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Effective Gene Delivery to Mesenchymal Stem Cells Based on the Reverse Transfection and Three-Dimensional Cell Culture System

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ABSTRACT

Purpose

To enhance the level and prolong the duration of gene expression for gene-engineered rat mesenchymal stem cells (MSCs) using non-viral vector.

Methods

A novel transfection system based on reverse transfection method and three-dimensional (3D) scaffold was developed. The reverse gene transfection system was evaluated for transfection efficiency compared to conventional methods. Collagen sponge and polyethylene terephthalate non-woven fabric were introduced as scaffolds to perform 3D culture with reverse transfection. pDNA coding TGFβ-1 was delivered to MSCs to assess its ability in inducing chondrogenesis with the 3D non-viral reverse transfection system.

Results

The reverse transfection method induced higher transgene levels than the conventional transfection in the presence of serum. The electric charge of the anionic gelatin plays an important role in this system by affecting the release pattern of the gene complexes and through the adsorption of serum protein to the substrate. During a long-time in vitro culture, MSCs cultured on 3D scaffolds exhibited a higher transgene expression level and more sustained transgene expression than those cultured and transfected on the two-dimensional substrate.

Conclusions

The combination of reverse transfection system with 3D cell culture scaffold benefits the cell proliferation and long-time gene transfection of MSCs.

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Abbreviations

BCA:

bicinchonic acid

CDI:

N, N’—carbonyldiimidazole

CPZ:

chlorpromazine

DMEM:

Dulbecco’s modified Eagle’s medium

MSCs:

mesenchymal stem cells

PEI:

poly (ethylene-imine)

PET:

polyethylene terephthalate

TGFβ-1:

transforming growth factor β-1

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ACKNOWLEDGMENTS

This work was financially supported by National Natural Science Foundation of China (30873173, 30973648 by Jian-Qing Gao, 81001410 by Yu-Lan Hu); Zhejiang Provincial Natural Science Foundation of China (R2090176 by Jian-Qing Gao); China-Japan Scientific Cooperation Program (81011140077 by Jian-Qing Gao and Yasuhiko Tabata); NSFC, China; JSPS, Japan; the Fundamental Research Funds for the Central Universities (by Jian-Qing Gao); the Foundation of Hangzhou Health Bureau (2009B05 by Gang Wang), and Zijin program of Zhejiang University (188020-544802[78] by Min Han). We thank Ms. Hong-Yu Lu for technical assistance.

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Authors and Affiliations

  1. Institute of Pharmaceutics, College of Pharmaceutical Sciences, Zhejiang University, 388 Yuhangtang Road, Hangzhou, 310058, People’s Republic of China

    Cai-Xia He, Ni Li, Yu-Lan Hu, Xiu-Mei Zhu, Hai-Jie Li, Min Han, Wen-Quan Liang & Jian-Qing Gao

  2. Department of Pharmacy, Zhejiang Provincial Corps Hospital of Chinese People’s Armed Police Forces, Jiaxing, 314000, People’s Republic of China

    Pei-Hong Miao & Zhong-Jie Hu

  3. Department of Pharmacy, First People’s Hospital of Hangzhou, Hangzhou, 310006, People’s Republic of China

    Gang Wang

  4. Department of Biomaterials, Field of Tissue Engineering Institute for Frontier Medical Sciences, Kyoto University, 53 Kawanara-cho, Shogoin, Sakyo-ku, 606-8507, Kyoto, Japan

    Yasuhiko Tabata

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  1. Cai-Xia He
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  2. Ni Li
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Correspondence to Jian-Qing Gao.

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He, CX., Li, N., Hu, YL. et al. Effective Gene Delivery to Mesenchymal Stem Cells Based on the Reverse Transfection and Three-Dimensional Cell Culture System. Pharm Res 28, 1577–1590 (2011). https://doi.org/10.1007/s11095-011-0390-0

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  • DOI: https://doi.org/10.1007/s11095-011-0390-0

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