A Modified Pre-plating Method for High-Yield and High-Purity Muscle Stem Cell Isolation From Human/Mouse Skeletal Muscle Tissues

Kiyoshi Yoshioka; Yasuo Kitajima; Narihiro Okazaki; Ko Chiba; Akihiko Yonekura; Yusuke Ono

doi:10.3389/fcell.2020.00793

A Modified Pre-plating Method for High-Yield and High-Purity Muscle Stem Cell Isolation From Human/Mouse Skeletal Muscle Tissues

Front Cell Dev Biol. 2020 Aug 13:8:793. doi: 10.3389/fcell.2020.00793. eCollection 2020.

Authors

Kiyoshi Yoshioka¹, Yasuo Kitajima¹, Narihiro Okazaki², Ko Chiba², Akihiko Yonekura², Yusuke Ono¹

Affiliations

¹ Department of Muscle Development and Regeneration, Institute of Molecular Embryology and Genetics, Kumamoto University, Kumamoto, Japan.
² Department of Orthopaedic Surgery, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki, Japan.

Abstract

Primary culture of skeletal muscle stem cells (MuSCs) is indispensable to study the dynamics of muscle regeneration and homeostasis. Here we describe the modified pre-plating method for isolating MuSCs in culture with greatly improved purity, yield, and procedure time. The protocol is based on the distinct adhesion characteristics of MuSCs. We reduced the procedure time to 2.5 days to obtain highly purified MuSCs through a newly employed re-plating step, which repeats incubation and cell-suspension. The re-plating step efficiently traps remaining fibroblastic cells, but not MuSCs, on a collagen-coated dish. Additionally, we confirmed that MuSCs can be isolated from small amounts of human/mouse muscle tissues, enabling us to perform experiments with amount-limited specimens. Thus, our method can be performed with basic laboratory equipment suitable for most facilities and without sophisticated MuSC handling techniques.

Keywords: isolation; muscle stem cell; pre-plating; satellite cell; skeletal muscle.