De Bari Cosimo, Dell'Accio Francesco, Luyten Frank P
University Hospitals, Katholieke Universiteit Leuven, Leuven, Belgium.
Arthritis Rheum. 2004 Jan;50(1):142-50. doi: 10.1002/art.11450.
We previously reported the identification in a nude mouse assay of molecular markers predictive of the capacity of articular cartilage-derived cells (ACDCs) to form ectopic stable cartilage that is resistant to vascular invasion and endochondral ossification. In the present study, we investigated whether in vitro-differentiated mesenchymal stem cells (MSCs) from the synovial membrane (SM) express the stable-chondrocyte markers and form ectopic stable cartilage in vivo.
Chondrogenesis was induced in micromass culture with the addition of transforming growth factor beta1 (TGFbeta1). After acquisition of the cartilage phenotype, micromasses were implanted subcutaneously into nude mice. Alternatively, cells were released enzymatically and either replated in monolayer or injected intramuscularly into nude mice. Marker analysis was performed by quantitative reverse transcription-polymerase chain reaction. Cell death was detected with TUNEL assay.
Cartilage-like micromasses and released cells expressed the stable-chondrocyte markers at levels comparable with those expressed by stable ACDCs. The released cells lost chondrocyte marker expression by 24 hours in monolayer and failed to form cartilage when injected intramuscularly into nude mice. Instead, myogenic differentiation was detected. When intact TGFbeta1-treated micromasses were implanted subcutaneously, they partially lost their cartilage phenotype and underwent cell death and neoangiogenesis within 1 week. At later time points (15-40 days), we retrieved neither cartilage nor bone, and human cells were not detectable.
The chondrocyte-like phenotype of human SM MSCs, induced in vitro under specific conditions, appears to be unstable and is not sufficient to obtain ectopic formation of stable cartilage in vivo. Studies in animal models of joint surface defect repair are necessary to evaluate the stability of the SM MSC chondrocyte-like phenotype within the joint environment.
我们之前报道了在裸鼠实验中鉴定出可预测关节软骨来源细胞（ACDCs）形成异位稳定软骨能力的分子标志物，该软骨对血管侵入和软骨内成骨具有抗性。在本研究中，我们调查了体外分化的滑膜间充质干细胞（MSCs）是否表达稳定软骨细胞标志物并在体内形成异位稳定软骨。
在微团培养中添加转化生长因子β1（TGFβ1）诱导软骨形成。获得软骨表型后，将微团皮下植入裸鼠体内。或者，通过酶解法释放细胞，将其重新接种到单层培养中或肌肉注射到裸鼠体内。通过定量逆转录-聚合酶链反应进行标志物分析。用TUNEL法检测细胞死亡。
软骨样微团和释放的细胞表达稳定软骨细胞标志物的水平与稳定ACDCs表达的水平相当。释放的细胞在单层培养24小时后失去软骨细胞标志物表达，肌肉注射到裸鼠体内时未能形成软骨。相反，检测到了肌源性分化。当完整的经TGFβ1处理的微团皮下植入时，它们在1周内部分失去软骨表型，发生细胞死亡和新生血管形成。在后期时间点（15 - 40天），我们既未检测到软骨也未检测到骨，且未检测到人类细胞。
在特定条件下体外诱导的人滑膜间充质干细胞的软骨样表型似乎不稳定，不足以在体内获得异位稳定软骨的形成。有必要在关节表面缺损修复的动物模型中进行研究，以评估滑膜间充质干细胞软骨样表型在关节环境中的稳定性。
