Collagen I Fibrous Substrates Modulate the Proliferation and Secretome of Estrogen Receptor-Positive Breast Tumor Cells in a Hormone-Restricted Microenvironment

Abstract

The fibril orientation of type I collagen has been shown to contribute to tumor invasion and metabolic changes. Yet, there is limited information about its impact on tumor cells' behavior in a restrictive growth environment. Restrictive growth environments are generated by the inhibition of a proliferation stimulus during therapy or as an inflammatory response to suppress tumor expansion. In this study, the impact of a type I collagen matrix orientation and fibrous architecture on cell proliferation and response to estrogen receptor (ER) therapy were examined using estrogen-dependent breast tumor cells (MCF-7 and T-47D) cultured in a hormone-restricted environment. The use of hormone-free culture media, as well as pharmacological inhibitors of ER, Tamoxifen, and Fulvestrant, were investigated as hormone restrictive conditions. Examination of cultures at 72 h showed that tumor cell proliferation was significantly stimulated (1.8-fold) in the absence of hormones on collagen fibrous substrates, but not on polycaprolactone fibrous substrates of equivalent orientation. ER inhibitors did not suppress cell proliferation on collagen fibrous substrates. The examination of reporter cells for ER signaling showed a lack of activity, thus confirming a shift toward an ER-independent proliferation mechanism. Examination of two selective inhibitors of α2β1 and α1β1 integrins showed that cell proliferation is suppressed in the presence of the α2β1 integrin inhibitor only, thereby indicating that the observed changes in tumor cell behavior are caused by a combination of integrin signaling and/or an intrinsic structural motif that is uniquely present in the collagen fibrils. Adjacent coculture studies on collagen substrates showed that tumor cells on collagen can stimulate the proliferation of cells on tissue culture plastic through soluble factors. The magnitude of this effect correlated with the increased surface anisotropy of the substrate. This sensing in fibril orientation was further supported by a differential expression pattern of secreted proteins that were identified on random and aligned orientation substrates. Overall, this study shows a new role for electrospun collagen I fibrous substrates by supporting a shift toward an ER-independent tumor cell proliferation mechanism in ER+ breast tumor cells.