ABSTRACT

Purpose

Resident articular stem cells isolated using a migratory assay called Migratory Chondroprogenitors (MCPs) have emerged as a promising cellular therapeutic for the treatment of cartilage pathologies. In-vivo studies using MCPs report their superiority over bone-marrow mesenchymal stem cells and chondrocytes for treating chondral defects. However, there is no consensus on their isolation protocol. This study aimed to compare four reported isolation methods of MCPs and identify the optimal and feasible protocol for future translational work.

Methods

Human MCPs isolated from osteoarthritic cartilage (n = 3) were divided into four groups: a) MCP1: 8–15 mm cartilage explants, b) MCP2: 8–10 mm explants digested in 0.1% collagenase for 2 hrs. and cultured c) MCP3: 1 mm cartilage explants and d) MCP 4: 25 mm explants with a X tear, 7-day culture, and trypsinization to release migrated cells. The MCPs were subjected to the following analysis: growth kinetics, surface marker expression, mRNA gene expression for markers of chondrogenesis and hypertrophy, and trilineage differentiation.

Results

MCPs isolated via the four methods showed similar surface marker profiles, chondrogenic (SOX-9, ACAN, COL2A1) and hypertrophic (COL1, RUNX2) gene expression. The migration time for the MCP3 group was the longest. The MCP1, MCP2, and MCP4 groups produced MCPs with comparable cellular expansion feasibility.

Conclusions

MCPs can be preferably isolated by the any of the three above methods based on the investigator’s discretion. In the case of small cartilage samples similar to the MCP3 group, the isolation of MCP is plausible, keeping in mind the additional time required.

中文翻译：

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人关节软骨迁移性软骨祖细胞分离培养方法的比较

摘要

目的

使用称为迁移软骨祖细胞（MCP）的迁移测定分离的驻留关节干细胞已成为治疗软骨病理学的有前途的细胞疗法。使用 MCP 的体内研究表明，它们在治疗软骨缺损方面优于骨髓间充质干细胞和软骨细胞。然而，他们的隔离方案尚未达成共识。本研究旨在比较四种报道的 MCP 分离方法，并为未来的转化工作确定最佳且可行的方案。

方法

从骨关节炎软骨中分离出的人 MCP（n  = 3）分为四组：a) MCP1：8–15 mm 软骨外植体，b) MCP2：8–10 mm 外植体在 0.1% 胶原酶中消化 2 小时。c) MCP3：1 mm 软骨外植体和 d) MCP 4：25 mm 外植体，X 形撕裂，培养 7 天，胰蛋白酶消化以释放迁移的细胞。对 MCP 进行以下分析：生长动力学、表面标记表达、软骨形成和肥大标记的 mRNA 基因表达以及三系分化。

结果

通过四种方法分离的 MCP 显示出相似的表面标记特征、软骨形成（SOX-9、ACAN、COL2A1）和肥大（COL1、RUNX2）基因表达。MCP3组的迁移时间最长。MCP1、MCP2 和 MCP4 组生产的 MCP 具有相当的细胞扩增可行性。

结论

根据研究者的判断，MCP 可以优选通过上述三种方法中的任何一种来分离。对于类似于 MCP3 组的小软骨样品，考虑到所需的额外时间，MCP 的分离是合理的。