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In vitro differentiation of human pluripotent stem cells into the B lineage using OP9-MS5 co-culture.

ABSTRACT: In vitro differentiation of human pluripotent stem cells (hPSCs) offers a genetically tractable system to examine the physiology and pathology of human tissue development and differentiation. We have used this approach to model the earliest stages of human B lineage development and characterize potential target cells for the in utero initiation of childhood B acute lymphoblastic leukemia. Herein, we detail critical aspects of the protocol including reagent validation, controls, and examples of surface markers used for analysis and cell sorting. For complete details on the use and execution of this protocol, please refer to Boiers et al. (2018).

SUBMITTER: Richardson SE

PROVIDER: S-EPMC8053808 | biostudies-literature | 2021 Jun

REPOSITORIES: biostudies-literature

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In vitro differentiation of human pluripotent stem cells into the B lineage using OP9-MS5 co-culture.

Richardson Simon E SE Ghazanfari Roshanak R Chhetri Jyoti J Enver Tariq T Böiers Charlotta C

STAR protocols 20210331 2

In vitro differentiation of human pluripotent stem cells (hPSCs) offers a genetically tractable system to examine the physiology and pathology of human tissue development and differentiation. We have used this approach to model the earliest stages of human B lineage development and characterize potential target cells for the in utero initiation of childhood B acute lymphoblastic leukemia. Herein, we detail critical aspects of the protocol including reagent validation, controls, and ...[more]

PMID: 33899010

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Project description:On the basis of their self-renewal capacity and their ability to differentiate into derivatives of all three germ layers, germ line-derived multipotent adult stem cells (maGSCs) from mouse testis might serve as one of preferable sources for pluripotent stem cells in regenerative medicine. In our study, we aimed for an efficient hepatic differentiation protocol that is applicable for both maGSCs and embryonic stem cells (ESCs). We attempted to accomplish this goal by using a new established co-culture system with OP9 stroma cells for direct differentiation of maGSCs and ESCs into hepatic cells. We found that the hepatic differentiation of maGSCs was induced by the OP9 co-culture system in comparison to the gelatin culture. Furthermore, we showed that the combination of OP9 co-culture with activin A resulted in the increased expression of endodermal and early hepatic markers Gata4, Sox17, Foxa2, Hnf4, Afp, and Ttr compared to differentiated cells on gelatin or on OP9 alone. Moreover, the hepatic progenitors were capable of differentiating further into mature hepatic cells, demonstrated by the expression of liver-specific markers Aat, Alb, Tdo2, Krt18, Krt8, Krt19, Cps1, Sek, Cyp7a1, Otc, and Pah. A high percentage of maGSC-derived hepatic progenitors (51% AFP- and 61% DLK1-positive) and mature hepatic-like cells (26% ALB-positive) were achieved using this OP9 co-culture system. These generated hepatic cells successfully demonstrated in vitro functions associated with mature hepatocytes, including albumin and urea secretion, glycogen storage, and uptake of low-density lipoprotein. The established co-culture system for maGSCs into functional hepatic cells might serve as a suitable model to delineate the differentiation process for the generation of high numbers of mature hepatocytes in humans without genetic manipulations and make germ line-derived stem cells a potential autologous and alternative cell source for hepatic transplants in metabolic liver disorders.

Dataset Information

In vitro differentiation of human pluripotent stem cells into the B lineage using OP9-MS5 co-culture.

Publications

<i>In vitro</i> differentiation of human pluripotent stem cells into the B lineage using OP9-MS5 co-culture.

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Dataset Information

In vitro differentiation of human pluripotent stem cells into the B lineage using OP9-MS5 co-culture.

Publications

&lt;i&gt;In vitro&lt;/i&gt; differentiation of human pluripotent stem cells into the B lineage using OP9-MS5 co-culture.

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OmicsDI is part of the ELIXIR infrastructure

Tweets

<i>In vitro</i> differentiation of human pluripotent stem cells into the B lineage using OP9-MS5 co-culture.