(723h) Elucidating the Roles of Chemical and Mechanical Stimuli in Cell Migration

Cell migration plays a critical role from morphogenesis to cell death.  For example, it plays a prominent role during gastrulation, tissue repair and in mediating the response.  In vivo, cells exhibit directed migration when presented with single or multiple stimuli that are chemical, mechanical, electrical, or optical in nature¹.  Prior reports have focused on investigating cellular migration only when a single cue is presented in the form of a gradient.  In the present study, our goal was to investigate migratory behavior when cells are presented with a choice between increasing substrate rigidity (mechanical) or higher protein concentration (chemical).  We have chosen to focus on this unique environment since it recapitulates the wound microenvironment in vivo.

Polyacrylamide (PAAM) hydrogels exhibiting chemical and mechanical profiles in opposing directions were created by photomasks and UV polymerization.  The Young’s modulus and collagen concentrations were modulated in a manner such that they increased in opposing directions.  AFM measurements were conducted on hydrated PAAM gels to calculate their elastic modulus.  The surface concentration of collagen was measured using an indirect ELISA assay².  PAAM gels were seeded with Balb/c 3T3 fibroblasts and time lapse microscopy was used to obtain phase contrast and fluorescence images.  Cellular parameters were measured using Nikon NIS Elements software.

Hydrogels were designed exhibiting mechanical gradient values ranging from 34-120kPa.  Initial studies were conducted to validate durotaxis on two different mechanical gradient profiles (119kPa/57kPa) and (95kPa/34kPa) while keeping the surface concentration of immobilized collagen constant.  We observed that greater than 85% of the cells on both mechanical gradient profiles exhibited a preference towards the rigid side of the substrate.  Hence, we concentrated on 119kPa-57kPa gels for future studies since statistically significant, higher displacement values were observed on these substrates.  Next, we designed hydrogels with opposing dual chemical and mechanical gradient profiles to investigate which cue would play a dominant role in directing cell locomotion.  The collagen concentration was increased either ~ 4-fold or 7-fold.  In dual gradient gels, 86% and 100% of cells exhibited directed migration towards the soft (57kPa) side of the gel when the collagen concentration increased 4-fold and 7-fold respectively.  A shallow (~ 4-fold) increase in immobilized collagen elicited statistically higher displacements and speeds in comparison to a steep (~ 7-fold) gradient. 

We have designed a novel dual gradient hydrogel substrate that can provide new insights into cellular locomotion.  Cellular locomotion on these interfaces demonstrated that durotaxis can be reversed when a chemical cue was presented in the opposing direction.  These results can be used to design novel biomaterials for interfacial tissue engineering.

References:

            (1)           Hale, N. A.; Yang, Y.; Rajagopalan, P. ACS Applied Materials & Interfaces 2010, 2, 2317.

            (2)        Gaudet, C.; Marganski, W. A.; Kim, S.; Brown, C. T.; Gunderia, V.; Dembo, M.; Wong, J. Y. Biophysical journal 2003, 85, 3329.