Background: Transforming growth factor beta (TGFβ) is important for the morphogenesis and secretory function of the mammary gland. It is one of the main activators of the epithelial-mesenchymal transition (EMT), a process important for tissue remodeling and regeneration. It also provides cells with the plasticity to form metastases during tumor progression. Noncancerous and cancer cells respond differently to TGFβ. However, knowledge of the cellular signaling cascades triggered by TGFβ in various cell types is still limited.
Methods: MCF10A (noncancerous, originating from fibrotic breast tissue) and MCF7 (cancer, estrogen receptor-positive) breast epithelial cells were treated with TGFB1 directly or through conditioned media from stimulated cells. Transcriptional changes (via RNA-seq) were assessed in untreated cells and after 1-6 days of treatment. Differentially expressed genes were detected with DESeq2 and the hallmark collection was selected for gene set enrichment analysis.
Results: TGFB1 induces EMT in both the MCF10A and MCF7 cell lines but via slightly different mechanisms (signaling through SMAD3 is more active in MCF7 cells). Many EMT-related genes are expressed in MCF10A cells at baseline. Both cell lines respond to TGFB1 by decreasing the expression of genes involved in cell proliferation: through the repression of MYC (and the protein targets) in MCF10A cells and the activation of p63-dependent signaling in MCF7 cells (CDKN1A and CDKN2B, which are responsible for the inhibition of cyclin-dependent kinases, are upregulated). In addition, estrogen receptor signaling is inhibited and caspase-dependent cell death is induced only in MCF7 cells. Direct incubation with TGFB1 and treatment of cells with conditioned media similarly affected transcriptional profiles. However, TGFB1-induced protein secretion is more pronounced in MCF10A cells; therefore, the signaling is propagated through conditioned media (bystander effect) more effectively in MCF10A cells than in MCF7 cells.
Conclusions: Estrogen receptor-positive breast cancer patients may benefit from high levels of TGFB1 expression due to the repression of estrogen receptor signaling, inhibition of proliferation, and induction of apoptosis in cancer cells. However, some TGFB1-stimulated cells may undergo EMT, which increases the risk of metastasis.
Keywords: Bystander effect; Cell cycle; Cell death; EMT; Estrogen signaling; MCF10A; MCF7; Mammary epithelial cells; TGFβ signaling.
Transforming growth factor beta (TGFβ) is a multifaceted cytokine that controls numerous physiological and pathological processes during development and carcinogenesis. Its best-known function is the activation of epithelial–mesenchymal transition (EMT), a process crucial for tissue remodeling and regeneration. During EMT, epithelial cells lose their connections to adjacent cells and acquire mesenchymal characteristics, such as migratory ability. Compared with cancer cells, normal (nontumorigenic) cells usually respond differently to TGFβ stimulation. Typically, TGFβ inhibits the proliferation of epithelial cells and may promote cell death. In cancer cells, TGFβ often promotes tumor progression. TheTGFβ-mediated induction of EMT increases cell mobility, which is associated with the formation of metastases and tumor invasion.In this work, we compared the transcriptional response of noncancerous (MCF10A) and cancerous (MCF7; estrogen receptor-positive) breast epithelial cells to direct stimulation by TGFB1 and its indirect effect through conditioned media. Some of TGFB1-induced changes, including inhibition of proliferation and induction of EMT, were similar in both cell lines. However, these changes were associated with different upstream signaling pathways. Other changes were more specific, such as disruption of estrogen-related signaling or induction of cell death in MCF7 cells. Direct incubation with TGFB1 and treatment of cells with conditioned media similarly affected target cells, indicating the presence of a bystander effect. Moreover, transcript profiling by RNA-seq revealed that the TGFβ signaling pathway is already active in untreated MCF10A cells, which may be due to their origination from a fibrotic lesion.
© 2024. The Author(s).