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Biologic response of human intervertebral disc cells to gene therapy cocktail

 Seong-Hwan Moon  ;  Kotaro Nishida  ;  Lars G. Gilbertson  ;  Hwan-Mo Lee  ;  Hyang Kim  ;  Ronald A. Hall  ;  Paul D. Robbins  ;  James D. Kang 
 SPINE, Vol.33(17) : 1850-1855, 2008 
Journal Title
Issue Date
Adenoviridae/genetics ; Adult ; Cells, Cultured ; Gene Transfer Techniques ; Genetic Therapy/methods* ; Genetic Vectors/administration & dosage ; Genetic Vectors/genetics ; Humans ; Intervertebral Disc/cytology* ; Intervertebral Disc/physiology* ; Middle Aged ; Proteoglycans/biosynthesis ; Proteoglycans/genetics
gene therapy ; intervertebral disc ; proteoglycan
STUDY DESIGN: In vitro experiment using human intervertebral disc (IVD) cells and adenovirus-therapeutic gene constructs. OBJECTIVE: To examine the biologic effect of "cocktail" therapeutic gene transfer to human IVD cells in three-dimensional cultures. SUMMARY OF BACKGROUND DATA: Gene therapy is regarded as a potential option for the treatment of degenerative disc disease. Although various anabolic genes have previously been introduced for this purpose, cocktail gene transfer of anabolic genes to IVD cells has never been attempted. METHODS: Human IVDs were harvested during surgical disc procedures and cultured. We prepared recombinant adenovirus constructs bearing the TGF-beta1 gene (Ad/TGF-beta1), the IGF-1 gene (Ad/IGF-1), and the BMP-2 gene (Ad/BMP-2). Transgene expression was detected by luciferase assays, enzyme linked immunosorbent assays, and Western blot analysis. Newly synthesized proteoglycan was measured by S-sulfate incorporation on Sephadex G-25 M in PD 10 columns. Human IVD cells were transduced by single, double, and triple combination of Ad/TGF-beta1, Ad/IGF-1, Ad/BMP-2 with an MOI of 75, then cultured three-dimensionally in alginate beads. RESULTS: Transgene expression was detected at 18 hours after viral transduction. IVD cultures with Ad/TGF-beta1, Ad/IGF-1, Ad/BMP-2 (MOI of 75) showed 2.9, 1.8, and 1.9 fold increases, respectively, in proteoglycan synthesis compared to control. Human IVD cultures with double gene combination (MOI of 75) showed 3.2 to 3.9 fold increases of proteoglycan synthesis. Lastly, Human IVD cultures with triple gene combination (TGF-beta1+IGF-1+BMP-2 genes with an MOI of 75) transfer demonstrated 4.7 fold increase in proteoglycan synthesis compared control. CONCLUSION: Combination or "cocktail" gene therapy offers a promising mechanism for maximizing matrixsynthesis with low dose of adenoviral mixtures, circumventing systemic, local toxic effect, and immune response.
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1. College of Medicine (의과대학) > Dept. of Orthopedic Surgery (정형외과학교실) > 1. Journal Papers
Yonsei Authors
Moon, Seong Hwan(문성환)
Lee, Hwan Mo(이환모) ORCID logo https://orcid.org/0000-0002-5405-3832
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