Type I collagen gels for assessing the combined effects of ligand concentration-dependent elasticity and fibril density on cells

Abstract Type I collagen (Col-I) is the most abundant fibrous protein in the extracellular matrix (ECM); its densification is involved in conditions including cancer, fibrosis, vascular diseases, and aging. Increased Col-I concentrations lead to increased ECM fibril density and stiffening; therefore, the effects of such changes in the microenvironment of cells require examination. Three types of Col-I gels were developed by introducing reagent-free radiation-induced crosslinking, while adjusting the distance between the Col-I monomers or fibrils. The adhesion area of HeLa human cervical cancer epithelial cells, human induced pluripotent stem (hiPS) cells, and 3T3-Swiss albino mouse embryonic fibroblasts, tended to decrease with increasing fibril density when the ligand concentration-dependent elasticity ( C - E ) was constant. In the absence of fibrils, the adhesion area of HeLa and 3T3-swiss cells increased with increasing C - E , whereas hiPS cells clustered regardless of C - E . When C - E and the fibril density increased simultaneously, the adhesion area of HeLa and 3T3-swiss cells remained unchanged due to the two opposing effects. Our results highlight the importance of evaluating the combined effects of the compositional, mechanical, and topographical cues of Col-I on cells. The developed Col-I gels will contribute to elucidating the role of Col-I in development, differentiation, regeneration, disease, and aging. Highlights Col-I concentration-dependent elasticity ( C - E ) and fibril density were separated. Adhesion areas of cells decreased with increasing fibril density at constant C - E . Adhesion areas of cells increased with increasing C - E in absence of Col-I fibrils. Adhesion areas remained unchanged when both C - E and fibril density increased. Structural, mechanical and topographical cues work in combination to affect cells. Graphical Abstract