Regulation of Epithelial-to-Mesenchymal Transition Using Biomimetic Fibrous Scaffolds

Abstract

Epithelial-to-mesenchymal transition (EMT) is a well-studied biological process that takes place during embryogenesis, carcinogenesis and tissue fibrosis. During EMT, the polarized epithelial cells with a cuboidal architecture adopt an elongated fibroblast-like morphology. This process is accompanied by the expression of many EMT-specific molecular markers. While the molecular mechanism leading to EMT has been well established, the effects of matrix topography and microstructure have not been clearly elucidated. Synthetic scaffolds mimicking the mesh-like structure of the basement membrane with an average fiber diameter of 0.5 μm and 5 μm were produced via electrospinning of poly(ε-caprolactone) (PCL) and were used to test the significance of fiber diameter on EMT. Cell-adhesive peptide motifs were conjugated to the fiber surface to facilitate cell attachment. Madin-Darby Canine Kidney (MDCK) cells grown on these substrates showed distinct phenotypes. On 0.5 μm substrates, cells grew as compact colonies with an epithelial phenotype. On 5 μm scaffolds, cells were more individually dispersed and appeared more fibroblastic. Upon addition of hepatocyte growth factor (HGF), an EMT inducer, cells grown on the 0.5 μm scaffold underwent pronounced scattering, as evidenced by the alteration of cell morphology, localization of focal adhesion complex, weakening of cell-cell adhesion, and upregulation of mesenchymal markers. By contrast, HGF did not induce a pronounced scattering of MDCK cells cultured on the 5.0 μm scaffold. Collectively, our results show that the alteration of the fiber diameter of proteins found in the basement membrane may create enough disturbances in epithelial organization and scattering that might have important implications in disease progression.

The impact of mir200 on extracellular matrix topography-guided epithelial-to-mesenchymal transition of prostate cancer cells

“…Saha et al [7] investigated the role of ECM fiber alignment on breast cancer EMT and further metastatic invasion using PCL fibers and showed that breast cancer cells increased their TGF-Beta 1 levels on aligned fibers as opposed to flat topographies. Similarly, Ravikrishnan et al [15] created an RGD functionalized fiber topography and investigated the role of fiber diameter on kidney epithelial cell (MDCK) EMT. They showed that the large diameter fibers foster MDCK cell EMT whereas small diameters promote epithelial cell cluster formation and preservation of epithelial morphology of MDCK cells.…”

“…[15] Briefly, PCL polymer solution was prepared and loaded into a syringe pump. For 5 µm diameter fibers 15% PCL dissolved in chloroform, and for 0.5 µm fibers 12% PCL dissolved in 7:3 DCM:DMF were used.…”

“…The PCL fiber mats were further conjugated with RGD (Arginine-Glycine and Aspartic acid) using surface amination and further crosslinking in the presence of a difunctional linker (sulfo-SMCC) as detailed before. [15] The RGD functional PCL fiber mats were then sterilized for further cell culture seeding. Flat PCL topographies created by spun were coated onto glass slides by 5% w/v PCL in chloroform solution and further annealed at 60 ̊C in order to fix onto the glass slides.…”

“…Using electrospinning technique polymeric fibers with controlled diameter can be reproduced which are structurally identical to the collagen fibers in prostate cancer ECM. [15] In this study, we developed an in vitro cancer model by creating two different fiber structures reminiscent of healthy and metastatic prostate microenvironment. The synthetic fiber structures were built to understand the effect of ECM fiber diameter on EMT behavior of LNCaP prostate carcinoma cell line.…”

50th Anniversary Perspective: Polymeric Biomaterials: Diverse Functions Enabled by Advances in Macromolecular Chemistry

“…207 Fiber-like scaffolds, designed to mimic fibrillar ECM proteins, have been shown to regulate cell migration, proliferation, and epithelial-to-mesenchymal transition (Figure 8A). 208, 209 Multiple processing strategies have been applied to develop microstructured materials, including photopatterning 210 and selective degradation, 211 as mentioned above. Such approaches yield materials that exhibit improved cell attachment and migration, 212 as well as enhanced tissue infiltration into scaffolds.…”