Engineering of a Microscale Niche for Pancreatic Tumor Cells Using Bioactive Film Coatings Combined with 3D-Architectured Scaffolds.

Two-photon polymerization has recently emerged as a promising technique to fabricate scaffolds for three-dimensional (3D) cell culture and tissue engineering. Here, we combined 3D-printed microscale scaffolds fabricated using two-photon polymerization with a bioactive layer-by-layer film coating. This bioactive coating consists of hyaluronic acid and poly(l-lysine) of controlled stiffness, loaded with fibronectin and bone morphogenic proteins 2 and 4 (BMP2 and BMP4) as matrix-bound proteins. Planar films were prepared using a liquid handling robot directly in 96-well plates to perform high-content studies of cellular processes, especially cell adhesion, proliferation, and BMP-induced signaling. The behaviors of two human pancreatic cell lines PANC1 (immortalized) and PAN092 (patient-derived cell line) were systematically compared and revealed important context-specific cell responses, notably in response to film stiffness and matrix-bound BMPs (bBMPs). Fibronectin significantly increased cell adhesion, spreading, and proliferation for both cell types on soft and stiff films; BMP2 increased cell adhesion and inhibited proliferation of PANC1 cells and PAN092 on soft films. BMP4 enhanced cell adhesion and proliferation of PANC1 and showed a bipolar effect on PAN092. Importantly, PANC1 exhibited a strong dose-dependent BMP response, notably for bBMP2, while PAN092 was insensitive to BMPs. Finally, we proved that it is possible to combine a microscale 3D Ormocomp scaffold fabricated using the two-photon polymerization technique with the bioactive film coating to form a microscale tumor tissue and mimic the early stages of metastatic cancer.