Cell encapsulation in photocrosslinked alginates: mechanical characterization and cell viability study

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ICE Krakow

ICE Krakow

ul. Marii Konopnickiej 17 30-302 Kraków


Le Visage, Catherine (Nantes Université, Oniris, Univ Angers, CHU Nantes, INSERM, Regenerative Medicine and Skeleton, RMeS, UMR 1229)


In the past few years, mesenchymal stromal cells (MSC) have been identified as a potential therapy to inflammatory diseases due to their ability to secrete immuno-modulatory and anti-inflammatory factors. Considering the risk of cell leakage outside the target site and the poor survival rate after a local injection, for example in the joint, cell encapsulation in hydrogels could overcome these limitations, and provide cells a suitable 3D microenvironment supporting their biological activities. Sodium alginate is the most frequent natural polymer used for cell encapsulation, and is crosslinked by addition of divalent cations such as Ca2+ (AlgCa). As such, a physiological environment with monovalent cations, such as Na+, might impair the stability of ionically crosslinked alginate. In this context, we hypothesized that MSC could be encapsulated in a more stable photocrosslinked methacrylated alginate (AlgMA) hydrogel.

Four degrees of methacrylation of AlgMA were tested (targeting degrees of substitution of 10, 25, 50 and 75%). Photopolymerisation of AlgMA was performed with a photo-initiator (Irgacure® 2959) under UV-A light exposition (365 nm), with concentrations of AlgMA and Irgacure of 2% and 0.05% (w/v), respectively. AlgMA solutions were exposed to UV-A light intensities of 8.9 mW/cm² (low) or 21.7 mW/cm² (high), and were compared to AlgCa hydrogels crosslinked with 100 mM CaCl2 and stored in 100 or 1.8 mM CaCl2 for 24 hours. The effect of the methacrylation degree on the stiffness of the photocrosslinked AlgMA gels was investigated, via compression tests (Microtester, CellScale). Hydrogel stability in Dulbecco’s Modified Eagled Medium (DMEM), ultrapure water and sodium citrate 80 mM, was assessed for AlgMA and AlgCa hydrogels with weight analysis during 15 days. Human adipose-derived MSC viability in AlgMA hydrogels was analyzed by LIVE/DEAD® assay for up to 15 days. All assays were performed on cylindrical hydrogel punches of 2 mm diameter and 1 mm height.

AlgMA hydrogels stiffness significantly increased with methacrylate substitution and duration of exposure to UV-A light, with no significant difference observed over 40 minutes of high intensity exposure, reaching 190 kPa. By comparison, the elastic moduli of AlgCa hydrogels were similar to the one of 50% and 75% AlgMA crosslinked at high UV-A intensity for only 6 and 2 minutes respectively (< 15 kPa). In ultrapure water the weight and the elastic modulus of the AlgMA hydrogels increased significantly, whereas they remained stable in culture medium for 15 days. AlgMA hydrogels did not dissolve in sodium citrate, unlike AlgCa hydrogels. The cell viability study showed that in all the AlgMA tested conditions, excepted 10% AlgMA, the encapsulated MSCs were viable (viability higher than 70%).

Human MSC have been encapsulated in alginates with four degrees of methacrylation. We confirmed that AlgMA hydrogels are more stable than AlgCa ones in a physiological environment containing sodium ions. All, excepted 10% AlgMA were suitable for cell encapsulation. Further in vitro experiments are under investigation to characterize the diffusion of immune-modulatory and anti-inflammatory factors and to determine whether encapsulated MSC in AlgMA hydrogels may be a relevant strategy to treat inflammatory diseases.


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