Regulation of hepatic stem/progenitor phenotype by microenvironment stiffness in hydrogel models of the human liver stem cell niche

Abstract

Human livers have maturational lineages of cells within liver acini, beginning periportally in stem cell niches, the canals of Hering, and ending in polyploid hepatocytes pericentrally and cholangiocytes in bile ducts. Hepatic stem cells (hHpSCs) in vivo are partnered with mesenchymal precursors to endothelia (angioblasts) and stellate cells, and reside in regulated microenvironments, stem cell niches, containing hyaluronans (HA). The in vivo hHpSC niche is modeled in vitro by growing hHpSC in two-dimensional (2D) cultures on plastic. We investigated effects of 3D microenvironment, mimicking the liver’s stem cell niche, on these hHpSCs by embedding them in HA-based hydrogels prepared with Kubota’s Medium (KM), a serum-free medium tailored for endodermal stem/progenitors. The KM-HA hydrogels mimicked the niches, matched diffusivity of culture medium, exhibited shear thinning and perfect elasticity under mechanical loading, and had predictable stiffness depending on their chemistry. KM-HA hydrogels, which supported cell attachment, survival and expansion of hHpSC colonies, induced transition of hHpSC colonies towards stable heterogeneous populations of hepatic progenitors depending on KM-HA hydrogel stiffness, as shown by both their gene and protein expression profile. These acquired phenotypes did not show morphological evidence of fibrotic responses. In conclusion, this study shows that the mechanical properties of the microenvironment can regulate differentiation in endodermal stem cell populations.

Introduction to In Situ Forming Hydrogels for Biomedical Applications

“…The free-standing ability of the gel is an important consideration for cell growth scaffolds. The stiffness of hydrogels has been reported to direct the differentiation of different cell types [79][80][81]. For drug delivery, hydrogels should preferentially reduce in viscosity upon injection and undergo rapid recovery upon removal of the stress to form the drug release gel depot.…”

Current Translational Challenges for Tissue Engineering: 3D Culture, Nanotechnology, and Decellularized Matrices

“…This signaling is referred to as mechanotransduction, and the response can range from instantaneous ion fluxes and biochemical pathways to gene and protein expression [54]. One important criterion to consider in designing a tissue scaffold is the moduli of local microenvironments, which can regulate hepatic stem and progenitor phenotypes, making substrate selections extremely important for any tissue experiment [55]. Measured values within native and decellularized scaffolds have indicated local differences in mechanical properties between tissue spaces and point toward innate topographical cues as relative for regulating differentiation and function of specific cell types within organs [56,57].…”

Sample, testing and analysis variables affecting liver mechanical properties: A review

“…Mechanical stiffness also contributes to driving the myofibroblastic differentiation of portal fibroblast [6] and hepatic stellate cells (HSCs) [7]. Moreover, hepatic stem and progenitor cells have also been shown to exhibit mechano-sensitivity and alter their behaviour on the basis of substrate stiffness [8]. Accurately characterising liver mechanical behaviour is thus relevant for both diagnostic purposes as well as for tissue engineering and in-vitro applications.…”