TY - JOUR
T1 - Single transcription factor efficiently leads human induced pluripotent stem cells to functional microglia
AU - Sonn, Iki
AU - Honda-Ozaki, Fumiko
AU - Yoshimatsu, Sho
AU - Morimoto, Satoru
AU - Watanabe, Hirotaka
AU - Okano, Hideyuki
N1 - Funding Information:
We would like to thank Drs. Shinya Yamanaka (Kyoto University), Minoru S.H. Ko and Yuhki Nakatake (Keio University), and Haruhiko Bito (University of Tokyo) for the kind gifts of the 201B7 iPSC line, the plasmid expressing SPI1 (PB-tet-PHS-SPI1, Id: 91215), and the plasmid expressing β-actin-GFP, respectively. We also thank Drs. Komei Fukushima, Hiroshi Kokubu (K Pharma, Inc.), Mitsuru Ishikawa, Kent Imaizumi, Seiji Ishii, and Chika Saegusa (Keio University) for their kind support and technical advice and all members of the H.O. Laboratory for their generous support for this study.
Funding Information:
This work was supported by the funding from JSPS KAKENHI Grant Number JP21J12528 to I.S., JP20H00485 and JP21H05273 to H.O., the Research Project for Practical Applications of Regenerative Medicine from the Japan Agency for Medical Research and Development (AMED) (grant no. 15bk0104027h0003, 16bk0104016h0004, and 17bk0104016h0005 to H.O.), and the Research Center Network for Realization Research Centers/Projects of Regenerative Medicine (the Program for Intractable Disease Research Utilizing Disease-specific iPS Cells and the Acceleration Program for Intractable Diseases Research Utilizing Disease-specific iPS Cells) from AMED (grant no. JP15bm0609003, JP16bm0609003, JP17bm0804003, JP18bm0804003, JP19bm0804003, JP20bm0804003, and JP21bm0804003 to H.O.).
Funding Information:
The Program for Initiative Research Projects from Keio University to H.O. F.O. is employed by K Pharma, Inc., and H.O. received research funding from K Pharma, Inc. The other authors declare that they have no competing interests.
Publisher Copyright:
© 2022, The Author(s).
PY - 2022/12
Y1 - 2022/12
N2 - Background: Microglia are innate immune cells that are the only residential macrophages in the central nervous system. They play vital physiological roles in the adult brain and during development. Microglia are particularly in the spotlight because many genetic risk factors recently identified for neurodegenerative diseases are largely expressed in microglia. Rare polymorphisms in these risk alleles lead to abnormal activity of microglia under traumatic or disease conditions. Methods: In the present study, to investigate the multifaceted functions of human microglia, we established a novel robust protocol to generate microglia from human induced pluripotent stem cells (hiPSCs) using a combination of cytokines and small chemicals essential for microglia ontogeny. Moreover, we highly enhanced the microglial differentiation efficiency by forcing the expression of PU.1, a crucial transcription factor for microglial development, during posterior mesoderm differentiation. Results: By our novel method, we demonstrated the generation of a greater number of hiPSC-derived microglia (hiMGLs, approximately 120-folds) than the prior methods (at most 40-folds). Over 90% of the hiMGLs expressed microglia-specific markers, such as CX3CR1 and IBA-1. Whole-transcriptome analysis revealed that these hiMGLs are similar to human primary microglia but differ from monocytes/macrophages. Furthermore, the specific physiological functions of microglia were confirmed through indices of lipopolysaccharide responsiveness, phagocytotic ability, and inflammasome formation. By co-culturing these hiMGLs with mouse primary neurons, we demonstrated that hiMGLs can regulate the activity and maturation of neurons. Conclusions: In this study, our new simple, rapid, and highly efficient method for generating microglia from hiPSCs will prove useful for future investigations on microglia in both physiological and disease conditions, as well as for drug discovery.
AB - Background: Microglia are innate immune cells that are the only residential macrophages in the central nervous system. They play vital physiological roles in the adult brain and during development. Microglia are particularly in the spotlight because many genetic risk factors recently identified for neurodegenerative diseases are largely expressed in microglia. Rare polymorphisms in these risk alleles lead to abnormal activity of microglia under traumatic or disease conditions. Methods: In the present study, to investigate the multifaceted functions of human microglia, we established a novel robust protocol to generate microglia from human induced pluripotent stem cells (hiPSCs) using a combination of cytokines and small chemicals essential for microglia ontogeny. Moreover, we highly enhanced the microglial differentiation efficiency by forcing the expression of PU.1, a crucial transcription factor for microglial development, during posterior mesoderm differentiation. Results: By our novel method, we demonstrated the generation of a greater number of hiPSC-derived microglia (hiMGLs, approximately 120-folds) than the prior methods (at most 40-folds). Over 90% of the hiMGLs expressed microglia-specific markers, such as CX3CR1 and IBA-1. Whole-transcriptome analysis revealed that these hiMGLs are similar to human primary microglia but differ from monocytes/macrophages. Furthermore, the specific physiological functions of microglia were confirmed through indices of lipopolysaccharide responsiveness, phagocytotic ability, and inflammasome formation. By co-culturing these hiMGLs with mouse primary neurons, we demonstrated that hiMGLs can regulate the activity and maturation of neurons. Conclusions: In this study, our new simple, rapid, and highly efficient method for generating microglia from hiPSCs will prove useful for future investigations on microglia in both physiological and disease conditions, as well as for drug discovery.
KW - Human pluripotent stem cells
KW - Microglia
KW - PU.1
KW - SPI1
UR - http://www.scopus.com/inward/record.url?scp=85133120306&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85133120306&partnerID=8YFLogxK
U2 - 10.1186/s41232-022-00201-1
DO - 10.1186/s41232-022-00201-1
M3 - Article
AN - SCOPUS:85133120306
SN - 1880-9693
VL - 42
JO - Inflammation and Regeneration
JF - Inflammation and Regeneration
IS - 1
M1 - 20
ER -
