Imunohistoquimica no diagnostico do disgerminoma e tumor do seio endodermico do ovario: expressao de citoceratina, alfa-fetoproteina e alfa-antitripsina / Immunohistochemistry in the diagnosis of dysgerminoma and endodermal sinus tumor of ovary: cytokeratin, alpha-fetoprotein and alpha-antitrypsin antibodies expression

Os autores estudaram sete casos de disgerminoma e tres de tumor do seio endodermico do ovario, no intuito de avaliar a utilidade da imunohistoquimica, empregando anticorpos anti-citoceratina, alfa-fetoproteina e alfa-1-antitripsina no diagnostico destes tumores. Os 10 tumores expressaram alfa-1-antitripsina. Nenhum disgerminoma expressou citoceratina enquanto todos os tumores do seio endodermico mostraram reacoes positivas. Os tres tumores do seio endodermico e um disgerminoma expressaram alfa-fetoproteina. Apos investigacao detalhada, este caso foi classificado como um tumor misto de celulas germinativas, tambem por corresponder a um mau prognostico e curta sobrevida, nao muito caracteristico de disgerminomas puros. Este estudo indica que a imunohistoquimica pode ser util no diagnostico destes tipos de tumor, principalmente quando ha areas suspeitas que podem levar a um diagnostico de um tumor misto de celulas germinativas.

Changes in glycolipid expression in human testicular tumor.

Glycolipids were extracted from testicular tumor tissues of 13 patients, and their pattern of expression compared with that of normal testicular tissue. The most conspicuous and consistent change in the tumor extracts was marked accumulation of CTH (ceramide trihexoside). Structural analysis by enzyme cleavage showed that CTH which accumulated in the tumor tissue was Gb3 (Gal alpha 1-4Gal beta 1-4Glc beta 1-Cer). Immunohistochemistry using anti-Gb3 monoclonal antibody (MAb) (1A4) also indicated massive accumulation of Gb3 in the tumor tissue. Gb3 may be a new marker of testicular tumors, especially seminomas, for which useful markers are so far lacking.

The abnormal occurrence and the differentiation-dependent distribution of N-acetyl and N-glycolyl species of the ganglioside GM2 in human germ cell tumors. A study with specific monoclonal antibodies.

Human primary germ cell tumors were analyzed for the presence of the ganglioside GM2 using three specific monoclonal antibodies which can distinguish the molecular species of the sialic acid moiety: the antibody MK1-16 is specific for N-acetyl GM2, MK2-34 is specific for N-glycolyl GM2, and MK1-17 detects both N-acetyl and N-glycolyl GM2. When the occurrence of the GM2 antigen was tested in 107 cases of human germ cell tumors by the immunohistochemical technique using these antibodies, seminoma was characterized as having the highest frequency of N-acetyl GM2 (89.4%, 42 of 47 cases) among germ cell tumors, followed by embryonal carcinoma (40.0%), and teratocarcinoma (26.6%). Compared with this, yolk sac tumors and choriocarcinoma had a much lower positive incidence of the N-acetyl GM2 antigen. On the other hand, the N-glycolyl GM2 antigen was not found at all in 47 cases of seminoma (0%), and the positive incidence was very low in embryonal carcinoma (6.6%), although considerably higher incidences were obtained with choriocarcinoma (25.0%), yolk sac tumor (22.2%), and teratocarcinoma (13.3%). The presence and molecular species of the GM2 antigens in these human germ cell tumors were also ascertained chemically by the thin-layer chromatography (TLC) immunostaining of the ganglioside fractions prepared from primary germ cell tumors. These results indicate that seminoma specifically contains N-acetyl GM2 and no N-glycolyl GM2, suggesting that N-acetyl GM2 could be a good marker for seminoma. On the other hand, non-seminomatous germ cell tumors were characterized by the presence of N-glycolyl GM2, one of the Hanganutziu-Deicher antigens (H-D antigens). Moreover, the positive occurrence of N-glycolyl GM2 correlated very well with the degree of differentiation of non-seminomatous germ cell tumors, i.e., the differentiated tumors such as yolk sac tumors, choriocarcinoma, and teratocarcinoma had a higher positive incidence of N-glycolyl GM2 type H-D antigen but a lower positive incidence of N-acetyl GM2 when compared with embryonal carcinoma, the most undifferentiated tumors among non-seminomatous germ cell tumors.

Keratin 7 is a marker for a subset of trophoblastic cells in human germ cell tumors.

Human testicular germ cell tumors were studied immunohistochemically with the monoclonal antibody to the 54-kd keratin polypeptide (keratin 7) to determine whether this antibody could be used selectively to identify trophoblastic cells. The antibody reacted with the intermediate filaments in the cytoplasm of some cells in choriocarcinoma cell lines, and in trophoblastic cells in mixed germ cell tumors and a seminoma. It did not react with classic seminoma cells, embryonal carcinoma, yolk sac carcinoma, or somatic tissues of mixed germ cell tumors. On the basis of these data we conclude that monoclonal antibody to keratin 7 is a marker for a subset of trophoblastic cells in human germ cell tumors.

[Mixed ovarian germinal neoplasia: adult embryonal carcinoma and dysgerminoma with trophoblasts]. / Neoplasia germinale ovarica complessa: carcinoma embrionale adulto e disgerminoma con trofoblasto.

Malignant mixed germ cell tumor of the ovary: embryonal carcinoma and dysgerminoma with trophoblastic cells. A case of ovarian neoplasia in a 19-years-old woman is described showing mixed germ cell tumor patterns. The main component is a solid embryonal carcinoma with mainly syncytial-like, highly anaplastic cells, displaying diffuse CK-immunoreactivities and scattered PLAP-positive cells. Many CK- and beta-HCG-positive syncytiotrophoblastic and intermediate trophoblastic cells are present. A second component is a dysgerminoma with lymphoid stroma and diffuse PLAP-cytomembrane immunoreactivities: rare cells, to be identified as intermediate trophoblast cells, are CK- and strongly beta-HCG-positive. Many syncytiotrophoblastic cells with a brisk CK- and beta-HCG-positivities are also noted. The embryonal carcinoma component is metastasized to the controlateral ovary, uterus and omentum. A complete immunohistochemical analysis is recommended to properly diagnose germ cell neoplasias of the ovary both for descriptive and prognostic-therapeutic purposes. The very rare presence in the same ovarian tumor of mixed patterns as adult embryonal carcinoma and dysgerminoma with trophoblastic cells, is stressed.

[Clinical study of testicular tumor: tumor tissue content of hCG in seminoma].

Positive cases of serum human chorionic gonadotropin (hCG) have been increased in patients with pure seminoma due to the progress of methods of measuring serum hCG. But the positive ratio and the value of serum hCG in pure seminoma cases are lower than those of non-seminoma cases. The relationship between the pathological type of testicular tumor and the hCG levels in both serum and tumor tissue was investigated, and clinical usefulness of the serum hCG levels as a tumor marker of pure seminoma was discussed. The materials were 19 pure seminomas, 14 non-seminomas and 11 normal testes as controls. HCG and hCG-beta in tumor tissue extracts and in the serum of blood from the spermatic and peripheral veins were measured by radioimmunoassay (RIA). The tissue content of hCG and hCG-beta were 276.7 +/- 136.1 mIU/g.tissue and 16.5 +/- 3.20 ng/g.tissue, respectively, in pure seminoma; 224,376 +/- 91,619 mIU/g.tissue and 4,608 +/- 1,817 ng/g.tissue, respectively, in non-seminoma including choriocarcinoma component; 1,807 +/- 1,428 mIU/g.tissue, and 37.0 +/- 11.2 ng/g.tissue, respectively, in non-seminoma including seminoma component but not including choriocarcinoma component; 20.4 +/- 3.1 mIU/g.tissue and 1.27 +/- 0.38 ng/g.tissue, respectively, in normal testes, (each M +/- SE). The tissue content of hCG-beta in pure seminoma is significantly higher than that in normal testis (p less than 0.01) and significantly lower than that in non-seminoma including choriocarcinoma component.(ABSTRACT TRUNCATED AT 250 WORDS)

[Testicular germinal tumors. Current therapeutic principles]. / Tumeurs germinales testiculaires. Principes thérapeutiques actuels.

The cure rate for germinal tumors of the testes are now very high, greater than 80%, all stages and histological types taken as a whole. This calls for a revision of therapeutic indications, in light of these results and modern staging methods. In localized stages, or tumors associated with a good prognosis, therapeutic de-escalation is in progress, in order to limit the side effects. In invasive forms or forms associated with a poor prognosis, the use of more aggressive methods, such as high dose chemotherapy, should further improve prognosis.

Alpha smooth muscle actin (alpha-SM actin) in normal human ovaries, in ovarian stromal hyperplasia and in ovarian neoplasms.

An immunohistochemical investigation of alpha-smooth muscle actin (alpha-SM actin) using the monoclonal anti-alpha-SM-1 antibody was carried out in 15 normal ovaries, in three ovaries with stromal hyperplasia and in 27 neoplastic ovaries. In selected cases the pattern of actin isoforms was examined by means of 2 D-gel electrophoresis. In addition, the tissues were stained for vimentin and desmin. In normal ovaries alpha-SM actin was found in the inner cortex and in the theca externa. In ovarian stromal hyperplasia expression of alpha-SM actin was minimal or absent. In primary and metastatic epithelial tumors there was positive stromal staining for alpha-SM actin, especially in the vicinity of epithelial elements. This tended to be more widespread in malignant neoplasms. Thecomas did not express alpha-SM-actin and could thus be differentiated from leiomyomas which stained intensely for alpha-SM actin. Only focal stromal staining of alpha-SM actin was observed in granulosa and germ cell tumors. In all the tissues studied blood vessels were strongly positive for alpha-SM actin. Desmin, although present in the stroma of most of the specimens, was less abundant than alpha-SM actin. We concluded that alpha-SM actin is a component of the normal human ovary where it may contribute to the contractility of its stroma. Its absence in the normal outer cortex and theca interna, and in stromal hyperplasia and thecoma implies that sex hormones do not constitute a stimulus for alpha-SM actin production in the ovary. Among neoplasms it is most widely represented in the stroma of epithelial tumors in which it may reflect stromal stimulation mediated by neoplastic epithelium.

[Sterility and tumors of the testis. Study of late exocrine and endocrine functions in stage I or IIA tumors]. / Stérilité et tumeurs du testicule. Etude des fonctions exocrine et endocrine à distance dans les tumeurs de stade I et IIA.

Spermatogenesis and plasma hormone levels (testosterone, follicle stimulating hormone and 17 beta estradiol) were studied in patients presenting stage I or IIA testicular tumors. Patients with a previous history of cryptorchidism or varicocele, and those who had received combined chemotherapy, were excluded. In 22 patients (10 seminomas and 12 non-seminoma tumors) a spermogram was obtained at the time of orchidectomy, before the procedure in 9 cases and immediately afterwards in 13. Hormone levels, together with tumor marker assay (alpha feto protein and beta HCG) were determined preoperatively. Exocrine and endocrine function were restudied after 3 years of follow up. There was an initial restudied after 3 years of follow up. There was an initial deterioration in spermatogenesis in 17 patients (77.8%) as shown by a sperm count less than 20 million per cm3, an ejaculate volume less than 1.5 cm3 and a motility after one hour of less than 60%. In this group 11 patients had a sperm count of less than 10 million per cm3. Endocrine anomalies discovered included an increase in serum beta HCG levels (7 cases), combined with a decrease in follicle stimulating hormone and an increase in 17 beta estradiol. After 3 years, only 5 of these 17 patients demonstrated fertile sperm (29.4%). The endocrine anomalies tended to regress after orchidectomy. While these endocrine anomalies were always accompanied by hypofertility, their absence was not synonymous with a normal spermogram. Thus the reestablishment of fertile sperm remains unlikely even in early stage testicular tumors.

DNA cytometry of pure dysgerminomas of the ovary.

The value of DNA cytometry for predicting the malignant potential of pure dysgerminomas of the ovary was investigated. Feulgen-thionin DNA image cytometry and propidium iodide DNA flow cytometry were performed in isolated nuclei from paraffin-embedded tissue of 25 dysgerminomas. Nineteen were in clinical stage Ia1, and six were in stage III (two in IIIa and four in IIIb). Eight patients showed recurrences during a 5-year follow-up period and one patient died of the disease. The DNA index (DI; the modal DNA value compared with that of normal cells) of the tumor cells was computed from the image and flow DNA histograms. Comparable DI values, ranging from 1.29 and 3.23, were found with both types of cytometry. No correlation was found with the clinical stage or the recurrence state. Furthermore, it was striking that, although values were found strongly deviating from the normal diploid content (DI = 1.0) suggesting an unfavorable prognosis, the survival rate was relatively high. It can be concluded that prediction of the clinical course of pure dysgerminomas by DNA cytometry does not seem feasible.

Experimental endocrinotherapy with tamoxifen of human ovarian dysgerminoma transplanted into nude mice.

The effect of Tamoxifen (TAM) on tumor growth was investigated experimentally using ovarian dysgerminoma serially transplanted into nude mice. The mice were divided into two groups according to estrogen receptor (ER) content of initially transplanted tumor; groups TAM-A and -B indicated ER rich and ER poor tumor, respectively. Their own control (CNT) was included in each group. After TAM treatment, the tumor size of TAM-A and -B groups was 150 fold and 350 fold that of initial tumor respectively, while that of group CNT was over 500 fold, suggesting that a strong anti-tumor effect of TAM was exerted especially on ER rich tumor. Histological changes following tamoxifen treatment such as a decline in the size of tumor cells and nuclei as well, and a decrease in the mitotic index were observed only in the TAM-A group. The concentration of ER and progesterone receptor (PR) in the cytosol of these tumors were measured. The ER levels in the initial tumor tissue of TAM-A and -B were 14.7 and 3.3 fmol/mg protein, respectively. The significant increase in PR content in TAM-A following TAM administration obviously indicated a so-called TAM-dependent PR induction effect. These results indicated that ovarian cancer containing ER, even though it originated from germ cells, would be responsible for antiestrogen therapy.

Clinical evaluation of basic fetoprotein in testicular cancer.

We attempted to determine the efficacy of basic fetoprotein as a marker in testicular cancer. The levels of serum basic fetoprotein were studied in 58 patients (31 with seminoma and 27 with nonseminoma). Elevated levels were observed in 22 seminoma (71 per cent) and 15 nonseminoma (56 per cent) patients, while the levels of other markers (beta-subunit of human chorionic gonadotropin, alpha-fetoprotein and lactic dehydrogenase) remained normal in 6 seminoma (19 per cent) and 2 nonseminoma (7 per cent) patients. The levels of basic fetoprotein changed in relation to the clinical courses and they elevated again in 3 of 4 patients with recurrence. The concentration of basic fetoprotein in testicular cancer tissue was significantly higher than in the normal testis. Histological localization of basic fetoprotein in testicular cancer tissue was demonstrated immunohistochemically. Thus, basic fetoprotein was considered to be a useful serum marker for testicular cancer.

Patterns of seminoma tissue markers and deletions.

Seminomas and control tissues were analyzed for several tumor markers. Very high levels of placental alkaline phosphatase (PLAP)-like enzyme levels were found in all 18 seminomas studied. The majority of the seminomas were of phenotype I, thus differing from palcental PLAP. The mean amount of enzyme protein as measured by monoclonal antibodies, was 100 times higher than in non-malignant tissues and 10 times lower than in placental tissue. The specific enzymatic activity in seminomas was about half of that observed in placenta. Similarly, the specific activity of PLAP-like enzymes in sera of patients with seminoma was only about half of that found in pregnancy sera. HCG was strongly elevated in 3 seminomas, but not obviously related to PLAP. Thirteen of the 17 pure seminomas had HCG over 100 IU/g, which was not seen in normal testes. Liver alkaline phosphatase (LAP) and intestinal alkaline phosphatase (IAP) were high in seminomatous tissues, the mean increases being 60-fold and 20-fold, respectively. The highest IAP levels were found in 2 yolk-sac tumors. Ferritin was moderately elevated in seminomas, but high in several control tissues. Carcinoembryonic antigen (CEA) was not elevated and alpha-fetoprotein (AFP) was not detected at all in pure seminomas. A decrease in carbohydrate antigen 50 (CA-50) content was noted in seminomas as compared to normal testes, yolk-sac tumors and choriocarcinomas. Defects in tumor-related enzymes may account for increase of PLAP and decrease of CA-50.

The differential diagnosis of yolk sac tumor and seminoma. Usefulness of cytokeratin, alpha-fetoprotein, and alpha-1-antitrypsin immunoperoxidase reactions.

Yolk sac tumor and seminoma may have a similar appearance in focal areas. Small biopsies may therefore be difficult to interpret. The authors studied 20 yolk sac tumors and 21 seminomas to investigate the utility of immunohistochemical markers in this differential diagnosis. All yolk sac tumors stained positively for cytokeratin (CK) but so did 43% of seminomas. The CK positivity of yolk sac tumors was generally more diffuse and intense, however, there was an overlap in the spectrum of intensity of CK positivity in yolk sac tumor and seminoma. Alpha-fetoprotein (AFP) was a less sensitive (55%) marker for yolk sac tumor than CK, but AFP was quite specific in this differential diagnosis because no seminoma stained for AFP. Alpha-1-antitrypsin was not a very useful marker because of poor sensitivity and specificity. The interpretation of light microscopic patterns remains of paramount importance in the differentiation of solid yolk sac tumor from seminoma.

Levels of alkaline phosphatase isozymes in human seminoma tissue.

The three human isozymes of alkaline phosphatases were quantitatively determined in normal testis and seminoma tissues. The highly selective assays were based on isozyme specific monoclonal antibodies. In the normal testis approximately 90% of the catalytic activity originates from the tissue unspecific alkaline phosphatase, and the remaining activity was due to trace expression of both intestinal (approximately 5%) and placental alkaline phosphatase (PLAP) or PLAP-like isozyme (approximately 5%). In homogenates of seminoma tissues, highly increased levels of all three isozymes were identified. Both the tissue unspecific alkaline phosphatase and PLAP-like enzymes displayed relative increases of 10- to 100-fold and intestinal alkaline phosphatase 2- to 10-fold compared with normal testis. This finding indicates that the entire genome coding for alkaline phosphatases may be activated in seminomas. The PLAP-like enzyme from seminoma cells comprises a heterogenous population of molecules demonstrating partial heat sensitivity and microheterogeneity upon starch gel electrophoresis in contrast to the pregnancy related PLAP. These findings have implications for the different PLAP assays used in the clinical monitoring of seminoma patients.

Molecular properties of F9 embryoglycan recognized by a unique antibody in sera from patients with germ cell tumors.

Human antibody against an embryoglycan present on a mouse teratocarcinoma cell line F9 was found in sera from 16 of 29 patients with embryonal carcinoma, yolk sac tumor, immature teratoma, and choriocarcinoma of gonadal and extragonadal origins by Farr assay. In contrast, none of the sera from patients (77 cases) with dysgerminoma, seminoma, germinoma, and mature teratoma or from patients (118 cases) with nongerm cell types of ovarian tumors contained this antibody. The antigenic embryoglycan was of high molecular weight (Mr greater than 70,000) on Sephacryl S300 column chromatography and carried binding sites for Grifonia simplicifolia agglutinin-1. The antigenic embryoglycan was also found in F9 cell-cultured medium. Absorption of patients' sera with synthetic Blood Group B trisaccharides failed to remove the antibody against F9 embryoglycan. None of these patients' sera showed higher hemagglutination titer to rabbit erythrocytes than the normal range. In contrast, alpha-galactosyl carbohydrates obtained from Ehrlich ascites tumor cells effectively inhibited the binding of patients' sera with F9 embryoglycan. These results indicate that the human antibody against F9 embryoglycan recognizes alpha-galactosyl structures that are distinct from B blood group antigen, but are cross-reactive with alpha-galactosyl structures on Ehrlich ascites cells.

Quantification of alpha-fetoprotein and beta-HCG in testis tumor patients.

Patients with testis tumor were investigated for serum and tissue levels of alpha-fetoprotein and beta-human chorionic gonadotropin (beta-HCG). The tissue immune peroxidase-antiperoxidase staining for the tumor marker was quantitated by computer-assisted immunohistophotometry and immuno-gamma ray histospectrometry. The results supported the general view that mostly polynuclear giant cells produce beta-HCG in 66% of nonseminoma cancer. This finding qualifies beta-HCG as relatively unspecific in the absence of chorioepithelial cells in the tumor. Discrepancies of tissue and serum beta-HCG values may be caused by deglycolysation of beta-HCG while penetrating the perivascular tissues. Alpha-fetoprotein (AFP) appears helpfully to discriminate the true seminoma cancer, which is constantly negative. Histologically pure seminoma which reacts for AFP therefore suggests sclerotic teratoma compartments. A constant finding is the significantly reduced synthesis rate of tumor markers in metastasis compared to primary tumor.