Several clinical situations require continuous glucocorticoid (GC) treatment during pregnancy. A well-known deleterious side effect of such treatment is the higher incidence of growth-restricted fetuses, for which a too shallow trophoblast invasion is presently hypothesised as the underlying cause. This study investigated whether the synthetic GC triamcinolone acetonide (TA) influences proliferation, invasion and endocrine activity of human trophoblast. BeWo and JEG-3 choriocarcinoma cell lines both express GC receptors (western blotting) and were used as models for human trophoblast. JAR devoid cells of GC receptor were used as negative control. The cells were cultured for 48 h without (control) or with 0.5, 5 and 50 mM TA. In the presence and absence of serum, proliferation was determined by cell counting and measuring the cell cycle regulating protein cyclin B1 (Western blotting); invasion was determined by a conventional Matrigel invasion assay and by measuring the secretion (ELISA) of matrix-metalloproteinases (MMP-2, MMP-9) into the culture medium; endocrine activity was assessed by measuring the levels of human chorionic gonadotropin (ELISA) into the culture medium. TA altered the number of viable and dead cells as well as cyclin B1 levels and, to a lesser extent, invasion of BeWo and JEG-3, with a strong influence of serum. BeWo and JEG-3 cells reacted differently and in most instances reverse. In the cell lines used as models of human trophoblast, TA alter some functions relevant to proliferation and invasion, and suggest that caution should be exercised when treating women with GCs during pregnancy.

Extravillous cytotrophoblasts are specialised epithelial cells of the placenta that proliferate or invade the maternal decidua. Little is known about the mechanisms that regulate these processes. Here the effects of several insulin and insulin-like growth factor-I (IGF-I) doses, either singly or in synergy with serum, on human chorionic gonadotropin-beta (hCG-beta) secretion (RIA), proliferation (cell counting, cyclin B(1) levels) and invasion [Matrigel invasion assay, secretion of matrix metalloproteinases (MMP) 2 and 9] were investigated. The choriocarcinoma cell lines BeWo, JAR and JEG-3 served as models for first trimester human trophoblasts. Both growth factors altered hCG-beta secretion and proliferation dependent on the cell line. Insulin stimulated proliferation in JAR cells and, to a lesser extent, in JEG-3 cells, and when cultured in serum-free medium, BeWo was not affected. Invasion was not affected although proMMP-2 levels in culture medium were altered under some conditions. A strong synergistic effect with serum was noted. In the presence of serum both growth factors reduced proliferation and invasion in a similar fashion. Since the cell models differ by their degree of differentiation, the data demonstrate that the effects of insulin and IGF-I strongly depend on serum and the degree of differentiation. It can be speculated that IGF-I can take on tasks of insulin in the regulation of trophoblast functions under conditions of insulinopenia.

2,2'-dihydroxy-6,6'-dinaphthyldisulfide (DDD) reacts with both protein thiol groups and with protein disulfides (Nöhammer 1977). By varying the pH of the DDD-reaction, as well as the reaction times, the complex reaction became specific with respect to the histochemical demonstration of protein-SH groups. Furthermore, the application of the histochemical DDD-reaction following quantitative blockade of the protein-SH groups enabled the demonstration of distinctive DDD-reactive disulfides. The specificity and the extent of the different histochemical DDD-staining methods were investigated by comparing macroscopically determined values of the protein-SH-contents, and the contents of the different kinds of disulfides in Ehrlich-ascites-tumor cells (EATC) (Modig 1968; Hofer 1975), with microspectrometrical values determined with the MCN-method of Nöhammer et al. (1981), and with microspectrometrical values measured on EATC after staining with the modified DDD-methods. Also, the method for the histochemical demonstration of protein-SH with DDD after the reduction of the disulfides with thioglycolate was investigated and conditions were found by which the protein-SH content could be determined quantitatively with DDD and Fast blue B after the reduction of the disulfides. With the aid of the MCN-method (Nöhammer et al. 1981), the intracellular disulfide interchange reaction was investigated, leading to pH-dependent changes of the SH-SS-ratio of fixed cells during their incubation in aqueous media. In addition the possibility of protein loss during the long incubation times of the fixed cells in the DDD-solutions was investigated. For the quantitative microscpecrometrical determination of the protein content of EATC the so-called tetrazonium-coupling method, optimized by Nöhammer (1978) and calibrated by Nöhammer et al. (1981), was used.

No abstract

Protein-thiol groups that react with dihydroxydinaphthyl disulphide during a 7 h incubation (so-called reactive protein thiols, PSH,) have been quantitatively measured on sections of human uterine cervix by microcytospectrophotometry.Measurements were made on areas (1 pmz) of epithelium and adjoining stroma in samples of normal cervix, and in samples obtained from patients with dysplasia, carcinoma-in-situ and invasive cancer. The ratio of PSH, in epithelium to stroma is substantially reduced in the pathological conditions compared with normal and in apparently normal adjacent areas. Such changes in PSH, are discussed in relation to the redox balance of the tissue, and free radical disturbances previously described.