Guiding Oligodendrocyte Precursor Cell Maturation With Urokinase Plasminogen Activator-Degradable Elastin-like Protein Hydrogels.

Demyelinating injuries and diseases, like multiple sclerosis, affect millions of people worldwide. Oligodendrocyte precursor cells (OPCs) have the potential to repair demyelinated tissues because they can both self-renew and differentiate into oligodendrocytes (OLs), the myelin producing cells of the central nervous system (CNS). Cell-matrix interactions impact OPC differentiation into OLs, but the process is not fully understood. Biomaterial hydrogel systems help to elucidate cell-matrix interactions because they can mimic specific properties of native CNS tissues in an in vitro setting. We investigated whether OPC maturation into OLs is influenced by interacting with a urokinase plasminogen activator (uPA) degradable extracellular matrix (ECM). uPA is a proteolytic enzyme that is transiently upregulated in the developing rat brain, with peak uPA expression correlating with an increase in myelin production in vivo. OPC-like cells isolated through the Mosaic Analysis with Double Marker technique (MADM OPCs) produced low-molecular-weight uPA in culture. MADM OPCs were encapsulated into two otherwise similar elastin-like protein (ELP) hydrogel systems: one that was uPA degradable and one that was nondegradable. Encapsulated MADM OPCs had similar viability, proliferation, and metabolic activity in uPA degradable and nondegradable ELP hydrogels. Expression of OPC maturation-associated genes, however, indicated that uPA degradable ELP hydrogels promoted MADM OPC maturation although not sufficiently for these cells to differentiate into OLs.

Impact of Elastin-like Protein Temperature Transition on PEG-ELP Hybrid Hydrogel Properties.

Biomaterial hydrogels are made by cross-linking either natural materials, which exhibit inherent bioactivity but suffer from batch-to-batch variations, or synthetic materials, which have a well-defined chemical structure but usually require chemical modification to exhibit bioactivity. Recombinant engineered proteins bridge the divide between natural and synthetic materials because proteins incorporate bioactive domains within the biopolymer backbone and have a well-defined amino acid structure and sequence. Recombinant engineered elastin-like proteins (ELPs) are modeled from the native tropoelastin sequence. ELPs are composed of repeating VPGxG penta-peptide sequences, where x is any guest residue except proline. ELPs undergo a lower critical solution temperature (LCST) transition above which they aggregate into a coacervate phase in aqueous solution. Here we show that the LCST transition impacts hydrogel microarchitecture which may serve as a useful design feature in engineering ELP-based hydrogels. We investigate how the ELP LCST transition contributes to the properties of hybrid poly(ethylene glycol) (PEG) and ELP (PEG-ELP) hydrogels. PEG-ELP hydrogels gelled below the LCST have a homogeneous distribution of ELP, while gelling above the LCST results in the formation of spherical ELP-rich regions within the bulk hydrogel. The ELP-rich microarchitecture is maintained when an amine-reactive cross-linker is incorporated during the gelation process. The formation of ELP-rich regions reduces PEG-ELP hydrogel bulk stiffness and increases optical density. Our characterizations of hydrogels created by using the LCST transition provide design criteria for incorporating microscale features. This may be a useful technique in understanding the role of localized bioactivity at the microscale level within hydrogel systems.