Epidermal growth factor receptors in spontaneous ovarian granulosa cell tumors of SWR-derived mice.

Epidermal growth factor (EGF) receptor binding properties were examined in spontaneous ovarian granulosa cell (GC) tumors from SWR and SWR-derived strains of mice. EGF binding was measured at room temperature in tissue homogenates from GC tumors and normal ovaries from adult randomly cycling mice. GC tumor tissue displayed significantly increased EGF binding and 2 receptor populations (R1 and R2). Normal ovarian tissue appeared to have only one receptor population with a dissociation constant (KD) similar to the R1 (high-affinity) receptor in GC tumors. In subsequent experiments, GC tumor and normal granulosa cells from immature mice were analyzed in primary cultures for EGF binding, immunofluorescence microscopy for receptors, and cell proliferation. After 24 hr in culture, the GC tumors bound 10-fold more EGF/micrograms protein than did normal granulosa cells. GC tumor cells, but not normal granulosa cells, showed specific immunofluorescence when reacted with a polyclonal antibody to mouse EGFR. During 96 hr in culture, GC tumor cells, but not normal cells, showed a significant proliferative response to EGF. In conclusion, the EGF binding capacity is markedly increased in GC tumor cells and the proliferation data suggest that this growth factor supports tumor growth in the SWR model system.

Immunohistochemistry of ovarian granulosa cell tumours. The value of tissue specific proteins and tumour markers.

Paraffin-embedded material of 47 ovarian tumours primarily diagnosed as granulosa cell tumours, including 2 cases of juvenile granulosa cell tumour, were studied immunohistochemically for the presence of intermediate filament proteins, epithelial membrane antigen and tumour markers. Forty-one lesions, including the 2 juvenile granulosa cell tumours, were vimentin positive, while keratin and epithelial membrane antigen expression could not be detected. Six tumours primarily diagnosed as poorly differentiated malignant granulosa cell tumours were vimentin negative, showed a mild to moderate positivity for keratin and intense positivity with the anti-epithelial membrane antigen antibody. These latter tumours were therefore classified as undifferentiated ovarian carcinomas, corresponding to their significantly poorer prognosis and shorter survival when compared with the granulosa cell tumours. Two of these six tumours were positive for carcino-embryonic antigen. Two small cell carcinomas of the ovary studied in addition expressed keratin in a proportion of tumour cells while no epithelial membrane antigen or vimentin was detectable. None of the tumours tested for alpha-fetoprotein, human chorionic gonadotrophin, human placental alkaline phosphatase and human placental lactogen, were positive. The data indicate the value of antibodies directed against intermediate filament proteins and epithelial membrane antigen to distinguish granulosa cell tumours from poorly differentiated carcinomas, a worthwhile distinction considering the much better prognosis of granulosa cell tumours.

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.

Correlation between steroid hormone receptors, histological and clinical parameters in ovarian carcinoma.

Estrogen (ER) and progesterone (PR) receptors in ovarian tumors of 62 patients (51 carcinomas, 11 benign tumors) were estimated by the dextran-coated charcoal method using Scatchard plot analyses. 63% of carcinomas were ER-positive (greater than 10 fmol/mg cytosol), 38% were PR-positive (greater than 25 fmol/mg cytosol), whereas in benign tumors only 45% were ER-positive and 36% were PR-positive. We found no statistically significant correlation between receptor content and stage of disease, menopausal status or age of the patient. The highest concentration of ER and PR was observed in patients between 61 and 70 years of age. Life table analysis for patients with advanced ovarian carcinomas showed no significant difference in survival time in the group with higher ER and PR content. This study also reports the results obtained in a group of patients with receptor-positive ovarian carcinomas treated with a combination of chemotherapy and antiestrogen therapy. In comparison to treatment with cytotoxic chemotherapy alone, no significant difference in the time of survival or duration of remission could be found.

Intermediate filaments cytokeratin and vimentin in ovarian sex cord-stromal tumours with correlative studies in adult and fetal ovaries.

The expression of the intermediate filaments cytokeratin and vimentin were studied immunohistochemically in a series of ovarian sex cord-stromal tumours (26 adult and juvenile granulosa cell tumours, 11 thecomas, six fibromas, three Sertoli-Leydig cell tumours and 1 sex cord tumour with annular tubules). Contrary to previous reports, granulosa cell tumours expressed cytokeratins as well as vimentin. Thecomas and fibromas expressed vimentin only. In Sertoli-Leydig cell tumours and the sex cord tumour with annular tubules, both cytokeratins and vimentin were detected. Correlative studies in adult ovaries showed that patterns of expression in non-neoplastic granulosa, thecal and stromal cells correspond to their neoplastic counterparts. Investigation of fetal ovaries demonstrated that these patterns of intermediate filament expression exist from relatively early stages of development. Ovarian surface epithelium and rete ovarii, like granulosa cells, co-expressed cytokeratin and vimentin. The demonstration of cytokeratins in granulosa cells and the reported presence of desmosomes and tonofilaments, suggests the epithelial nature of these cells although not clarifying their histogenesis. The presence of both these intermediate filaments in granulosa and Sertoli-Leydig cell tumours as well as in some ovarian carcinomas which may mimic them, limits their value in differential diagnosis between these tumour groups.

Desmosomal plaque-associated vimentin filaments in human ovarian granulosa cell tumors of various histologic patterns.

Proteins of intermediate-sized filaments and desmosomal plaques (desmoplakins) of four human ovarian granulosa cell tumors were studied by immunofluorescence and immunoelectron microscopy and by two-dimensional gel electrophoresis of microdissected tissue samples. All tumor cells, irrespective of their specific histologic patterns, contained both vimentin and desmoplakins. Cytokeratin-positive structures were absent or very scant in most tumor regions, but more common in trabecular, insular, macro- and microfollicular structures. Biochemical analysis revealed the presence of Cytokeratin Polypeptides 8 and 18. Desmin filaments, neurofilaments, and glial filaments were not detected. Immunoelectron microscopy showed vimentin filaments attached to desmoplakin-positive plaques of desmosomes. These results indicate that granulosa cell tumors contain true desmosomes, which are associated primarily with vimentin filaments. This phenomenon has so far only been described in meningiomas and in blastema cells of nephroblastomas. Our observations suggest that in most neoplastic granulosa cells one epithelial feature, ie, cytokeratin expression, is greatly reduced, whereas desmosomes are still formed in appreciable frequencies. This unusual constellation of cytoskeletal elements in granulosa cell tumors may be useful in the differential diagnosis from other ovarian neoplasms, especially undifferentiated carcinomas. The importance of the use of antibodies specific for exclusively desmosomal proteins in classifying morphologically ill-defined junctional structures (eg, "rudimentary junctions," "primitive junctions," "desmosome-like junctions") is emphasized.

Androgenic juvenile granulosa cell tumour. Case report.

An androgenic juvenile granulosa cell tumour of the ovary in a postmenarcheal girl is presented. Distinctive histological features of this variant are stressed and virilizing effects are explained by an immunohistochemical study of hormones produced by granulosa and theca cells.

Estradiol and progesterone receptors in normal ovary and ovarian tumors.

Specific estradiol (ER) and progesterone receptors (PR) in the tumor cytosol of 54 patients with primary ovarian cancer were examined. Also 30 benign ovarian neoplasms and 19 normal ovaries were analysed. Of the malignant tumors, 46% contained ER and 41% PR. The ER content of most malignant tumors was 3-30 fmoles/mg protein; three tumors revealed an ER content over 30 fmoles/mg protein. The PR content of most malignant tumors was 8-80 fmoles/mg protein. Three endometrioid adenocarcinomas and three serous cystadenocarcinomas had a PR level over 100 fmoles/mg protein. ER and PR were simultaneously detected in 26% of the malignancies, whereas 39% of them were receptor negative. No significant differences were found between premenopausal and postmenopausal women, as regards the presence of steroid receptors. Carcinomas were significantly more often ER-positive than were benign tumors. The ER content of both the benign neoplasms and the normal ovary was low, less than or equal to 15 fmoles/mg protein, whereas higher PR levels were found in these two groups. The presence of steroid receptors in malignant ovarian tumors can possibly be used as an indicator of hormone dependency of these malignancies.

[Estrogen and progesterone receptors in malignant ovarian neoplasms]. / Ostrogen- und Progesteronrezeptoren in malignen ovarialtumoren.

Estrogen receptors (ER) and progesterone receptors (PR) were evaluated in 173 primary ovarian cancers and in 6 ovarian metastases. In epithelial ovarian carcinomas 63% had ER and 46% PR. Almost all granulosa cell tumours were receptor-positive, while sarcomas, dysgerminomas, and teratomas lacked ER and PR. Both receptors were found less often in tumours of the histological grade I than in those of grade II and III. During the development of metastases and during chemotherapy there was a loss of PR in 27% and 53% of the cases, respectively, while the amount of ER remained more or less constant. In addition to ovarian cancers ER and PR were present in carcinomas of the fallopian tube as well. ER-negative and especially PR-negative tumours seemed to respond better to chemotherapy than receptor-positive carcinomas. The possible significance of ER and PR with regard to the success of an endocrine treatment is discussed.

Steroid-receptor proteins in nonepithelial malignancies of the ovary.

Tissue samples from 20 patients with nonepithelial ovarian malignancies were assayed for cytosol estrogen (ERc) and in selected samples progestin-receptor (PRc) proteins. The nine germ cell malignancies assayed were low in ERc. One recurrent granulosa cell tumor had an ERc concentration of 36 fmole/mg cytosol protein (fmole/mg c.p.) while three other stromal tumors were low in ERc. Three of seven mixed mesodermal tumors had a high ERc concentration (34-91 fmole/c.p.). Two primary and one recurrent granulosa cell tumors and one endodermal sinus tumor had borderline to elevated PRc levels (18-254 fmole/c.p.). Four of seven mixed mesodermal tumors had borderline to slightly elevated PRc concentrations (13-22 fmole/c.p.). The low ERc in germ cell malignancies of the ovary is consistent with previous clinical observations that the growth of these tumors is not influenced by removal of the contralateral ovary. The high ERc levels detected in three of seven mixed mesodermal ovarian tumors and one of three granulosa cell tumors suggest that hormone-related therapy should be investigated in the management of patients with these malignancies.

Ovarian granulosa cell tumors--immunohistochemical localization of estradiol and ultrastructure, with functional correlations.

The authors studied immunohistochemical localization of estradiol and ultrastructure in 11 ovarian granulosa cell tumors in an effort to establish the cellular source of increased estrogen in patients with granulosa cell tumors. Estradiol was identified by the immunoperoxidase method in 10 tumors, and staining was confined to scattered groups of granulosa cells or isolated granulosa cells. The majority of tumor cells in all 5 cases examined by electron microscopy were undifferentiated cells, but in 2 cases, a small percentage of tumor cells contained mitochondria with tubular cristae and well-developed smooth endoplasmic reticulum. Interpreted in conjunction with in vitro studies of granulosa cells, the results suggest that neoplastic granulosa cells may resemble ovarian follicular granulosa cells of all stages of maturation and are capable of estrogen production.

The ultrastructure of a feminizing granulosa-theca tumor.

A 13-year-old girl with evidence of hyperestrinism had a right ovarian tumor removed. This neoplasm was composed predominantly of granulosa cells and exhibited some changes of luteinization. The neoplastic cells displayed an investment of basement membrane material which was closely related to fibrillar collagen within the tumor. Scattered throughout the neoplasms were small numbers of cells with charcteristics of the theca and a few cells had cytoplasmic features suggestive of smooth muscle. The ultrastructural findings of eight similar tumors have been described. In none is the basement membrane material fully appreciated; nevertheless, the production of this extracellular material is a major and distinctive feature of granulosa-theca tumors.