A modified preplate technique for efficient isolation and proliferation of mice muscle-derived stem cells
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We modified an existing protocol to develop a more efficient method to acquire and culture muscle-derived stem cells (MDSCs) and compared the characteristics of cells obtained from the two methods. This method is based on currently used multistep enzymatic digestion and preplate technique. During the replating process, we replaced the traditional medium with isolation medium to promote fibroblast-like cell adherence at initial replating step, which shortened the purifying duration by up to 4 days. Moreover, we modified the culture container to provide a stable microenvironment that promotes MDSC adherence. We compared the cell morphology, growth curve and the expression of specific markers (Sca-1, CD34, PAX7 and Desmin) between the two cell groups separately obtained from the two methods. Afterwards, we compared the neural differentiation capacity of MDSCs with other muscle-derived cell lineages. The protocol developed here is a fast and effective method to harvest and purify MDSCs from mice limb skeletal muscle.
KeywordsRegenerative medicine Cell and tissue culture Myogenic stem cells In vitro culture
ZX and LY performed the biopsy and cells culture, HL and WF took charge of the immunofluorescence and flow cytometry, LC and JZ accomplished Statistical analysis. ZX drafted the manuscript. Co-corresponding authors XY and ZQ designed and guided the study. All authors have read and approved the final manuscript.
This work was supported by the National Natural Science Foundation of China (81571921 and 81671908), the Union Youth Science and Research Fund (3332015155), the Science Fund of Plastic Surgery Hospital, CAMS, PUMC (Q2015013) and the Innovation Fund of Graduate Students, CAMS, PUMC (2017-1002-1-10).
Compliance with ethical standards
Conflict of interest
The authors declare they have no competing interests.
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