Crosslink density of a biomimetic poly(HEMA)-based hydrogel influences growth and proliferation of attachment dependent RMS 13 cells

The mechanical properties of biomaterials have profound consequences on cellular and host responses, however, the underlying mechanisms remain poorly understood. Presented are findings that confirm a clear relationship between the elastic modulus, the bulk-to-bound water ratio and the adaptive attachment of attachment dependent cells. We show that biomimetic hydrogels possessing no specific integrin binding motifs but that are of lower elastic modulus and lower bulk-to-bound water ratio, preferentially support cell attachment. Anchorage-dependent human muscle fibroblasts (RMS 13) were cultured on tetraethylene glycol (TEGDA) cross-linked poly(2-hydroxyethyl methacrylate) [poly(HEMA)]-based biomimetic hydrogels containing phosphorylcholine (PC) (1 mol%) and dimethylamino amino ethyl methacrylate (DMAEMA), a 3° amine (5 mol%), as well as on silicone and agarose controls. Changes in the cross-link density (1 to 12 mol%) of the hydrogel produced a monotonic reduction in the glass transition temperature, T _g (131.8 °C at 1 mol% TEGDA to 110.4 °C at 12 mol% TEGDA), but an exponential increase in the bulk-to-bound water (4.25 at 1 mol% to 27.04 at 12 mol%) that exactly parallels the increase in elastic modulus as measured by nano-indentation AFM (152 ± 62 kPa at 1 mol% TEGDA to 1777 ± 1152 kPa at 12 mol% TEGDA). Enumeration, MTT assay and fluorescence microscopy following 4, 8 and 12 days of culture confirms that short-term cell viability and long term proliferation were favored on low cross-link density, low modulus hydrogels and that cells were preferentially attached to low cross-link density hydrogels. Bound water is central to the adaptive attachment of attachment dependent cells.