Center of Experimental Orthopaedics, Saarland University Medical Center, Homburg, Germany.
Am J Sports Med. 2013 Jan;41(1):194-202. doi: 10.1177/0363546512465840. Epub 2012 Nov 19.
Basic fibroblast growth factor (FGF-2) is a powerful stimulator of fibroblast proliferation and type I/III collagen production.
Overexpression of FGF-2 via direct recombinant adeno-associated virus (rAAV) vector-mediated gene transfer enhances the healing of experimental lesions to the human anterior cruciate ligament (ACL).
Controlled laboratory study.
rAAV vectors carrying a human FGF-2 sequence or the lacZ marker gene were applied to primary human ACL fibroblasts in vitro and to intact or experimentally injured human ACL explants in situ to evaluate the efficacy and duration of transgene expression and the potential effects of FGF-2 treatment upon the proliferative, metabolic, and regenerative activities in these systems.
Sustained, effective dose-dependent lacZ expression was achieved in all systems tested (up to 96% ± 2% in vitro and 80%-85% in situ for at least 30 days). rAAV allowed for continuous FGF-2 production both in vitro and in the intact ACL in situ (32.7 ± 1.4 and 33.1 ± 0.8 pg/mL/24 h, respectively, ie, up to 41-fold more than in the controls at day 30; always P ≤ .001), leading to significantly and durably enhanced levels of proliferation and type I/III collagen production vis-à-vis lacZ (at least 3- and 4-fold increases at day 30, respectively; always P ≤ .001). Most notably, rAAV FGF-2 promoted a significant, long-term production of the factor in experimental ACL lesions (92.7 ± 3.9 pg/mL/24 h, ie, about 5-fold more than in the controls; P ≤ .001) associated with enhanced levels of proliferation and type I/III collagen synthesis (at least 2- and 4-fold increases at day 30, respectively; always P ≤ .001). Remarkably, the FGF-2 treatment allowed for a decrease in the amplitude of such lesions possibly because of the increased expression in contractile α-smooth muscle actin, ligament-specific transcription factor scleraxis, and nuclear factor-κB for proliferation and collagen deposition, which are all markers commonly induced in response to injury.
Efficient, stable FGF-2 expression via rAAV enhances the healing of experimental human ACL lesions by activating key cellular and metabolic processes.
This approach has potential value for the development of novel, effective treatments for ligament reconstruction.
碱性成纤维细胞生长因子（FGF-2）是一种强大的成纤维细胞增殖和 I/III 型胶原产生刺激物。
通过直接重组腺相关病毒（rAAV）载体介导的基因转移过表达 FGF-2，可增强实验性前交叉韧带（ACL）损伤的愈合。
对照实验室研究。
rAAV 载体携带人 FGF-2 序列或 lacZ 标记基因，用于体外原代人 ACL 成纤维细胞和完整或实验性损伤的人 ACL 外植体原位，以评估转基因表达的有效性和持续时间，以及 FGF-2 处理对这些系统中增殖、代谢和再生活性的潜在影响。
在所测试的所有系统中均实现了持续、有效的剂量依赖性 lacZ 表达（体外高达 96%±2%，原位至少 30 天为 80%-85%）。rAAV 允许在体外和完整的 ACL 原位中持续产生 FGF-2（分别为 32.7±1.4 和 33.1±0.8 pg/mL/24 h，即第 30 天比对照组多 41 倍；始终 P≤0.001），导致增殖和 I/III 型胶原产生的水平显著且持久增加与 lacZ 相比（第 30 天分别增加至少 3 倍和 4 倍；始终 P≤0.001）。值得注意的是，rAAV FGF-2 可促进实验性 ACL 损伤中因子的长期大量产生（92.7±3.9 pg/mL/24 h，即比对照组多约 5 倍；P≤0.001），同时伴有增殖和 I/III 型胶原合成水平的增加（第 30 天分别增加至少 2 倍和 4 倍；始终 P≤0.001）。值得注意的是，FGF-2 治疗可能会降低这种损伤的幅度，这可能是由于收缩性α-平滑肌肌动蛋白、韧带特异性转录因子 Scleraxis 和增殖和胶原沉积的核因子-κB 的表达增加，这些都是对损伤的常见反应诱导的标志物。
通过 rAAV 实现高效、稳定的 FGF-2 表达可通过激活关键的细胞和代谢过程来增强实验性人 ACL 损伤的愈合。
这种方法对于开发新型有效的韧带重建治疗方法具有潜在价值。
