Bioactive poly(ethylene glycol) hydrogels to recapitulate the HSC niche and facilitate HSC expansion in culture

Abstract

Hematopoietic stem cells (HSCs) have been used therapeutically for decades, yet their widespread clinical use is hampered by the inability to expand HSCs successfully in vitro. In culture, HSCs rapidly differentiate and lose their ability to self-renew. We hypothesize that by mimicking aspects of the bone marrow microenvironment in vitro we can better control the expansion and differentiation of these cells. In this work, derivatives of poly(ethylene glycol) diacrylate hydrogels were used as a culture substrate for hematopoietic stem and progenitor cell (HSPC) populations. Key HSC cytokines, stem cell factor (SCF) and interferon-γ (IFNγ), as well as the cell adhesion ligands RGDS and connecting segment 1 were covalently immobilized onto the surface of the hydrogels. With the use of SCF and IFNγ, we observed significant expansion of HSPCs, ∼97 and ∼104 fold respectively, while maintaining c-kit(+) lin(-) and c-kit(+) Sca1(+) lin(-) (KSL) populations and the ability to form multilineage colonies after 14 days. HSPCs were also encapsulated within degradable poly(ethylene glycol) hydrogels for three-dimensional culture. After expansion in hydrogels, ∼60% of cells were c-kit(+), demonstrating no loss in the proportion of these cells over the 14 day culture period, and ∼50% of colonies formed were multilineage, indicating that the cells retained their differentiation potential. The ability to tailor and use this system to support HSC growth could have implications on the future use of HSCs and other blood cell types in a clinical setting.

Endosteal-like extracellular matrix expression on melt electrospun written scaffolds

“…Tissue-engineered approaches to the HSC niche are frequently based on hydrogels [32,[37][38][39][40]. However, the use of solid scaffolds with higher mechanical properties allows more complex approaches.…”

Challenges and Opportunities to Harnessing the (Hematopoietic) Stem Cell Niche

“…Presentation of extracellular components or growth factors via surface immobilization of material substrates have revealed that HSC-matrix interactions significantly influence HSC adhesion [59], spreading [31], proliferation [60, 61], and differentiation [62]. HSCs show preferential adhesion to several extracellular matrix (ECM) proteins and proteoglycans [28, 59].…”

“…When whole bone marrow cells were cultured on plastic culture plates with physically adsorbed tropoelastin, the number of HSCs in the population significantly increased compared to the negative control, suggesting that changes in surface elasticity contributed to the observed HSC expansion [60]. Similarly, hematopoietic stem and progenitor cells (HSPCs) cultured on polyethylene glycol diacrylate (PEGDA) gel surfaces modified with SCF, IFNγ, and RGD showed considerably more expansion compared to unmodified counterparts, emphasizing the importance of sustained growth factor presentation in HSC growth [61]. In another study, magnetic microbeads functionalized with notch ligand delta-like ligand 4 (DLL4) via streptavidin-biotin conjugation facilitated HSC differentiation to T cell precursors when they were added to HSC co-cultures with stromal cells [62].…”

“…For instance, immobilized biomolecules mimicking membrane-bound isoforms could amplify desired output over soluble isoforms by providing sustained ligand-receptor interactions leading to prolonged receptor activation [69, 71]. As such, immobilized stem cell factor (SCF; a key cytokine known to be involved in HSC survival and maintenance [72]) was shown to enhance adhesion and proliferation activity of primary HSCs and hematopoietic cell lines compared to soluble SCF [61, 71, 73–75]. Additionally, temporal regulation of biomolecular presentation (e.g., availability, diffusive transport) has been found to be important in modulating HSC activities [50, 70, 76–78].…”

“…Here, we describe several recent developments in 3D constructs for recreating HSC bone marrow niche, which have been critically linked to HSC activities including adhesion [92], proliferation [61, 73, 92–94], and engraftment [92]. A wide range of synthetic 3D HSC niches utilize co-culture with other cells, typically a variation of mesenchymal or stromal cells, in order to promote increased cell survivability and growth support [1].…”