EFFECT OF INHIBITOR OF DIFFERENTIATION 1 GENE TRANSFECTION ON BONE MORPHOGENETIC PROTEIN 2 PROMOTING CHONDROGENIC GENE EXPRESSIONS OF RABBIT INTERVERTEBRAL CARTILAGE ENDPLATE CELLS_Chinese Journal of Reparative and Reconstructive Surgery

Authors：

ZHOUQiang ,  QUANZhengxue , LUOXiaoji , TANGKe , ZHOUXu , ZHANGYuan , LEITao

Department of Orthopaedics, the First Affiliated Hospital of Chongqing Medical University, Chongqing, 400016, P. R. China;

Corresponding?author：

QUANZhengxue, Email: quanzx18@126.com

Keywords：

Bone morphogenetic protein 2; Inhibitor of differentiation 1; Cartilage endplate cells; Gene transfection; Rabbit

DOI：

10.7507/1002-1892.20150330

Video：

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Objective To study the effect of inhibitor of differentiation 1 (Id1) gene transfection on bone morphogenetic protein 2 (BMP-2) promoting the expressions of collagen type Ⅱ (COL Ⅱ) and aggrecan (ACAN) in intervertebral cartilage endplate cells (EPCs). Methods EPCs were harvested from the New Zealand white rabbits, the 2nd generation EPCs were used for experiment. The transfection efficiency of green fluorescent protein blank lentivirus, high expression of Id1 lentivirus, RNA interference (RNAi) Id1 lentivirus transfection in the EPCs were observed by the fluorescence microscopy, real-time fluorescence quantitative PCR, and Western blot. Blank vector, single BMP-2 gene, BMP-2 and Id1 genes were transfected into EPCs, respectively. The cell morphology and the expressions of COL Ⅱ and ACAN in each group were observed. Results Lentiviral transfection had no significant effect on the cell morphology. The EPCs were effectively transfected by the high expression Id1 lentivirus and RNAi Id1 lentivirus; the expression of Id1 mRNA was also significantly interfered. The expressions of COL Ⅱ and ACAN mRNA and synthesis of COL Ⅱ and ACAN protein were significantly higher in BMP-2 lentivirus and high expression Id1 lentivirus groups than control group (P<0.05). The expression of COL Ⅱ and ACAN protein were down regulated in the cartilage endplate cells when the expression of Id1 gene was decreased (P<0.05). Conclusion Up-regulation of Id1 gene expression can enhance the effects of BMP-2 on the synthesis of COL Ⅱ and ACAN in EPCs.

Citation： ZHOUQiang, QUANZhengxue, LUOXiaoji, TANGKe, ZHOUXu, ZHANGYuan, LEITao. EFFECT OF INHIBITOR OF DIFFERENTIATION 1 GENE TRANSFECTION ON BONE MORPHOGENETIC PROTEIN 2 PROMOTING CHONDROGENIC GENE EXPRESSIONS OF RABBIT INTERVERTEBRAL CARTILAGE ENDPLATE CELLS. Chinese Journal of Reparative and Reconstructive Surgery, 2015, 29(12): 1547-1552. doi: 10.7507/1002-1892.20150330 Copy

1.	Malandrino A, Lacroix D, Hellmich C, et al. The role of endplate poromechanical properties on the nutrient availability in theintervertebral disc. Osteoarthritis Cartilage, 2014, 22(7):1053-1060.
2.	Gu W, Zhu Q, Gao X, et al. Simulation of the progression of intervertebral disc degeneration due to decreased nutritional supply. Spine (Phila Pa 1976), 2014, 39(24):E1411-1417.
3.	Tim Yoon S, Su Kim K, Li J, et al. The effect of bone morphogenetic protein-2 on rat intervertebral disc cells in vitro. Spine (Phila Pa 1976), 2003, 28(16):1773-1780.
4.	Peng Y, Kang Q, Luo Q, et al. Inhibitor of DNA binding/differentiation helix-loop-helix proteins mediate bone morphogenetic protein-induced osteoblast differentiation of mesenchymal stem cells. J Biol Chem, 2004, 279(31):32941-32949.
5.	Du Y, Yip H. Effects of bone morphogenetic protein 2 on Id expression and neuroblastoma cell differentiation. Differentiation, 2010, 79(2):84-92.
6.	Urban JP, Smith S, Fairbank JC. Nutritionof theintervertebral disc. Spine (Phila Pa 1976), 2004, 29(23):2700-2709.
7.	Roberts S, Menage J, Urban JP. Biochemical and structural properties of the cartilage end-plate and its relation to the intervertebral disc. Spine (Phila Pa 1976), 1989, 14(2):166-174.
8.	Aigner T, Gresk-Otter KR, Fairbank JC, et al. Variation with age in the pattern of type X collagen expression in normal and scoliotic human intervertebral discs. Calcif Tissue Int, 1998, 63(3):263-268.
9.	徐宏光, 楊名, 王弘, 等. 頸椎軟骨終板ANK的表達變化. 中華醫學雜志, 2010, 90(17):1192-1196.
10.	M??tt? JH, Wadge S, MacGregor A, et al. Vertebral endplate (modic) change is an independent risk factor for episodes of severe and disabling low back pain. Spine (Phila Pa 1976), 2015, 40(15):1187-1193.
11.	Neidlinger-Wilke C, Boldt A, Brochhausen C, et al. Molecular interactions between human cartilaginous endplates and nucleus pulposus cells:a preliminary investigation. Spine (Phila Pa 1976), 2014, 39(17):1355-1364.
12.	Wu Y, Cisewski S, Sachs BL, et al. Effect of cartilage endplate on cell based disc regeneration:a finite element analysis. Mol Cell Biomech, 2013, 10(2):159-182.
13.	Norton JD, Deed RW, Craggs G, et al. Id helix-loop-helix proteins in cell growth and differentiation. Trends Cell Biol, 1998, 8(2):58-65.
14.	胡有谷, 陳伯華. 椎間盤退變基因治療進展. 中國骨與關節外科, 2013, 6(Suppl 1):11-16.
15.	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.
16.	Li J, Yoon ST, Hutton WC. Effect of bone morphogenetic protein-2(BMP-2) on matrix production, other BMPs, and BMP receptors in rat intervertebral disc cells. J Spinal Disord Tech, 2004, 17(5):423-428.
17.	Kim DJ, Moon SH, Kim H, et al. Bone morphogenetic protein-2 facilitates expression of chondrogenic, not osteogenic, phenotype of human intervertebral disc cells. Spine (Phila Pa 1976), 2004, 28(24):2679-2684.
18.	Neumann AJ, Gardner OF, Williams R, et al. Human articular cartilage progenitor cells are responsive to mechanical stimulation and adenoviral-mediated over expression of bone-morphogenetic protein 2. PLoS One, 2015, 10(8):e0136229.
19.	Peng Y, Kang Q, Luo Q, et al. Inhibitor of DNA binding/differentiation helix-loop-helix proteins mediate bone morphogenetic protein-induced osteoblast differentiation of mesenchymal stem cells. J Biol Chem, 2004, 279(31):32941-32949.
20.	Desprez PY, Lin CQ, Thomasset N, et al. A novel pathway for mammary epithelial cell invasion induced by the helix-loop-helix protein Id-1. Mol Cell Biol, 1998, 18(8):4577-4588.
21.	Ling MT, Lau TC, Zhou C, et al. Overexpression of Id-1 in prostate cancer cells promotes angiogenesis through the activation of vascular endothelial growth factor VEGF. Carcinogenesis, 2005, 26(10):1668-1676.
22.	Sakurai D, Tsuchiya N, Yamaguchi A, et al. Crucial role of in hibitor of DNA binding/differentiation in the vascular endothelial growth factor-induced activation and angiogenic processes of human endothelial cells. J Immunology, 2004, 173(9):5801-5809.

1. Malandrino A, Lacroix D, Hellmich C, et al. The role of endplate poromechanical properties on the nutrient availability in theintervertebral disc. Osteoarthritis Cartilage, 2014, 22(7):1053-1060.
2. Gu W, Zhu Q, Gao X, et al. Simulation of the progression of intervertebral disc degeneration due to decreased nutritional supply. Spine (Phila Pa 1976), 2014, 39(24):E1411-1417.
3. Tim Yoon S, Su Kim K, Li J, et al. The effect of bone morphogenetic protein-2 on rat intervertebral disc cells in vitro. Spine (Phila Pa 1976), 2003, 28(16):1773-1780.
4. Peng Y, Kang Q, Luo Q, et al. Inhibitor of DNA binding/differentiation helix-loop-helix proteins mediate bone morphogenetic protein-induced osteoblast differentiation of mesenchymal stem cells. J Biol Chem, 2004, 279(31):32941-32949.
5. Du Y, Yip H. Effects of bone morphogenetic protein 2 on Id expression and neuroblastoma cell differentiation. Differentiation, 2010, 79(2):84-92.
6. Urban JP, Smith S, Fairbank JC. Nutritionof theintervertebral disc. Spine (Phila Pa 1976), 2004, 29(23):2700-2709.
7. Roberts S, Menage J, Urban JP. Biochemical and structural properties of the cartilage end-plate and its relation to the intervertebral disc. Spine (Phila Pa 1976), 1989, 14(2):166-174.
8. Aigner T, Gresk-Otter KR, Fairbank JC, et al. Variation with age in the pattern of type X collagen expression in normal and scoliotic human intervertebral discs. Calcif Tissue Int, 1998, 63(3):263-268.
9. 徐宏光, 楊名, 王弘, 等. 頸椎軟骨終板ANK的表達變化. 中華醫學雜志, 2010, 90(17):1192-1196.
10. M??tt? JH, Wadge S, MacGregor A, et al. Vertebral endplate (modic) change is an independent risk factor for episodes of severe and disabling low back pain. Spine (Phila Pa 1976), 2015, 40(15):1187-1193.
11. Neidlinger-Wilke C, Boldt A, Brochhausen C, et al. Molecular interactions between human cartilaginous endplates and nucleus pulposus cells:a preliminary investigation. Spine (Phila Pa 1976), 2014, 39(17):1355-1364.
12. Wu Y, Cisewski S, Sachs BL, et al. Effect of cartilage endplate on cell based disc regeneration:a finite element analysis. Mol Cell Biomech, 2013, 10(2):159-182.
13. Norton JD, Deed RW, Craggs G, et al. Id helix-loop-helix proteins in cell growth and differentiation. Trends Cell Biol, 1998, 8(2):58-65.
14. 胡有谷, 陳伯華. 椎間盤退變基因治療進展. 中國骨與關節外科, 2013, 6(Suppl 1):11-16.
15. 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.
16. Li J, Yoon ST, Hutton WC. Effect of bone morphogenetic protein-2(BMP-2) on matrix production, other BMPs, and BMP receptors in rat intervertebral disc cells. J Spinal Disord Tech, 2004, 17(5):423-428.
17. Kim DJ, Moon SH, Kim H, et al. Bone morphogenetic protein-2 facilitates expression of chondrogenic, not osteogenic, phenotype of human intervertebral disc cells. Spine (Phila Pa 1976), 2004, 28(24):2679-2684.
18. Neumann AJ, Gardner OF, Williams R, et al. Human articular cartilage progenitor cells are responsive to mechanical stimulation and adenoviral-mediated over expression of bone-morphogenetic protein 2. PLoS One, 2015, 10(8):e0136229.
19. Peng Y, Kang Q, Luo Q, et al. Inhibitor of DNA binding/differentiation helix-loop-helix proteins mediate bone morphogenetic protein-induced osteoblast differentiation of mesenchymal stem cells. J Biol Chem, 2004, 279(31):32941-32949.
20. Desprez PY, Lin CQ, Thomasset N, et al. A novel pathway for mammary epithelial cell invasion induced by the helix-loop-helix protein Id-1. Mol Cell Biol, 1998, 18(8):4577-4588.
21. Ling MT, Lau TC, Zhou C, et al. Overexpression of Id-1 in prostate cancer cells promotes angiogenesis through the activation of vascular endothelial growth factor VEGF. Carcinogenesis, 2005, 26(10):1668-1676.
22. Sakurai D, Tsuchiya N, Yamaguchi A, et al. Crucial role of in hibitor of DNA binding/differentiation in the vascular endothelial growth factor-induced activation and angiogenic processes of human endothelial cells. J Immunology, 2004, 173(9):5801-5809.

Chinese Journal of Reparative and Reconstructive Surgery

EFFECT OF INHIBITOR OF DIFFERENTIATION 1 GENE TRANSFECTION ON BONE MORPHOGENETIC PROTEIN 2 PROMOTING CHONDROGENIC GENE EXPRESSIONS OF RABBIT INTERVERTEBRAL CARTILAGE ENDPLATE CELLS

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