Abstract
Three-dimensional (3D) cell culture is regarded as a more physiologically relevant method of growing cells in the laboratory compared to traditional monolayer cultures. Recently, the application of polystyrene-based scaffolds produced using polyHIPE technology (porous polymers derived from high internal phase emulsions) for routine 3D cell culture applications has generated very promising results in terms of improved replication of native cellular function in the laboratory. These materials, which are now available as commercial scaffolds, are superior to many other 3D cell substrates due to their high porosity, controllable morphology, and suitable mechanical strength. However, until now there have been no reports describing the surface-modification of these materials for enhanced cell adhesion and function. This study, therefore, describes the surface functionalization of these materials with galactose, a carbohydrate known to specifically bind to hepatocytes via the asialoglycoprotein receptor (ASGPR), to further improve hepatocyte adhesion and function when growing on the scaffold. We first modify a typical polystyrene-based polyHIPE to produce a cell culture scaffold carrying pendent activated-ester functionality. This was achieved via the incorporation of pentafluorophenyl acrylate (PFPA) into the initial styrene (STY) emulsion, which upon polymerization formed a polyHIPE with a porosity of 92% and an average void diameter of 33 μm. Histological analysis showed that this polyHIPE was a suitable 3D scaffold for hepatocyte cell culture. Galactose-functionalized scaffolds were then prepared by attaching 2′-aminoethyl-β-d-galactopyranoside to this PFPA functionalized polyHIPE via displacement of the labile pentafluorophenyl group, to yield scaffolds with approximately ca. 7–9% surface carbohydrate. Experiments with primary rat hepatocytes showed that cellular albumin synthesis was greatly enhanced during the initial adhesion/settlement period of cells on the galactose-functionalized material, suggesting that the surface carbohydrates are accessible and selective to cells entering the scaffold. This porous polymer scaffold could, therefore, have important application as a 3D scaffold that offers enhanced hepatocyte adhesion and functionality.
Highlights
Substantial evidence exists to support three-dimensional (3D) cell culture as a more physiologically relevant growth environment compared to that with conventional monolayer cultures.[1−4] Cells cultured in 3D more closely mimic their native morphology, unlike monolayer cultures in which cells are often flattened into a two-dimensional (2D) shape
Several groups have employed porous polymers derived from high internal phase emulsions as scaffolds for 3D cell culture.[6−11] In particular, polystyrene-based polyHIPE scaffolds have shown very promising results with a range of different cell types due to their controllable morphology, high porosity and suitable mechanical properties.[12−16] These materials are available as commercial 3D scaffolds (AlvetexScaffold by Reinnervate) and have already been adopted by a broad range of research groups.[17,18]
We show that a polystyrene-based polyHIPE containing pentafluorophenyl acrylate (PFPA) can be produced with a suitable morphology to support 3D cell growth
Summary
Substantial evidence exists to support three-dimensional (3D) cell culture as a more physiologically relevant growth environment compared to that with conventional monolayer cultures.[1−4] Cells cultured in 3D more closely mimic their native morphology, unlike monolayer cultures in which cells are often flattened into a two-dimensional (2D) shape. Biomacromolecules neighbors, in turn increasing cellular communication that is important in regulating normal cell function. Recognizing these advantages associated with a 3D growth environment, many researchers require practical technologies that can enable routine 3D cell culture in the laboratory.[5]. Cells in vivo are surrounded by a complex extracellular matrix that contributes to cell anchorage and function They receive a plethora of biochemical cues from molecules such as carbohydrates and proteins that serve to regulate normal cell behavior. Being able to mimic some of these biological interactions on the surface of synthetic 3D scaffolds is an attractive prospect.[19] achieving this for polystyrene-based polyHIPEs is challenging. Postpolymerization modifications of polystyrene are possible but often require harsh reaction conditions due to the inert nature of the polymer.[20,21]
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