Institute of Sports Medicine, Peking University Third Hospital, Beijing, China.
Arthroscopy. 2013 Dec;29(12):2001-2011.e2. doi: 10.1016/j.arthro.2013.09.076.
To evaluate the effect of poly(lactic-co-glycolic acid) (PLGA) nanoparticles delivering pDC316-BMP4-EGFP plasmid into rabbit adipose-derived stem cells (ADSCs) in vitro and chondrogenesis of the bone morphogenetic protein 4 (BMP-4)--transfected ADSCs seeded onto poly(L-lactic-co-glycolic acid) (PLLGA) scaffold in a rabbit model.
Cell viability and transfection efficiency of PLGA nanoparticles were measured by Cell Counting Kit-8 (Dojindo, Kumamoto, Japan) and flow cytometry. The BMP-4 and chondrogenesis markers were detected by real-time polymerase chain reaction and enzyme-linked immunosorbent assay. Thirty rabbits (60 knees) with full-thickness cylinder articular cartilage defects (diameter, 4.5 mm; depth, 0.8 mm) on the femoral trochlea were divided into a group in which the BMP-4--transfected ADSCs were seeded onto PLLGA scaffold and implanted into the defects (group ABNP), a group with untransfected ADSCs seeded onto scaffold (group ABP), and a group with a scaffold without cells (group P). Outcomes were evaluated by histology, Rudert score, Pineda score, and scanning electronic microscopy by 2 blinded observers at weeks 6 and 12 postoperatively. Statistical analyses were performed with analysis of variance and the Kruskal-Wallis test. The statistical significance level was set at P < .05.
The expression of chondrogenesis-related genes and proteins was significantly increased in BMP-4--transfected ADSCs in vitro (P < .05). The cell viability was 79.86% ± 5.04% after 24 hours. The transfection efficiency was 25.86% ± 4.27% after 72 hours. Defects in group ABNP showed the best in vivo cartilage regeneration. At week 12, the Rudert scores in group ABNP (7.00 ± 1.75) were better than those in group ABP (6.00 ± 2.00) or group P (5.00 ± 1.75) (P < .05), as were the Pineda scores (2.50 ± 3.00, 5.00 ± 2.00, and 6.00 ± 1.75, respectively; P < .001).
BMP-4 plasmid can be successfully delivered into ADSCs by PLGA nanoparticles and promoted in vitro chondrogenesis. When compared with the control cells, BMP-4--transfected ADSCs seeded onto PLLGA scaffold significantly improve in vivo chondrogenesis in a rabbit articular defect model.
PLGA nanoparticles and BMP-4 have potential for gene therapy in the treatment of chondral defects of the knee.
评估聚（丙交酯-共-乙交酯）（PLGA）纳米粒将 pDC316-BMP4-EGFP 质粒递送至兔脂肪来源干细胞（ADSCs）体外的效果，以及骨形态发生蛋白 4（BMP-4）转染的 ADSCs 接种到聚（L-丙交酯-共-乙交酯）（PLLA）支架上在兔模型中的软骨生成。
通过细胞计数试剂盒（Dojindo，熊本，日本）和流式细胞术测量 PLGA 纳米粒的细胞活力和转染效率。通过实时聚合酶链反应和酶联免疫吸附试验检测 BMP-4 和软骨生成标志物。30 只（60 只膝关节）兔的股骨滑车有全层圆柱状关节软骨缺损（直径 4.5mm；深度 0.8mm），分为将 BMP-4 转染的 ADSCs 接种到 PLLGA 支架上并植入缺损的组（ABNP 组）、未转染 ADSCs 接种到支架上的组（ABP 组）和无细胞支架组（P 组）。术后 6 周和 12 周通过组织学、鲁德特评分、品田评分和扫描电子显微镜由 2 名盲法观察者进行评估。采用方差分析和 Kruskal-Wallis 检验进行统计学分析。统计显著性水平设为 P<.05。
体外 BMP-4 转染的 ADSCs 中软骨生成相关基因和蛋白的表达显著增加（P<.05）。转染后 24 小时细胞活力为 79.86%±5.04%。转染后 72 小时的转染效率为 25.86%±4.27%。ABNP 组的体内软骨再生效果最佳。在第 12 周，ABNP 组的鲁德特评分（7.00±1.75）优于 ABP 组（6.00±2.00）或 P 组（5.00±1.75）（P<.05），品田评分也是如此（2.50±3.00、5.00±2.00 和 6.00±1.75，分别；P<.001）。
PLGA 纳米粒可成功将 BMP-4 质粒递送至 ADSCs 并促进体外软骨生成。与对照细胞相比，BMP-4 转染的 ADSCs 接种到 PLLGA 支架上可显著改善兔关节软骨缺损模型中的体内软骨生成。
PLGA 纳米粒和 BMP-4 具有治疗膝关节软骨缺损的基因治疗潜力。
Zotero 插件