Partitioning microfluidic channels with hydrogel to construct tunable 3-D cellular microenvironments

Abstract

Accurate modeling of the cellular microenvironment is important for improving studies of cell biology in vitro. Here, we demonstrate a flexible method for creating a cellular microenvironment in vitro that allows i) controlled spatial distribution (patterning) of multiple types of cells within three-dimensional (3-D) matrices of a biologically-derived, thermally-curable hydrogel (Matrigel) and ii) application of gradients of soluble factors, such as cytokines, across the hydrogel. The technique uses laminar flow to divide a microchannel into multiple subchannels separated by microslabs of hydrogel. It does not require the use of UV light or photoinitiators, and is compatible with cell culture in the hydrogel. This technique makes it possible to design model systems to study cellular communication mediated by the diffusion of soluble factors within 3-D matrices. Such factors can originate either from secretions of neighboring cells patterned within the microchannel, or from an external source-e.g., a solution of growth factors injected into a subchannel. This method is particularly useful for studying cells such as those of the immune system, which are often weakly adherent and difficult to position precisely with standard systems for cell culture. We demonstrated this application by co-culturing two types of macrophage-like cells (BAC1.2F5 and LADMAC cell lines) within spatially separated regions of a slab of hydrogel. This pair of cell lines represents a simple model system for intercellular communication: the LADMAC cells produce colonystimulating factor 1 (CSF-1), which is required by the BAC cells for survival.

Hydrogel discs for digital microfluidics

“…25,28,31 For example, microchannel systems are well suited for very low ($nl) sample volumes. This property makes microchannel enzyme microreactors useful for analytical applications, but such systems are often not easily adapted for use for preparative applications.…”

“…[22][23][24] There has been great enthusiasm for combining hydrogels with microchannel-based labon-a-chip systems. [25][26][27][28][29][30][31][32][33][34] For example, microfluidic systems are useful for reagent delivery and collection of products from hydrogel-based enzyme microreactors. 25 Likewise, microfluidic systems for culturing cells encapsulated in hydrogel structures are useful because fluidic ports are a) Lindsey K. Fiddes and Vivienne N. Luk contributed equally to this work.…”

“…easily incorporated for nutrient delivery and waste removal. 28 These advances are promising and useful; however, the plumbing complexities incurred with controlling many different reagents simultaneously make the combination of microchannels and hydrogels an imperfect match for all applications. Moreover, in continuous flow microchannel systems, analytes generated by enzymes or cells immobilized in hydrogels can diffuse away from the gels, complicating the analysis.…”

A Digital Microfluidic Approach to Proteomic Sample Processing

“…[201][202][203] There has been great enthusiasm for combining hydrogels with microchannel-based lab-on-achip systems. [204][205][206][207][208][209][210][211][212] For example, microfluidic systems are useful for reagent delivery and collection of products from hydrogel-based enzyme microreactors. 204 Likewise, microfluidic systems for culturing cells encapsulated in hydrogel structures are useful because fluidic ports are easily incorporated for nutrient delivery and waste removal.…”

“…204 Likewise, microfluidic systems for culturing cells encapsulated in hydrogel structures are useful because fluidic ports are easily incorporated for nutrient delivery and waste removal. 207 These advances are promising and useful; however, the plumbing complexities incurred with controlling many different reagents simultaneously make the combination of microchannels and hydrogels an imperfect match for all applications. Moreover, in continuous flow microchannel systems, analytes generated by enzymes or cells immobilized in hydrogels can diffuse away from the gels, complicating the analysis.…”