Canonical Wnt signaling activation by chimeric antigen receptors for efficient cardiac differentiation from mouse embryonic stem cells

Inflammation and Regeneration

Research Article

Yasuyuki S. Kida¹  Yuka Akagi²  Teruyuki Nagamune³  Masahiro Kawahara⁴  Tomoaki Ishida⁵  Masato Nakahara⁵  Dai Ihara⁶  Yukihiro Harada⁶  Osamu Nakagawa⁷  Koji Hasegawa⁸  Takahiro Sogo⁹  Tomoe Ueyama^1,10  Tasuku Tsukamoto^1,10  Shu Nakao^1,11  Teruhisa Kawamura^1,11 

[1] Cellular and Molecular Biotechnology Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), Central 5-41, 1-1-1 Higashi, 305-8565, Tsukuba, Ibaraki, Japan;School of Integrative & Global Majors, University of Tsukuba, 1-1-1 Tennoudai, 305-8572, Tsukuba, Ibaraki, Japan;Cellular and Molecular Biotechnology Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), Central 5-41, 1-1-1 Higashi, 305-8565, Tsukuba, Ibaraki, Japan;Tsukuba Life Science Innovation Program (T-LSI), School of Comprehensive Human Sciences, University of Tsukuba, 1-1-1 Tennoudai, 305-8572, Tsukuba, Ibaraki, Japan;Department of Bioengineering, Graduate School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, 113-8656, Tokyo, Japan;Department of Bioengineering, Graduate School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, 113-8656, Tokyo, Japan;Laboratory of Cell Vaccine, Center for Vaccine and Adjuvant Research, National Institutes of Biomedical Innovation, Health, and Nutrition (NIBIOHN), 7-6-8 Saito-Asagi, 567-0085, Ibaraki City, Osaka, Japan;Department of Biomedical Sciences, College of Life Sciences, Ritsumeikan University, 1-1-1 Noji-higashi, 525-8577, Kusatsu, Shiga, Japan;Department of Biomedical Sciences, College of Life Sciences, Ritsumeikan University, 1-1-1 Noji-higashi, 525-8577, Kusatsu, Shiga, Japan;Department of Molecular Physiology, National Cerebral and Cardiovascular Center Research Institute, 6-1 Kishibe-Shimmachi, Suita, 564-8565, Osaka, Japan;Department of Molecular Physiology, National Cerebral and Cardiovascular Center Research Institute, 6-1 Kishibe-Shimmachi, Suita, 564-8565, Osaka, Japan;Division of Translational Research, Kyoto Medical Center, National Hospital Organization, 1-1 Mukaihata-cho, Fukakusa, Fushimi-ku, 612-8555, Kyoto, Japan;Ritsumeikan Global Innovation Research Organization, Ritsumeikan University, 1-1-1 Noji-higashi, 525-8577, Kusatsu, Shiga, Japan;Ritsumeikan Global Innovation Research Organization, Ritsumeikan University, 1-1-1 Noji-higashi, 525-8577, Kusatsu, Shiga, Japan;Department of Biomedical Sciences, College of Life Sciences, Ritsumeikan University, 1-1-1 Noji-higashi, 525-8577, Kusatsu, Shiga, Japan;Ritsumeikan Global Innovation Research Organization, Ritsumeikan University, 1-1-1 Noji-higashi, 525-8577, Kusatsu, Shiga, Japan;Department of Biomedical Sciences, College of Life Sciences, Ritsumeikan University, 1-1-1 Noji-higashi, 525-8577, Kusatsu, Shiga, Japan;Division of Translational Research, Kyoto Medical Center, National Hospital Organization, 1-1 Mukaihata-cho, Fukakusa, Fushimi-ku, 612-8555, Kyoto, Japan;

关键词: Canonical Wnt signaling;    Cardiac differentiation;    Chimeric antigen receptor;    Pluripotent stem cells;    Regenerative medicine;   

DOI  :  10.1186/s41232-023-00258-6

 received in 2021-08-27, accepted in 2023-01-15,  发布年份 2023

来源: Springer

【 摘 要 】

BackgroundCanonical Wnt signaling is involved in a variety of biological processes including stem cell renewal and differentiation, embryonic development, and tissue regeneration. Previous studies reported the stage-specific roles of the Wnt signaling in heart development. Canonical Wnt signal activation by recombinant Wnt3a in the early phase of differentiation enhances the efficiency of myocardial cell production from pluripotent stem cells. However, the hydrophobicity of Wnt proteins results in high cost to produce the recombinant proteins and presents an obstacle to their preparation and application for therapeutics, cell therapy, or molecular analysis of Wnt signaling.MethodsTo solve this problem, we generated an inexpensive molecule-responsive differentiation-inducing chimeric antigen receptor (designated as diCAR) that can activate Wnt3a signaling. The extracellular domains of low-density-lipoprotein receptor-related protein 6 (LRP6) and frizzeled-8 (FZD8) were replaced with single-chain Fv of anti-fluorescein (FL) antibody, which can respond to FL-conjugated bovine serum albumin (BSA-FL) as a cognate ligand. We then analyzed the effect of this diCAR on Wnt signal activation and cardiomyocyte differentiation of mouse embryonic stem cells in response to BSA-FL treatment.ResultsEmbryonic stem cell lines stably expressing this paired diCAR, named Wnt3a-diCAR, showed TCF/β-catenin-dependent transactivation by BSA-FL in a dose-dependent manner. Treatment with either Wnt3a recombinant protein or BSA-FL in the early phase of differentiation revealed similar changes of global gene expressions and resulted in efficient myocardial cell differentiation. Furthermore, BSA-FL-mediated signal activation was not affected by a Wnt3a antagonist, Dkk1, suggesting that the signal transduction via Wnt3a-diCAR is independent of endogenous LRP6 or FZD8.ConclusionWe anticipate that Wnt3a-diCAR enables target-specific signal activation, and could be an economical and powerful tool for stem cell-based regeneration therapy.

【 授权许可】

CC BY   
© The Author(s) 2023

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