Collagen V Presented at the Cell-Substrate Interface Mimics Substrate Stiffening to Enhance Fibroblast Traction, Adhesion Turnover, and Persistent Migration

Abstract

Collagen type V (Col-V) is a minor fibrillar collagen that regulates collagen fibril assembly, and its haploinsufficiency causes classical Ehlers-Danlos syndrome, in which wound healing is delayed. Whether Col-V also changes how fibroblasts adhere to, pull on, and migrate across a collagen-presenting surface has not been tested directly. Here, we covalently coupled Col-I or Col-V to soft (2.7 kPa) silicone substrates and quantified the mechanical response of NIH/3T3 fibroblasts. On Col-V, cells generated about 1.5-fold higher traction, spread more, and formed smaller focal adhesions that each carried more force. Adhesions on Col-V assembled and disassembled faster. Single fibroblasts on Col-V migrated more slowly but more persistently, and fibroblast monolayers closed a 500 micrometer gap faster. Raising substrate stiffness to 13 kPa reproduced this phenotype on Col-I, including higher traction, larger spread area, and smaller focal adhesions, indicating that Col-V presentation produces an adhesion state that otherwise requires a stiffer substrate. In separate reconstituted collagen hydrogels, co-assembly with 10 to 20% Col-V yielded thinner, straighter fibrils, and 20% Col-V increased the indentation modulus of the gel. These results identify the collagen type presented at the cell-material interface as a design parameter that can partly substitute for bulk stiffness in controlling fibroblast traction and migration, and they suggest Col-V as a tunable component of collagen-based biomaterials.