Signal Crosstalk Promoted Proliferative Lesions in Mouse Mammary Glands As a Consequence of ET-1 Overexpression

Abstract

Objective: As a distinct cellular signaling model, a receptor crosstalk between G protein-coupled receptors (GPCR) and epidermal growth factor receptor (EGFR) has been demonstrated in various in vitro studies. In addition, recent in vitro studies had focused on the signaling pathways of endothelin-1 (ET-1) in the pathophysiology of cancer. Accordingly, a growing interest in the analysis of the receptor crosstalk between ET-1 receptors and EGFR and functional consequences of EGFR activation of proliferative diseases evoked us to the analysis of this phenomenon in vivo. Materials and Methods: We performed a comparative study between ET-1 transgenic mice and control mice during the late pregnancy (n=7), early lactation (n=6) the mid of lactation (n=10) and involution day 14 (n=7) periods. Hematoxylin and eosin (HE)- stained parallel sections from mammary glands were microscopically examined. The key signal proteins (ETAR, ETBR, ERK1/2, pEGFR) in transactivation of EGFR were analyzed employing Western blot techniques. Genes (amphiregulin, TGFa, EGF, HB-EGF, ADAM 17) known to play an important role in these activities were analyzed using real-time PCR (RT-PCR) techniques. Results: ET-1 transgenic mice exhibited hyperproliferative lesions (lactational hyperplasia) during the middle of the lactation period. Our RT-PCR analyses showed a prominent up regulation of amphiregulin and ADAM 17 in ET-1 transgenic mice. Moreover, we found higher EGFR and ERKs activations in the transgenic mammary glands. Conclusion: This study highlights a causative effect of upregulated ET-1 gene expression on the induction of proliferative lesions via EGFR transactivation in mammary glands. Further, ET-1 overexpression induced an upregulation of amphiregulin and ADAM17 expressions in the transgenic mammary glands. These results suggests that the enhanced ET-1 gene expression and its receptors might have a crucial role in proliferative diseases maintaining EGFR activation.