INCORPORATING PTGF-β1/CALCIUM PHOSPHATE NANOPARTICLES WITH FIBRONECTIN INTO 3-DIMENSIONAL COLLAGEN/CHITOSAN SCAFFOLDS: EFFICIENT, SUSTAINED GENE DELIVERY TO STEM CELLS FOR CHONDROGENIC DIFFERENTIATION
INCORPORATING PTGF-β1/CALCIUM PHOSPHATE NANOPARTICLES WITH FIBRONECTIN INTO 3-DIMENSIONAL COLLAGEN/CHITOSAN SCAFFOLDS: EFFICIENT, SUSTAINED GENE DELIVERY TO STEM CELLS FOR CHONDROGENIC DIFFERENTIATION
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DOI：
10.22203/ecm.v023a06
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发表时间：
2012-01-01
影响因子：
3.1
通讯作者：
Yu, Jiangnan
中科院分区：
文献类型：
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作者：
Cao, Xia;Deng, Wenwen;Yu, Jiangnan
本研究的目的是制备基于胶原/壳聚糖支架的3维纳米颗粒基因递送系统（3D-NGDS），其中质粒转化生长因子β1（TGF-β1）/磷酸钙纳米颗粒与纤连蛋白（FN）混合用于转染间充质干细胞（MSC）。使用扫描电子显微镜来表征 3 维胶原/壳聚糖支架的微观结构。对 MSC 培养物中 TGF-β 1 浓度进行量化的分析显示，转染 3D-NGDS 的 MSC 在较长时间内表现出非常高水平的 TGF-β 1，在两周内保持约 10 ng/mL 的 TGF-β 1 浓度，并在第 6 天观察到最高水平（12.6 ng/mL）。对 II 型胶原蛋白的免疫组织化学分析显示，从第 9 天到第 15 天，在 3D-NGDS 转染的 MSC 中观察到的 II 型胶原蛋白的产生量比通过 Lipofectamine 2000 方法转染的 MSC 中高得多。 3D-NGDS 的糖胺聚糖含量与用 TGF-β 1 以及 TGF-β 1 加地塞米松处理的那些相当，并且显着高于用游离质粒和 Lipofectamine 2000 处理的那些。通过 ELISA 观察到第 21 天时 3D-NGDS 的 I 型胶原表达受到显着抑制。这些结果表明，用3D-NGDS转染无需地塞米松即可在体外成功诱导MSC软骨形成分化。总之，3D-NGDS 可以发展成为临床试验中将外源核酸转移到 MSC 的一种有前途的替代方法。
The objective of this study was to prepare a 3-dimensional nanoparticle gene delivery system (3D-NGDS) based on collagen/chitosan scaffolds, in which plasmid transforming growth factor beta 1 (TGF-beta 1)/calcium phosphate nanoparticles mixed with fibronectin (FN) were used to transfect mesenchymal stem cells (MSCs). Scanning electron microscopy was used to characterise the microstructure of 3-dimensional collagen/chitosan scaffolds. An analysis performed to quantify the TGF-beta 1 concentrations in MSC cultures revealed that the MSCs transfected with the 3D-NGDS showed remarkably high levels of TGF-beta 1 over long periods, retaining a concentration of TGF-beta 1 of approximately 10 ng/mL within two weeks, with the highest level (12.6 ng/mL) being observed on the 6th day. An immunohistochemistry analysis for collagen type II revealed that much higher production of collagen II from the 9th to 15th day was observed in the 3D-NGDS-transfected MSCs than that in MSCs transfected by the Lipofectamine 2000 method. The glycosaminoglycan content of the 3D-NGDS was comparable to those treated with TGF-beta 1 as well as TGF-beta 1 plus dexamethasone, and was significantly higher than those treated with free plasmid and Lipofectamine 2000. A remarkable type I collagen expression inhibition of the 3D-NGDS at day 21 was observed via ELISA. These results suggested that transfection with the 3D-NGDS could successfully induce MSC chondrogenic differentiation in vitro without dexamethasone. In summary, the 3D-NGDS could be developed into a promising alternative method to transfer exogenous nucleic acid to MSCs in clinical trials.