Amphiregulin acts as an autocrine growth factor in two human polarizing colon cancer lines that exhibit domain selective EGF receptor mitogenesis

Abstract

Summary Colonic enterocytes, like many epithelial cells in vivo, are polarized with functionally distinct apical and basolateral membrane domains. The aims of this study were to characterize the endogenous epidermal growth factor (EGF)-like ligands expressed in two polarizing colon cancer cell lines, HCA-7 Colony 29 (HCA-7) and Caco-2, and to examine the effects of cell polarity on EGF receptor-mediated mitogenesis. HCA-7 and Caco-2 cells were grown on plastic, or as a polarized monolayer on Transwell filters. Cell proliferation was measured by 3 H-thymidine incorporation and EGF receptor (EGFR) binding was assessed by Scatchard analysis. EGFR ligand expression was determined by Northern blot analysis, reverse transcription polymerase chain reaction, metabolic labelling and confocal microscopy. We found that amphiregulin (AR) was the most abundant EGFR ligand expressed in HCA-7 and Caco-2 cells. AR was localized to the basolateral surface and detected in basolateral-conditioned medium. Basolateral administration of neutralizing AR antibodies significantly reduced basal DNA replication. A single class of high-affinity EGFRs was detected in the basolateral compartment, whereas the apical compartment of polarized cells, and cells cultured on plastic, displayed two classes of receptor affinity. Basolateral administration of transforming growth factor alpha (TGF-α) or an EGFR neutralizing antibody also resulted in a dose-dependent stimulation or attenuation, respectively, of DNA replication. However, no mitogenic response was observed when these agents were added to the apical compartment or to confluent cells cultured on plastic. We conclude that amphiregulin acts as an autocrine growth factor in HCA-7 and Caco-2 cells, and EGFR ligand-induced proliferation is influenced by cellular polarity.

Protease-activated Receptor 2 in Colon Cancer

“…Moreover, we could measure the increase of TGF-␣ concentration in the culture medium of PAR2 agonist-stimulated HT-29 cells. Amphiregulin, another EGF-R ligand, has been shown to also be expressed in colon cancer (28,29). Involvement of amphiregulin in PAR2-mediated EGF-R transactivation in HT-29 cells is unlikely for the following reasons: (i) incubation of HT-29 cells with neutralizing antibody against TGF-␣ in the culture medium completely blocks PAR2-induced cell proliferation; and (ii) neutralizing antibodies against bioactive amphiregulin did not alter the HT-29 cell proliferation induced by PAR2 agonists.…”

Inhibition of Apoptosis by Amphiregulin via an Insulin-like Growth Factor-1 Receptor-dependent Pathway in Non-small Cell Lung Cancer Cell Lines

“…AR, an EGF-related growth factor produced by epithelial cells, functions as an autocrine/paracrine cell proliferation factor (42,43). Several lines of evidence suggest that AR is an autocrine growth factor in human carcinoma cells (44 -51) and tumorigenicity of a human breast epithelial cell line is abolished by treatment with AR antisense (49).…”

Epidermal growth factor receptor mutation type iii transfected into a small cell lung cancer cell line is predominantly localized at the cell surface and enhances the malignant phenotype

“…4 In human malignancies overexpression or amplification of the EGFR has been shown to have an important role as activation of the EGFR initiates a cascade of phosphorylations that results in DNA synthesis and cell replication. [5][6][7][8] Therefore, presence or overexpression of the EGFR could give malignant cells a growth advantage. Furthermore, it has been shown that presence of the EGFR results in invasive tumors, higher stages of disease or disease progression.…”