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Nonviral gene transfer to human meniscal cells. Part I: transfection analyses and cell transplantation to meniscus explants

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Abstract

Purpose

Our aim was to evaluate whether nonviral vectors can genetically modify primary human juvenile and adult meniscal fibrochondrocytes at low toxicity in vitro and to test the hypothesis that transfected human meniscal fibrochondrocytes transplanted into longitudinal defects and onto human medial meniscus explant cultures are capable of expressing transgene products in vitro.

Methods

Eighteen nonviral gene transfer systems were examined to identify the best suited method for an efficient transfection of primary cultures of juvenile and adult human meniscal fibrochondrocytes using luciferase and lacZ reporter gene constructs and then transplanted to meniscus explant cultures.

Results

Gene transfer systems FuGENE 6, GeneJammer, TurboFectin 8, calcium phosphate co-precipitates and GeneJuice led to minimal toxicity in both cell types. Nanofectin 2 and JetPEI resulted in maximal luciferase activity in both cell types. Maximal transfection efficiency based on X-gal staining following lacZ gene transfer was achieved using Lipofectamine 2000, revealing a mean transfection efficiency of 8.6 % in human juvenile and of 8.4 % in adult meniscal fibrochondrocytes. Transfected, transplanted meniscal fibrochondrocytes adhered to the meniscal tissue and continued to express the transgene for at least five days following transfection.

Conclusions

Nonviral gene transfer systems are safe and capable of transfecting both juvenile and adult human meniscal fibrochondrocytes, which, when transplanted to meniscal tissue in vitro, permit the expression of selected transgenes to be maintained. These results are of value for combining gene therapy and cell transplantation approaches as a means to enhance meniscal repair.

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References

  1. Aagaard H, Verdonk R (1999) Function of the normal meniscus and consequences of meniscal resection. Scand J Med Sci Sports 9(3):134–140

    Article  CAS  PubMed  Google Scholar 

  2. Al-Dosari MS, Gao X (2009) Nonviral gene delivery: principle, limitations, and recent progress. AAPS J 11(4):671–681. doi:10.1208/s12248-009-9143-y

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  3. Borovecki F, Pecina-Slaus N, Vukicevic S (2007) Biological mechanisms of bone and cartilage remodelling–genomic perspective. Int Orthop 31(6):799–805. doi:10.1007/s00264-007-0408-8

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  4. Chatain F, Robinson AH, Adeleine P, Chambat P, Neyret P (2001) The natural history of the knee following arthroscopic medial meniscectomy. Knee Surg Sports Traumatol Arthrosc 9(1):15–18

    Article  CAS  PubMed  Google Scholar 

  5. Cucchiarini M, Schetting S, Terwilliger EF, Kohn D, Madry H (2009) rAAV-mediated overexpression of FGF-2 promotes cell proliferation, survival, and alpha-SMA expression in human meniscal lesions. Gene Ther 16(11):1363–1372. doi:10.1038/gt.2009.91

    Article  CAS  PubMed  Google Scholar 

  6. Day CS, Kasemkijwattana C, Menetrey J, Floyd SS Jr, Booth D, Moreland MS, Fu FH, Huard J (1997) Myoblast-mediated gene transfer to the joint. J Orthop Res 15(6):894–903

    Article  CAS  PubMed  Google Scholar 

  7. Elsler S, Schetting S, Schmitt G, Kohn D, Madry H, Cucchiarini M (2012) Effective, safe nonviral gene transfer to preserve the chondrogenic differentiation potential of human mesenchymal stem cells. J Gene Med 14(7):501–511. doi:10.1002/jgm.2644

    Article  CAS  PubMed  Google Scholar 

  8. Englund M, Guermazi A, Lohmander LS (2009) The meniscus in knee osteoarthritis. Rheum Dis Clin North Am 35(3):579–590. doi:10.1016/j.rdc.2009.08.004

    Article  PubMed  Google Scholar 

  9. Evans CH, Ghivizzani SC, Robbins PD (2004) Orthopaedic gene therapy. Clin Orthop 429:316–329

    Article  PubMed  Google Scholar 

  10. Goto H, Shuler FD, Lamsam C, Moller HD, Niyibizi C, Fu FH, Robbins PD, Evans CH (1999) Transfer of lacZ marker gene to the meniscus. J Bone Joint Surg Am 81(7):918–925

    CAS  PubMed  Google Scholar 

  11. Greis PE, Bardana DD, Holmstrom MC, Burks RT (2002) Meniscal injury: I. Basic science and evaluation. J Am Acad Orthop Surg 10(3):168–176

    PubMed  Google Scholar 

  12. Heiligenstein S, Cucchiarini M, Laschke MW, Bohle RM, Kohn D, Menger MD, Madry H (2011) Evaluation of nonbiomedical and biomedical grade alginates for the transplantation of genetically modified articular chondrocytes to cartilage defects in a large animal model in vivo. J Gene Med 13(4):230–242. doi:10.1002/jgm.1557

    Article  CAS  PubMed  Google Scholar 

  13. Ivkovic A, Marijanovic I, Hudetz D, Porter RM, Pecina M, Evans CH (2011) Regenerative medicine and tissue engineering in orthopaedic surgery. Front Biosci (Elite Ed) 3:923–944

    Google Scholar 

  14. Ivkovic A, Pascher A, Hudetz D, Maticic D, Jelic M, Dickinson S, Loparic M, Haspl M, Windhager R, Pecina M (2010) Articular cartilage repair by genetically modified bone marrow aspirate in sheep. Gene Ther 17(6):779–789. doi:10.1038/gt.2010.16

    Article  CAS  PubMed  Google Scholar 

  15. Lind M, Bünger C (2005) Orthopaedic applications of gene therapy. Int Orthop 29(4):205–209. doi:10.1007/s00264-005-0650-x

    Article  PubMed Central  PubMed  Google Scholar 

  16. Lu H, Dai Y, Lv L, Zhao H (2014) Chitosan-graft-polyethylenimine/DNA nanoparticles as novel non-viral gene delivery vectors targeting osteoarthritis. PLoS One 9(1):e84703. doi:10.1371/journal.pone.0084703

    Article  PubMed Central  PubMed  Google Scholar 

  17. Madry H, Cucchiarini M, Kaul G, Kohn D, Terwilliger EF, Trippel SB (2004) Menisci are efficiently transduced by recombinant adeno-associated virus vectors in vitro and in vivo. Am J Sports Med 32(8):1860–1865

    Article  PubMed  Google Scholar 

  18. Madry H, Cucchiarini M, Stein U, Remberger K, Menger MD, Kohn D, Trippel SB (2003) Sustained transgene expression in cartilage defects in vivo after transplantation of articular chondrocytes modified by lipid-mediated gene transfer in a gel suspension delivery system. J Gene Med 5(6):502–509

    Article  CAS  PubMed  Google Scholar 

  19. Madry H, Kaul G, Cucchiarini M, Stein U, Zurakowski D, Remberger K, Menger MD, Kohn D, Trippel SB (2005) Enhanced repair of articular cartilage defects in vivo by transplanted chondrocytes overexpressing insulin-like growth factor I (IGF-I). Gene Ther 12(14):1171–1179

    Article  CAS  PubMed  Google Scholar 

  20. Madry H, Luyten FP, Facchini A (2012) Biological aspects of early osteoarthritis. Knee Surg Sports Traumatol Arthrosc 20(3):407–422. doi:10.1007/s00167-011-1705-8

    Article  PubMed  Google Scholar 

  21. Madry H, Orth P, Cucchiarini M (2011) Gene therapy for cartilage repair. Cartilage 2(3):201–225. doi:10.1007/s00167-010-1071-y

    Article  CAS  Google Scholar 

  22. Madry H, Trippel SB (2000) Efficient lipid-mediated gene transfer to articular chondrocytes. Gene Ther 7(4):286–291

    Article  CAS  PubMed  Google Scholar 

  23. Madry H, Zurakowski D, Trippel SB (2001) Overexpression of human insulin-like growth factor-I promotes new tissue formation in an ex vivo model of articular chondrocyte transplantation. Gene Ther 8(19):1443–1449

    Article  CAS  PubMed  Google Scholar 

  24. McDermott ID, Amis AA (2006) The consequences of meniscectomy. J Bone Joint Surg Br 88(12):1549–1556. doi:10.1302/0301-620X.88B12.18140

    Article  CAS  PubMed  Google Scholar 

  25. Nabel EG, Plautz G, Nabel GJ (1992) Transduction of a foreign histocompatibility gene into the arterial wall induces vasculitis. Proc Natl Acad Sci U S A 89(11):5157–5161

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  26. Nakata K, Shino K, Hamada M, Mae T, Miyama T, Shinjo H, Horibe S, Tada K, Ochi T, Yoshikawa H (2001) Human meniscus cell: characterization of the primary culture and use for tissue engineering. Clin Orthop (391 Suppl):S208-S218

  27. Narita A, Takahara M, Sato D, Ogino T, Fukushima S, Kimura Y, Tabata Y (2012) Biodegradable gelatin hydrogels incorporating fibroblast growth factor 2 promote healing of horizontal tears in rabbit meniscus. Arthroscopy 28(2):255–263. doi:10.1016/j.arthro.2011.08.294

    Article  PubMed  Google Scholar 

  28. Orth P, Weimer A, Kaul G, Kohn D, Cucchiarini M, Madry H (2008) Analysis of novel nonviral gene transfer systems for gene delivery to cells of the musculoskeletal system. Mol Biotechnol 38(2):137–144

    Article  CAS  PubMed  Google Scholar 

  29. Pecina M, Jelic M, Ivkovic A, Hudetz D (2006) Gene therapy applications in orthopaedics. Int Orthop 30(3):215–216. doi:10.1007/s00264-005-0047-x

    Article  PubMed Central  PubMed  Google Scholar 

  30. Shieh A, Bastrom T, Roocroft J, Edmonds EW, Pennock AT (2013) Meniscus tear patterns in relation to skeletal immaturity: children versus adolescents. Am J Sports Med 41(12):2779–2783. doi:10.1177/0363546513504286

    Article  PubMed  Google Scholar 

  31. Steinert AF, Palmer GD, Capito R, Hofstaetter JG, Pilapil C, Ghivizzani SC, Spector M, Evans CH (2007) Genetically enhanced engineering of meniscus tissue using ex vivo delivery of transforming growth factor-beta 1 complementary deoxyribonucleic acid. Tissue Eng 13(9):2227–2237. doi:10.1089/ten.2006.0270

    Article  CAS  PubMed  Google Scholar 

  32. Stilli S, Marchesini Reggiani L, Marcheggiani Muccioli GM, Cappella M, Donzelli O (2011) Arthroscopic treatment for symptomatic discoid lateral meniscus during childhood. Knee Surg Sports Traumatol Arthrosc 19(8):1337–1342. doi:10.1007/s00167-011-1440-1

    Article  PubMed  Google Scholar 

  33. van Beuningen HM, Glansbeek HL, van der Kraan PM, van den Berg WB (2000) Osteoarthritis-like changes in the murine knee joint resulting from intra-articular transforming growth factor-beta injections. Osteoarthritis Cartilage 8(1):25–33

    Article  PubMed  Google Scholar 

  34. Verdonk R (2011) The meniscus: past, present and future. Knee Surg Sports Traumatol Arthrosc 19(2):145–146. doi:10.1007/s00167-010-1333-8

    Article  PubMed  Google Scholar 

  35. Yang HY, Vonk LA, Licht R, van Boxtel AM, Bekkers JE, Kragten AH, Hein S, Varghese OP, Howard KA, Öner FC, Dhert WJ, Creemers LB (2013) Cell type and transfection reagent-dependent effects on viability, cell content, cell cycle and inflammation of RNAi in human primary mesenchymal cells. Eur J Pharm Sci 53:35–44. doi:10.1016/j.ejps.2013.12.006

    Article  PubMed  Google Scholar 

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Acknowledgments

We thank the members of the Department of Orthopaedic Surgery for providing human meniscal tissue.

Conflict of interest

The authors declare that they have no conflict of interest. The authors alone are responsible for the content and writing of the paper.

Authors’ contributions

HPL participated in the design of the study, carried out the transfection and cell viability studies and performed the statistical analysis. GK participated in its design, carried out the transplantation studies and performed the statistical analysis. MC participated in the study design and draft. HM conceived and designed the study, co-ordinated it and drafted the manuscript. All authors read and approved the final manuscript.

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Authors and Affiliations

  1. Center of Experimental Orthopaedics, Saarland University, 66421, Homburg, Germany

    Hsiao-Ping Lee, Gunter Kaul, Magali Cucchiarini & Henning Madry

  2. Department of Orthopaedic Surgery, Saarland University, 66421, Homburg, Germany

    Henning Madry

Authors
  1. Hsiao-Ping Lee
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  2. Gunter Kaul
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  3. Magali Cucchiarini
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  4. Henning Madry
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Corresponding author

Correspondence to Henning Madry.

Additional information

Presented in part at the 55th Annual Meeting of the Orthopaedic Research Society, 2009, Las Vegas, NV, USA

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Lee, HP., Kaul, G., Cucchiarini, M. et al. Nonviral gene transfer to human meniscal cells. Part I: transfection analyses and cell transplantation to meniscus explants. International Orthopaedics (SICOT) 38, 1923–1930 (2014). https://doi.org/10.1007/s00264-014-2410-2

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  • DOI: https://doi.org/10.1007/s00264-014-2410-2

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