metallothionein and Choriocarcinoma

Proper functioning of trophoblastic cells is essential for maintenance of the placenta and development of the embryo/fetus. Exposure of trophoblasts to toxic exogenous factors, such as cadmium (Cd), perturbs placental function and affects fetal outcome. Cellular responses to Cd exposure include induction of the metal-binding protein, metallothionein (MT), and initiation of apoptosis. To analyze the functional relationship between cellular MT levels and apoptosis in trophoblasts, we have examined the effects of DNA transfection-mediated alterations in MT levels on trophoblastic function and apoptosis, with and without Cd exposure, using the trophoblast-like JEG-3 human choriocarcinoma cell line. JEG-3 cells stably transfected with human MT-IIa cDNA expression constructs, in either sense or antisense orientation, were unchanged in human chorionic gonadotropin (hCG) production or expression of the apoptotic markers, bcl-2 and CPP-32. However, MT overexpression significantly prolonged the recovery time of intracellular Ca flux, whereas reduced basal MT increased the incidence of apoptosis as determined by morphology and terminal deoxynucleotidyl end labeling (TUNEL) staining. Upon Cd exposure, a dose-dependent decrease in hCG secretion was seen in all JEG-3 cultures, without any correlation to basal MT expression. Basal MT levels, however, significantly affected the extent of apoptosis, the incidence being inversely related to basal MT level. These results suggest that while MT does not ameliorate heavy-metal induced perturbation of some trophoblastic functions, its expression is critical for protection of these cells from Cd-induced apoptosis and could act to maintain placental integrity in cases of maternal Cd exposure.

Topics: Apoptosis; Cadmium; Caspase 3; Caspases; Choriocarcinoma; Chorionic Gonadotropin; Culture Media, Conditioned; DNA Damage; Dose-Response Relationship, Drug; Female; Gene Expression Regulation, Enzymologic; Humans; In Situ Nick-End Labeling; Metallothionein; Proto-Oncogene Proteins c-bcl-2; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Transfection; Trophoblasts; Tumor Cells, Cultured

Maternal exposure to cadmium (Cd) during pregnancy has been linked to low fetal birthweight, which may be attributed to placental damage and/or dysfunction in nutrient transport. Previous studies have suggested that Cd is accumulated in the placenta, and that placental transport of calcium (Ca) and zinc (Zn) is perturbed by Cd. To investigate the mechanism of Cd perturbation of Ca transport, we used JEG-3, a human choriocarcinoma cell line which exhibits trophoblastic properties, to analyse Cd effects in vitro. Treatment with Cd at low, physiologically relevant concentrations (e.g. 0.04 microM) did not result in obvious changes in cell morphology or integrity, whereas higher concentrations (> or = 0.16 microM) affected cell integrity. With lower concentrations of Cd treatment for 24 h, activities of cellular Ca uptake and transport, and Ca2+ binding were decreased, and intracellular [Ca2+] ([Ca2+]i) profile was also altered; however, membrane-associated Ca(2+)-activated ATPase activity remained relatively unchanged. Interestingly, cellular Ca uptake activity was unaffected by short-term (30 min) Cd pretreatment. The 24-h Cd treatment also resulted in elevated expression of the metal-binding protein, metallothionein, whereas the expression of a trophoblast-specific cytosolic Ca(2+)-binding protein (HCaBP) was drastically reduced. These results strongly suggest that Cd exposure significantly compromises the Ca handling ability of trophoblastic cells; this effect is probably not due to perturbations in Ca channel or membrane Ca pump activities, but rather a consequence of alterations in subcellular, cytosolic Ca2+ binding activities.

Topics: Adenosine Triphosphatases; Biological Transport; Cadmium; Calcium; Calcium-Binding Proteins; Cell Line; Cell Survival; Choriocarcinoma; Enzyme Activation; Female; Gene Expression; Humans; Metallothionein; Pregnancy; RNA, Messenger; Trophoblasts; Tumor Cells, Cultured

The induction of metallothionein (MT) in a cell line derived from a malignant trophoblastic tumor (JAr cells) was demonstrated using the Cd/heme radioassay following exposure to Cd or Zn. Cd at an optimal concentration of 1 microM produced a 30-fold increase in MT following a 24 hr incubation. Induction by Cd was both time and dose dependent, with a significant increase in MT noted as early as 3 hr, with levels continuing to increase up to 24 hr. Zn was also quite effective in inducing MT synthesis in this cell line. Exposure to 80 microM Zn for 24 hr produced a 70-fold increase in MT. Although Cd was a more potent inducer of MT, exposure to Zn resulted in a greater magnitude of induction. The magnitude of MT induction in JAr cells was much greater than that seen in cultured trophoblasts from term chorion laeve. The degree of induction seen in this cell line makes it an interesting model for the study of MT's role in trophoblast function. MT induction in trophoblasts may reflect a protective mechanism against heavy metal toxicity and/or an integral aspect of normal zinc homeostasis.

Topics: Cadmium; Cell Line; Choriocarcinoma; Female; Humans; Metallothionein; Pregnancy; Time Factors; Trophoblasts; Zinc