Estrogens and alkylphenols promote proliferation of the seminoma-like TCam-2 cell line through ERα36-dependent pathways

Hussein Ajj (1) , Sophie Pinel (1) , Stéphane Flament (1) , Amand Chesnel (1) , Hélène Dumond (1)

1 SIGRETO - Signalisation, Génomique et Recherche Translationnelle en Oncologie

Background Seminoma, originated from carcinoma in situ cells (CIS), is one of the main causes of cancer in young men. Postpubertal development of these testicular germ cell tumors suggests a hormone-sensitive way of CIS cell proliferation induction probably stimulated by lifelong exposure to endocrine disruptors. In a first step to understand the mechanisms underlying the deleterious effects of endocrine disrupting compounds on germ cells, we aimed to decipher the estrogen-dependent transduction pathways in TCam2 cells. Then, we began to assess the effects of a [4tert-octyl + 4-nonylphenol] mix on testicular germ cell tumors in vitro and in vivo. Material and methods In this study, we used the unique seminoma TCam-2 cell line which do not express the canonical ER66 estrogen receptor but ERα36, a truncated isoform retaining the DNA-binding, partial dimerisation and ligand-binding domains and a specific C-term 27 aa sequence. Cells were exposed to either estradiol at concentrations in the range of those detected in an adult human testis or to a [4tert-octyl + 4-nonylphenol] mix used at low doses - i.e. those found in food and drinking water. In vitro, we performed cell proliferation assays, siRNA- or shRNA-directed knockdown, microarray-directed gene targeting and signaling pathways identification after short term (1h) or mid-term (24h or 48h) treatment. We also addressed the question of TCam-2 derived tumor growth in xenografted Nude mice treated with the [4tert-octyl + 4-nonylphenol] mix. Results We demonstrate in vitro that estradiol and the alkylphenol mix trigger TCam-2 cell proliferation through ER36-dependent pathways. We establish that estradiol can activate GPER-cAMP/PKA signaling pathway. Stable ERα36 knockdown indicates that ERα36 is (i) necessary for cell proliferation (ii) a downstream target of estradiol-activated GPER/PKA/CREB pathway, (iii) required for estradiol-dependent EGFR expression. The [4tert-octyl + 4-nonylphenol] mix signaling pathway is clearly ER36 dependent but seems to be partially non-estrogenic. Finally, we show that the [4tert-octyl + 4-nonylphenol] mix stimulates tumor growth in TCam-2 xenografted Nude mice. Conclusions Our results highlight the functional role of ERα36 in context of seminoma cell proliferation and the importance of testing ERα36 in vivo as a possible marker for endocrine disruptor susceptibility.

Domaines: Cancer

Hussein Ajj, Sophie Pinel, Stéphane Flament, Amand Chesnel, Hélène Dumond. Estrogens and alkylphenols promote proliferation of the seminoma-like TCam-2 cell line through ERα36-dependent pathways. Colloque International du Programme national de recherche sur les perturbateurs endocriniens, PNRPE, Dec 2012, Paris, France. pp.PS1-8. ⟨hal-00840939⟩