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Scalable Production and Cryostorage of Organoids Using Core-Shell Decoupled Hydrogel Capsules.

ABSTRACT: Organoids, organ-mimicking multicellular structures derived from pluripotent stem cells or organ progenitors, have recently emerged as an important system for both studies of stem cell biology and development of potential therapeutics; however, a large-scale culture of organoids and cryopreservation for whole organoids, a prerequisite for their industrial and clinical applications, has remained a challenge. Current organoid culture systems relying on embedding the stem or progenitor cells in bulk extracellular matrix (ECM) hydrogels (e.g., Matrigel™) have limited surface area for mass transfer and are not suitable for large-scale productions. Here, we demonstrate a capsule-based, scalable organoid production and cryopreservation platform. The capsules have a core-shell structure where the core consists of Matrigel™ that supports the growth of organoids, and the alginate shell form robust spherical capsules, enabling suspension culture in stirred bioreactors. Compared with conventional, bulk ECM hydrogels, the capsules, which could be produced continuously by a two-fluidic electrostatic co-spraying method, provided better mass transfer through both diffusion and convection. The core-shell structure of the capsules also leads to better cell recovery after cryopreservation of organoids probably through prevention of intracellular ice formation.

SUBMITTER: Lu YC

PROVIDER: S-EPMC5870136 | biostudies-literature | 2017 Dec

REPOSITORIES: biostudies-literature

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Scalable Production and Cryostorage of Organoids Using Core-Shell Decoupled Hydrogel Capsules.

Lu Yen-Chun YC Fu Dah-Jiun DJ An Duo D Chiu Alan A Schwartz Robert R Nikitin Alexander Yu AY Ma Minglin M

Advanced biosystems 20171122 12

Organoids, organ-mimicking multicellular structures derived from pluripotent stem cells or organ progenitors, have recently emerged as an important system for both studies of stem cell biology and development of potential therapeutics; however, a large-scale culture of organoids and cryopreservation for whole organoids, a prerequisite for their industrial and clinical applications, has remained a challenge. Current organoid culture systems relying on embedding the stem or progenitor cells in bul ...[more]

PMID: 29607405

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Project description:BackgroundOrganoids are three-dimensional in vitro-grown cell clusters that recapitulate key features of native organs. In regenerative medicine, organoid technology represents a promising approach for the replacement of severely damaged organs, such as the pancreas in patients with type 1 diabetes. Isolation human pancreas organoids (hPOs) in chemically defined serum-free culture media would be a major milestone for this approach.MethodsStarting from discarded pancreatic tissues, we developed a large-scale process for obtaining clinically relevant quantities of undifferentiated organoids, obviating enzymatic digestion and operator-dependent pancreatic ducts picking steps. hPO identity was characterized by molecular and flow cytometry analysis.ResultsThis work demonstrates that it is possible to obtain a large-scale production of organoids. We introduced some innovations in the isolation, expansion, and freezing of hPOs from five donors. First of all, the choice of the starting material (islet-depleted pancreas) that allows obtaining a high quantity of hPOs at low passages. On the other hand, we introduced mechanical dissociation and we eliminated the picking step to exclude the operator-depending steps, without affecting the success of the culture (100% success rate). Another important improvement was to replace R-spondin-1 (Rspo1) conditioned medium with Rspo1 recombinant molecule to obtain a well-defined composition of the expansion medium. Finally, we implemented a GMP-compliant freezing protocol. hPOs showed exponential growth with diameter and area that increased three- and eight-fold in 7 days, respectively. Immunophenotypic profile and gene expression analysis revealed that hPOs were composed of ductal (82.33 ± 8.37%), acinar (2.80 ± 1.25%) cells, and pancreatic progenitors (5.81 ± 2.65%).ConclusionThis work represents a milestone for a GMP-compliance hPO production and, ultimately, their clinical application as a type 1 diabetes therapy.

Dataset Information

Scalable Production and Cryostorage of Organoids Using Core-Shell Decoupled Hydrogel Capsules.

Publications

Scalable Production and Cryostorage of Organoids Using Core-Shell Decoupled Hydrogel Capsules.

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