Biofunctionalized Hydrogel Microscaffolds Promote 3D Hepatic Sheet Morphology

Abstract

Development of artificial tissues providing the proper geometrical, mechanical, and environmental cues for cells is highly coveted in the field of tissue engineering. Recently, microfabrication strategies in combination with other chemistries have been utilized to capture the architectural complexity of intricate organs, such as the liver, in in vitro platforms. Here it is shown that a biofunctionalized poly (ethylene glycol) (PEG) hydrogel scaffold, fabricated using a sphere-template, facilitates hepatic sheet formation that follows the microscale patterns of the scaffold surface. The design takes advantage of the excellent diffusion properties of porous, uniform 3D hydrogel platforms, and the enhanced-cell-extracellular matrix interaction with the display of conjugated collagen type I, which in turn elicits favorable Huh-7.5 response. Collectively, the experimental findings and corresponding simulations demonstrate the importance of biofunctionalized porous scaffolds and indicate that the microscaffold shows promise in liver tissue engineering applications and provides distinct advantages over current cell sheet and hepatocyte spheroid technologies.

Long-term culture of human liver tissue with advanced hepatic functions

“…For this, we were inspired by the 3D hexagonally arrayed liver lobules -the functional units of human liver -that collectively make up the human liver organ, and we created a network of interconnected hexagonally arrayed spherical cavities within a biocompatible hydrogel, into which primary human liver fetal cell mixtures were seeded. We coated the scaffold with extracellular matrix (ECM) proteins to facilitate cell attachment upon seeding and subsequent cell growth, as evidenced thus far with tumor-derived liver cells (16,17). The architectural features of the scaffold then enable the cells to self-assemble into a 3D configuration.…”

Spheroid Formation of Hepatocarcinoma Cells in Microwells: Experiments and Monte Carlo Simulations

“…The structure and size of the corresponding scaffolds vary in a broad range from two-dimensional (2D) arrays of sub-millimeter wells to complex 3D structures aiming at mimicking specific organs [2, 3]. Chemically, the scaffolds are often fabricated by using natural hydrogels [2], synthetic polymers [1], or combination of such materials [4]. Cells growing in scaffolds typically aggregate.…”

“…Such models were used to scrutinize the limitations in the nutrient supply and oxygen transport in porous scaffolds on the coarse-grained level without or with explicit description of single pores (see e.g. references [4, 17, 18] and [18, 19], respectively, and references therein). MC simulations, based often on the lattice approximation and describing evolution of an ensemble of individual cells, are efficient in the situations with complex geometry and/or in the cases when the focus is on aggregation of cells (as in our present study).…”

“…MC simulations, based often on the lattice approximation and describing evolution of an ensemble of individual cells, are efficient in the situations with complex geometry and/or in the cases when the focus is on aggregation of cells (as in our present study). The available generic 2D and 3D MC simulations have been focused on the growth and differentiation of stem cells [20], cell seeding [21], and formation of cell sheets [4]. Related theoretical studies concern stem-cell niches [22–25] and scaffold-less biofabrication [26].…”

“…These cells belonging to a human hepatocarcinoma cell line are widely used as a liver cell model for the exploration of HCV infection [27]. Earlier, we observed the formation of Huh-7.5 cell spheroids in PEG-based hydrogels [28] and multilayer cell sheets in a biofunctionalized 3D scaffold [4, 29]. Our present work is focused on the same cells and has three novel ingredients.…”