29-kDa FN-f inhibited autophagy through modulating localization of HMGB1 in human articular chondrocytes

  • Hwang, Hyun Sook (Division of Rheumatology, Department of Internal Medicine, Hallym University Sacred Heart Hospital) ;
  • Choi, Min Ha (Division of Rheumatology, Department of Internal Medicine, Hallym University Sacred Heart Hospital) ;
  • Kim, Hyun Ah (Division of Rheumatology, Department of Internal Medicine, Hallym University Sacred Heart Hospital)
  • Received : 2018.03.19
  • Accepted : 2018.05.04
  • Published : 2018.10.31


Fibronectin fragments found in the synovial fluid of patients with osteoarthritis (OA) induce the catabolic responses in cartilage. Nuclear high-mobility group protein Box 1 (HMGB1), a damage-associated molecular pattern, is responsible for the regulation of signaling pathways related to cell death and survival in response to various stimuli. In this study, we investigated whether changes induced by 29-kDa amino-terminal fibronectin fragment (29-kDa FN-f) in HMGB1 expression influences the pathogenesis of OA via an HMGB1-modulated autophagy signaling pathway. Human articular chondrocytes were enzymatically isolated from articular cartilage. The level of mRNA was measured by quantitative real-time PCR. The expression of proteins was examined by western blot analysis, immnunofluorescence assay, and enzyme-linked immunosorbent assay. Interaction of proteins was evaluated by immunoprecipitation. The HMGB1 level was significantly lower in human OA cartilage than in normal cartilage. Although 29-kDa FN-f significantly reduced the HMGB1 expression at the mRNA and protein levels 6 h after treatment, the cytoplasmic level of HMGB1 was increased in chondrocytes treated with 29-kDa FN-f, which significantly inhibited the interaction of HMGB1 with Beclin-1, increased the interaction of Bcl-2 with Beclin-1, and decreased the levels of Beclin-1 and phosphorylated Bcl-2. In addition, the level of microtubule-associated protein 1 light chain 3-II, an autophagy marker, was down-regulated in chondrocytes treated with 29-kDa FN-f, whereas the effect was antagonized by mTOR knockdown. Furthermore, prolonged treatment with 29-kDa FN-f significantly increased the release of HMGB1 into the culture medium. These results demonstrated that 29-kDa FN-f inhibits chondrocyte autophagy by modulating the HMGB1 signaling pathway.


Autophagy;Chondrocyte;Fibronectin fragment;HMGB1;Osteoarthritis


Supported by : Ministry of Health & Welfare, National Research Foundation (NRF), Hallym University


  1. Song B, Song H, Wang W et al (2017) Beclin 1 overexpression inhibits chondrocyte apoptosis and downregulates extracellular matrix metabolism in osteoarthritis. Mol Med Rep 16, 3958-3964
  2. Carames B, Hasegawa A, Taniguchi N, Miyaki S, Blanco FJ and Lotz M (2012) Autophagy activation by rapamycin reduces severity of experimental osteoarthritis. Ann Rheum Dis 71, 575-581
  3. Zhang T, Liu J, Zheng X, Zhang B and Xia C (2018) Different roles of Akt and mechanistic target of rapamycin in serumdependent chondroprotection of human osteoarthritic chondrocytes. Int J Mol Med 41, 977-984
  4. Hwang HS, Park IY, Kim DW, Choi SY, Jung YO and Kim HA (2015) PEP-1-FK506BP12 inhibits matrix metalloproteinase expression in human articular chondrocytes and in a mouse carrageenan-induced arthritis model. BMB Rep 48, 407-412
  5. Yusein-Myashkova S, Ugrinova I and Pasheva E (2016) Non-histone protein HMGB1 inhibits the repair of damaged DNA by cisplatin in NIH-3T3 murine fibroblasts. BMB Rep 49, 99-104
  6. Magna M and Pisetsky DS (2015) The Role of Cell Death in the Pathogenesis of SLE: Is Pyroptosis the Missing Link? Scand J Immunol 82, 218-224
  7. Heinola T, de Grauw JC, Virkki L et al (2013) Bovine chronic osteoarthritis causes minimal change in synovial fluid. J Comp Pathol 148, 335-344
  8. Wang H, Bloom O, Zhang M et al (1999) HMG-1 as a late mediator of endotoxin lethality in mice. Science 285, 248-251
  9. Almonte-Becerril M, Navarro-Garcia F, Gonzalez-Robles A, Vega-Lopez MA, Lavalle C and Kouri JB (2010) Cell death of chondrocytes is a combination between apoptosis and autophagy during the pathogenesis of Osteoarthritis within an experimental model. Apoptosis 15, 631-638
  10. Zhang Y, Vasheghani F, Li YH et al (2015) Cartilagespecific deletion of mTOR upregulates autophagy and protects mice from osteoarthritis. Ann Rheum Dis 74, 1432-1440
  11. Chang J, Wang W, Zhang H, Hu Y, Wang M and Yin Z (2013) The dual role of autophagy in chondrocyte responses in the pathogenesis of articular cartilage degeneration in osteoarthritis. Int J Mol Med 32, 1311-1318
  12. Tang D, Kang R, Livesey KM et al (2010) Endogenous HMGB1 regulates autophagy. J Cell Biol 190, 881-892
  13. Katsara O, Attur M, Ruoff R, Abramson SB and Kolupaeva V (2017) Increased Activity of the Chondrocyte Translational Apparatus Accompanies Osteoarthritic Changes in Human and Rodent Knee Cartilage. Arthritis Rheumatol 69, 586-597
  14. Tchetina EV, Poole AR, Zaitseva EM et al (2013) Differences in Mammalian target of rapamycin gene expression in the peripheral blood and articular cartilages of osteoarthritic patients and disease activity. Arthritis 2013, 461486
  15. Ding L, Buckwalter JA and Martin JA (2017) DAMPs Synergize with Cytokines or Fibronectin Fragment on Inducing Chondrolysis but Lose Effect When Acting Alone. Mediators Inflamm 2017, 2642549
  16. Yu C, Yu X, Zhu HW et al (2016) Expression pattern of HMGB1 and its association with autophagy in acute necrotizing pancreatitis. Mol Med Rep 14, 5507-5513
  17. Rosenberg JH, Rai V, Dilisio MF, Sekundiak TD and Agrawal DK (2017) Increased expression of damageassociated molecular patterns (DAMPs) in osteoarthritis of human knee joint compared to hip joint. Mol Cell Biochem 436, 59-69
  18. Yasuda T (2006) Cartilage destruction by matrix degradation products. Mod Rheumatol 16, 197-205
  19. Homandberg GA, Wen C and Hui F (1998) Cartilage damaging activities of fibronectin fragments derived from cartilage and synovial fluid. Osteoarthritis Cartilage 6, 231-244
  20. Fang F and Jiang D (2016) IL-1beta/HMGB1 signalling promotes the inflammatory cytokines release via TLR signalling in human intervertebral disc cells. Biosci Rep 36,
  21. Park JS, Gamboni-Robertson F, He Q et al (2006) High mobility group box 1 protein interacts with multiple Toll-like receptors. Am J Physiol Cell Physiol 290, C917-924
  22. Stros M (2010) HMGB proteins: interactions with DNA and chromatin. Biochim Biophys Acta 1799, 101-113
  23. Taniguchi N, Kawakami Y, Maruyama I and Lotz M (2018) HMGB proteins and arthritis. Hum Cell 31, 1-9
  24. Javaherian K, Liu JF and Wang JC (1978) Nonhistone proteins HMG1 and HMG2 change the DNA helical structure. Science 199, 1345-1346
  25. Min HJ, Ko EA, Wu J et al (2013) Chaperone-like activity of high-mobility group box 1 protein and its role in reducing the formation of polyglutamine aggregates. J Immunol 190, 1797-1806
  26. Xie DL, Hui F, Meyers R and Homandberg GA (1994) Cartilage chondrolysis by fibronectin fragments is associated with release of several proteinases: stromelysin plays a major role in chondrolysis. Arch Biochem Biophys 311, 205-212
  27. Xie DL, Meyers R and Homandberg GA (1992) Fibronectin fragments in osteoarthritic synovial fluid. J Rheumatol 19, 1448-1452
  28. Homandberg GA and Hui F (1996) Association of proteoglycan degradation with catabolic cytokine and stromelysin release from cartilage cultured with fibronectin fragments. Arch Biochem Biophys 334, 325-331
  29. Pichika R and Homandberg GA (2004) Fibronectin fragments elevate nitric oxide (NO) and inducible NO synthetase (iNOS) levels in bovine cartilage and iNOS inhibitors block fibronectin fragment mediated damage and promote repair. Inflamm Res 53, 405-412
  30. Hwang HS, Park SJ, Cheon EJ, Lee MH and Kim HA (2015) Fibronectin fragment-induced expression of matrix metalloproteinases is mediated by MyD88-dependent TLR-2 signaling pathway in human chondrocytes. Arthritis Res Ther 17, 320
  31. Lee SA, Kwak MS, Kim S and Shin JS (2014) The role of high mobility group box 1 in innate immunity. Yonsei Med J 55, 1165-1176
  32. Marquez RT and Xu L (2012) Bcl-2:Beclin 1 complex: multiple, mechanisms regulating autophagy/apoptosis toggle switch. Am J Cancer Res 2, 214-221
  33. Davalos AR, Kawahara M, Malhotra GK et al (2013) p53-dependent release of Alarmin HMGB1 is a central mediator of senescent phenotypes. J Cell Biol 201, 613-629
  34. Jeon OH, Kim C, Laberge RM et al (2017) Local clearance of senescent cells attenuates the development of post-traumatic osteoarthritis and creates a pro-regenerative environment. Nat Med 23, 775-781
  35. Goldring MB and Goldring SR (2007) Osteoarthritis. J Cell Physiol 213, 626-634
  36. Kim HA, Cho ML, Choi HY et al (2006) The catabolic pathway mediated by Toll-like receptors in human osteoarthritic chondrocytes. Arthritis Rheum 54, 2152-2163
  37. Rahman MA and Rhim H (2017) Therapeutic implication of autophagy in neurodegenerative diseases. BMB Rep 50, 345-354
  38. Loeser RF (2009) Aging and osteoarthritis: the role of chondrocyte senescence and aging changes in the cartilage matrix. Osteoarthritis Cartilage 17, 971-979
  39. Carames B, Olmer M, Kiosses WB and Lotz MK (2015) The relationship of autophagy defects to cartilage damage during joint aging in a mouse model. Arthritis Rheumatol 67, 1568-1576
  40. Carames B, Taniguchi N, Otsuki S, Blanco FJ and Lotz M (2010) Autophagy is a protective mechanism in normal cartilage, and its aging-related loss is linked with cell death and osteoarthritis. Arthritis Rheum 62, 791-801
  41. Carames B, Taniguchi N, Seino D, Blanco FJ, D'Lima D and Lotz M (2012) Mechanical injury suppresses autophagy regulators and pharmacologic activation of autophagy results in chondroprotection. Arthritis Rheum 64, 1182-1192
  42. Wang CC, Lee CH, Peng YJ, Salter DM and Lee HS (2015) Platelet-Rich Plasma Attenuates 30-kDa Fibronectin Fragment-Induced Chemokine and Matrix Metalloproteinase Expression by Meniscocytes and Articular Chondrocytes. Am J Sports Med 43, 2481-2489