EXPRESSION AND FUNCTION OF HIGH MOBILITY GROUP BOX CHROMOSOMAL PROTEIN 1 IN SYNOVIOCYTES OF PATIENTS WITH OSTEOARTHRITIS_Chinese Journal of Reparative and Reconstructive Surgery

Authors：

 SONGDengxin ¹ , CAOYun ¹ , DINGXu ¹ , HEXiaowen ¹ , HUANGTao ¹ , ZHAOYi ¹ , QIJun ²

1. Department of Orthopedics, Baoshan Branch of Shanghai First People's Hospital, Shanghai, 200940, P. R. China;
2. Department of Orthopedics, Tongji Hospital, Huazhong University of Science and Technology;

Corresponding author：

SONGDengxin, Email: dxsong7576@hotmail.com

Keywords：

Osteoarthritis; Synoviocyte; Small interfering RNA; High mobility group box chromosomal protein 1

DOI：

10.7507/1002-1892.20150296

Video：

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Objective To explore the pathological role of high mobility group box chromosomal protein 1 (HMGB1) in osteoarthritis (OA) by comparing the difference of HMGB1 in the synoviocytes between OA and normal knees. Methods Synoviocyte lines from OA and normal knees were collected and cultured. Immunohistochemistry and Western blot were applied to identify the difference of HMGB1 between the OA and normal synoviocyte lines. The eukaryotic expression vector containing human Pgenesil-1/HMGB1 small interfering RNA (siRNA) were constructed and identified. The synoviocyte lines were transfected with the eukaryotic expression vector of Pgenesil-1/HMGB1 siRNA (Pgenesil-1/HMGB1 siRNA group) and with Pgenesil-1 plasmid (Pgenesil-1 group) and were not transfected as a control (untransfected group). Western blot was applied to identify the difference of HMGB1 among groups, and the levels of interleukin 1β (IL-1β) and tumor necrosis factor α (TNF-α) protein synthesis in the supernatants were measured by ELISA. Results Primary knee synoviocytes cultured in vitro were fibroblast-like cells with longspindle shape. The immunohistochemistry and immunofluorescence results showed positive staining for HMGB1 in cytoplasm and weak positive staining in the nucleus in the OA synoviocyte line, but positive staining for HMGB1 in the nucleus and weak positive staining in the cytoplasm in the synoviocyte line of normal knee. The level of HMGB1 in the OA synoviocytes (0.687±0.025) was significantly higher than that of normal synoviocytes (0.172±0.030) (t=32.159, P=0.000) by Western blot. The recombinant plasmid Pgenesil-1/HMGB1 siRNA was successfully constructed. The expression of HMGB1 protein in Pgenesil-1/HMGB1 siRNA group (0.134±0.048) was significantly lower than that of Pgenesil-1 group (0.581±0.032) and untransfected group (0.514±0.069) (P<0.05). ELISA results showed that IL-1β and TNF-α in supernatants of Pgenesil-1/HMGB1 siRNA group were significantly lower than those of Pgenesil-1 group and untransfected group (P<0.05). Conclusion The up-regulated expression and expressed location (from nucleus to cytoplasm) of HMGB1 in the synoviocyte are closely related to OA. The siRNA targeting inhibition of HMGB1 gene expression can obviously inhibit IL-1β and TNF-α in supernatants of the OA synoviocyte line and delayed the inflammation of OA.

Citation： SONGDengxin, CAOYun, DINGXu, HEXiaowen, HUANGTao, ZHAOYi, QIJun. EXPRESSION AND FUNCTION OF HIGH MOBILITY GROUP BOX CHROMOSOMAL PROTEIN 1 IN SYNOVIOCYTES OF PATIENTS WITH OSTEOARTHRITIS. Chinese Journal of Reparative and Reconstructive Surgery, 2015, 29(11): 1376-1380. doi: 10.7507/1002-1892.20150296 Copy

1.	那键, 刘艺, 马克勇, 等. 老年性骨关节炎的分子生物学机制及治疗展望. 中国老年学杂志, 2010, 30(20):3035-3037.
2.	Goldring MB, Otero M, Plumb DA, et al. Roles of inflammatory and anabolic cytokines in cartilage metabolism:signals and multiple effectors converge upon MMP-13 regulation in osteoarthritis. Eur Cell Mater, 2011, 21:202-220.
3.	Goldring MB. Chondrogenesis, chondrocyte differentiation, and articular cartilage metabolism in health and osteoarthritis. Ther Adv Musculoskelet Dis, 2012, 4(4):269-285.
4.	Loeser RF, Goldring SR, Scanzello CR, et al. Osteoarthritis:a disease of the joint as an organ. Arthritis Rheum, 2012, 64(6):1697-1707.
5.	任红革, 李振浩, 李国振, 等. 趋化因子13在膝关节骨关节炎滑膜中的表达研究. 中国修复重建外科杂志, 2015, 29(1):39-42.
6.	de Seny D, Cobraiville G, Charlier E, et al. Apolipoprotein-A1 as a damage-associated molecular patterns protein in osteoarthritis:ex vivo and in vitro pro-inflammatory properties. PLoS One, 2015, 10(4):e0122904.
7.	王凤龙, 江建明, 肖军, 等. 白细胞介素-18在骨关节炎滑膜细胞中的表达及意义. 中华风湿学杂志, 2010, 14(4):260-262.
8.	García-Armandis I, Guillén MI, Gomar F, et al. High mobility group box 1 potentiates the pro-inflammatory effects of interleukin-1β in osteoarthritic synoviocytes. Arthritis Res Ther, 2010, 12(4):R165.
9.	Yang H, Tracey KJ. Targeting HMGB1 in inflammation. Biochim Biophys Acta, 2010, 1799(1-2):149-156.
10.	Mosquera JA. Role of the receptor for advancedglycation end products (RAGE) in inflammation. Invest Clin, 2010, 51(2):257-268.
11.	Kosaka T, Fukui R, Matsui M, et al. RAGE, receptor of advanced glycation endoproducts, negatively regulates chondrocytes differentiation. PLoS One, 2014, 9(9):e108819.
12.	Nogueira-Machado JA, de Oliveira Volpe CM. HMGB-1 as a target for inflammation controlling. Recent Pat Endocr Metab Immune Drug Discov, 2012, 6(3):201-209.
13.	Terada C, Yoshida A, Nasu Y, et al. Gene expression and localization of high-mobility group box chromosomal protein-1(HMGB-1) in human osteoarthritic cartilage. Acta Med Okayama, 2011, 65(6):369-377.
14.	Huang LF, Yao YM, Meng HD. The effect of high mobility group box-1 protein on immune function of human T lymphocytes in vitro. Chin Crit CareMed, 2008, 20(1):7-13.
15.	Shen S, Guo J, Luo Y, et al. Functional proteomics revealed IL-1β amplifies TNF downstream protein signals in human synoviocytes in a TNF-independent manner. Biochem Biophys Res Commun, 2014, 450(1):538-544.
16.	Lee SG, Lee EJ, Park WD, et al. Anti-inflammatory and antiosteoarthritis effects of fermented Achyranthes japonica Nakai. J Ethnophar-macol, 2012, 142(3):634-641.
17.	Ding X, Zhang Y, Huang Y, et al. Cadherin-11 involves in synovitis and increases the migratory and invasive capacity of fibroblastlike synoviocytes of osteoarthritis. Int Immunopharmacol, 2015, 26(1):153-161.

1. 那键, 刘艺, 马克勇, 等. 老年性骨关节炎的分子生物学机制及治疗展望. 中国老年学杂志, 2010, 30(20):3035-3037.
2. Goldring MB, Otero M, Plumb DA, et al. Roles of inflammatory and anabolic cytokines in cartilage metabolism:signals and multiple effectors converge upon MMP-13 regulation in osteoarthritis. Eur Cell Mater, 2011, 21:202-220.
3. Goldring MB. Chondrogenesis, chondrocyte differentiation, and articular cartilage metabolism in health and osteoarthritis. Ther Adv Musculoskelet Dis, 2012, 4(4):269-285.
4. Loeser RF, Goldring SR, Scanzello CR, et al. Osteoarthritis:a disease of the joint as an organ. Arthritis Rheum, 2012, 64(6):1697-1707.
5. 任红革, 李振浩, 李国振, 等. 趋化因子13在膝关节骨关节炎滑膜中的表达研究. 中国修复重建外科杂志, 2015, 29(1):39-42.
6. de Seny D, Cobraiville G, Charlier E, et al. Apolipoprotein-A1 as a damage-associated molecular patterns protein in osteoarthritis:ex vivo and in vitro pro-inflammatory properties. PLoS One, 2015, 10(4):e0122904.
7. 王凤龙, 江建明, 肖军, 等. 白细胞介素-18在骨关节炎滑膜细胞中的表达及意义. 中华风湿学杂志, 2010, 14(4):260-262.
8. García-Armandis I, Guillén MI, Gomar F, et al. High mobility group box 1 potentiates the pro-inflammatory effects of interleukin-1β in osteoarthritic synoviocytes. Arthritis Res Ther, 2010, 12(4):R165.
9. Yang H, Tracey KJ. Targeting HMGB1 in inflammation. Biochim Biophys Acta, 2010, 1799(1-2):149-156.
10. Mosquera JA. Role of the receptor for advancedglycation end products (RAGE) in inflammation. Invest Clin, 2010, 51(2):257-268.
11. Kosaka T, Fukui R, Matsui M, et al. RAGE, receptor of advanced glycation endoproducts, negatively regulates chondrocytes differentiation. PLoS One, 2014, 9(9):e108819.
12. Nogueira-Machado JA, de Oliveira Volpe CM. HMGB-1 as a target for inflammation controlling. Recent Pat Endocr Metab Immune Drug Discov, 2012, 6(3):201-209.
13. Terada C, Yoshida A, Nasu Y, et al. Gene expression and localization of high-mobility group box chromosomal protein-1(HMGB-1) in human osteoarthritic cartilage. Acta Med Okayama, 2011, 65(6):369-377.
14. Huang LF, Yao YM, Meng HD. The effect of high mobility group box-1 protein on immune function of human T lymphocytes in vitro. Chin Crit CareMed, 2008, 20(1):7-13.
15. Shen S, Guo J, Luo Y, et al. Functional proteomics revealed IL-1β amplifies TNF downstream protein signals in human synoviocytes in a TNF-independent manner. Biochem Biophys Res Commun, 2014, 450(1):538-544.
16. Lee SG, Lee EJ, Park WD, et al. Anti-inflammatory and antiosteoarthritis effects of fermented Achyranthes japonica Nakai. J Ethnophar-macol, 2012, 142(3):634-641.
17. Ding X, Zhang Y, Huang Y, et al. Cadherin-11 involves in synovitis and increases the migratory and invasive capacity of fibroblastlike synoviocytes of osteoarthritis. Int Immunopharmacol, 2015, 26(1):153-161.

Chinese Journal of Reparative and Reconstructive Surgery

EXPRESSION AND FUNCTION OF HIGH MOBILITY GROUP BOX CHROMOSOMAL PROTEIN 1 IN SYNOVIOCYTES OF PATIENTS WITH OSTEOARTHRITIS

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