Mazandaran University of Medical Sciences, School of Medicine, Department of Immunology, Molecular and Cell Biology Research Center - Sari, Iran.
Mazandaran University of Medical Sciences, Orthopedic Research Center - Sari, Iran.
Rev Assoc Med Bras (1992). 2021 Nov;67(11):1654-1658. doi: 10.1590/1806-9282.20210706.
Cell culture technology has become a popular method in the field of cell biology, pharmacology, and medical researches. Primary cells represent the normal physiological condition of human cells. Fibroblasts are the most common native cells of connective tissue that play a crucial role in the entire pathogenesis of various disorders, such as rheumatoid arthritis (RA). Fibroblast-like synoviocytes (FLSs), which overlie the loose connective tissue of the synovial sublining, are known to be the central mediators of joint damage. The most routine approach for the isolation of FLS is an enzymatic digestion of synovial tissue. This experimental study is designed to introduce an easy, fast, and high-throughput method compared with enzymatic digestion for isolation of FLS.
The synovial tissue and synovial fluid (SF) samples were collected from eight patients with RA who underwent routine knee replacement surgery. Synovial tissue was incubated with collagenase VIII enzyme, while SF was washed with a similar volume of phosphate-buffered saline. The cells were further subcultured and stored based on the standard protocols. The purity of isolated synoviocytes was confirmed using flow cytometry analysis.
Isolation of FLS from SF was more successful with a faster rate, 3-5 days after culture. The morphological assessment and flow cytometry analysis confirmed the purity of SF-derived cells in passage 4.
SF could be a more accessible source of FLS than synovial tissue. Obtaining primary FLS from SF is a simple, fast, and cost-effective way to have a large-scale cell during a short time.
细胞培养技术已成为细胞生物学、药理学和医学研究领域中一种流行的方法。原代细胞代表了人类细胞的正常生理状态。成纤维细胞是结缔组织中最常见的天然细胞，在各种疾病的整个发病机制中起着至关重要的作用，如类风湿关节炎（RA）。滑膜成纤维细胞（FLS）覆盖在滑膜下疏松结缔组织上，已知是关节损伤的中心介质。分离 FLS 的最常规方法是对滑膜组织进行酶消化。本实验研究旨在介绍一种与酶消化相比，更简单、快速和高通量的方法来分离 FLS。
从 8 名接受常规膝关节置换手术的 RA 患者中收集滑膜组织和滑液（SF）样本。用胶原酶 VIII 酶孵育滑膜组织，而 SF 则用相似体积的磷酸盐缓冲盐水冲洗。根据标准方案进一步进行细胞传代培养和储存。使用流式细胞术分析确认分离的滑膜细胞的纯度。
SF 中 FLS 的分离成功率更高，培养 3-5 天后即可成功。形态学评估和流式细胞术分析证实了第 4 代 SF 来源细胞的纯度。
SF 可能是比滑膜组织更容易获得 FLS 的来源。从 SF 中获得原代 FLS 是一种简单、快速且具有成本效益的方法，可以在短时间内获得大量细胞。
