Mouse Fap Stable Cell Line - TrampC2

Alterations towards a permissive stromal microenvironment provides important clues for tumor growth, invasion, and metastasis. In this study, fibroblast activation protein (FAP), a serine protease selectively produced by tumor-associated fibroblasts in more than 90% of epithelial tumors, was used as a platform to study tumor-stroma interactions. The researchers utilized a FAP-overexpressing fibroblast cell line to synthesize an in vivo-like three-dimensional (3D) matrix system, which was used as a matrix landscape to study matrix-induced cancer cell behavior.

The researchers observed that FAP remodeled the ECM by modulating protein levels as well as increasing the levels of fibronectin and collagen fiber organization. FAP-dependent changes in ECM structure/composition promoted tumor invasion along a characteristic parallel fiber orientation, as demonstrated by enhanced directionality and velocity of pancreatic cancer cells on FAP+ stroma. This phenotype could be reversed by inhibiting FAP enzymatic activity during matrix production, resulting in ECM disorganization and impeding tumor invasion. Furthermore, the tumor invasion phenotype induced by FAP+ stroma was mediated by β1-integrin/FAK.

To establish stable FAP-expressing fibroblasts, the mouse fap gene was cloned under the Tet inducible system. FAP expression was induced within 24 hours after Dox treatment, and its expression was clearly maintained for at least 10 days (Figure 1A). Parental NIH-3T3 cells and FAP+ fibroblasts cultured in the absence of Dox did not show detectable FAP induction, so these fibroblasts could be used as negative controls. Indeed, ECM fibers produced by FAP+ fibroblasts were arranged in parallel patterns compared to FAP- matrices (Figure 1B). These patterns were reminiscent of tumor-associated patterns previously observed in vitro. Furthermore, by measuring the relative orientation angles of the fibers, the organization level of the various fibronectin fibers produced by the fibroblasts could be quantified. The average percentage of parallel fibers with an orientation within ±100 of the pattern angle was determined using MetaMorph software. The measured percentages were 27% and 45% for FAP- and FAP+ 3D ECM, respectively.

Figure 1. Matrix fiber organization is increased in FAP + matrices. (Lee H O, et al. 2011)