Autocrine stimulation of IGF1 in estrogen-induced growth of endometrial carcinoma cells: involvement of the mitogen-activated protein kinase pathway followed by up-regulation of cyclin D1 and cyclin E.

To examine estrogen-induced growth mechanisms of endometrial carcinoma, we investigated the estrogen-induced activation of the mitogen-activated protein kinase (MAPK) pathway and cell cycle regulators. Estradiol (E(2)) treatment at concentrations of 10(-8) M and 10(-6) M to estrogen receptor (ER)-positive endometrial carcinoma Ishikawa cells for 24 h resulted in increased cell proliferation by 20% and 28% respectively. The E(2)-induced proliferation was associated with the activation of extracellular signal-regulated kinase (MAPK)3/1 and up-regulation of cyclin D1 and E, which were suppressed by the addition of an MAP2K inhibitor (U0126) or an ER antagonist (ICI 182 780). Then, our screening for estrogen-inducible growth factors identified that IGF1 was up-regulated remarkably by E(2). Immunoprecipitation using conditioned medium of Ishikawa cells after E(2) treatment confirmed the E(2)-induced secretion of IGF1 protein. Treatment with recombinant IGF1 stimulated cell proliferation in a dose-dependent fashion, in association with MAPK3/1 phosphorylation and up-regulation of cyclin D1 and E. These IGF1-induced responses were suppressed by treatment with MAP2K inhibitor or anti-IGF1 receptor antibody. Immunohistochemical staining confirmed the expression of activated MAPK3/1 in normal proliferative phase endometria and endometrial carcinomas, indicating the involvement of this pathway in actively proliferating endometrial tissues in vivo. These findings suggest that E(2)-induced proliferation of endometrial carcinoma cells is mediated by the MAPK3/1 pathway via autocrine stimulation of IGF1.