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CLC number: R681.5

On-line Access: 2016-01-06

Received: 2015-07-30

Revision Accepted: 2015-11-30

Crosschecked: 2015-12-16

Cited: 2

Clicked: 3418

Citations:  Bibtex RefMan EndNote GB/T7714

 ORCID:

Gang Feng

http://orcid.org/0000-0002-2814-4319

Xu-wei Luo

http://orcid.org/0000-0002-2626-0631

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Journal of Zhejiang University SCIENCE B 2016 Vol.17 No.1 P.30-42

http://doi.org/10.1631/jzus.B1500182


Adenovirus-mediated GDF-5 promotes the extracellular matrix expression in degenerative nucleus pulposus cells


Author(s):  Xu-wei Luo, Kang Liu, Zhu Chen, Ming Zhao, Xiao-wei Han, Yi-guang Bai, Gang Feng

Affiliation(s):  Research Institute of Tissue Engineering and Stem Cells, Nanchong Central Hospital and the Second Clinical Institute of North Sichuan Medical College, Nanchong 637000, China

Corresponding email(s):   fenggangncch@163.com

Key Words:  Intervertebral disc, Degeneration, Growth and differentiation factor-5 (GDF-5), Adenovirus, Gene therapy, Nucleus pulposus


Xu-wei Luo, Kang Liu, Zhu Chen, Ming Zhao, Xiao-wei Han, Yi-guang Bai, Gang Feng. Adenovirus-mediated GDF-5 promotes the extracellular matrix expression in degenerative nucleus pulposus cells[J]. Journal of Zhejiang University Science B, 2016, 17(1): 30-42.

@article{title="Adenovirus-mediated GDF-5 promotes the extracellular matrix expression in degenerative nucleus pulposus cells",
author="Xu-wei Luo, Kang Liu, Zhu Chen, Ming Zhao, Xiao-wei Han, Yi-guang Bai, Gang Feng",
journal="Journal of Zhejiang University Science B",
volume="17",
number="1",
pages="30-42",
year="2016",
publisher="Zhejiang University Press & Springer",
doi="10.1631/jzus.B1500182"
}

%0 Journal Article
%T Adenovirus-mediated GDF-5 promotes the extracellular matrix expression in degenerative nucleus pulposus cells
%A Xu-wei Luo
%A Kang Liu
%A Zhu Chen
%A Ming Zhao
%A Xiao-wei Han
%A Yi-guang Bai
%A Gang Feng
%J Journal of Zhejiang University SCIENCE B
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%P 30-42
%@ 1673-1581
%D 2016
%I Zhejiang University Press & Springer
%DOI 10.1631/jzus.B1500182

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T1 - Adenovirus-mediated GDF-5 promotes the extracellular matrix expression in degenerative nucleus pulposus cells
A1 - Xu-wei Luo
A1 - Kang Liu
A1 - Zhu Chen
A1 - Ming Zhao
A1 - Xiao-wei Han
A1 - Yi-guang Bai
A1 - Gang Feng
J0 - Journal of Zhejiang University Science B
VL - 17
IS - 1
SP - 30
EP - 42
%@ 1673-1581
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PB - Zhejiang University Press & Springer
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DOI - 10.1631/jzus.B1500182


Abstract: 
Objective: To construct a recombinant adenovirus vector-carrying human growth and differentiation factor-5 (GDF-5) gene, investigate the biological effects of adenovirus-mediated GDF-5 (Ad-GDF-5) on extracellular matrix (ECM) expression in human degenerative disc nucleus pulposus (NP) cells, and explore a candidate gene therapy method for intervertebral disc degeneration (IDD). Methods: Human NP cells of a degenerative disc were isolated, cultured, and infected with Ad-GDF-5 using the AdEasy-1 adenovirus vector system. On Days 3, 7, 14, and 21, the contents of the sulfated glycosaminoglycan (sGAG), deoxyribonucleic acid (DNA) and hydroxyproline (Hyp), synthesis of proteoglycan and collagen II, gene expression of collagen II and aggrecan, and NP cell proliferation were assessed. Results: The adenovirus was an effective vehicle for gene delivery with prolonged expression of GDF-5. Biochemical analysis revealed increased sGAG and Hyp contents in human NP cells infected by Ad-GDF-5 whereas there was no conspicuous change in basal medium (BM) or Ad-green fluorescent protein (GFP) groups. Only cells in the Ad-GDF-5 group promoted the production of ECM, as demonstrated by the secretion of proteoglycan and up-regulation of collagen II and aggrecan at both protein and mRNA levels. The NP cell proliferation was significantly promoted. Conclusions: The data suggest that Ad-GDF-5 gene therapy is a potential treatment for IDD, which restores the functions of degenerative intervertebral disc through enhancing the ECM production of human NP cells.

生长分化因子-5(GDF-5)促进人退行性变髓核细胞外基质表达情况的研究

目的:研究腺病毒介导的GDF-5对人退行性变椎间盘 髓核细胞生长和细胞外基质表达的影响,探索椎间盘退行性变基因治疗的新途径。
创新点:首次验证了GDF-5对人退行变的髓核细胞的生 长和细胞外基质的分泌均具有明显的促进作用,为椎间盘退行性变疾病早期基因治疗提供了新的途径。
方法:利用腺病毒介导的GDF-5转染人退变的髓核细胞,设空白对照组、阴性对照组(绿色荧光蛋白(GFP)组)和实验组(GDF-5组)三个组,3、7、14和21天四个时间点。在预定的时间点,通过蛋白质免疫印迹(Western blotting)、番红-O染色、免疫组化染色、酶联免疫法定量检测(ELISA)、逆转录聚合酶链式反应(RT-PCR)和细胞外基质的定量分析等手段,验证GDF-5对退变髓核细胞外基质表达的影响,同时对GDF-5对髓核细胞生长情况的促进作用进行了表征。
结论:Ad-GDF-5能成功转染人退变的髓核细胞(图1),能有效地促进人退变髓核细胞细胞外基质Aggrecan和II型胶原(Collagen II)分泌(图4~7),RT-PCR的结果表明Aggrecan和Collagen II基因表达也得到了明显的增强(图8)。同时GDF-5对人退变的髓核细胞的生长也有一定的促进作用(图9)。因此,Ad-GDF-5是椎间盘退行性变早期基因治疗的新途径。

关键词:椎间盘;退行性变;生长分化因子-5;髓核;腺病毒;基因治疗

Darkslateblue:Affiliate; Royal Blue:Author; Turquoise:Article

Reference

[1]Bucher, C., Gazdhar, A., Benneker, L.M., et al., 2013. Nonviral gene delivery of growth and differentiation factor 5 to human mesenchymal stem cells injected into a 3D bovine intervertebral disc organ culture system. Stem Cells Int., 2013:326828.

[2]Chubinskaya, S., Hurtig, M., Rueger, D.C., 2007. OP-1/BMP-7 in cartilage repair. Int. Orthop., 31(6):773-781.

[3]Costello, D.J., O'Keeffe, G.W., Hurley, F.M., et al., 2012. Transplantation of novel human GDF5-expressing CHO cells is neuroprotective in models of Parkinson’s disease. J. Cell. Mol. Med., 16(10):2451-2460.

[4]Cui, M., Wan, Y., Anderson, D.G., et al., 2008. Mouse growth and differentiation factor-5 protein and DNA therapy potentiates intervertebral disc cell aggregation and chondrogenic gene expression. Spine J., 8(2):287-295.

[5]Daans, M., Luyten, F.P., Lories, R.J., 2011. GDF5 deficiency in mice is associated with instability-driven joint damage, gait and subchondral bone changes. Ann. Rheum. Dis., 70(1):208-213.

[6]Elkasrawy, M.N., Hamrick, M.W., 2010. Myostatin (GDF-8) as a key factor linking muscle mass and bone structure. J. Musculoskelet. Neuronal. Interact., 10(1):56-63.

[7]Ellman, M.B., An, H.S., Muddasani, P., et al., 2008. Biological impact of the fibroblast growth factor family on articular cartilage and intervertebral disc homeostasis. Gene, 420(1):82-89.

[8]Eskola, P.J., Lemmelä, S., Kjaer, P., et al., 2012. Genetic association studies in lumbar disc degeneration: a systematic review. PLoS ONE, 7(11):e49995.

[9]Farndale, R.W., Buttle, D.J., Barrett, A.J., 1986. Improved quantitation and discrimination of sulphated glycosaminoglycans by use of dimethylmethylene blue. Biochim. Biophys. Acta, 883(2):173-177.

[10]Feng, G., Wan, Y., Balian, G., et al., 2008. Adenovirus-mediated expression of growth and differentiation factor-5 promotes chondrogenesis of adipose stem cells. Growth Factors, 26(3):132-142.

[11]Feng, G., Wan, Y., Shen, F.H., et al., 2009. Nucleus pulposus explant culture model. J. Orthop. Res., 27(6):814-819.

[12]Freemont, A.J., 2009. The cellular pathobiology of the degenerate intervertebral disc and discogenic back pain. Rheumatology, 48(1):5-10.

[13]Hanley, E.N.Jr., Herkowitz, H.N., Kirkpatrick, J.S., et al., 2010. Debating the value of spine surgery. J. Bone Joint Surg. Am., 92(5):1293-1304.

[14]Hogan, M., Girish, K., James, R., et al., 2011. Growth differentiation factor-5 regulation of extracellular matrix gene expression in murine tendon fibroblasts. J. Tissue Eng. Regen. Med., 5(3):191-200.

[15]Hua, G., Haiping, Z., Baorong, H., et al., 2013. Effect of ulinastatin on the expression of iNOS, MMP-2, and MMP-3 in degenerated nucleus pulposus cells of rabbits. Connect. Tissue Res., 54(1):29-33.

[16]Kaneyama, S., Nishida, K., Takada, T., et al., 2008. Fas ligand expression on human nucleus pulposus cells decreases with disc degeneration processes. J. Orthop. Sci., 13(2):130-135.

[17]Kim, J.S., Ellman, M.B., An, H.S., et al., 2012. Lactoferricin mediates anabolic and anti-catabolic effects in the intervertebral disc. J. Cell. Physiol., 227(4):1512-1520.

[18]Kim, Y.J., Sah, R.L., Doong, J.Y., et al., 1988. Fluorometric assay of DNA in cartilage explants using Hoechst 33258. Anal. Biochem., 174(1):168-176.

[19]Larson III, J.W., Levicoff, E.A., Gilbertson, L.G., et al., 2006. Biologic modification of animal models of intervertebral disc degeneration. J. Bone Joint Surg. Am., 88(Suppl. 2):83-88.

[20]Li, X., Leo, B.M., Beck, G., et al., 2004. Collagen and proteoglycan abnormalities in the GDF-5-deficient mice and molecular changes when treating disk cells with recombinant growth factor. Spine, 29(20):2229-2234.

[21]Liang, H., Ma, S.Y., Feng, G., et al., 2010. Therapeutic effects of adenovirus-mediated growth and differentiation factor-5 in a mice disc degeneration model induced by annulus needle puncture. Spine J., 10(1):32-41.

[22]Masuda, K., 2008. Biological repair of the degenerated intervertebral disc by the injection of growth factors. Eur. Spine J., 17(Suppl. 4):441-451.

[23]Melrose, J., Shu, C., Young, C., et al., 2012. Mechanical destabilization induced by controlled annular incision of the intervertebral disc dysregulates metalloproteinase expression and induces disc degeneration. Spine, 37(1):18-25.

[24]Moore, Y.R., Dickinson, D.P., Wikesjö, U.M., 2010. Growth/ differentiation factor-5: a candidate therapeutic agent for periodontal regeneration? A review of pre-clinical data. J. Clin. Periodontol., 37(3):288-298.

[25]Nishida, K., Kang, J.D., Suh, J.K., et al., 1998. Adenovirus-mediated gene transfer to nucleus pulposus cells. Implications for the treatment of intervertebral disc degeneration. Spine, 23(22):2437-2442.

[26]Nishida, K., Kang, J.D., Gilbertson, L.G., et al., 1999. 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 β1 encoding gene. Spine, 24(23):2419-2425.

[27]Pfirrmann, C.W., Metzdorf, A., Zanetti, M., et al., 2001. Magnetic resonance classification of lumbar intervertebral disc degeneration. Spine, 26(17):1873-1878.

[28]Phillips, F.M., Slosar, P.J., Youssef, J.A., et al., 2013. Lumbar spine fusion for chronic low back pain due to degenerative disc disease: a systematic review. Spine, 38(7):E409-E422.

[29]Saiga, K., Furumatsu, T., Yoshida, A., et al., 2010. Combined use of bFGF and GDF-5 enhances the healing of medial collateral ligament injury. Biochem. Biophys. Res. Commun., 402(2):329-334.

[30]Shimer, A.L., Chadderdon, R.C., Gilbertson, L.G., et al., 2004. Gene therapy approaches for intervertebral disc degeneration. Spine, 29(23):2770-2778.

[31]Smith, L.J., Nerurkar, N.L., Choi, K.S., et al., 2011. Degeneration and regeneration of the intervertebral disc: lessons from development. Dis. Model. Mech., 4(1):31-41.

[32]Sobajima, S., Kim, J.S., Gilbertson, L.G., et al., 2004. Gene therapy for degenerative disc disease. Gene Ther., 11(4):390-401.

[33]Stoyanov, J.V., Gantenbein-Ritter, B., Bertolo, A., et al., 2011. Role of hypoxia and growth and differentiation factor-5 on differentiation of human mesenchymal stem cells towards intervertebral nucleus pulposus-like cells. Eur. Cell. Mater., 21:533-547.

[34]Walsh, A.J., Bradford, D.S., Lotz, J.C., 2004. In vivo growth factor treatment of degenerated intervertebral discs. Spine, 29(2):156-163.

[35]Wang, H., Kroeber, M., Hanke, M., et al., 2004. Release of active and depot GDF-5 after adenovirus-mediated overexpression stimulates rabbit and human intervertebral disc cells. J. Mol. Med., 82(2):126-134.

[36]Williams, F.M., Popham, M., Hart, D.J., et al., 2011. GDF5 single-nucleotide polymorphism rs143383 is associated with lumbar disc degeneration in Northern European women. Arthritis Rheum., 63(3):708-712.

[37]Zaidi, S.H., Huang, Q., Momen, A., et al., 2010. Growth differentiation factor 5 regulates cardiac repair after myocardial infarction. J. Am. Coll. Cardiol., 55(2):135-143.

[38]Zhang, Y.G., Sun, Z.M., Liu, J.T., et al., 2009. Features of intervertebral disc degeneration in rat’s aging process. J. Zhejiang Univ.-Sci. B (Biomed. & Biotechnol.), 10(7):522-527.

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