One-Step Generation and Purification of Cell-Encapsulated Hydrogel Microsphere With an Easily Assembled Microfluidic Device.

Tao Zhang,Cong Liu,Wen-Fei Dong,Minxuan Sun,Haiwen Li,Zhiqiang Zhang,Keming Jiang,Hong Zhang,Ru Wang,Qian Mei,Wuping Zhou,Yuanshuai Zhou

doi:10.3389/fbioe.2021.816089

Abstract

Cell-laden hydrogel microspheres with uniform size show great potential for tissue repair and drug screening applications. Droplet microfluidic systems have been widely used for the generation of cell-laden hydrogel microspheres. However, existing droplet microfluidic systems are mostly based on complex chips and are not compatible with well culture plates. Moreover, microspheres produced by droplet microfluidics need demulsification and purification from oil, which requires time and effort and may compromise cell viability. Herein, we present a simple one-step approach for producing and purifying hydrogel microspheres with an easily assembled microfluidic device. Droplets were generated and solidified in the device tubing. The obtained hydrogel microspheres were then transferred to a tissue culture plate filled with cell culture media and demulsified through evaporation of the oil at 37°C. The removal of oil caused the gelled microspheres to be released into the cell culture media. The encapsulated cells demonstrated good viability and grew into tumor spheroids in 12–14 days. Single cell-laden hydrogel microspheres were also obtained and grown into spheroid in 14 days. This one-step microsphere generation method shows good potential for applications in automated spheroid and organoid cultures as well as drug screening, and could potentially offer benefits for translation of cell/microgel technologies.

Highlights

Spheroids are three-dimensional (3D) spherical cellular aggregates that allow cells to interact with neighboring cells and their extracellular matrix
To generate and purify cell-encapsulated hydrogel microspheres in one step, we developed an assembled microfluidic platform
Cell viability was evaluated after encapsulation, and the results show that the HCT116 cells maintained good viability and formed cell clusters in the hydrogel microspheres after 7–14 days of culturing (Figures 4D–F).This microfluidic system was further verified with the U87 tumor cell line

Summary

Introduction

Spheroids are three-dimensional (3D) spherical cellular aggregates that allow cells to interact with neighboring cells and their extracellular matrix. Spheroids have become the most common and versatile three-dimensional model since they were first used in cell cultures in the 1950s (Dyson, 2019; Boucherit et al, 2020; Sivakumar et al, 2020; Decarli et al, 2021; Roper et al, 2021). Scaffold-free approaches are easy to perform and can be applied to various cell types (Franchi-Mendes et al, 2021). Microfabrication techniques such as photolithography (Du et al, 2008; Brandenberg et al, 2020) and micromolding (Tekin et al, 2011; Tao et al, 2019) have been used to create spheroids, allowing the volume and shape of spheroids to be precisely controlled. Most spheroid fabrication approaches require an excessive amount of time and effort, and producing uniform spheroids with a high throughput and in a reproducible manner is challenging

Methods

Results

Conclusion