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CD82 is a marker to isolate β cell precursors from human iPS cells and plays a role for the maturation of β cells.

ABSTRACT: Generation of pancreatic β cells from pluripotent stem cells is a key technology to develop cell therapy for insulin-dependent diabetes and considerable efforts have been made to produce β cells. However, due to multiple and lengthy differentiation steps, production of β cells is often unstable. It is also desirable to eliminate undifferentiated cells to avoid potential risks of tumorigenesis. To isolate β cell precursors from late stage pancreatic endocrine progenitor (EP) cells derived from iPS cells, we have identified CD82, a member of the tetraspanin family. CD82⁺ cells at the EP stage differentiated into endocrine cells more efficiently than CD82^- EP stage cells. We also show that CD82⁺ cells in human islets secreted insulin more efficiently than CD82^- cells. Furthermore, knockdown of CD82 expression by siRNA or inhibition of CD82 by monoclonal antibodies in NGN3⁺ cells suppressed the function of β cells with glucose-stimulated insulin secretion, suggesting that CD82 plays a role in maturation of EP cells to β cells.

SUBMITTER: Watanabe A

PROVIDER: S-EPMC8100138 | biostudies-literature | 2021 May

REPOSITORIES: biostudies-literature

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CD82 is a marker to isolate β cell precursors from human iPS cells and plays a role for the maturation of β cells.

Watanabe Ami A Tanaka Anna A Koga Chizuko C Matsumoto Masahito M Okazaki Yasushi Y Kin Tatsuya T Miyajima Atsushi A

Scientific reports 20210505 1

Generation of pancreatic β cells from pluripotent stem cells is a key technology to develop cell therapy for insulin-dependent diabetes and considerable efforts have been made to produce β cells. However, due to multiple and lengthy differentiation steps, production of β cells is often unstable. It is also desirable to eliminate undifferentiated cells to avoid potential risks of tumorigenesis. To isolate β cell precursors from late stage pancreatic endocrine progenitor (EP) cells derived from iP ...[more]

PMID: 33953224

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Project description:BackgroundBrain microvascular endothelial cells (BMECs) play a major role in the blood-brain barrier (BBB), and are critical for establishing an in vitro BBB model. Currently, iPSC-derived BMECs (iBMECs) have been used to construct in vitro BBB models with physiological barrier functions, such as high trans-endothelial electrical resistance (TEER) and expression of transporter proteins. However, the relatively low p-glycoprotein (P-gp) level and a decrease in the efflux ratio of its substrates in iBMECs suggest their immature nature. Therefore, more mature iBMECs by optimizing the differentiation induction protocol is beneficial for establishing a more reliable in vitro BBB model for studying central nervous system (CNS) drug transport.MethodsTo identify human brain endothelial cell fate-inducing factors, HUVEC was transfected with Zic3A-, Zic3B-, and Sox18-expressing lentivirus vector. Since SOX18 was found to induce BMEC properties, we used a Dox-inducible Tet-on system to express SOX18 during iBMEC differentiation and explored the impact of SOX18 expression on iBMEC maturation.ResultsSox18-mediated iBMECs achieved a higher TEER value than normal iBMECs (> 3000 Ω cm2). From day 6 to day 10 (d6-10 group), the iBMECs with SOX18 expression expressed a series of tight junction markers and showed upregulation of Mfsd2a, a specific marker of the BBB. The d6-10 group also expressed SLC2A1/Glut1 at levels as high as normal iBMECs, and upregulated ABCB1/P-gp and ABCC1/MRP1 expression. Moreover, Sox18-mediated iBMECs showed higher viability than normal iBMECs after puromycin treatment, indicating that SOX18 expression could upregulate P-gp activity in iBMECs.ConclusionsInducible SOX18 expression in iBMECs gained BBB phenotypes, including high TEER values and upregulation of tight junction-related genes, endothelial cell (EC) markers, BBB transporters, and higher cell viability after treatment with puromycin. Collectively, we provide a differentiation method for the maturation of human iPS cell-derived BMECs with SOX18 expression, describing its contribution to form an in vitro BBB model for CNS drug transport studies.

Dataset Information

CD82 is a marker to isolate β cell precursors from human iPS cells and plays a role for the maturation of β cells.

Publications

CD82 is a marker to isolate β cell precursors from human iPS cells and plays a role for the maturation of β cells.

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