Lectin binding in tissues from hydatidiform mole, invasive mole and choriocarcinoma to concanavalin-A, wheat germ agglutinin and peanut agglutinin.

A light microscopic analysis of lectin receptors in normal placenta and trophoblastic disease was performed utilizing biotinylated Concanavalin-A (Con-A), wheat germ agglutinin (WGA), and peanut agglutinin (PNA), in conjunction with an avidin-biotin peroxidase complex. Hydatidiform mole, invasive mole and choriocarcinoma exhibited increased receptors to Con-A and WGA compared to normal placenta. Increased reactivity to Con-A and WGA was associated merely with increased growth and proliferation of trophoblasts rather than a malignant transformation. Normal placenta, partial and complete mole generally showed moderate to strong binding with PNA after neuraminidase treatment, while invasive mole and choriocarcinoma (11 of 15 cases) generally showed minimal to absent reaction with PNA. Heterogeneity of PNA binding in choriocarcinoma was manifested by the presence of PNA reactivity in the trophoblast membrane in 2 cases wherein no prior neuraminidase treatment was given. This suggests that in some malignant trophoblasts, there is absence of sialic acid in the terminal cell surface carbohydrate groups resulting in the exposure of N-acetylgalactoseamine.

[Prediction of the malignant tendency of hydatidiform mole by determination of DNA content].

The cellular DNA content of paraffin-embedded molar tissue blocks obtained from the first curettage was measured by flow cytometry in 64 cases of hydatidiform mole. Of these 64 cases 32 developed malignant changes and 32 remained persistently benign during a period of follow-up of 0.5 to 7.5 years. The proportion of aneuploid was 59.4% in patients demonstrating malignant changes and 34.4% in those with persistent benign diseases. The difference was statistically significant (P less than 0.05). This preliminary result suggests that DNA aneuploid may be used as a parameter for the prediction of possible malignant changes and selecting of indications for prophylactic chemotherapy.

[The DNA content in the cell nuclei of trophoblastic tumors]. / Soderzhanie DNK v iadrakh kletok opukholei trofoblasta.

The DNA content in cytotrophoblast (CTB) and syncytiotrophoblast (STB) cell nuclei was assayed in tissue sections of 7 hydatidiform moles (HM) and 27 choriocarcinomas (CH). The procedure involved Feulgen's reaction and scanning cytophotometry. The analysis of summarized histograms showed the DNA distribution in CTB cell nuclei, on the one hand, and that in STB, on the other, to differ significantly in both the tumors. The HM studied cases were referred to as two subtypes on the basis of such parameters as modal class value, its ploidy and degree of nuclear poly- and heteroploidy of CTB and STB. These characteristics were used to identify three patterns of CH. A pronounced modal class (2c--4c) was typical of type 1. A wider range of modal class (2c--10c or 4c--8c) was observed in type 2. Type 3 of tumor was characterized by a pronounced polyploidy with the absence of the modal class. The analysis of individual CTB and histograms showed no significant differences between HM and CH with respect to the DNA content. An increase in the share of highly polyploid cells was associated with a shorter survival of patients.

Inhibin: a new circulating marker of hydatidiform mole?

OBJECTIVE: To define the concentrations of inhibin in serum and tissue of patients with hydatidiform mole and assess their value as a clinical marker of the condition. DESIGN: Prospective study of new patients with hydatidiform mole, comparison of paired observations, and case-control analysis. SETTING: A university hospital, two large public hospitals, and a private women's clinic in Japan. PATIENTS: Seven consecutive referred patients seen over four months with newly diagnosed complete hydatidiform mole, including one in whom the mole was accompanied by viable twin fetuses (case excluded from statistical analysis because of unique clinical features). All patients followed up for six months after evacuation of molar tissue. END POINT: Correlation of serum inhibin concentrations with trophoblastic disease. MEASUREMENTS AND MAIN RESULTS: Serum concentrations of inhibin, human chorionic gonadotrophin, and follicle stimulating hormone were compared before and seven to 10 days after evacuation of the mole. Before evacuation the serum inhibin concentrations (median 8.3 U/ml; 95% confidence interval 2.4 to 34.5) were significantly greater than in 21 normal women at the same stage of pregnancy (2.8 U/ml; 2.1 to 3.6), and inhibin in molar tissue was also present in high concentrations (578 U/ml cytosol; 158 to 1162). Seven to 10 days after evacuation inhibin concentrations in serum samples from the same patients declined significantly to values (0.4 U/ml; 0.1 to 1.4) similar to those seen in the follicular phase of normal menstrual cycles. None of the four patients whose serum inhibin concentrations were 0.4 U/ml or less after evacuation developed persistent trophoblastic disease. Though serum human chorionic gonadotrophin concentrations declined after evacuation (6.6 x 10(3) IU/l; 0.8 x 10(3) to 32.6 x 10(3], they remained far higher than in non-pregnant women. Serum follicle stimulating hormone concentrations remained suppressed. CONCLUSIONS: In this small study serum inhibin concentrations higher than those found in the early follicular phase one to two weeks after evacuation of a hydatidiform mole seemed to be specific for persistent trophoblastic disease. Further data are needed to confirm these promising results.

Immunohistochemical studies of fetal trophoblast and maternal decidua in hydatidiform mole and choriocarcinoma.

Immunohistochemical techniques have been used to investigate the expression of fetal trophoblast antigens and the maternal leucocytic response in molar pregnancy and choriocarcinoma. The antigenic phenotype of morphologically defined trophoblast populations in complete, partial and invasive moles was analogous to that in normal pregnancy. All trophoblast phenotypes described in normal pregnancy were also identified in choriocarcinoma, suggesting that extensive differentiation into heterogeneous subgroups occurs in malignant trophoblast. The maternal leucocytic infiltrate in molar pregnancy consisted of T lymphocytes and class II MHC-positive macrophages. CD2-positive, CD3-negative lymphocytes were identified in molar decidua but not in uterine tissue in choriocarcinoma. Similarly, endometrial granulocytes were present in molar decidua but not in choriocarcinoma; these cells were associated with decidualization rather than with fetal trophoblast.

[Characteristics of the production of chorionic gonadotropin by the cells of trophoblastic tumors (immunohistochemical research)]. / Osobennosti produktsii khorionicheskogo gonadotropina kletkami trofoblasticheskikh opukholei (immunogistokhimicheskoe issledovanie).

An immunohistochemical investigation of chorionic gonadotropin (CGT) in normal placental, hydatidiform mole and choriocarcinoma tissues was undertaken. Cyto- and syncytiotrophoblast both were shown to produce the hormone in normal and tumor trophoblast; the rate of synthesis increased in proportion to the degree of cell elements differentiation, except for very large elements of syncytiotrophoblast with numerous pycnotic nuclei which were found to contain either only traces of response to CGT or none at all. The percentage of syncytiotrophoblast--produced hormone progressively increased in the succession of the placenta--hydatidiform mole--choriocarcinoma. No significant difference in the levels of immunohistochemically--identified hormone was found in the hydatidiform mole and choriocarcinoma.

Partial hydatidiform moles: deoxyribonucleic acid content and course.

Partial moles are either diploid (46 chromosomes) or triploid (69 chromosomes). In 35 cases ploidy was measured by flow cytometry for nuclear deoxyribonucleic acid quantitation, indicating approximate chromosome number. Six of the 35 were triploid (17%) and the balance was diploid (83%). No complications occurred in the triploid group, while five of 25 (20%) diploid cases with evaluable follow-up had nonfatal sequelae. Complications included persistent mole treated by recurettage and four cases requiring chemotherapy (human chorionic gonadotropin titer plateau, clinical metastasis, overt choriocarcinoma, and placental site trophoblastic tumor). A combined morphologic and genetic classification of partial moles is recommended for identification and risk assessment. Deoxyribonucleic acid cell cycle fractions of the diploid partial moles may be helpful in the identification of patients at high risk for complications. A proliferation index greater than 3.6 separated the cases with sequelae from most of the uncomplicated cases (sensitivity 100%, specificity 95%).

Villous alpha-fetoprotein in molar and non-molar abortuses.

The alpha-fetoprotein (AFP) content of chorionic villi from elective and spontaneous abortuses, diploid androgenetic and paternally derived triploid conceptuses was measured by a radioimmunoassay technique and related to the presence of fetal parts and chromosome constitution. AFP values above 10 kIU/l were detected in villi from all chromosomally normal conceptuses, except for one without fetal parts, and also in nine out of 13 triploid conceptuses and 19 out of 27 diploid androgenetic conceptuses. Early fetal demise may explain villous AFP concentrations below 10 kIU/l in one karyotypically normal and four triploid abortuses. The finding of immunoreactive, genuine AFP in diploid androgenetic molar conceptuses, assumed to be anembryonic, indicates AFP synthesis by abnormal trophoblastic tissue and intravesicular accumulation of AFP.

Differing concentrations of human chorionic gonadotropin and prolactin in the cyst fluid of hydatidiform mole and in amniotic fluid.

The concentrations of human chorionic gonadotropin and prolactin in the cyst fluid of molar tissue were compared with that of normal amniotic fluid. Molar fluid was aspirated from the vesicles of molar tissue in eight women (duration of amenorrhea 14.1 +/- 1 week, mean +/- SEM). Amniotic fluid was obtained at amniocentesis in 24 women (mean duration of amenorrhea 15.9 +/- 0.2 week). Hormones were measured by specific radioimmunoassay. The concentrations (mean +/- SEM) of human chorionic gonadotropin in molar fluid and amniotic fluid were 581,829 +/- 112,581 and 3187 +/- 505 mlU/ml (p less than 0.001), respectively. For prolactin the levels in molar fluid and amniotic fluid were 44 +/- 24 and 1962 +/- 313 ng/ml (p less than 0.001), respectively. These data demonstrate that molar fluid contains 182-fold higher levels of human chorionic gonadotropin and 45-fold lower levels of prolactin than amniotic fluid obtained from normal pregnant women with a similar duration of amenorrhea. In addition to altered endocrine function of the trophoblast, there may be altered prolactin secretion from the decidua in molar pregnancy as compared with normal pregnancy. Further research is required to evaluate prolactin production from decidua of molar pregnancy.

Thyrotropic activity of acidic isoelectric variants of human chorionic gonadotropin from trophoblastic tumors.

Previous studies have indicated that hCG has a weak intrinsic thyroid-stimulating activity. Differences in the molecular composition and biological activity of hCG in patients with trophoblastic diseases and pregnant women occur, but are not well defined. Therefore, we have studied the effect of serum samples and purified hCG preparations from patients with trophoblastic diseases on T3 release from human and porcine thyroid slices in vitro. We examined 30 serum samples from 13 patients with nonseminomatous testicular germ cell tumors, 3 from women with choriocarcinoma, and 5 from patients with hydatidiform moles. In all but 1 serum sample from the tumor patients, but in none of 11 serum samples of pregnant women, T3-releasing activity was found. Two patients with testicular cancer and 1 patient with molar pregnancy experienced episodes of frank hyperthyroidism. Isoelectric focusing on polyacrylamide gels of tumor sera (n = 15) revealed substantial amounts of acidic isoelectric variants, pI 3.3-3.9, which were only barely detectable in pregnancy sera. The percentage of acidic hCG variants with pI 3.3-4.0 to total hCG with pI 3.3-5.2, as determined by hCG (+hCG beta) RIA of the eluted fractions of polyacrylamide gel isoelectric focussing, varied from 12-45% in sera of tumor patients and from 0-4% in pregnant sera. We purified the acidic variants of hCG with pI 3.6-3.8 (hCGav) from the urine of our patients. The beta-subunit of purified hCGav had a slightly higher mol wt (35,750) than that of hCG CR 119 (34,190) on polyacrylamide gel electrophoresis. The hCGav showed a dose-dependant stimulation of T3 release and cAMP generation from human thyroid slices, whereas the other hCG fractions on isoelectric focussing had no thyrotropic effect in similar dose levels. The TSH-like activity of hCGav could be roughly estimated as 10 mIU TSH/IU hCGav. Anti-hCG (+hCG beta) antiserum, but not anti-hTSH antiserum, neutralized the biological activity of hCGav. These findings strongly suggest that acidic hCG variants act as functional stimulators of the human thyroid in vitro. Since these molecular variants of hCG can exist in patients with trophoblastic diseases in significant amounts, they could be responsible for some cases of hyperthyroidism in trophoblastic diseases.

[A study of the properties of human chorionic gonadotropin (hCG) from total hydatidiform mole with low hCG immunoactivity].

A study was conducted to define biochemical characteristics of human chorionic gonadotropin (hCG) in total hydatidiform mole patients with low serum and urinary hCG levels and to inquire into the reason for its low immunological hCG level by comparing with the hCG of molar patients with a high hCG level. Both hCGs purified from chorionic tissue of molar patients with low and high hCG levels have essentially identical physicochemical, immunological and biological properties. They have low levels of biological activity and practically identical amino acid and carbohydrate compositions. It appears that moles of patients with high levels as well as those with low levels of immunoreactivity produce this purified form of hCG. Extracts of molar tissue from patients with high levels of hCG immunoreactivity also contained other substances with apparent immunoreactivity which can be found in small amounts in tissue extracts from patients who excreted low levels of hCG immunoreactivity. In contrast, extracts of molar tissue and urine from patients with low hCG levels contained hCG-like substances which cannot be detected with the ordinary commercial hCG immunoassay kits. These differences may account for the different levels excreted by these two types of patients.

Eutopic and ectopic macromolecular human placental lactogen.

Macromolecular forms of human placental lactogen have received little attention because it has been thought that such forms either compose only a small fraction of total immunoactive placental lactogen or are merely laboratory artifacts. We examined serum and placental tissue from women with normal pregnancy (first and third trimesters), serum and tissue from women with eutopic tumors (mole and choriocarcinoma), and serum from men with ectopic placental lactogen production. Samples were chromatographed on dextran gel (Sephadex G-100), and placental lactogen was measured in the fractions by radioimmunoassay. In all specimens examined, immunoactive placental lactogen was found at the void volume of the column (molecular weight greater than 150,000 daltons). This macromolecular placental lactogen comprised less than 4% of total placental lactogen in the third trimester, in mole, and in 16 of 18 first-trimester samples but was significantly higher, up to 19%, in the malignant cases. In two first-trimester placental extracts (but not in their matched sera) macromolecular placental lactogen was the dominant (greater than 45% of the total placental lactogen) immunoactive species. Authentic monomeric placental lactogen was not converted to macromolecular placental lactogen by repeated freezing and thawing. Third-trimester placental macromolecular placental lactogen was unstable; only 13% remained at the void on rechromatography. First-trimester placental macromolecular placental lactogen, on the other hand, was stable to rechromatography. The behavior of immunochemical dilutions of macromolecular placental lactogen from first-trimester placenta was similar to that of monomeric placental lactogen in the same sample. Macromolecular placental lactogen is probably not artifact, and it can comprise a large fraction of the total immunoactive placental lactogen in certain conditions.

Detection of incompletely sialylated human chorionic gonadotropin by peanut agglutinin in choriocarcinoma.

Human chorionic gonadotropin (hCG) in normal pregnancy and hydatidiform mole did not bind to the peanut agglutinin (PNA)-Sepharose column, whereas significant amount of hCG in choriocarcinoma adsorbed to the column. Therefore, PNA-Sepharose affinity chromatography is an effective tool for the detection of desialylated hCG and may be useful also for the diagnosis of choriocarcinoma.

Polyamines and nucleic acids in gestational trophoblastic tumors.

To test the relationship between tumor malignancy and content and distribution of polyamines and nucleic acids, 2 forms of human gestational trophoblastic tumors were examined: the hydatidiform mole (self-limited form) and the human chorio-carcinoma (invasive form) xenografted into nude mice. The results indicate that there are 2 significant differences between the choriocarcinoma and the mole: 1) the choriocarcinoma is characterized by increased polyamine and nucleic acid levels, 2) tissues differ in their putrescine:spermidine and spermidine:spermine ratios. There is an increase in polyamines in the urine of mole-bearing patients over that of normal controls. The correlation between putrescine and spermidine with the chorionic gonadotropin indicates that these 2 polyamines reflect the biological activity of the mole.

[Dynamics of various peptide hormone receptors in human placental cell membranes].

The action of peptide hormone receptors on the cell membranes and the cyclic-AMP in its post receptor system dynamics was studied. [Results] Receptors for hCG-LH, hPL-hGH, ACTH and Insulin were found in the placental cell membranes but receptors for FSH and PRL were not found. The possibility was suggested that the cyclic-AMP was the second messenger in the receptor system for hCG-LH, hPL and ACTH. The differences between normal placental cell membranes and hydatidiform mole were studied from the point of view of the receptor level. Receptors on molar trophoblast for hCG-LH, hPL-hGH and ACTH increased about two fold to three fold over those on normal trophoblast. The presence of receptors for hCG-LH, hPL and ACTH was confirmed also in GCH-1 (gestational choriocarcinoma cell line). The possibility is shown that the placenta has an auto regularity system in its own tissue throughout the various peptide hormones receptors such as hCG-LH and hPL.

Human chorionic gonadotropin levels in complete and partial hydatidiform moles and in nonmolar abortuses.

The rates of regression of human chorionic gonadotropin (hCG) in patients with complete hydatidiform moles, partial hydatidiform moles, and nonmolar abortions were compared. No difference in rates of regression was found among the three groups, but levels of hCG immediately after uterine evacuation were significantly higher in the group with complete hydatidiform moles. Differences in the time required for hCG levels to become undetectable were attributed to the difference in the degree of initial elevation of hCG.

Complete hydatidiform moles combine maternal mitochondria with a paternal nuclear genome.

The parental origin of mitochondria in hydatidiform moles has been investigated by analysis of genetic variants of mtDNA restriction enzyme patterns. In six complete moles the mtDNA was found to be maternal in origin, with no contribution from the sperm mitochondria, while the nuclear genome was shown to be exclusively paternal in five cases. The occurrence of mtDNA variation in the healthy population was investigated using white blood cells and placentae, and the most common variation occurred at the Ava II restriction sites. The variants exhibited by molar mtDNA were the same as those found in material from healthy individuals.

Immunocytochemical localization of placental lactogen and chorionic gonadotropin in the normal placenta and trophoblastic tumors, with emphasis on intermediate trophoblast and the placental site trophoblastic tumor.

This report presents preliminary observations on the immunocytochemical localization of human chorionic gonadotropin (hCG) and human placental lactogen (hPL) in placental site trophoblastic tumors, hydatidiform moles, and choriocarcinomas and compares the findings with those of a similar immunocytochemical analysis of the placenta at various stages of development. In addition to cytotrophoblast (CT) and syncytiotrophoblast (ST), a third form of trophoblast designated "intermediate trophoblast" (IT) is present during normal pregnancy and in trophoblastic disease. Intermediate trophoblastic cells are mononucleate, larger than CT, and contain more abundant eosinophilic cytoplasm, resulting in a partial resemblance to ST. Intermediate trophoblast has distinctive immunocytochemical features that distinguish it from CT and ST. The localization of hPL and hCG in both IT and ST varies with the age of the placenta, with the type of trophoblastic neoplasm, and from one specimen to another within each category of tumor. Syncytiotrophoblast may contain both hormones in large amounts, whereas IT contains hPL predominantly and hCG focally. Cytotrophoblast is devoid of hCG and hPL except in choriocarcinoma, which may show focal weak staining for hCG. Immunocytochemical identification of hCG and hPL has proved helpful in clarifying the histogenesis of trophoblastic neoplasms and may also be of value in establishing their diagnosis and in determining their prognosis.