TNFα induces NFκB/p50 in association with the growth and morphogenesis of normal and transformed rat mammary epithelial cells

Abstract

In contrast to the cytotoxic or cytostatic effect of TNFα on many breast cancer cell lines, TNFα stimulates growth and morphogenesis of normal rat mammary epithelial cells (MEC). The present studies were carried out to determine whether there are intrinsic differences between normal and malignant MEC which may explain the differing responsiveness to TNFα. Freshly isolated rat MEC organoids from normal mammary gland or 1-methyl-1-nitrosourea-induced mammary tumors were treated with TNFα for 21 days. Unexpectedly, TNFα stimulated growth and morphogenesis of both normal and transformed MEC in primary culture, although in transformed cells its effects were delayed and the majority of the colonies were histologically abnormal, with multiple cell layers and no lumen. Since NFκB is a key mediator of TNFα action and has been implicated in carcinogenesis, the expression of the p50, p52, p65, and c-rel NFκB proteins in normal and transformed MEC was determined. Expression of p52 was significantly reduced in tumor cells, and p50 was absent, although its putative precursor, p105 was abundant. There were no changes in the levels of p65 or c-rel. TNFα induced a pronounced and sustained increase of a p50 homodimeric NFκB/DNA complex in both normal and transformed MEC. However, in transformed MEC, NFκB binding was initially undetectable but then increased in response to TNFα. Thus, NFκB expression and DNA binding activity are altered during mammary carcinogenesis. In addition, the significant increase in NFκB/p50 DNA-binding was temporally coincident with TNFα-induced growth and morphogenesis, suggesting that it may play a significant role in both normal development and carcinogenesis.