Abstract
The purpose of this study was to develop a fibrin gel system capable of serving as a three dimensional scaffold for the chondrogenesis of rabbit bone marrow mesenchymal stem cells (BM-MSCs) and to examine the effect of two fibrinolytic inhibitors, aprotinin and aminohexanoic acid, on this system. Rabbit BM-MSCs were obtained from the tibias and femurs of New Zealand white rabbits. After chondrogenic potential of BM-MSCs was verified by pellet culture, 2 x 106 cells were pelleted and suspended in fibrinogen (80mg/ml) and then mixed with equal parts of thrombin (5 IU/ml). The specimen were then divided into four groups: aprotinin control (with aprotinin); aprotinin + transforming growth factor (TGF-beta) (with aprotinin and TGF-beta 1); amino control (with aminohexanoic acid); and amino+TGF-beta (with aminohexanoic acid and TGF- beta1). Each of these groups was further divided into three groups depending on the concentration of the inhibitor. Both of the aprotinin groups received 0.0875, 0.175, or 0.35 TIU/ml of aprotinin and both of the aminohexanoic acid groups were supplemented with 2, 4, or 8 mg/ml of aminohexanoic acid. The gels were harvested and analyzed at 7, 14, and 21 days. All of the aprotinin+TGF-beta groups exhibited a significantly higher aggrecan gene expression than control groups whereas only the amino+TGF-รข group treated with 8mg/ml was significantly higher than those of the control groups. In addition, the 0.0875 and 0.175 TIU/ml aprotinin+TGF-beta groups exhibited significantly higher levels of expression than the 2 and 4 mg/ml amino+TGF-beta groups. There were no significant differences among the different concentrations of aprotinin or aminohexanoic acid with or without the treatment of TGF-beta. Similar trends were also seen when the glycosaminoglycan (GAG) content was measured and analyzed. These findings suggest that fibrin gels are a suitable environment for the chondrogenesis of BM-MSCs and that aprotinin in combination with TGF-beta1 is the optimal condition for stimulating BM-MSCs to differentiate into chondrocytes.