Institutes of Immunology and Pathology, Rikshospitalet University Hospital, 0027 Oslo, Norway.
Knee Surg Sports Traumatol Arthrosc. 2010 Oct;18(10):1407-16. doi: 10.1007/s00167-009-1017-4. Epub 2009 Dec 18.
Treatment of focal lesions of the articular cartilage of the knee using chondrocytes in a hyaluronic acid (HA) scaffold is already being investigated in clinical trials. An alternative may be to use mesenchymal stem cells (MSC). We have compared articular chondrocytes with MSC from human bone marrow (BM) and adipose tissue (AT), all cultured in HA scaffolds, for their ability to express genes and synthesize proteins associated with chondrogenesis. The cells were expanded in monolayer cultures. After seeding into the scaffold, the chondrocytes were maintained in medium, while the two MSC populations were given a chondrogenic differentiation medium. Chondrogenesis was assessed by real-time RT-PCR for chondrocyte-associated genes, by immunohistochemistry and by ELISA for collagens in the supernatant. Redifferentiation of the dedifferentiated chondrocytes in the HA scaffold was shown by a modest increase in type II collagen mRNA (COL2A1) and reduction in COL1A1. BM-MSC expressed 600-fold higher levels of COL2A1 than chondrocytes after 3 weeks in the scaffold. The levels of aggrecan (AGC1) and COL1A1 were similar for chondrocyte and BM-MSC scaffold cultures, while COL10A1 was higher in the BM-MSC. AT-MSC expressed levels of COL2A1 and COL1A1 similar to chondrocytes, but less AGC1 and COL10A1. Surprisingly, little collagen II protein was observed in the scaffold. Instead, collagen II was found in the culture medium. Chondrogenesis in HA scaffolds was more efficient using BM-MSC than AT-MSC or chondrocytes. Some of the secreted collagen II escaped entrapment in the extracellular space and was detected in the culture medium.
使用透明质酸 (HA) 支架中的软骨细胞治疗膝关节关节软骨局灶性病变已在临床试验中进行研究。另一种选择可能是使用间充质干细胞 (MSC)。我们比较了关节软骨细胞与来自人骨髓 (BM) 和脂肪组织 (AT) 的 MSC，所有细胞均在 HA 支架中培养，以评估它们表达与软骨生成相关的基因和合成蛋白质的能力。细胞在单层培养物中扩增。接种到支架中后，将软骨细胞维持在培养基中，而两种 MSC 群体则给予软骨分化培养基。通过实时 RT-PCR 评估软骨细胞相关基因、免疫组织化学和上清液中的胶原蛋白酶联免疫吸附试验 (ELISA) 评估软骨生成。在 HA 支架中，通过适度增加 II 型胶原 mRNA (COL2A1) 和降低 COL1A1，来证明去分化软骨细胞的再分化。BM-MSC 在支架中培养 3 周后，COL2A1 的表达水平比软骨细胞高 600 倍。软骨细胞和 BM-MSC 支架培养物的 AGC1 和 COL1A1 水平相似，而 BM-MSC 的 COL10A1 水平较高。AT-MSC 的 COL2A1 和 COL1A1 表达水平与软骨细胞相似，但 AGC1 和 COL10A1 水平较低。令人惊讶的是，支架中几乎没有观察到 II 型胶原蛋白蛋白。相反，在培养基中发现了 II 型胶原蛋白。与 AT-MSC 或软骨细胞相比，HA 支架中的 BM-MSC 软骨生成效率更高。一些分泌的 II 型胶原蛋白逃脱了细胞外空间的捕获，并在培养基中检测到。
