Studying adhesion and migration of HESC cells on linear stiffness gradient hydrogels

ALEJANDRA M FERNANDEZ; LAURA FERNANDEZ; E ELIZABETH SAMANIEGO ONOFRE; ROSSANA RAMHORST; SILVINA PONCE DAWSON; LORENA SIGAUT; LIA I PIETRASANTA

Córdoba

Congreso; LI Reunión Anual de la Sociedad Argentina de Biofísica; 2023

Sociedad Argentina de Biofísica

In many processes, such as embryonic development, tumor progression, and immune response against pathogens, we observed durotaxis, in which cells prefer moving towards areas of increased rigidity. Cells can actively sense and detect the rigidity of their environment through focal adhesions, which guide their movement. To study cell adhesion and migration with respect to the rigidity of the environment, we produced polyacrylamide hydrogels with a linear stiffness gradient using polyacrylamide (PAA) gels, following the method described by Hadden et al. (2017). This substrate is nontoxic, biocompatible, cost-effective, and highly reproducible. Once the hydrogel with the gradient was prepared, the substrate surface was functionalized with the extracellular matrix protein fibronectin, and human endometrial stromal cells (HESC) were seeded onto it.The mechanical properties of the substrate and cells were determined using atomic force microscopy (AFM) and force spectroscopy (FS). The presence of the stiffness gradient was confirmed by fitting the approximation curves from hundreds of force-distance curves and determining the Young´s modulus at various points along the hydrogel. As the indenter used has a blunt pyramidal geometry, we applied the correction model described in the work of Lin et al. (2007), which adapts the pointed pyramidal tip model [3]. Moreover, we examined the adhesion and migration processes of HESC cells to a substrate with a stiffness gradient using optical microscopy. The analysis of the cell trajectories allowed us to calculate the instantaneous speed and direction of cells from multiple regions of the substrate. References[1] Hadden W. J. et al. (2017). PNAS, 114: 5647-5652.[2] Lin D. C. et al. (2007). J. Biomech Eng., 129: 430-440. [3] Bilodeau G. (1992). ASME J. Appl. Mech., 59: 519–523.Acknowledgments CONICET, UBA, ANPCyT.