Nitrogen Plasma-Modified Silk Fibroin Films Promote Wettability and Airway Epithelial Cell Growth

The regulation of cell behavior by surface properties is a central principle for bioengineering tissue scaffolds. The effects of surface wettability on airway epithelial cells are particularly instrumental to functional epithelium formation but are not well understood, thus restricting bioengineered airway graft efficacy. Here, we demonstrate that nitrogen low-temperature plasma (N2 LTP) treatment enhances the surface wettability of two types of 3D-printed silk fibroin films (opaque and transparent) and promotes the growth of primary human bronchial epithelial cells (HBECs) in submerged culture over 28 days, thereby establishing a meaningful correlation between surface wettability and HBEC growth. The improved surface wettability is characterized by a reduction in water contact angle (WCA) from approximately 60° to 30°. In addition, the N2 LTP-induced surface modification is largely decoupled from the bulk chemical properties of silk fibroin films, as characterized by Fourier transform infrared and Raman spectroscopies, thus allowing independent control over surface and bulk properties. Furthermore, the N2 LTP treatment significantly increases cell density and decreases cell contour area toward the formation of a monolayer, highlighting the important role of surface wettability in mediating the adhesion and growth of HBECs. For example, following N2 LTP treatment, the cell contour area of opaque silk fibroin films on day 14 exhibited a significant decrease from 802.7 ± 53.5 to 584.4 ± 62.2 μm2 (P < 0.001), accompanied by a concurrent increase in cell density from 1111.9 ± 102.2 to 1479.3 ± 152.8 cells/mm2 (P < 0.001). Finally, both the 3D printing and the N2 LTP modification take place at room temperature, representing a mild processing strategy for devising silk fibroin scaffolds. This work offers critical insights into the nitrogen plasma modification of silk fibroin and surface-mediated behavior of epithelial cells, representing an emerging avenue for airway graft development.