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Pharmaceutical Research

, Volume 28, Issue 6, pp 1338–1348 | Cite as

Optimal Construction and Delivery of Dual-Functioning Lentiviral Vectors for Type I Collagen-Suppressed Chondrogenesis in Synovium-Derived Mesenchymal Stem Cells

  • Feng Zhang
  • Yongchang Yao
  • Ruijie Zhou
  • Kai Su
  • Fudiman Citra
  • Dong-An Wang
Research Paper

ABSTRACT

Purpose

This study aims to deliver both transforming growth factor β3 (TGF-β3) and shRNA targeting type I collagen (Col I) by optimal construction and application of various dual-functioning lentiviral vectors to induce Col I-suppressed chondrogenesis in synovium-derived mesenchymal stem cells (SMSCs).

Methods

We constructed four lentiviral vectors (LV-1, LV-2, LV-3 and LV-4) with various arrangements of the two expression cassettes in different positions and orientations. Col I inhibition efficiency and chondrogenic markers were assessed with qPCR, ELISA and staining techniques. Among the four vectors, LV-1 has two distant and reversely oriented cassettes, LV-2 has two distant and same-oriented cassettes, LV-3 has two proximal and reversely oriented cassettes, and LV-4 has two proximal and same-oriented cassettes. Col I and chondrogenic markers, including type II collagen (Col II), aggrecan and glycosaminoglycan (GAG), were examined in SMSCs cultured in 3-D alginate hydrogel.

Results

All of the four vectors showed distinct effects in Col I level as well as diverse inductive efficiencies in upregulation of the cartilaginous markers. Based on real-time PCR results, LV-1 was optimal towards Col I-suppressed chondrogenesis.

Conclusion

LV-1 vector is competent to promote Col I-suppressed chondrogenesis in SMSCs.

KEY WORDS

dual-functioning lentiviral vector synovial mesenchymal stem cells (SMSCs) TGF-β3 type I collagen chondrogenesis 

ABBREVIATIONS

Col I

type I collagen

Col II

type II collagen

LV-dual

recombinant lentiviral vector that expresses both shRNA targeting type I collagen and TGF- β3

LV-T

recombinant lentiviral vector that specifically expresses TGF-β3

shRNA

short hairpin RNA

SMSC

synovium-derived mesenchymal stem cell

TGF-β3

transforming growth factor-β3

Notes

ACKNOWLEDGEMENTS

This research was financially supported by AcRF Tier 1 Grant RG64/08, Ministry of Education (MoE), and NMRC/EDG/1001/2010, Singapore.

Supplementary material

11095_2010_305_MOESM1_ESM.docx (375 kb)
Esm 1 (DOCX 374 kb)

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Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Feng Zhang
    • 1
  • Yongchang Yao
    • 1
    • 2
    • 3
  • Ruijie Zhou
    • 1
  • Kai Su
    • 1
  • Fudiman Citra
    • 1
  • Dong-An Wang
    • 1
    • 3
  1. 1.Division of Bioengineering School of Chemical and Biomedical EngineeringNanyang Technological UniversitySingaporeRepublic of Singapore
  2. 2.School of Materials Science and EngineeringSouth China University of TechnologyGuangzhouChina
  3. 3.Key Laboratory of Specially Functional Materials and Advanced Manufacturing Technology, Ministry of EducationSouth China University of TechnologyGuangzhouChina

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