通过Percoll密度梯度离心法从幼鼠中分离和纯化卫星细胞。

Isolation and Purification of Satellite Cells from Young Rats by Percoll Density Gradient Centrifugation.

作者信息

Matsuyoshi Yuji, Akahoshi Mariko, Nakamura Mako, Tatsumi Ryuichi, Mizunoya Wataru

机构信息

Department of Bioresource Sciences, Faculty of Agriculture, Kyushu University, Fukuoka, Japan.

出版信息

Methods Mol Biol. 2019;1889:81-93. doi: 10.1007/978-1-4939-8897-6_6.

DOI:10.1007/978-1-4939-8897-6_6

PMID:30367410

Abstract

Satellite cells (SCs) are myogenic stem cells that play an important role in skeletal muscle regeneration and hypertrophy. Primary cultures of SCs are useful to analyze cell functions; however, it is difficult to obtain highly pure SCs from young rats with the conventional procedures. The purpose of this study is to establish a purification method for SC isolation from young rats and quantitatively evaluate the purification procedure employing Percoll, a common research tool to purify cells. We elucidated the purity of SCs collected by Percoll density gradient centrifugation using real-time RT-qPCR and immunocytochemistry for desmin. Percoll treatment increased the purity of SCs isolated from young rats to nearly 90%, which was comparable to that achieved with the conventional method using middle-aged rats.

摘要

卫星细胞（SCs）是肌源性干细胞，在骨骼肌再生和肥大过程中发挥重要作用。SCs的原代培养有助于分析细胞功能；然而，采用传统方法从幼鼠中获取高度纯化的SCs却很困难。本研究的目的是建立一种从幼鼠中分离SCs的纯化方法，并使用用于纯化细胞的常用研究工具Percoll对纯化过程进行定量评估。我们通过实时RT-qPCR和针对结蛋白的免疫细胞化学方法，阐明了经Percoll密度梯度离心收集的SCs的纯度。Percoll处理将从幼鼠中分离出的SCs的纯度提高到了近90%，这与使用中年大鼠的传统方法所达到的纯度相当。