Case 1. Peritumoral GCNIS

Clinical case. A 31-year-old patient presented with a right testicular mass evolving for the last 3 months. After orchiectomy, it was diagnosed as seminoma. The figures correspond to the peritumoral parenchyma.

Pathological findings. Peritumoral parenchyma shows an intratubular germinal neoplasm, mixed atrophy (seminiferous tubes with spermatogenesis next to others with Sertoli cell-only), Sertoli cell nodules, and infiltration of the rete testis. The tubules with GCNIS are characterized by their smaller diameter, absence or marked decrease of their lumina, thickening of the wall, and presence, most of the time, of only two types of cells, the tumor cells similar to gonocytes and Sertoli cells. The GCNIS cells are large cells located over the basement membrane. The Sertoli cells show small-sized spherical nuclei and appear displaced towards the lumen. Sertoli cell nodules are formed by conglomerates of seminiferous tubules that usually contain only Sertoli cells but in this case they are infiltrated by GCNIS cells. The rete testis also shows a pagetoid infiltration by GCNIS cells. Among the most common immunohistochemical techniques to confirm, the presence of GCNIS are: placental alkaline phosphatase (PLAP), OCT3/4, and D2-40.

Comments. Identification of GCNIS cells in the adult is easy due to the nuclear characteristics of the cells and their disposition over the basement membrane. In case of doubt, most of the times due to poor fixation of the surgical specimen, immunohistochemical techniques are very useful. The tumor cells express the same markers as embryonic germ cells and seminoma cells. GCNIS is the neoplasm from which seminomas, embryonal carcinoma, teratomas postpubertal-type, yolk sac tumor postpubertal-type, and choriocarcinoma originate. It is present in the adjacent non-tumoral parenchyma of 90% of seminomas and in almost all non-seminomatous tumors. The findings observed in the peritumoral parenchyma in this patient (GCNIS, Sertoli cell nodules, mixed atrophy) suggest, as is common in many testes with germ cell tumors, that the testicle carries a primary anomaly, triggered during fetal life, which has altered the estrogen/androgen ratio, a situation known as testicular dysgenesis.

Case 2. GCNIS and Infertility

Clinical case. A 36-year-old patient with oligozoospermia. His testicles were of normal size. Serum determinations of FSH, LH, and testosterone were normal. There was no history of cryptorchidism. The testicular ultrasound examination showed a slight decrease of the testicular size and normal echogenicity. A bilateral testicular biopsy was performed.

Pathological findings. The right testicle showed mixed atrophy. The images correspond to the left testicle. The sample included part of four lobules. The tubules of one lobule show slight ectasis and complete spermatogenesis. Another lobule is formed by smaller tubules showing GCNIS cells which are easily identifiable by their cytological characteristics. This finding is also supported immunohistochemically by nuclear immunostaining for OCT3/4 and plasma membrane reactivity for PLAP. The remaining two lobules have complete spermatogenesis, but in the same tubule there is a large number of GCNIS cells in the basal compartment next to spermatogonia. These cells are also observed in the seminiferous tubules filled with sperm. Some tubules contain megalospermatocytes.

Comments. Four important lessons are drawn from this case: First, the testicular biopsy can offer us a surprise. Do not forget that in 0.6–0.8% of biopsies performed due to infertility there is a GCNIS. Second, if the fixation is not correct, the cytological characteristics of the tumor cells are distorted and the diagnosis is difficult, even more if tubules with tumor cells and spermatogenesis that may have a normal size predominate. Third, GCNIS is initially focal, adopts a lobular distribution, but later on it adopts a pagetoid growth pattern and can be extended to 100% of the seminiferous tubules. Finally, fourth, do not confuse GCNIS cells either with hypertrophic spermatogonia or with megalospermatocytes.

Case 3. Seminoma

Clinical case. A 41-year-old patient who consulted for a non-painful enlargement of the left testicle for more than 6 months. Tumor markers were negative. The tumor extension study was also negative.

Pathological findings. Surgical specimen that corresponds to a radical orchiectomy. The testicle measures 6.9 × 5.6 × 4 cm. The section shows a tumor that occupies 4/5 of the testicle. It is lobed, bulging from the surrounding parenchyma, grayish with areas of necrosis. Histologically, it is constituted by a proliferation of cells in sheets interrupted by thin septa with lymphocytes. The tumor cells are characterized for having a broad, clear, or slightly eosinophilic cytoplasm, and crisp plasma membranes. The nucleus is central and shows one or two nucleoli. Mitoses are very abundant. In the septa, the presence of groups of macrophages and sarcoid granulomas is frequent. GCNIS and tubules with preserved spermatogenesis are observed in the peritumoral parenchyma. The epididymis and the spermatic cord show no alterations.

Comments. The macroscopic aspect of a lobed mass that protrudes from the parenchyma at the sectioned surface is suggestive of seminoma. Conventional histology, both in regard to tumor cells and to the presence of fine septa with lymphocytes and granulomas, is characteristic. The ultrastructure shows that the cell population is not homogeneous. The differences are not in the nuclei that resemble those of GCNIS (irregular nuclear membrane, several large nucleoli) but in the cytoplasm that varies from full of glycogen to complete absence of it. In the cytoplasm, the presence of annular lamellae stands out, a fact that can occur in other tumors, but that in seminoma is a constant. In the past, the high number of mitoses was considered typical of the so-called “anaplastic seminoma,” an obsolete term without prognostic interest. Seminomas represent 50% of all germ cell tumors of the testicle, and the age of presentation is 10 years older than that of the rest of germ cell tumors.

Case 4. Seminoma with Intense Lymphoid Infiltrates and Granulomatous Reaction

Clinical case. A 40-year-old patient with a several-month-old non-painful enlargement of his right testicle. Tumor markers were negative.

Pathological findings. The surgical specimen comprises testicle, epididymis, and 9 cm of spermatic cord. The testicle measures 8 × 6.5 × 5 cm. The sectioned surface shows a tumor that comprises most of the parenchyma. It is a lobed grayish tumor with areas of necrosis and hemorrhage. Histologically, it is constituted by a proliferation of cells with clear cytoplasm that are arranged in sheets or cords. The interstitium shows a dense lymphoid infiltrate that forms lymphoid follicles. In other fields, the infiltrate is disposed between the cords of tumor cells until erasing all tumor structures. Several tumor nodules show a granulomatous infiltrate with plenty uni- or multinucleated histiocytes and areas of intense fibrosis with microliths. The epididymis and the spermatic cord show no alterations. The peritumoral parenchyma also shows marked tubular atrophy with abundant lymphoid infiltrates.

Comments. The intensity of the lymphoid infiltrates is very variable from one seminoma to another, and even in different areas within the same seminoma. The most abundant lymphoid cells are the T lymphocytes but it is not uncommon to observe other cells such as plasma cells and eosinophils. The largest accumulations are observed around the vessels. The presence of important infiltrates with lymphoid follicles as in this case is rare. Another peculiarity of this case is the intense granulomatous reaction composed by histiocytes that are arranged in clusters with or without giant cells, which can make observation of the tumor cells difficult. The marked fibrosis with hyalinization that separates the cords of the tumor cells is also noteworthy. In tumors in which this pattern predominates, immunohistochemistry may be necessary to rule out a granulomatous orchitis. The presence of microliths, both in the tumor and in the peritumoral atrophic testicular parenchyma, can also be the expression of an underlying testicular dysgenesis.

Case 5. Seminoma with a Tubular Pattern

Clinical case. A 36-year-old man who sought consultation for an increase in the size of his right testicle. Testicular tumor markers were negative. Radical orchiectomy was performed.

Pathological findings. The surgical specimen weighs 42 g. The testicle measures 4.5 × 3.5 × 2.5 cm. A well-defined tumor takes two-thirds of the testicular parenchyma, with a uniform and bright surface, without areas of necrosis or hemorrhage. The tumor presents a tubular pattern consistent in lace with focal pseudoglandular changes. In the periphery, a cord-like growth predominates. The cells, with a spherical nucleus, a voluminous nucleolus, and clear cytoplasm, were supported by very thin septa. The tumor lacks lymphocytic infiltrates or a granulomatous reaction. There are both pagetoid growth and eosinophilic cytoplasmic inclusions in the epithelial cells at the level of the rete testis.

Comments. Tubular pattern is a rare growth pattern in seminomas. It can raise a differential diagnosis with Sertoli cell tumors, and in some cases with other tumors such as carcinoid tumor and yolk sac tumor. It is noteworthy that in Sertoli cell tumors the septa separating the cellular cords are highly hyalinized, that tubular formations usually show lumina of rigid contours, and that they show no pagetoid infiltration of the rete testis or GCNIS in the neighboring parenchyma. In case of doubt, immunohistochemistry confirms the diagnosis (immunoreaction for PLAP, 0CT3/4 support the diagnosis of seminoma; immunoexpression for cytokeratins, inhibin and calretinin that of Sertoli cell tumor; neuroendocrine markers such as synaptophysin, chromogranin, and CD56 support the diagnosis of carcinoid tumor; and finally, AFP and glypican-3 are positive in yolk sac tumor).

Case 6. Seminoma with an Intertubular Growth Pattern

Clinical case. A 34-year-old patient who presented with infertility (severe oligozoospermia). His left testicle was slightly enlarged. The ultrasonography showed a 1-cm hypoechoic tumor in the lower pole. Tumor markers only showed a slightly elevated beta-HCG.

Pathological findings. The tumor, located in the lower pole of the testicle, is well delimited and corresponds to a seminoma with syncytiotrophoblast cells. GCNIS is observed in the tubules of several testicular lobules. In the parenchyma distant from the tumor, which presents a mixed atrophy and diffuse hyperplasia of Leydig cells, an infiltration in rows or cords of tumor cells stands out. These cells are introduced among peritubular myoid cells both in the tubules with spermatogenesis and in Sertoli cell-only tubules and also infiltrate the rete testis.

Comments. Seminomas usually destroy the seminiferous tubules and only in a third of the cases in the periphery of a tumor nodule a localized intertubular growth is observed. Ulbright in 1999 reported 12 cases of seminoma that showed this type of growth in the absence of a discernable clinical mass.

The intertubular seminoma has been defined (Browne 2005) as the presence of intertubular growth not contiguous to the main tumor and presents at least three high-power fields away from the tumor mass. The tumor frequently infiltrates the rete testis and its behavior is more aggressive. It can raise a differential diagnosis with the intertubular growth of some metastases of prostate adenocarcinoma.

Case 7. Plasmacytoid Seminoma with Syncytiotrophoblastic Cells

Clinical case. A 36-year-old patient presented with a painless left testicle enlargement for the last 6 months. The tumor markers were: AFP negative, beta-hCG 129 mUI/L.

Pathological findings. The testis measures 8.2 × 7.1 × 6.8 cm. It shows a tumor that only respected half of the testicular parenchyma at the lower pole. When sectioned, the tumor shows a grayish encephaloid surface with small hemorrhagic areas and necrosis. A mild hydrocele is also present. Histologically, a sheet growth pattern is observed of cells with plasmocytoid appearance (broad eosinophilic cytoplasm, eccentric nucleus). There are small clusters of perivascular lymphocytes. In the vicinity of the vessels, there are isolated syncytial cells. The tumor does not infiltrate testicular mediastinum, epididymis, or spermatic cord. Plasmocytoid-like cells are PLAP positive while syncytial cells show immunoexpression for beta-hCG.

Comments. The plasmacytoid aspect of some seminomas has been related to the proximity of their cells to necrotic areas. But there are tumors in which the cells are clearly separated so we should consider that this is one of the most varied patterns a seminoma can show. The differential diagnosis is basically posed with primary or secondary testicular plasmacytomas, metastasis of plasmacytoid carcinomas, and plasmacytoid melanoma. Immunohistochemistry may be essential. Of note in this case is the presence of syncytial cells distributed regularly within the tumor. This finding is reported in 10–20% of seminomas. The prognosis of plasmacytoid seminoma is similar to that of the other seminoma variants.

Case 8. Seminoma with a Microcystic Pattern, Signet-Ring Cells, and Early Embryonal Carcinoma

Clínical case. A 42-year-old infertile male with a clinical history of right cryptorchidism descended at 10 years of age and later removal of testicular remnants was referred to the Urology department. He also had a medical history of gonorrhea, condylomata acuminata, and molluscum contagiosum. He sought consultation for slow and progressive increase of his left hemiscrotum since 2–3 months ago. No other symptoms were observed. Testicular serum markers were negative.

Pathological findings. The radical orchioepididectomy specimen weighed 61 g. The testicle measured 7 × 5 × 5 cm. A tumor was observed upon sectioning that was taking the greater part of the testicle; it had a whitish nodular surface with central hemorrhagic areas. The epididymis, the testicular covers, and the spermatic cord were free of infiltration. Histologically, the tumor comprises two types of cells. The first cell type form most of the neoplasm. It consists of a proliferation in sheets of medium-sized cells separated by thin septa of connective tissue with few lymphocytes. The nuclei of these tumor cells vary from spherical to triangular; a large nucleolus stands out in the spherical ones, and a marked hyperchromatism is noticeable in the triangular cells. The cytoplasm, in general, is eosinophilic. Focally, groups of tumor cells show signet-ring morphology. The tumor adopts a microcystic pattern, mainly in these areas. The second cell type is arranged around the vessels. These cells have higher pleomorphism, irregular karyotheca with bizarre nucleoli and heterochromatin granules. The immunohistochemical study shows in the first cell type positive expression for PLAP and in the second cell type positivity for cytokeratins and CD30.

Comments. Most of the tumor is a plasmacytoid seminoma with signet-ring cells. Immunostaining for PLAP and absence of cytokeratins staining confirm this diagnosis. The second component is constituted by perivascular islets of cells that express cytokeratins and CD30, which is identified as a differentiation from embryonal carcinoma. It is a very uncommon case. On the one hand, it is a seminoma in which microcystic, plasmocytoid, and ring-ring cell patterns are combined. On the other hand, an embryonal carcinoma is developing in the form of perivascular nests. The role of immunohistochemistry is essential for the correct identification of these cases.

Case 9. Seminoma with Syncytiotrophoblastic Cells

Clinical case. A 35-year-old patient complaining of a progressive increase of his right testicle over the last 4 months. The testicular markers were: AFP normal, beta-hCG 210 mUI/mL. Radical orchiectomy was performed.

Pathological findings. The testicle averages 6.3 × 5.9 × 5 cm. The epididymis was highly thickened with a maximum thickness of 3 cm at the level of the body and tail. The spermatic cord showed no alterations. When sectioned, the testicle shows a whitish tumor that takes its central part in continuity with the epididymal tumor. Histologically, the pattern of tumor growth is in sheets. Tumor cells have spherical nuclei, central nucleoli, and pale cytoplasm with crisp cytoplasmic membranes. There are multinucleated cells of eosinophilic cytoplasm among them. There are lymphocytes and sarcoid granulomas in the septa that score the tumor. Surrounding the tumor there are several groups of seminiferous tubules filled with tumor cells. Most of these tumor cells, both with growth in sheets or with intratubular growth, are positive for PLAP immunostaining. Multinucleated giant cells observed among the tumor cells are beta-HCG positive.

Comments. The presence of syncytiotrophoblast cells is relatively frequent in all tumors derived from GCNIS. Its frequency is estimated in seminomas at about 10 and 20%. What is very variable is the number of cells and their distribution within the tumor. In this tumor, they are very abundant in some fields. Syncytiotrophoblast cells are easily differentiated from other multinucleated giant cells that are often present in these tumors. They are located between the cells of the seminoma, the nuclei are frequently arranged in mulberry-like clusters, and vacuoles can be observed in their cytoplasm. Frequently, the syncytiotrophoblast cells are associated with small hemorrhages. The second type of giant cells that can be seen in the seminoma are Langhans-type cells. These cells are usually located in the septa and are accompanied by other inflammatory cells forming granulomas. Syncytiotrophoblast cells are responsible for the presence of high beta-HCG levels in these patients so their serum determination can be useful for monitoring them.

Case 10. Embryonal Carcinoma with Syncytiotrophoblastic Cells

Clinical case. A 27-year-old patient with a rapidly growing painless tumor in the left testicle. Tumor markers: only beta-hCG was slightly elevated, the rest were normal. Retroperitoneal metastases were observed in the extension study.

Pathological findings. The surgical specimen of radical orchiectomy consisted of a testicle measuring 5.8 × 4.5 × 4.2 cm. The section showed a well-defined reddish central mass with hemorrhagic areas. The epididymis and the spermatic cord were normal. Histologically, the tumor is separated from the parenchyma by connective tissue with a dense lymphoid infiltrate. Tumor cells adopt solid growth in some areas, and papillary and microcystic patterns in others. Tumor cells show significant anisocytosis and anisocariosis, and frequent mitosis. They infiltrate the testicular mediastinum and permeate the lymphatic vessels. Among these cells, multinucleated giant cells with large cytoplasmic vacuoles are occasionally observed. The tumor cells are positive for CD30 and OCT3/4 and negative for glypican-3 and AFP. In the peritumoral parenchyma, there is GCNIS and mixed atrophy.

Comments. This case can be considered the prototype of an embryonal carcinoma with solid growth in some areas, papillary growth in other areas and microcystic or glandular growth still in other areas. The presence of syncytiotrophoblast cells is also frequent (60% of cases). The presence of intratubular necrosis and the extensive infiltration of the lymphatic vessels that justifies, as in this case, the great tendency to metastasize, are also characteristic. The tumor in this patient does not pose the usual differential diagnoses of embryonal carcinoma such as seminoma and yolk sac tumor. The ultrastructure and immunohistochemistry support the diagnosis. Pure embryonal carcinoma is a rare tumor (less than 3% of all germ cell tumors). Embryonal carcinomas do not show AFP rise. If this were found, the identification of a yolk sac tumor should put us in the alert. In these cases, it would be a mixed tumor.

Case 11. Embryonal Carcinoma with Lymphoid Stroma

Clinical case. A 18-year-old patient who presented with progressive painless increase of the left testicle in the last 3 months. Ultrasonography showed that the left testicle was taken by a 5 × 3 × 3 cm mass with hypoechoic areas alternating with hyperechoic areas that focally encompasses the epididymis. The tumor has left the lower pole free. Testicular tumor markers were negative.

Pathological findings. The testicle size is 5 × 4 × 4 cm. It shows a 3.2-cm tumor located in the upper pole and extending to the epididymis. The tumor is whitish-gray with hemorrhagic areas. Histologically, the cells are arranged in nests separated by thick septa that show dense lymphoid infiltrates. The tumor nests are made up of cells with great nuclear pleomorphism, spherical nuclei next to other triangular or folded with different sizes. The cytoplasm is generally eosinophilic but there are areas where it is clear or vacuolated. The lymphoid infiltrate shows no sarcoid granulomas and in some fields it is so dense that the tumor cells are barely distinguishable. The cells are keratin positive and express CD30. Placental alkaline phosphatase is positive in isolated cells. GCNIS is observed in most of the tubules in the rest of the parenchyma, as well as tubular hyalinization in the remaining tubules.

Comments. Histologically, the first diagnostic impression was that of a seminoma because of its frequency and because of the abundant lymphoid stroma, and the focal presence of clear cytoplasm cells. The study at higher magnification makes it possible to specify, both in the cells of the eosinophilic cytoplasm, and in those of the clear cytoplasm, the enormous pleomorphism of the nuclei with the presence of important notches and some large cells compared to the relative good sphericity of the nuclei of seminoma cells. The immunoexpression for CD30 and cytokeratins AE1/AE3 facilitates the diagnosis. Pure embryonal carcinoma only accounts for 2–3% of germ cell tumors and only 7% of embryonal carcinomas show a dense lymphoid infiltrate. In view of these data, it is not surprising that the diagnosis was fast.

Case 12. Embryonal Carcinoma with Blastocyst-Like Pattern

Clinical case. A 36-year-old patient with painless enlargement of the left testicle since the last 7 months. Tumor markers (AFP and beta-hCG) were negative.

Pathological findings. The testicle size is 9 × 8 × 8 cm. The sectioned surface was mottled with whitish nodular areas, hemorrhagic areas, and necrotic areas. A small peripheral crescent of testicular parenchyma was recognized. Histologically, it was a mixed germ cell tumor with teratoma of the postpubertal-type, seminoma and embryonal carcinoma in similar proportions. The images have been selected from the embryonal carcinoma component only. The embryonal carcinoma showed solid, papillary and cystic areas, the latter predominating. Inside the cysts, originating in the epithelium that covers them, there are multiple vesicular formations. The lining of the vesicles is flat with vacuolated cytoplasm cells except at a pole where the cells are cubic or cylindrical. Different vesicles resembling blastocysts are anastomosed through this area. All cells are positive for CD30 and cytokeratins. The cells express D2-40 on the outer surface of the vesicles.

Comments. Less than 1% of embryonal carcinomas, whether presented as pure forms or as part of a combined tumor, show this peculiar type of pattern. The knowledge of the existence of this pattern within embryonal carcinomas is important. In particular, it must be distinguished from a cystic pattern of yolk sac tumor. The absence of yolk sac markers such as AFP and glypican-3 support the diagnosis.