Recombinant collagen scaffolds as substrates for human neural stem/progenitor cells

Abstract

Adhesion to the microenvironment profoundly affects stem cell functions, including proliferation and differentiation, and understanding the interaction of stem cells with the microenvironment is important for controlling their behavior. In this study, we investigated the effects of the integrin binding epitopes GFOGER and IKVAV (natively present in collagen I and laminin, respectively) on human neural stem/progenitor cells (hNSPCs). To test the specificity of these epitopes, GFOGER or IKVAV were placed within the context of recombinant triple-helical collagen III engineered to be devoid of native integrin binding sites. HNSPCs adhered to collagen that presented GFOGER as the sole integrin-binding site, but not to IKVAV-containing collagen. For the GFOGER-containing collagens, antibodies against the β1 integrin subunit prevented cellular adhesion, antibodies against the α1 subunit reduced cell adhesion, and antibodies against α2 or α3 subunits had no significant effect. These results indicate that hNSPCs primarily interact with GFOGER through the α1β1 integrin heterodimer. These GFOGER-presenting collagen variants also supported differentiation of hNSPCs into neurons and astrocytes. Our findings show, for the first time, that hNSPCs can bind to the GFOGER sequence, and they provide motivation to develop hydrogels formed from recombinant collagen variants as a cell delivery scaffold. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 1363-1372, 2018.

Extracellular Matrix‐Based Strategies for Immunomodulatory Biomaterials Engineering

“…Decellularized matrices have been commercialized for clinical use and show unique immunomodulatory effects based on source tissue and physical characteristics, but these materials remain heterogeneous and subject to contamination with cell products, effectively limiting engineering methods to finely control the immune response. Single ECM materials, on the other hand, have more precise composition and thus consistent cellular responses, but require sophisticated molecular engineering tools to tune their biochemistry and immunomodulatory effects …”

“…Given the dependence of cellular interactions with collagen on its structure and exposed ligands, future studies engineering the tertiary and quaternary structure of collagens may provide further control over immune responses using these biomaterials. In addition, the design of cell interaction sites and non‐native cysteines into recombinant, full‐length human collagen III has been an approach for examining their ability to induce neural stem/progenitor cell differentiation and myofibroblast growth . Similarly, a potential route for future studies could be the use of such recombinant strategies to specifically prescribe non‐native functionality (e.g., non‐native cell interaction sites, degradation kinetics, and mechanical properties) into collagen; this approach could then examine the impact of specific molecular modifications of collagen on immune cells and enable the tuning of collagen's immunomodulatory effects, as part of the larger wound‐healing response.…”

Tailoring Collagen to Engineer the Cellular Microenvironment

“…The full‐length collagen variant genes were introduced into CEN/ARS plasmids and transformed into BYα2β2. Expression of collagen was induced, the yeast was harvested, and the resulting collagen variants were purified according to previously described protocols . Yeasts cells were mechanically disrupted using a Thermo Electron French Press and digested with pepsin.…”

“…Expression of collagen was induced, the yeast was harvested, and the resulting collagen variants were purified according to previously described protocols. [18,33] Yeasts cells were mechanically disrupted using a Thermo Electron French Press and digested with pepsin. Cell debris and digested proteins were removed by ultracentrifugation.…”

Effect of Laminin Derived Peptides IKVAV and LRE Tethered to Hyaluronic Acid on hiPSC Derived Neural Stem Cell Morphology, Attachment and Neurite Extension

“…Laminin α1 chain derived Ile-Lys-Val-Ala-Val (IKVAV) promotes neural differentiation and axon extension [24,25] in addition to altering MMP 2 and 9 expression through calcium (Ca 2+ )-dependent integrin signaling mechanisms [26]. However, IKVAV does not always support cellular attachment [27,28]. Due to this and other shortcomings, additional peptides are often used to supplement IKVAV's biological effects to achieve the desired biological response [29][30][31].…”