Advances in Clinical and Experimental Medicine

Title abbreviation: Adv Clin Exp Med
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ISSN 1899–5276 (print)
ISSN 2451-2680 (online)
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Advances in Clinical and Experimental Medicine

2020, vol. 29, nr 6, June, p. 639–647

doi: 10.17219/acem/121509

Publication type: original article

Language: English

License: Creative Commons Attribution 3.0 Unported (CC BY 3.0)

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In vivo delivery of MMP3-shRNA and Sox9 lentivirus cocktail enhances matrix synthesis to prevent lumbar disc degeneration

Zheng Zhao1,A,B,C,D,E,F, Siyuan Li2,A,B,C,D,F, Hui Huang1,A,B,F, Jing Fang1,B,C,F, Huawei Wei1,B,C,F, Yongming Xi1,A,B,C,D,E,F

1 Department of Orthopedics, Affiliated Hospital of Qingdao University, China

2 Department of Orthopedics, Shandong Provincial Third Hospital, Jinan, China


Background. Intervertebral disc degeneration (IDD) is characterized by increased proteolytic degradation of the extracellular matrix (ECM), leading to a loss of collagen II and proteoglycan in the nucleus pulposus (NP). Although MMP3 has been reported to play a central role in disc degeneration, it is still unknown whether gene therapy targeting MMP3 can inhibit IDD.
Objectives. To investigate whether lentivirus-mediated MMP3 knockdown is capable of attenuating IDD. More importantly, we also explored whether combined gene therapy that simultaneously antagonizes MMP3 and overexpresses Sox9 can synergistically inhibit IDD and induce augmented matrix reconstitution in the degenerative NP.
Material and Methods. We performed direct injection of lentiviral vectors LV-MMP3-shRNA and/or LV-Sox9 into rabbit lumbar discs. The animals were scanned using magnetic resonance imaging (MRI) at 8, 12 and 24 weeks after the operation. We also evaluated the gene expression and synthesis of NP matrix components, including collagen II, aggrecan and proteoglycan.
Results. The MRI scans showed remarkable needle-puncture-induced progressive IDD in animals injected with PBS or 10^7 viral particles (VP) of the control virus. In contrast, injection of 10^7 VP of LV-MMP3-shRNA or LV-Sox9 substantially inhibited IDD. MMP3 knockdown or Sox9 overexpression stimulated collagen II and aggrecan expression, as well as proteoglycan synthesis. Notably, the injection of a cocktail of LV-MMP3-shRNA and LV-Sox9 (5 × 10^6 VP each) greatly delayed the development of IDD and induced the highest levels of collagen II and proteoglycan production, indicating a synergistic effect in ECM induction.
Conclusion. Our results suggest that gene therapy targeting MMP3 is an efficient way to delay IDD. Combined gene therapy possesses a stronger capacity to induce matrix components in degenerative NP tissue than single-gene delivery.

Key words

gene therapy, disc degeneration, SOX9, MMP3

References (22)

  1. Hodgkinson T, Shen B, Diwan A, Hoyland JA, Richardson SM. Therapeutic potential of growth differentiation factors in the treatment of degenerative disc diseases. JOR Spine. 2019;2(1):e1045.
  2. Lyu FJ, Cheung KM, Zheng Z, Wang H, Sakai D, Leung VY. IVD progenitor cells: A new horizon for understanding disc homeostasis and repair. Nat Rev Rheumatol. 2019;15(2):102–112.
  3. Gruber HE, Hoelscher GL, Ingram JA, Hanley EN Jr. Matrix metalloproteinase-26, a novel MMP, is constitutively expressed in the human intervertebral disc in vivo and in vitro. Exp Mol Pathol. 2012;92(1):59–63.
  4. Eser B, Eser O, Yuksel Y, et al. Effects of MMP-1 and MMP-3 gene polymorphisms on gene expression and protein level in lumbar disc herniation. Genet Mol Res. 2016;15(3). doi:10.4238/gmr.15038669
  5. Lv FJ, Peng Y, Lim FL, et al. Matrix metalloproteinase 12 is an indicator of intervertebral disc degeneration co-expressed with fibrotic markers. Osteoarthr Cartil. 2016;24(10):1826–1836.
  6. Zigouris A, Alexiou GA, Batistatou A, Voulgaris S, Kyritsis AP. The role of matrix metalloproteinase 9 in intervertebral disc degeneration. J Clin Neurosci. 2011;18(10):1424–1425.
  7. Bachmeier BE, Nerlich A, Mittermaier N, et al. Matrix metalloproteinase expression levels suggest distinct enzyme roles during lumbar disc herniation and degeneration. Eur Spine J. 2009;18(11):1573–1586.
  8. Lefebvre V, Angelozzi M, Haseeb A. SOX9 in cartilage development and disease. Curr Opin Cell Biol. 2019;61:39–47.
  9. Sive JI, Baird P, Jeziorsk M, Watkins A, Hoyland JA, Freemont AJ. Expression of chondrocyte markers by cells of normal and degenerate intervertebral discs. Mol Pathol. 2002;55(2):91–97.
  10. Gruber HE, Norton HJ, Ingram JA, Hanley EN Jr. The SOX9 transcription factor in the human disc: Decreased immunolocalization with age and disc degeneration. Spine (Phila Pa 1976). 2005;30(6):625–630.
  11. Sobajima S, Kim JS, Gilbertson LG, Kang JD. Gene therapy for degenerative disc disease. Gene Ther. 2004;11(4):390–401.
  12. Nishida K, Kang JD, Gilbertson LG, et al. Modulation of the biologic activity of the rabbit intervertebral disc by gene therapy: An in vivo study of adenovirus-mediated transfer of the human transforming growth factor beta 1 encoding gene. Spine (Phila Pa 1976). 1999;24(23):2419–2425.
  13. Nishida K, Suzuki T, Kakutani K, et al. Gene therapy approach for disc degeneration and associated spinal disorders. Eur Spine J. 2008;17(Suppl 4):459–466.
  14. Ren S, Liu Y, Ma J, et al. Treatment of rabbit intervertebral disc degeneration with co-transfection by adeno-associated virus-mediated SOX9 and osteogenic protein-1 double genes in vivo. Int J Mol Med. 2013;32(5):1063–1068.
  15. Masuda K, Aota Y, Muehleman C, et al. A novel rabbit model of mild, reproducible disc degeneration by an anulus needle puncture: Correlation between the degree of disc injury and radiological and histological appearances of disc degeneration. Spine (Phila Pa 1976). 2005;30(1):5–14.
  16. Kwon YJ. A minimally invasive rabbit model of progressive and reproducible disc degeneration confirmed by radiology, gene expression, and histology. J Korean Neurosurg Soc. 2013;53(6):323–330.
  17. Lei T, Zhang Y, Zhou Q, et al. A novel approach for the annulus needle puncture model of intervertebral disc degeneration in rabbits. Am J Transl Res. 2017;9(3):900–909.
  18. Vo NV, Hartman RA, Yurube T, Jacobs LJ, Sowa GA, Kang JD. Expression and regulation of metalloproteinases and their inhibitors in intervertebral disc aging and degeneration. Spine J. 2013;13(3):331–341.
  19. Newell N, Carpanen D, Evans JH, Pearcy MJ, Masouros SD. Mechanical function of the nucleus pulposus of the intervertebral disc under high rates of loading. Spine (Phila Pa 1976). 2019;44(15):1035–1041.
  20. Paul R, Haydon RC, Cheng H, et al. Potential use of Sox9 gene therapy for intervertebral degenerative disc disease. Spine (Phila Pa 1976). 2003;28(8):755–763.
  21. Moon SH, Nishida K, Gilbertson LG, et al. Biologic response of human intervertebral disc cells to gene therapy cocktail. Spine (Phila Pa 1976). 2008;33(17):1850–1855.
  22. Luo H, Wang C, Liu M, et al. Inhibition of SOX9 promotes inflammatory and immune responses of dental pulp. J Endod. 2018;44(5):792–799.