Maeda Satoshi, Matsumoto Masaya, Segawa Kotaro, Iwamoto Kaori, Nakamura Norimasa
TWOCELLS Co., Ltd, 1-6-10 Deshio, Minami-ku, Hiroshima, 734-0001 Japan.
Department of Orthopaedics, Osaka University Graduate School of Medicine, 2-2 Yamadaoka, Suita, Osaka, 565-0871 Japan.
Cytotechnology. 2024 Oct;76(5):595-612. doi: 10.1007/s10616-024-00637-y. Epub 2024 Jul 2.
Synovial mesenchymal stem cells (sMSCs) have great potential for cartilage repair, but their therapeutic design to avoid adverse effects associated with unknown factors remains a challenge. In addition, because long-term preservation is indispensable to maintain high quality levels until implantation, it is necessary to reduce their fluctuations. This study aimed to investigate the properties and feasibility of novel scaffold-free tissue-engineered constructs using serum-free media and to develop long-term preservation methods. sMSCs were cultured in serum-free media, seeded at high density in a monolayer, and finally developed as a sheet-like construct called "gMSC1". The properties of frozen gMSC1 (Fro-gMSC1) were compared with those of refrigerated gMSC1 (Ref-gMSC1) and then examined by their profile. Chondrogenic differentiation potential was analyzed by quantitative real-time polymerase chain reaction and quantification of glycosaminoglycan content. Xenografts into the cartilage defect model in rats were evaluated by histological staining. gMSC1 showed nearly similar properties independent of the preservation conditions. The animal experiment demonstrated that the defect could be filled with cartilage-like tissue with good integration to the adjacent tissue, suggesting that gMSC1 was formed and replaced the cartilage. Furthermore, several chondrogenesis-related factors were significantly secreted inside and outside gMSC1. Morphological analysis of Fro-gMSC1 revealed comparable quality levels to those of fresh gMSC1. Thus, if cryopreserved, gMSC1, with no complicated materials or processes, could have sustained cartilage repair capacity. gMSC1 is a prominent candidate in novel clinical practice for cartilage repair, allowing for large quantities to be manufactured at one time and preserved for a long term by freezing.
The online version contains supplementary material available at 10.1007/s10616-024-00637-y.
滑膜间充质干细胞（sMSCs）在软骨修复方面具有巨大潜力，但其治疗设计要避免与未知因素相关的不良反应仍是一项挑战。此外，由于长期保存对于在植入前维持高质量水平必不可少，因此有必要减少其波动。本研究旨在探讨使用无血清培养基的新型无支架组织工程构建体的特性和可行性，并开发长期保存方法。将sMSCs在无血清培养基中培养，以高密度接种成单层，最终发育成一种称为“gMSC1”的片状构建体。将冷冻的gMSC1（Fro-gMSC1）的特性与冷藏的gMSC1（Ref-gMSC1）的特性进行比较，然后通过其特征进行检测。通过定量实时聚合酶链反应和糖胺聚糖含量定量分析软骨形成分化潜能。通过组织学染色评估大鼠软骨缺损模型中的异种移植物。gMSC1显示出几乎相似的特性，与保存条件无关。动物实验表明，缺损部位可被类似软骨的组织填充，并与相邻组织良好整合，这表明gMSC1形成并替代了软骨。此外，gMSC1内外均显著分泌几种与软骨形成相关的因子。Fro-gMSC1的形态学分析显示其质量水平与新鲜gMSC1相当。因此，如果进行冷冻保存，gMSC1无需复杂的材料或工艺，就可能具有持续的软骨修复能力。gMSC1是软骨修复新临床实践中的一个突出候选者，能够一次性大量制造并通过冷冻长期保存。
在线版本包含可在10.1007/s10616-024-00637-y获取的补充材料。
