagnosis should be ruled out before making a diagnosis of mature teratoma. Readers are referred to the monograph by Zajicek⁶⁶ for descriptions of pure teratoma We reported the cytologic features in a case of epidermoid cyst.⁸³

Sex cord-gonadal stromal tumors

Leydig cell tumor (Fig. 13.41)^60,61,66,71

Fig. 13.41 Leydig cell tumor of testis

Isolated naked nuclei with a prominent nucleolus, mild anisokaryosis. (DQ, HP).

Criteria for diagnosis

♦ Highly cellular smears,

♦ Single cells on a background of granular eosinophilic cytoplasmic material,

♦ Round nuclei, frequently naked, showing some anisokaryosis and one to three nucleoli,

♦ Chromatin finely granular and evenly distributed,

♦ Intact cells have poorly defined borders, abundant eosinophilic, finely granular cytoplasm and eccentric nuclei,

♦ Cytoplasm may be spongy as a result of lipid accumulation,

♦ A spindle cell pattern may be observed,

♦ Nuclear pseudoinclusions and Reinke’s crystals (intra- and extracytoplasmic) may be seen,

♦ Staining for inhibin-alpha may be helpful, but this marker also stains other sex cord-stromal tumors. PLAP and OCT 3/4 are negative.

Numerous Leydig cells may also be obtained from testicles of patients with Klinefelter’s syndrome and from the so-called testicular tumor of the adrenal genital syndrome, but the clinical setting is different.

The finding of intracytoplasmic lipofuscin pigment and several intracytoplasmic as well as intranuclear Reinke’s crystals serve to clinch the diagnosis on FNA.⁷¹ Cytologic features do not permit the separation of benign from malignant Leydig cell tumors.

Hematopoietic tumors

The clinicopathological features of primary testicular and paratesticular lymphomas have been described by Al-Abbadi et al.⁸⁵

A malignant lymphoid infiltrate found in FNA of the testis raises three possibilities with different clinical implications: a terminal testicular leukemic or lymphomatous infiltration, the initial manifestation of a previously undiagnosed lymphoma, and a primary testicular lymphoma.

The distinction between testicular lymphoma, seminoma, embryonal carcinoma and granulomatous orchitis is easier in cytologic specimens than in tissue sections. Misinterpretation can occur, especially in cases presenting at an age similar to that for TGCT. Appropriate immunostaining may be of help.

FNA has been used for early detection of testicular involvement and relapse of acute lymphoblastic leukemia in children.^86,87 Granulocytic sarcoma may involve the testis; only rarely has this entity been diagnosed by FNA.⁸⁸

Secondary tumors of the testis

Rarely, metastatic carcinoma may simulate a primary testicular tumor. The origin of the cancer is most often in the lung or the prostate. We have seen a gastric signet ring cell carcinoma metastatic to the testis. A clinicopathological analysis of 26 cases has been recently published.⁸⁹

Tumors of paratesticular structures

Adenomatoid tumor ( Fig. 13.42)⁹⁰

Fig. 13.42 Adenomatoid tumor

Clusters of plump spindle cells resembling mesothelial cells; abundant well-defined cytoplasm; a few vacuoles; mild nuclear atypia (A, DQ; B, Pap, HP).

Criteria for diagnosis

♦ Pink, finely granular, or amorphous proteinaceous background,

♦ Moderate to abundant cellularity,

♦ Monolayered sheets, naked nuclei, well-defined multilayered clusters of small monotonous cells; vaguely glandular pattern occasionally,

♦ Abundant, poorly defined cytoplasm,

♦ Pink–clear paranuclear vacuoles,

♦ Intercellular windows visible but not prominent,

♦ Round to oval, eccentric nuclei; one small nucleolus; finely reticular, evenly distributed chromatin,

♦ Scanty stromal fragments.

The clinicocytologic presentation of adenomatoid tumors is unique and easily recognized by FNA cytology, which permits its differentiation from primary testicular neoplasms.⁹⁰ However, the diagnosis of adenomatoid tumor can be problematic in an atypical location as it may be mistaken for seminoma.²⁴

Mesothelioma (Fig. 13.43)

We have studied two cases displaying cytological features similar to those found in pleural effusions. The main differential diagnoses are longstanding hydrocele with reactive mesothelial cells forming numerous three-dimensional balls, and metastastic carcinoma. FNA findings in intrascrotal mesothelioma have been described in a few cases, some with a well-documented history of asbestos exposure.^82,91,92 The immunohistochemical profile of malignant mesotheliomas of the tunica vaginalis has been reported.⁹³

Fig. 13.43 Mesothelioma

Numerous malignant morules. Notice only one malignant cell population on a background of erythrocytes and inflammatory cells (Pap, IP).

Other tumors

Liposarcoma and leiomyosarcoma are the most common sarcomas of the scrotum in adults, paratesticular rhabdomyosarcoma in children.¹² A few case reports with FNA have been published.^19,67,94,95 The cytology of a congenital malignant rhabdoid tumor has been described.⁹⁶ Paratesticular sarcomas may mimic a primary testicular tumor clinically and ultrasonographically. FNA is helpful to rule out a TGCT.^19,67,94,96

The cytologic findings of a testicular papillary serous cystadenoma have recently been described.⁹⁷

A case of primary melanoma of scrotum in 55-yr-old man diagnosed by FNA has been reported.⁹⁸

Malignant tumors

The primary diagnosis of endometrial adenocarcinoma is made by other means, but FNB may be used to confirm tumor recurrence or metastasis (Fig. 13.49).¹²²

Fig. 13.49 Intermediate-grade endometrial adenocarcinoma

Poorly cohesive malignant epithelial cells; some microglandular groups (MGG, HP).

(Courtesy Dr. J. Klijanienko, Curie Institute, France.)

The cytological findings in uterine leiomyosarcoma are of a spindle cell sarcoma. Smooth muscle differentiation is suggested by a dense, eosinophilic cytoplasm and truncated nuclei, but a type-specific diagnosis should be supported by immunostaining (see Chapter 15). High cellularity, reduced cell cohesion, nuclear pleomorphism, chromatin abnormalities and the presence of necrosis suggest malignancy. Mitotic counts are unreliable in FNB samples.²² Leiomyosarcoma with a round cell epithelioid pattern can be mistaken for carcinoma unless immune marker studies are added (Fig. 13.50).

Fig. 13.50 Uterine leiomyosarcoma

(A) Epithelioid pattern; dispersed, pleomorphic malignant cells of predominantly rounded shape, pale fragile cytoplasm (MGG, HP); (B) Corresponding tissue section; note abnormal mitotic figure (H&E, IP).

FNA smears of endometrial stromal sarcoma are highly cellular.^123,124 The cells are plump spindled, eosinophilic with anastomosing cytoplasmic processes. In low-grade tumors (endolymphatic stromal myosis), the cells are relatively small and uniform with bland nuclei, and form cohesive clusters. In high-grade tumors, cells are mainly dispersed and there is no architectural pattern. Nuclei are oval, vesicular, enlarged and moderately pleomorphic, with granular chromatin and small nucleoli. Mitotic figures are frequently observed (Fig. 13.51). Positive staining for CD10 is helpful in the distinction from other spindle cell lesions.

Fig. 13.51 Endometrial stromal sarcoma

(A) Poorly cohesive plump spindle cells with pale ovoid nuclei and small nucleoli; note anastomosing strands of eosinophilic cytoplasm and a few mitoses (H&E, IP); (B) Corresponding tissue section (H&E, IP).

Coexistence of malignant glandular epithelial cells and malignant mesenchymal cells in smears from a uterine or ovarian tumor suggests a malignant mixed Mullerian tumor or carcinosarcoma. A few cases diagnosed by FNB have been reported.^125,126 Mesenchymal elements are not often demonstrated in metastatic deposits of this tumor.²⁷

Cystadenoma, serous and mucinous^104,105

Cyst fluid from benign serous cystadenoma of ovary is usually poor in cells, and any cells present are often degenerate or show shrinkage artifacts making a type-specific diagnosis difficult. Cell blocks are often helpful. Well-preserved cells form papillary aggregates, have small, dark, bland nuclei and may have vacuolated cytoplasm (Fig. 13.54). However, papillary tufts of mesothelial cells from the broad ligament closely resemble those of a serous cystadenoma, and may appear atypical if reactive. This possibility must be kept in mind when examining ovarian aspirates. Tall columnar cells with basal nuclei and empty-looking cytoplasm, and viscous mucus in the background suggest mucinous cystadenoma (Fig. 13.55).

Fig. 13.54 Benign serous ovarian cystadenoma

Intraoperative cyst fluid aspiration. Sheets and papillary clusters of small, round epithelial cells and small relatively uniform nuclei (Pap, HP).

Fig. 13.55 Benign mucinous ovarian cystadenoma

Sheets and strips of columnar epithelial cells with abundant pale vacuolated cytoplasm and uniform bland nuclei. Background mucus seen in other parts of smear (Pap, HP).

Serous cystadenocarcinoma (Fig. 13.58)

Fig. 13.58 Serous cystadenocarcinoma

(A) Papillary tissue fragment of crowded atypical glandular cells; large overlapping nuclei; prominent nucleoli; scanty pale cytoplasm (Pap, HP); (B) Cell block. Papillary tumor fragments, psammoma bodies( upper right) (H & E, HP).

The tumor cells generally have a moderate amount of cyanophilic (Pap) cytoplasm, which may be vacuolated but stains negative for mucin. A papillary microarchitectural pattern is not always obvious. Finger-like aggregates of cells with a well-defined border on three sides are more commonly seen than true papillary structures with stromal cores. The microarchitectural pattern is best shown in cell blocks (Fig. 13.58B). Psammoma bodies showing concentric lamellation are a helpful sign when present.

Mucinous cystadenocarcinoma (Fig. 13.59)

Fig. 13.59 Mucinous cystadenocarcinoma

Clusters of pleomorphic malignant glandular cells with a background of mucus (DQ, HP).

Smears are usually cell rich. However, the sample may consist only of mucinous secretion with few cells, mainly mucinophages or bland-looking, spindly epithelial cells similar to those of pseudomyxoma peritonei. The cells resemble those of mucinous cystadenoma but show a variable degree of nuclear enlargement and crowding, an increased N : C ratio and nuclear criteria of malignancy, more obvious in less well-differentiated cancers. The background is of thick, stringy mucus, which stains bluish-violet with MGG. Distinction from metastatic adenocarcinoma may be difficult.

Endometrioid adenocarcinoma (Fig. 13.60)

The cytological features of endometrioid carcinoma are not very specific.²³ The cells have an eosinophilic granular cytoplasm and the microarchitectural pattern is glandular, cribriform and partly solid. Mucin is absent or sparse. It may be difficult to distinguish from serous adenocarcinoma, but the main differential diagnosis is metastatic adenocarcinoma, particularly of colonic origin.

Fig. 13.60 Endometrioid carcinoma

Loose cluster of malignant glandular cells with an acinar pattern and some nuclear palisading (MGG, HP).

Immunocytochemistry is helpful in the differential diagnosis between primary ovarian carcinoma and metastatic adenocarcinoma. Primary ovarian carcinoma is usually CK7+, CK20− and CDX2−, whereas colonic cancer is CK7−, CK20+ and CDX2+.

Clear cell carcinoma (Fig. 13.61)

Clear cell carcinoma is an adenocarcinoma in which the cells have abundant, pale, finely granular and vacuolated cytoplasm containing glycogen. The cells form papillary clusters with globoid stromal cores of hyaline, basement membrane-like material (Fig. 13.61B).¹³³ These structures have been called ‘raspberry bodies’ and are of diagnostic significance in ascitic fluid. ‘Hobnail’ cells, as seen in histological sections, may be recognizable also in smears. Nuclear pleomorphism can be relatively mild. The smear background appears ‘tigroid’ and intracytoplasmic glycogen can be demonstrated by PAS-staining of air-dried smears.¹³⁴

Fig. 13.61 Clear cell carcinoma

Papillary clusters of cells with abundant, pale, finely vacuolated cytoplasm. Globular cores of hyaline stromal material (A, MGG, IP; B, H&E, HP)

(Courtesy Dr. J. Philips, Sydney, NSW.)

Anaplastic ovarian carcinoma

Anaplastic carcinoma has a very poor prognosis. FNB may allow preoperative radiotherapy for this category of tumors. Anaplastic carcinomas are predominantly solid and aspirates are usually highly cellular. Some evidence of glandular differentiation may be seen. Nuclei are large, pleomorphic and obviously malignant. Large intracytoplasmic, hyaline eosinophilic inclusions similar to those seen in hepatocellular carcinoma may be found.

Metastatic carcinoma to ovary

The possibility of metastatic origin must be considered in all poorly differentiated malignancies. Differential cytokeratin staining is of some help in this distinction. Smears from Krukenberg tumors (ovarian metastasis from gastrointestinal adenocarcinoma) contain signet ring cells, single or small clusters, with intracytoplasmic mucin vacuoles. Cells are usually few due to the abundant fibrous stroma. A CNB may be necessary to provide a confident diagnosis. Malignant melanoma not uncommonly metastasises to the ovary. Criteria for diagnosis are described elsewhere (see Chapter 14).

Granulosa cell tumor (Figs 13.62-13.65)^135,136

Fig. 13.62 Granulosa cell tumor

Clusters of cells with some microfollicular groupings; uniformly small round nuclei some with folds; scanty, poorly defined cytoplasm. The globular structures represent the contents of Call-Exner bodies (A, DQ, HP; B, H&E, HP).

Fig. 13.63 Granulosa cell tumor, diffuse pattern

(A) Loose cluster of cells with uniform, ovoid nuclei, some with grooves (Pap, HP); (B) Corresponding tissue section (H&E, IP).

Fig. 13.64 Malignant granulosa cell tumor

Clustered and dispersed cells; suggestion of follicular pattern; pleomorphic nuclei; coarse chromatin; prominent nucleoli; fragile cytoplasm (A, MGG; B, Pap, HP).

Fig. 13.65 Metastasis of granulosa cell tumor to lung

FNB of solitary 2-cm nodule upper lobe left lung in woman age 35 with past history of oophorectomy; cell-rich smear of small cells with round or ovoid uniform nuclei and indistinct cytoplasm; some microfollicular groups (A, DQ, HP; B, immunostaining for inhibin, IP).

Fine needle aspiration smears from granulosa cell tumors of follicular type are highly cellular of both single cells and syncytial aggregates. A follicular pattern is often discernible. In some cases, rounded bodies, 30–100 microns in diameter, of non-cellular material, are present. These probably represent the contents of Call-Exner bodies. They are brightly purple, fibrillar in MGG-stained smears (Fig. 13.62), pale eosinophilic and structureless in H&E. Nuclei are rounded and do not vary much in size. Nuclear grooves (‘coffee bean nuclei’) are frequent, best seen in alcohol-fixed Pap smears. Nuclear chromatin is finely granular and evenly distributed. Nucleoli are small and indistinct. The cells have pale vacuolated cytoplasm with indistinct cell borders. Smears from tumors of the diffuse type are less characteristic. Plump spindle cells predominate, nuclear grooves are present, but there is no characteristic microarchitectural pattern (Fig. 13.63). Immunostaining is helpful: tumor cells are positive for inhibin and vimentin, negative for cytokeratin.

The FNA findings were similar in two examples of malignant granulosa cell tumor but smears were even more cellular and there was marked nuclear pleomorphism. Nuclear chromatin was irregular and nucleoli were prominent (Fig. 13.64). FNA confirmation of granulosa cell tumor supported by markers is possible also in distant metastatic tumor deposits (Fig. 13.65).

Other gonadal stromal tumors of the ovary are rare and there are few reports of FNB diagnosis.^137,138