Hydrogel Encapsulation Facilitates Rapid‐Cooling Cryopreservation of Stem Cell‐Laden Core–Shell Microcapsules as Cell–Biomaterial Constructs

Abstract

Core-shell structured stem cell microencapsulation in hydrogel has wide applications in tissue engineering, regenerative medicine, and cell-based therapies because it offers an ideal immunoisolative microenvironment for cell delivery and 3D culture and differentiation. Long-term storage of such microcapsules as cell-biomaterial constructs by cryopreservation is an enabling technology for their wide distribution and ready availability for clinical transplantation. However, most of the existing studies focused on cryopreservation of separated cells or cells in microcapsules without a core-shell structure (i.e., hydrogel beads). The goal of this study is to achieve cryopreservation of stem cells encapsulated in core-shell microcapsules as cell-biomaterial constructs or biocomposites. To this end, a capillary microfluidics-based core-shell alginate hydrogel encapsulation technology is developed to facilitate cryopreservation of porcine adipose-derived stem cells (pADSCs) laden microcapsules with very low concentration (2 mol L−1) of cell membrane penetrating cryoprotective agents (CPAs) by suppressing ice formation. This may provide a low-CPA and cost-effective approach for vitreous cryopreservation of “ready-to-use” stem cell-biomaterial constructs, facilitating their off-the-shelf availability and widespread applications.

Effects of Cryopreservation on Cell Metabolic Activity and Function of Biofabricated Structures Laden with Osteoblasts

“…Cryopreservation is the use of very low temperature to preserve living cells and tissues in a quiescent status for a long period, without losing their viability, activity, and function [14]. Several works have been devoted to the cryopreservation of engineered biological substitutes [15][16][17][18][19][20][21][22][23][24][25], but there is still the need for information on the cryopreservation of biofabricated osteoblast constructs, a paramount goal addressed in this work. Pioneer studies demonstrate that post-thawing osteoblast viability was better maintained (40-50%) when cryopreserved in dimethyl sulfoxide (DMSO) than in other cryoprotectant additives [26,27].…”

Advanced Biotechnology for Cell Cryopreservation

“…Droplet-based techniques find wide applications in various fields, such as inkjet printing, emulsion polymerization, and DNA arraying, owing to their high efficiency and low cost [80][81][82][83][84]. Introduction of this advanced technology into cell cryopreservation created a series of novel vitrification protocols that are characterized by lower CPA concentration and higher cooling and warming rates.…”

Cell‐Surface Engineering for Advanced Cell Therapy

“…[107] Cells encapsulated in core-shell structures have been reported to improvee ncapsulation, immune protection,c ryopreservation,t ransplantation, and cell viability. [108,109] Core-shell-encapsulated cells have usually been fabricated using droplet-based microfluidic techniques because they offer severala dvantages,s uch as controllable thickness and size with al ow size distribution, and high homogeneity. [110] However,t hey usually require specialized facilities,i ncluding lithography techniques; moreover,t he durability of the elastomer (polydimethylsiloxane, PDMS)c ommonly used to construct the device as well as the biocompatibility of the cytotoxic chemicals used in their construction, such as oil, alcohols and acids, have been questioned.…”

“…Encapsula-tion of porcineA DSCs in core-shell microcapsules comprising ac ell suspension solution and alginate was shown to reduce ice crystal formation around the plasma membrane, thereby loweringt he concentrationo fc ell cryoprotective agent (CPA) required for cryopreservation by the vitrification method. [109] Considering that high concentrations of CPAc an induce cell senescence,m etabolic changes and osmotic imbalance, resulting in cytotoxicity andc ellular injury,t he cytoprotective effect of cell encapsulation likely reflects not only the reduction in ice crystal formation, but also the reduced concentration of CPAu sed for cryopreservation.A pplication of this cytoprotective effect for cryopreservation could maximize the availability of cell-based therapiesb yp reserving cell aggregates in an optimized"ready-to-use" state.…”