The extracellular matrix presents spatially varying physical cues that can influence cell behavior in many processes. Physical gradients within hydrogels that mimic the heterogenous mechanical microenvironment are useful to study the impact of these cues on cellular responses. Therefore, simple and reliable techniques to create such gradient hydrogels are highly desirable. This work demonstrates the fabrication of stiffness gradient Gellan gum (GG) hydrogels by applying a temperature gradient across a microchannel containing hydrogel precursor solution. Thermophoretic migration of components within the precursor solution generates a concentration gradient that mirrors the temperature gradient profile, which translates into mechanical gradients upon crosslinking. Using this technique, GG hydrogels with stiffness gradients ranging from 20 to 90 kPa over 600µm are created, covering the elastic moduli typical of moderately hard to hard tissues. MC3T3 osteoblast cells are then cultured on these gradient substrates, which exhibit preferential migration and enhanced osteogenic potential toward the stiffest region on the gradient. Overall, the thermophoretic approach provides a non-toxic and effective method to create hydrogels with defined mechanical gradients at the micron scale suitable forin vitrobiological studies and potentially tissue engineering applications.
Keywords: hydrogels; microfluidics; osteoblasts; stiffness gradients; thermophoresis.
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