piperidines and Choriocarcinoma

Evidence has accumulated showing that vasoactive peptides, such as endothelin-1, adrenomedullin and urotensin-II, are expressed in various kinds of tumour cells. In the present study, the expression of endothelin-1 and endothelin receptors was studied in eight human tumour cell lines: T98G (glioblastoma), IMR-32 and NB69 (neuroblastoma), BeWo (choriocarcinoma), SW-13 (adrenocortical carcinoma), DLD-1 (colonic carcinoma), HeLa (cervical carcinoma) and VMRC-RCW (renal carcinoma). Reverse transcriptase-PCR showed expression of endothelin-1 mRNA in seven out of the eight cell lines, the exception being BeWo cells. ET(A) receptor mRNA was expressed in T98G, IMR-32 and NB69 cells, but weakly in the other cells. ET(B) receptor mRNA was expressed in IMR-32, NB69 and BeWo cells, but only weakly in T98G and HeLa cells. Immunoreactive endothelin was detected in the culture media of six out of the eight cell lines, but not in that of IMR-32 or BeWo cells. Treatment of T98G cells with an anti-endothelin-1 antibody or an anti-adrenomedullin antibody for 24 h decreased cell numbers to approx. 84% and 90% of control respectively. Treatment with the ET(A) receptor antagonist BQ-610 (1 microM) significantly decreased cell number to about 90% of control, whereas the ET(B) receptor antagonist BQ-788 had no significant effect. On the other hand, exogenously added endothelin-1, adrenomedullin or urotensin-II (0.1 microM) had no significant effects on cell number. These results suggest that endothelin-1 acts as a paracrine or autocrine growth stimulator in tumours. The effect of endothelin-1 on tumour growth appears to be mediated by the ET(A) receptor.

Topics: Adrenal Cortex Neoplasms; Adrenomedullin; Antibodies, Monoclonal; Cell Division; Choriocarcinoma; Colonic Neoplasms; Endothelin Receptor Antagonists; Endothelin-1; Glioblastoma; Growth Substances; HeLa Cells; Humans; Kidney Neoplasms; Neuroblastoma; Oligopeptides; Peptides; Piperidines; Receptor, Endothelin A; Receptor, Endothelin B; RNA, Messenger; Tumor Cells, Cultured; Urotensins; Vasodilator Agents

BeWo choriocarcinoma cells were cultured onto solid microcarrier beads, packed into syringe barrels and superfused. The unidirectional choline uptake across the microvillous membrane of the cells was measured by a rapid single-circulation paired-tracer dilution procedure using methyl[3H]choline with D-[14C]mannitol as the extracellular reference molecule. Choline influx was saturable with a K(t) of 214+/-15 microM and a V(max) of 45.29+/-0.94 nmol/min/mg of cell protein. Uptake of labelled choline was partially inhibited by nicotine, strongly inhibited by hemicolinium-3, and was reduced by about 50 per cent in sodium-free perfusates. A range of agents was added to the stirrer flasks 24 h prior to the experiments to determine if intracellular or extracellular levels of choline or its metabolic product, acetylcholine, regulated choline uptake. Pre-incubation with 2 mM choline reduced the choline maximal uptake by half, while pre-incubation with 100 microM alpha-NETA [2-(alpha-naphthoyl)ethyltrimethyl-ammonium] reduced the influx by 77 per cent. Choline influx was also reduced to about half in the presence of 100 microM vesamicol, bethanecol or neostigmine. It is concluded that BeWo cells possess a choline transporter similar to that described in isolated cytotrophoblasts and syncytiotrophoblast microvillous membrane preparations, and that uptake appeared to be regulated by both intracellular and extracellular concentrations of choline and acetylcholine. Therefore, these cells provide a novel model for studying the role of acetylcholine in human placenta.

Topics: Bethanechol; Biological Transport; Choline; Choriocarcinoma; Drug Carriers; Female; Hemicholinium 3; Humans; Microvilli; Naphthalenes; Neostigmine; Nicotine; Physostigmine; Piperidines; Quaternary Ammonium Compounds; Trophoblasts; Tumor Cells, Cultured; Uterine Neoplasms

Epidemiologic studies suggest that women who smoke have lower endogenous estrogen than nonsmokers. To explore the possible link between cigarette smoking and decreased endogenous estrogens, we have examined the effects of constituents of tobacco on estrogen production in human choriocarcinoma cells and term placental microsomes. In choriocarcinoma cell cultures, nicotine, cotinine (a major metabolite of nicotine), and anabasine (a minor component of cigarette tobacco) all inhibited androstenedione conversion to estrogen in a dose-dependent fashion. Removal of nicotine, cotinine, and anabasine from the culture medium resulted in the complete reversal of the inhibition of aromatase. In the choriocarcinoma cell cultures, a supraphysiologic concentration of androstenedione (73 microM) in the culture medium blocked the inhibition of aromatase caused by nicotine, cotinine, and anabasine. In preparations of term placental microsomes, nicotine, cotinine, and anabasine inhibited the conversion of testosterone to estrogen. Kinetic analysis demonstrated the inhibition to be competitive with respect to the substrate. These findings suggest that some nicotinic alkaloids directly inhibit aromatase. This mechanism may explain, in part, the decreased estrogen observed in women who smoke.

Topics: Anabasine; Androstenedione; Aromatase Inhibitors; Carbachol; Cells, Cultured; Choriocarcinoma; Cotinine; Decamethonium Compounds; Dose-Response Relationship, Drug; Estradiol; Female; Hexamethonium Compounds; Humans; In Vitro Techniques; Kinetics; Microsomes; Nicotine; Piperidines; Placenta; Pregnancy; Pyrrolidinones; Smoking; Succinylcholine; Trophoblasts; Uterine Neoplasms