Integration of Hydrogel Microparticles With Three-Dimensional Liver Progenitor Cell Spheroids

Abstract

The study of the liver progenitor cell microenvironment has demonstrated the important roles of both biochemical and biomechanical signals in regulating the progenitor cell functions that underlie liver morphogenesis and regeneration. While controllable twodimensional in vitro culture systems have provided key insights into the effects of growth factors and extracellular matrix composition and mechanics on liver differentiation, it remains unclear how microenvironmental signals may differentially affect liver progenitor cell responses in a three-dimensional (3D) culture context. In addition, there have only been limited efforts to engineer 3D culture models of liver progenitor cells through the tunable presentation of microenvironmental stimuli. We present an in vitro model of 3D liver progenitor spheroidal cultures with integrated polyethylene glycol hydrogel microparticles for the internal presentation of modular microenvironmental cues and the examination of the combinatorial effects with an exogenous soluble factor. In particular, treatment with the growth factor TGFβ1 directs differentiation of the spheroidal liver progenitor cells toward a biliary phenotype, a behavior which is further enhanced in the presence of hydrogel microparticles. We further demonstrate that surface modification of the hydrogel microparticles with heparin influences the behavior of liver progenitor cells toward biliary differentiation. Taken together, this liver progenitor cell culture system represents an approach for controlling the presentation of microenvironmental cues internalized within 3D spheroidal aggregate cultures. Overall, this strategy could be applied toward the engineering of instructive microenvironments that control stem and progenitor cell differentiation within a 3D context for studies in tissue engineering, drug testing, and cellular metabolism.

Optimization of 3D-aggregated spheroid model (3D-ASM) for selecting high efficacy drugs

“…These methods show low throughput and low reproducibility due to the difficulty of changing clean culture media and the problem of spheroid loss. There is also a method for aggregating and culturing cells in 3D using mesh-type structures such as aggrewell 6 and matrix-free microwell systems 7 . However, this method is a scaffold-free type and aggregated cells in media easily moved in media.…”

Liver Regeneration and Cell Transplantation for End-Stage Liver Disease

“…A recent study suggested an in vitro model of 3D LPCs spheroidal cultures with integrated polyethylene glycol hydrogel microparticles for the internal presentation of modular microenvironmental cues, which could be applied to the engineering of instructive microenvironments of transplanted LPCs. In this study, modification of the microparticles with TGF-β1 or heparin influences the behavior of LPCs toward BECs differentiation [ 64 ]. However, until now there has been limited data concerning engineering of 3D culture models of LPCs through the tunable presentation of microenvironmental stimuli.…”

Microtissue Geometry and Cell‐Generated Forces Drive Patterning of Liver Progenitor Cell Differentiation in 3D

“…[ 22 ] As in other organ systems, 3D culture of liver more closely mimics in vivo conditions, and we have observed that differentiation of BMEL aggregates can also be affected by modifying the 3D environment. [ 23 ] To date, the relationships between geometry, mechanical signaling, and liver progenitor fate specification in 3D has not yet been characterized.…”