A practical guide to hydrogels for cell culture

Abstract

There is growing appreciation of the role that the extracellular environment plays in regulating cell behavior. Mechanical, structural, and compositional cues, either alone or in concert, can drastically alter cell function. Biomaterials, and particularly hydrogels, have been developed and implemented to present defined subsets of these cues for investigating countless cellular processes as a means of understanding morphogenesis, aging, and disease. Although most scientists concede that standard cell culture materials (tissue culture plastic and glass) do a poor job of recapitulating native cellular milieus, there is currently a knowledge barrier for many researchers in regard to the application of hydrogels for cell culture. Here, we introduce hydrogels to those who may be unfamiliar with procedures to culture and study cells with these systems, with a particular focus on commercially available hydrogels.

Figure 1. Cell culture atop 2D hydrogels

(a) Conventional 2D culture on super-physiologically stiff plastic or glass substrates leads to cells displaying aberrant phenotypes. (b) Culturing cells on 2D hydrogel films has some of the same disadvantages as conventional methods, but permits user-defined control of the substrate stiffness and adhesive ligand presentation. Human mesenchymal stem cells (MSCs) cultured on increasingly stiff 2D substrates display increasing spread area. From left: 1 kPa polyacrylamide (PA), 11 kPa PA, 34 kPa PA, and glass (~ GPa). Scale bar: 10 μm. Images modified from with permission. (c) Substrate stiffness (y-axis) and adhesive ligand type (x-axis) combine to regulate MSC morphology. Human MSCs spread more with increasing stiffness, but cells on laminin-coated hydrogels are smaller compared to other ECM protein coatings. Images modified from with permission. Scale bar: 50 μm.

Figure 2. 3D hydrogels for cell culture

(a) 3D hydrogels can be engineered to present a more realistic microenvironment to cells. Hydrogel design variables are indicated. (b) Mouse MSCs cultured in 3D alginate hydrogels display rounded morphology regardless of substrate stiffness. Left panel: 5 kPa, right panel: 110 kPa. Images modified from with permission. (c) Bovine dermal fibroblasts encapsulated in 3D collagen hydrogels spread at low stiffness (< 1 kPa). Image modified from with permission. (d) Human MSCs cultured in a hyaluronic acid (HA) hydrogel are restricted from spreading regardless of substrate stiffness (shown here ~ 4 kPa). Image modified from with permission. (e) Human MSCs cultured within a HA hydrogel with equivalent stiffness to (d) but crosslinked with MMP-degradable crosslinkers permits cells to locally remodel their environment, generate tractions, and spread. Image modified from with permission. (f) Human foreskin fibroblast spreading and migration speed is influenced by collagen fibril size. Image modified from with permission. Scale bars: 10 μm.