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Recent studies have highlighted the role of external biophysical cues on cell morphology and function. In particular, substrate geometry and rigidity in two dimensions has been shown to impact cell growth, death, differentiation, and motility. Knowledge of how these physical cues affect cell function in three dimensions is critical for successful development of novel regenerative therapies. In this work, the effect of discrete micromechanical cues in three-dimensional (3D) system on cell proliferation, gene expression, and extracellular matrix synthesis was investigated. Poly(ethylene glycol) dimethacrylate hydrogel microrods were fabricated using photolithography and suspended in gel to create a 3D culture with microscale cues of defined mechanical properties in the physiological range (2-50 kPa). These microrods significantly affected fibroblast proliferation, matrix production, and gene expression. Cultures with stiff microrods reduced fibroblast proliferation and downregulated expression of key extracellular matrix proteins involved in scar tissue formation. In addition, the contractility marker alpha smooth muscle actin and adhesion molecule integrin alpha3 were also significantly downregulated. Cultures with soft microrods had no significant difference on fibroblast proliferation and expression of Cyclin D1, alpha smooth muscle actin, and integrin alpha3 compared to cultures with no microrods. Here, we present a new platform of potentially injectable microrods with tunable elasticity; in addition, we show that cell proliferation and gene expression are influenced by discrete physical cues in 3D.

Microtopographical Cues in 3D Attenuate Fibrotic Phenotype and Extracellular Matrix Deposition: Implications for Tissue Regeneration