Molecular Biology Reports

, Volume 46, Issue 2, pp 2493–2504 | Cite as

Bacterial TIR domain-derived peptides inhibit innate immune signaling and catabolic responses in chondrocyte

  • Lei Hong
  • Shijie Wang
  • Jinpeng Guo
  • Xin Yin
  • Qianjin Yu
  • Mingjuan Yang
  • Yufei WangEmail author
  • Yuehua KeEmail author
  • Wenfeng LiEmail author
Original Article


Osteoarthritis (OA) is a degenerative joint disease, in which low-grade inflammation plays an important role at the initiating step. Low-doses of LPS-induced inflammation in the plasma activate chondrocytes and promote the secretion proinflammatory cytokines, leading to secondary inflammation. Blocking OA­associated TLR activation is a promising strategy for the development of suitable therapies. Here, we want to find some bacteria-derived peptides that can block TLR signaling in chondrocytes more efficiently. Based on previous studies, we screened 12 TIR domain-derived peptides for their effects on NF-кB activation induced by LPS, IL-1β or TNF-α in murine ATDC-5 cells. We evaluated their effects on LPS-induced cytokine expression and secretion. Among them, two bacteria-derived peptides, TcpC-DD and TcpB-DD, showed the most potent inhibitory activities. In comparison with TcpB-DD, TcpC-DD exhibited broader TLR-inhibitory specificity during inflammation in chondrocytes. Furthermore, both TcpC-DD and TcpB-DD displayed strong inhibition of LPS- and IL-1β-induced catabolic reactions in chondrocytes. However, only TcpC-DD exhibited obvious suppression of TNF-α-induced catabolism. In conclusion, we identified two novel inhibitory peptides that modulate catabolism in chondrocytes and innate immune responses, and these peptides could be used to develop novel therapeutic strategies for OA.


Decoy peptide Chondrocyte Innate immune signaling TLR pathway Catabolic responses 


Author contributions

LH, SW, JG, YK, and WL designed the project. LH, SJ, XY, QY, XY, YW and YK performed the experiments. LH, SW, JG and WL analyzed the data. All authors were involved in writing and editing the paper. YW, YK and WL supervised all aspects of this work.


This work was financially supported by Beijing Municipal Natural Science Foundation (7174330), National Natural Science Foundation of China (30672133, 81671977, and 81301405) and National Key Research and Development Program of China (2016YFC1202905, 2018ZX10713003, 2018ZX10732401).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

11033_2019_4627_MOESM1_ESM.docx (88 kb)
Figure S1. Effects of TIR domain-derived decoy peptides on TNF-α induced mRNA expression. ATDC-5 cells were pretreated with peptides at 20 µM for 30 min and then stimulated with TNF-α at 10 ng/ml for 1 h. Next, cytoplasmic mRNA expression of TNF-α (A), IL-1β (B), IFN-β (C) and IL-6 (D) were measured and normalized to the expression of GAPDH. The results represent the mean values from triplicate experiments. Significant differences, compared to the TB-0 group, are indicated by asterisks (*P < 0.05, Student’s t-test). Figure S2. TcpC-DD inhibited TNF-α-induced cytokine secretion. ATDC-5 cells were transfected with the siRNA against MyD88 for 24h, then were treated with peptides at 20 µM for 30 min and then stimulated with TNF-α at 10 ng/ml for 24 h. Supernatants were collected and analyzed for IL-1β (A), IFN-β (B), and IL-6 (C) by ELISA. IL-1β was detected in cell lysates. The results represent the mean values from triplicate experiments. Significant differences, compared to the TB-0 group, are indicated by asterisks (*P < 0.05, Student’s t-test). (DOCX 87 KB)


  1. 1.
    Poulet B, Staines KA (2016) New developments in osteoarthritis and cartilage biology. Curr Opin Pharmacol 28:8–13CrossRefGoogle Scholar
  2. 2.
    Berenbaum F (2013) Osteoarthritis as an inflammatory disease (osteoarthritis is not osteoarthrosis!), Osteoarthritis and cartilage/OARS. Osteoarthr Res Soc 21(1):16–21CrossRefGoogle Scholar
  3. 3.
    Sellam J, Berenbaum F (2010) The role of synovitis in pathophysiology and clinical symptoms of osteoarthritis, Nature reviews. Rheumatology 6(11):625–635Google Scholar
  4. 4.
    Goldring MB, Otero M (2011) Inflammation in osteoarthritis. Curr Opin Rheumatol 23(5):471–478CrossRefGoogle Scholar
  5. 5.
    Gomez R, Villalvilla A, Largo R, Gualillo O, Herrero-Beaumont G (2015) TLR4 signalling in osteoarthritis–finding targets for candidate DMOADs. Nat Rev Rheumatol 11(3):159–170CrossRefGoogle Scholar
  6. 6.
    Kim HA, Cho ML, Choi HY, Yoon CS, Jhun JY, Oh HJ, Kim HY (2006) The catabolic pathway mediated by Toll-like receptors in human osteoarthritic chondrocytes. Arthritis Rheum 54(7):2152–2163CrossRefGoogle Scholar
  7. 7.
    Sillat T, Barreto G, Clarijs P, Soininen A, Ainola M, Pajarinen J, Korhonen M, Konttinen YT, Sakalyte R, Hukkanen M, Ylinen P, Nordstrom DC (2013) Toll-like receptors in human chondrocytes and osteoarthritic cartilage. Acta Orthop 84(6):585–592CrossRefGoogle Scholar
  8. 8.
    Huang Z, Kraus VB (2016) Does lipopolysaccharide-mediated inflammation have a role in OA? Nat Rev Rheumatol 12(2):123–129CrossRefGoogle Scholar
  9. 9.
    Mullen LM, Chamberlain G, Sacre S (2015) Pattern recognition receptors as potential therapeutic targets in inflammatory rheumatic disease. Arthritis Res Ther 17:122CrossRefGoogle Scholar
  10. 10.
    Piao W, Shirey KA, Ru LW, Lai W, Szmacinski H, Snyder GA, Sundberg EJ, Lakowicz JR, Vogel SN, Toshchakov VY (2015) A decoy peptide that disrupts TIRAP recruitment to TLRs is protective in a murine model of influenza. Cell Rep 11(12):1941–1952CrossRefGoogle Scholar
  11. 11.
    Piao W, Ru LW, Piepenbrink KH, Sundberg EJ, Vogel SN, Toshchakov VY (2013) Recruitment of TLR adapter TRIF to TLR4 signaling complex is mediated by the second helical region of TRIF TIR domain. Proc Natl Acad Sci USA 110(47):19036–19041CrossRefGoogle Scholar
  12. 12.
    Piao W, Vogel SN, Toshchakov VY (2013) Inhibition of TLR4 signaling by TRAM-derived decoy peptides in vitro and in vivo. J Immunol 190(5):2263–2272CrossRefGoogle Scholar
  13. 13.
    Toshchakov VY, Szmacinski H, Couture LA, Lakowicz JR, Vogel SN (2011) Targeting TLR4 signaling by TLR4 Toll/IL-1 receptor domain-derived decoy peptides: identification of the TLR4 Toll/IL-1 receptor domain dimerization interface. J Immunol 186(8):4819–4827CrossRefGoogle Scholar
  14. 14.
    Toshchakov VY, Fenton MJ, Vogel SN (2007) Cutting Edge: differential inhibition of TLR signaling pathways by cell-permeable peptides representing BB loops of TLRs. J Immunol 178(5):2655–2660CrossRefGoogle Scholar
  15. 15.
    Couture LA, Piao W, Ru LW, Vogel SN, Toshchakov VY (2012) Targeting Toll-like receptor (TLR) signaling by Toll/interleukin-1 receptor (TIR) domain-containing adapter protein/MyD88 adapter-like (TIRAP/Mal)-derived decoy peptides. J Biol Chem 287(29):24641–24648CrossRefGoogle Scholar
  16. 16.
    Snyder GA, Cirl C, Jiang J, Chen K, Waldhuber A, Smith P, Rommler F, Snyder N, Fresquez T, Durr S, Tjandra N, Miethke T, Xiao TS (2013) Molecular mechanisms for the subversion of MyD88 signaling by TcpC from virulent uropathogenic Escherichia coli. Proc Natl Acad Sci USA 110(17):6985–6990CrossRefGoogle Scholar
  17. 17.
    Ve T, Williams SJ, Kobe B (2015) Structure and function of Toll/interleukin-1 receptor/resistance protein (TIR) domains. Apoptosis 20(2):250–261CrossRefGoogle Scholar
  18. 18.
    Narayanan KB, Park HH (2015) Toll/interleukin-1 receptor (TIR) domain-mediated cellular signaling pathways. Apoptosis 20(2):196–209CrossRefGoogle Scholar
  19. 19.
    Gay NJ, Symmons MF, Gangloff M, Bryant CE (2014) Assembly and localization of Toll-like receptor signalling complexes. Nat Rev Immunol 14(8):546–558CrossRefGoogle Scholar
  20. 20.
    Ke Y, Li W, Wang Y, Yang M, Guo J, Zhan S, Du X, Wang Z, Yang M, Li J, Li W, Chen Z (2016) Inhibition of TLR4 signaling by Brucella TIR-containing protein TcpB-derived decoy peptides. Int J Med Microbiol IJMM 306(6):391–400CrossRefGoogle Scholar
  21. 21.
    Li W, Ke Y, Wang Y, Yang M, Gao J, Zhan S, Xinying D, Huang L, Li W, Chen Z, Li J (2016) Brucella TIR-like protein TcpB/Btp1 specifically targets the host adaptor protein MAL/TIRAP to promote infection. Biochem Biophys Res Commun 477(3):509–514CrossRefGoogle Scholar
  22. 22.
    Yadav M, Zhang J, Fischer H, Huang W, Lutay N, Cirl C, Lum J, Miethke T, Svanborg C (2010) Inhibition of TIR domain signaling by TcpC: MyD88-dependent and independent effects on Escherichia coli virulence. PLoS Pathog 6(9):e1001120CrossRefGoogle Scholar
  23. 23.
    Verstrepen L, Bekaert T, Chau TL, Tavernier J, Chariot A, Beyaert R (2008) TLR-4, IL-1R and TNF-R signaling to NF-kappaB: variations on a common theme. Cell Mol Life Sci 65(19):2964–2978CrossRefGoogle Scholar
  24. 24.
    Takeuchi O, Kawai T, Muhlradt PF, Morr M, Radolf JD, Zychlinsky A, Takeda K, Akira S (2001) Discrimination of bacterial lipoproteins by Toll-like receptor 6. Int Immunol 13(7):933–940CrossRefGoogle Scholar
  25. 25.
    Ozinsky A, Underhill DM, Fontenot JD, Hajjar AM, Smith KD, Wilson CB, Schroeder L, Aderem A (2000) The repertoire for pattern recognition of pathogens by the innate immune system is defined by cooperation between toll-like receptors. Proc Natl Acad Sci USA 97(25):13766–13771CrossRefGoogle Scholar
  26. 26.
    Diebold SS, Kaisho T, Hemmi H, Akira S, Reis e Sousa C (2004) Innate antiviral responses by means of TLR7-mediated recognition of single-stranded RNA. Science 303(5663):1529–1531CrossRefGoogle Scholar
  27. 27.
    Martel-Pelletier J, Wildi LM, Pelletier JP (2012) Future therapeutics for osteoarthritis. Bone 51(2):297–311CrossRefGoogle Scholar
  28. 28.
    Goldring MB, Berenbaum F (2015) Emerging targets in osteoarthritis therapy. Curr Opin Pharmacol 22:51–63CrossRefGoogle Scholar
  29. 29.
    Chevalier X, Eymard F, Richette P (2013) Biologic agents in osteoarthritis: hopes and disappointments. Nat Rev Rheumatol 9(7):400–410CrossRefGoogle Scholar
  30. 30.
    Dinarello CA, Simon A, van der Meer JW (2012) Treating inflammation by blocking interleukin-1 in a broad spectrum of diseases. Nat Rev Drug Discov 11(8):633–652CrossRefGoogle Scholar
  31. 31.
    Nair A, Kanda V, Bush-Joseph C, Verma N, Chubinskaya S, Mikecz K, Glant TT, Malfait AM, Crow MK, Spear GT, Finnegan A, Scanzello CR (2012) Synovial fluid from patients with early osteoarthritis modulates fibroblast-like synoviocyte responses to toll-like receptor 4 and toll-like receptor 2 ligands via soluble CD14. Arthritis Rheum 64(7):2268–2277CrossRefGoogle Scholar
  32. 32.
    Miller MA, McTernan PG, Harte AL, Silva NF, Strazzullo P, Alberti KG, Kumar S, Cappuccio FP (2009) Ethnic and sex differences in circulating endotoxin levels: a novel marker of atherosclerotic and cardiovascular risk in a British multi-ethnic population. Atherosclerosis 203(2):494–502CrossRefGoogle Scholar
  33. 33.
    Lorenz W, Buhrmann C, Mobasheri A, Lueders C, Shakibaei M (2013) Bacterial lipopolysaccharides form procollagen-endotoxin complexes that trigger cartilage inflammation and degeneration: implications for the development of rheumatoid arthritis. Arthritis Res Ther 15(5):R111CrossRefGoogle Scholar
  34. 34.
    Lew WY, Bayna E, Molle ED, Dalton ND, Lai NC, Bhargava V, Mendiola V, Clopton P, Tang T (2013) Recurrent exposure to subclinical lipopolysaccharide increases mortality and induces cardiac fibrosis in mice. PLoS ONE 8(4):e61057CrossRefGoogle Scholar
  35. 35.
    Alaidarous M, Ve T, Casey LW, Valkov E, Ericsson DJ, Ullah MO, Schembri MA, Mansell A, Sweet MJ, Kobe B (2014) Mechanism of bacterial interference with TLR4 signaling by Brucella Toll/interleukin-1 receptor domain-containing protein TcpB. J Biol Chem 289(2):654–668CrossRefGoogle Scholar
  36. 36.
    Snyder GA, Deredge D, Waldhuber A, Fresquez T, Wilkins DZ, Smith PT, Durr S, Cirl C, Jiang J, Jennings W, Luchetti T, Snyder N, Sundberg EJ, Wintrode P, Miethke T, Xiao TS (2014) Crystal structures of the Toll/Interleukin-1 receptor (TIR) domains from the Brucella protein TcpB and host adaptor TIRAP reveal mechanisms of molecular mimicry. J Biol Chem 289(2):669–679CrossRefGoogle Scholar
  37. 37.
    Vyncke L, Bovijn C, Pauwels E, Van Acker T, Ruyssinck E, Burg E, Tavernier J, Peelman F (2016) Reconstructing the TIR Side of the Myddosome: a Paradigm for TIR-TIR Interactions. Structure 24(3):437–447CrossRefGoogle Scholar
  38. 38.
    Rana RR, Simpson P, Zhang M, Jennions M, Ukegbu C, Spear AM, Alguel Y, Matthews SJ, Atkins HS, Byrne B (2011) Yersinia pestis TIR-domain protein forms dimers that interact with the human adaptor protein MyD88. Microb Pathog 51(3):89–95CrossRefGoogle Scholar
  39. 39.
    Newman RM, Salunkhe P, Godzik A, Reed JC (2006) Identification and characterization of a novel bacterial virulence factor that shares homology with mammalian Toll/interleukin-1 receptor family proteins. Infect Immun 74(1):594–601CrossRefGoogle Scholar
  40. 40.
    Cirl C, Wieser A, Yadav M, Duerr S, Schubert S, Fischer H, Stappert D, Wantia N, Rodriguez N, Wagner H, Svanborg C, Miethke T (2008) Subversion of Toll-like receptor signaling by a unique family of bacterial Toll/interleukin-1 receptor domain-containing proteins. Nat Med 14(4):399–406CrossRefGoogle Scholar

Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  1. 1.Department of Orthopedics, Forth Medical CenterGeneral Hospital of the Chinese People’s Liberation ArmyBeijingPeople’s Republic of China
  2. 2.College of Bioscience and BioengineeringHebei University of Science and TechnologyShijiazhuangPeople’s Republic of China
  3. 3.Institute of Disease Control and PreventionBeijingPeople’s Republic of China
  4. 4.Department of Laboratory Medicine, The Third Medical CenterGeneral Hospital of the Chinese People’s Liberation ArmyBeijingPeople’s Republic of China

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