Kawamura Koichiro, Chu Constance R, Sobajima Satoshi, Robbins Paul D, Fu Freddie H, Izzo Nicholas J, Niyibizi Christopher
Department of Orthopaedic Surgery, University of Pittsburgh, Pittsburgh, PA, USA.
Exp Hematol. 2005 Aug;33(8):865-72. doi: 10.1016/j.exphem.2005.05.010.
The objective of the present study was to investigate the potential of application of growth factor genes to induce chondrogenic differentiation of human-derived mesenchymal stem cells (MSCs). The growth factor genes evaluated in the present study were transforming growth factor 1 (TGF-beta1) and insulin-like growth factor 1 (IGF-1).
Human MSCs were transduced with the adenoviral vectors carrying either TGF-beta1 or IGF-1 (AdTGF-beta1 and AdIGF-1 respectively) or a combination of both growth factor genes at different multiplicities of infection (MOI) and were then made into pellets. Pellets were also made from nontransduced cells and maintained in culture medium supplemented with 10 ng/mL of TGF-beta1. At specified time points, histological analysis, cartilage matrix gene expression, and immunofluorescence were performed to determine the extent of chondrogenic differentiation.
MSCs transduced with the AdTGF-beta1 demonstrated robust chondrogenic differentiation, while those made from AdIGF-1 did not. AdTGF-beta1 pellets demonstrated aggrecan gene expression as early as day 3 of pellet culture, while type II collagen gene expression was detected by day 10 of culture. The AdIGF-1, alone or in combination with TGF-beta1 pellets, did not show any type II collagen gene expression at any time point. By immunofluoresecence, type X collagen was distributed throughout the matrix in TGF-beta1 protein pellets while the growth factor gene pellets displayed scant staining.
The results suggest that sustained administration of TGF-beta1 may be more effective in suppressing terminal differentiation than intermittent dosing and thus effective for cartilage repair.
本研究的目的是探讨生长因子基因在诱导人源间充质干细胞（MSCs）软骨分化方面的应用潜力。本研究中评估的生长因子基因是转化生长因子1（TGF-β1）和胰岛素样生长因子1（IGF-1）。
用人骨髓间充质干细胞转导携带TGF-β1或IGF-1的腺病毒载体（分别为AdTGF-β1和AdIGF-1）或两种生长因子基因的组合，感染复数不同，然后制成微球。未转导的细胞也制成微球，并在补充有10 ng/mL TGF-β1的培养基中培养。在特定时间点，进行组织学分析、软骨基质基因表达和免疫荧光检测，以确定软骨分化程度。
用AdTGF-β1转导的MSCs表现出强烈的软骨分化，而用AdIGF-1转导的则没有。AdTGF-β1微球在微球培养第3天就显示出聚集蛋白聚糖基因表达，而在培养第10天检测到II型胶原基因表达。AdIGF-1单独或与TGF-β1微球联合使用，在任何时间点都未显示出任何II型胶原基因表达。通过免疫荧光，X型胶原分布在TGF-β1蛋白微球的整个基质中，而生长因子基因微球染色较少。
结果表明，持续给予TGF-β1在抑制终末分化方面可能比间歇给药更有效，因此对软骨修复有效。
