Clinical Oral Investigations

, Volume 23, Issue 12, pp 4205–4212 | Cite as

Expression of Toll-like receptors 2 and 4 and its association with matrix metalloproteinases in symptomatic and asymptomatic apical periodontitis

  • A. Fernández
  • A. M. Cárdenas
  • J. Astorga
  • P. Veloso
  • A. Alvarado
  • P. Merino
  • D. Pino
  • D. Reyes-Court
  • M. Hernández
Original Article


To determine Toll-like receptors (TLR)2 and TLR4 expression levels and associate them with matrix metalloproteinases (MMPs) in asymptomatic apical periodontitis (AAP), symptomatic apical periodontitis (SAP), and healthy controls. Apical tissue/lesion samples were obtained from chronic AAP (n = 35) and SAP (n = 29), and healthy periodontal ligament (HPL, n = 10) with indication of tooth extraction, respectively. mRNA expression levels of TLR2, TLR4, MMP-1, MMP-2, MMP-8, and MMP-13 were determined by real-time reverse-transcription polymerase chain reaction. The data were analyzed with Kruskal-Wallis and Dunn’s pot hoc test (p < 0.05). The correlation coefficient was obtained using the Spearman correlation (p < 0.05). TLR2, MMP-1, MMP-2, and MMP-13 mRNA levels were the highest in SAP followed by AAP and controls (p < 0.05). TLR4 and MMP-8 were over expressed in AAP and SAP compared to HPL (p < 0.05). TLR2 positively correlated with TLR4, MMP-1, MMP-8, and MMP-13 in SAP (p < 0.05). TLR2 and TLR4 are overexpressed in apical lesions versus healthy periodontal ligament and correlate with collagenolytic MMPs. Particularly, TLR2 is overexpressed in SAP in association with MMP-1, MMP-8, and MMP-13. Our results suggest that the activation of TLR2 along with MMP overexpression might contribute to SAP clinical presentation and progression. TLRs, MMPs, and their interaction can explain the clinical presentations and evolution of apical periodontitis and might represent key targets for new diagnostic and treatment approaches.


Matrix metalloproteinases Periapical periodontitis Toll-like receptor 2 Toll-like receptor 4 



We thank Peter Gebicke-Haerter and Andrés Tittarelli for their valuable support with bioanalyzer equipment. Alejandra Fernández thanks the fellowship CONICYT 21181377, from the Chilean Government.


The work was supported by FONDECYT grant no. 1160741.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards.

Informed consent

Informed consent was obtained from all individual participants included in the study.


  1. 1.
    Menzies RA, Reiter AM, Lewis JR (2014) Assessment of apical periodontitis in dogs and humans: a review. J Vet Dent 31(1):8–21. CrossRefPubMedGoogle Scholar
  2. 2.
    Croitoru IC, CraiToiu S, Petcu CM, Mihailescu OA, Pascu RM, Bobic AG, Agop Forna D, CraiToiu MM (2016) Clinical, imagistic and histopathological study of chronic apical periodontitis. Romanian J Morphol Embryol 57(2 Suppl):719–728Google Scholar
  3. 3.
    Salinas-Munoz M, Garrido-Flores M, Baeza M, Huaman-Chipana P, Garcia-Sesnich J, Bologna R, Vernal R, Hernandez M (2017) Bone resorptive activity in symptomatic and asymptomatic apical lesions of endodontic origin. Clin Oral Investig 21(8):2613–2618. CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Desai SV, Love RM, Rich AM, Seymour GJ (2011) Toll-like receptor 2 expression in refractory periapical lesions. Int Endod J 44(10):907–916. CrossRefPubMedGoogle Scholar
  5. 5.
    Takeuchi O, Hoshino K, Kawai T, Sanjo H, Takada H, Ogawa T, Takeda K, Akira S (1999) Differential roles of TLR2 and TLR4 in recognition of gram-negative and gram-positive bacterial cell wall components. Immunity 11(4):443–451CrossRefGoogle Scholar
  6. 6.
    de Oliveira Rde C, Beghini M, Borges CR, Alves PM, de Araujo MS, Pereira SA, Rodrigues V Jr, Rodrigues DB (2014) Higher expression of galectin-3 and galectin-9 in periapical granulomas than in radicular cysts and an increased toll-like receptor-2 and toll-like receptor-4 expression are associated with reactivation of periapical inflammation. J Endod 40(2):199–203. CrossRefPubMedGoogle Scholar
  7. 7.
    Kassem A, Henning P, Lundberg P, Souza PP, Lindholm C, Lerner UH (2015) Porphyromonas gingivalis stimulates bone resorption by enhancing RANKL (receptor activator of NF-kappaB ligand) through activation of toll-like receptor 2 in osteoblasts. J Biol Chem 290(33):20147–20158. CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Itoh K, Udagawa N, Kobayashi K, Suda K, Li X, Takami M, Okahashi N, Nishihara T, Takahashi N (2003) Lipopolysaccharide promotes the survival of osteoclasts via Toll-like receptor 4, but cytokine production of osteoclasts in response to lipopolysaccharide is different from that of macrophages. J Immunol 170(7):3688–3695CrossRefGoogle Scholar
  9. 9.
    Tang L, Zhou XD, Wang Q, Zhang L, Wang Y, Li XY, Huang DM (2011) Expression of TRAF6 and pro-inflammatory cytokines through activation of TLR2, TLR4, NOD1, and NOD2 in human periodontal ligament fibroblasts. Arch Oral Biol 56(10):1064–1072. CrossRefPubMedGoogle Scholar
  10. 10.
    Barreiros D, Nelson PF, Paula-Silva FWG, Oliveira KMH, Lucisano MP, Rossi A, Silva LAB, Kuchler EC, Silva RAB (2018) MMP2 and MMP9 are associated with apical periodontitis progression and might be modulated by TLR2 and MyD88. Braz Dent J 29(1):43–47. CrossRefPubMedGoogle Scholar
  11. 11.
    Cavalla F, Hernández-Ríos P, Sorza T, Biguetti C, Hernández M (2017) Matrix metalloproteinases as regulators of periodontal inflammation. Int J Mol Sci 18(2):440CrossRefGoogle Scholar
  12. 12.
    Paula-Silva FW, da Silva LA, Kapila YL (2010) Matrix metalloproteinase expression in teeth with apical periodontitis is differentially modulated by the modality of root canal treatment. J Endod 36(2):231–237. CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Lin SK, Chiang CP, Hong CY, Lin CP, Lan WH, Hsieh CC, Kuo MY (1997) Immunolocalization of interstitial collagenase (MMP-1) and tissue inhibitor of metalloproteinases-1 (TIMP-1) in radicular cysts. J Oral Pathol Med 26(10):458–463CrossRefGoogle Scholar
  14. 14.
    Mundi Burgos V, Dezerega Piwonka A, Osorio Alfaro C, Dutzan Muñoz N, Franco Martínez ME, Ortega Pinto AV, Hernandez Rios M (2011) Inmunodetección de metaloproteinasas de matriz extracelular (MMPs) -2, -9, -13, -14 en lesiones apicales asociadas con periotontitis apical asintomática. Rev Clin Periodoncia Implantol Rehabil Oral 4(1):17–21CrossRefGoogle Scholar
  15. 15.
    Baeza M, Garrido M, Hernandez-Rios P, Dezerega A, Garcia-Sesnich J, Strauss F, Aitken JP, Lesaffre E, Vanbelle S, Gamonal J, Brignardello-Petersen R, Tervahartiala T, Sorsa T, Hernandez M (2016) Diagnostic accuracy for apical and chronic periodontitis biomarkers in gingival crevicular fluid: an exploratory study. J Clin Periodontol 43(1):34–45. CrossRefPubMedGoogle Scholar
  16. 16.
    Pereira Faustino IS, Azevedo RS, Takahama A Jr (2016) Metalloproteinases 2 and 9 immunoexpression in periapical lesions from primary endodontic infection: possible relationship with the histopathological diagnosis and the presence of pain. J Endod 42(4):547–551. CrossRefPubMedGoogle Scholar
  17. 17.
    Agarwal S, Misra R, Aggarwal A (2010) Induction of metalloproteinases expression by TLR ligands in human fibroblast like synoviocytes from juvenile idiopathic arthritis patients. Indian J Med Res 131:771–779PubMedGoogle Scholar
  18. 18.
    Lisboa RA, Andrade MV, Cunha-Melo JR (2013) Toll-like receptor activation and mechanical force stimulation promote the secretion of matrix metalloproteinases 1, 3 and 10 of human periodontal fibroblasts via p38, JNK and NF-kB. Arch Oral Biol 58(6):731–739. CrossRefPubMedGoogle Scholar
  19. 19.
    Gutmann JL, Baumgartner JC, Gluskin AH, Hartwell GR, Walton RE (2009) Identify and define all diagnostic terms for periapical/periradicular health and disease states. J Endod 35(12):1658–1674. CrossRefPubMedGoogle Scholar
  20. 20.
    Glickman GN (2009) AAE consensus conference on diagnostic terminology: background and perspectives. J Endod 35(12):1619–1620. CrossRefPubMedGoogle Scholar
  21. 21.
    Buonavoglia A, Latronico F, Pirani C, Greco MF, Corrente M, Prati C (2013) Symptomatic and asymptomatic apical periodontitis associated with red complex bacteria: clinical and microbiological evaluation. Odontology 101(1):84–88. CrossRefPubMedGoogle Scholar
  22. 22.
    Hou L, Sasaki H, Stashenko P (2000) Toll-like receptor 4-deficient mice have reduced bone destruction following mixed anaerobic infection. Infect Immun 68(8):4681–4687CrossRefGoogle Scholar
  23. 23.
    Rider D, Furusho H, Xu S, Trachtenberg AJ, Kuo WP, Hirai K, Susa M, Bahammam L, Stashenko P, Fujimura A, Sasaki H (2016) Elevated CD14 (cluster of differentiation 14) and toll-like receptor (TLR) 4 signaling deteriorate periapical inflammation in TLR2 deficient mice. Anat Rec (Hoboken) 299(9):1281–1292. CrossRefGoogle Scholar
  24. 24.
    Cavalla F, Reyes M, Vernal R, Alvarez C, Paredes R, Garcia-Sesnich J, Infante M, Farina V, Barron I, Hernandez M (2013) High levels of CXC ligand 12/stromal cell-derived factor 1 in apical lesions of endodontic origin associated with mast cell infiltration. J Endod 39(10):1234–1239. CrossRefPubMedGoogle Scholar
  25. 25.
    Garrido M, Dezerega A, Bordagaray MJ, Reyes M, Vernal R, Melgar-Rodriguez S, Ciuchi P, Paredes R, Garcia-Sesnich J, Ahumada-Montalva P, Hernandez M (2015) C-reactive protein expression is up-regulated in apical lesions of endodontic origin in association with interleukin-6. J Endod 41(4):464–469. CrossRefPubMedGoogle Scholar
  26. 26.
    Hernández P, Mäntylä P, Tervahartiala T, Sorsa T, Hernández M (2012) Análisis de MMPs en fluidos orales en el diagnóstico complementario de las enfermedades periodontales. Rev Clin Periodoncia Implantol Rehabil Oral 5(3):150–153CrossRefGoogle Scholar
  27. 27.
    Sbardella D, Fasciglione GF, Gioia M, Ciaccio C, Tundo GR, Marini S, Coletta M (2012) Human matrix metalloproteinases: an ubiquitarian class of enzymes involved in several pathological processes. Mol Asp Med 33:119–208. CrossRefGoogle Scholar
  28. 28.
    Buzoglu HD, Unal H, Ulger C, Mert S, Kucukyildirim S, Er N (2009) The zymographic evaluation of gelatinase (MMP-2 and -9) levels in acute and chronic periapical abscesses. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 108(5):e121–e126. CrossRefPubMedGoogle Scholar
  29. 29.
    Letra A, Ghaneh G, Zhao M, Ray H Jr, Francisconi CF, Garlet GP, Silva RM (2013) MMP-7 and TIMP-1, new targets in predicting poor wound healing in apical periodontitis. J Endod 39(9):1141–1146. CrossRefPubMedGoogle Scholar
  30. 30.
    Ahmed GM, El-Baz AA, Hashem AA, Shalaan AK (2013) Expression levels of matrix metalloproteinase-9 and gram-negative bacteria in symptomatic and asymptomatic periapical lesions. J Endod 39(4):444–448. CrossRefPubMedGoogle Scholar
  31. 31.
    Kawasaki Y, Xu ZZ, Wang X, Park JY, Zhuang ZY, Tan PH, Gao YJ, Roy K, Corfas G, Lo EH, Ji RR (2008) Distinct roles of matrix metalloproteases in the early- and late-phase development of neuropathic pain. Nat Med 14(3):331–336. CrossRefPubMedPubMedCentralGoogle Scholar
  32. 32.
    Corotti MV, Zambuzzi WF, Paiva KB, Menezes R, Pinto LC, Lara VS, Granjeiro JM (2009) Immunolocalization of matrix metalloproteinases-2 and -9 during apical periodontitis development. Arch Oral Biol 54(8):764–771. CrossRefPubMedGoogle Scholar
  33. 33.
    Hernandez M, Dutzan N, Garcia-Sesnich J, Abusleme L, Dezerega A, Silva N, Gonzalez FE, Vernal R, Sorsa T, Gamonal J (2011) Host-pathogen interactions in progressive chronic periodontitis. J Dent Res 90(10):1164–1170. CrossRefPubMedGoogle Scholar
  34. 34.
    Hadziabdic N, Kurtovic-Kozaric A, Pojskic N, Sulejmanagic N, Todorovic L (2016) Gene-expression analysis of matrix metalloproteinases 1 and 2 and their tissue inhibitors in chronic periapical inflammatory lesions. J Oral Pathol Med 45(3):224–230. CrossRefPubMedGoogle Scholar
  35. 35.
    Trombone AP, Cavalla F, Silveira EM, Andreo CB, Francisconi CF, Fonseca AC, Letra A, Silva RM, Garlet GP (2016) MMP1-1607 polymorphism increases the risk for periapical lesion development through the upregulation MMP-1 expression in association with pro-inflammatory milieu elements. J Appl Oral Sci 24(4):366–375. CrossRefPubMedPubMedCentralGoogle Scholar
  36. 36.
    Shin SJ, Lee W, Lee JI, Baek SH, Kum KY, Shon WJ, Bae KS (2011) Matrix metalloproteinase-8 and substance P levels in gingival crevicular fluid during endodontic treatment of painful, nonvital teeth. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 112(4):548–554. CrossRefPubMedGoogle Scholar
  37. 37.
    Kuula H, Salo T, Pirila E, Tuomainen AM, Jauhiainen M, Uitto VJ, Tjaderhane L, Pussinen PJ, Sorsa T (2009) Local and systemic responses in matrix metalloproteinase 8-deficient mice during porphyromonas gingivalis-induced periodontitis. Infect Immun 77(2):850–859. CrossRefPubMedGoogle Scholar
  38. 38.
    Hernandez M, Valenzuela MA, Lopez-Otin C, Alvarez J, Lopez JM, Vernal R, Gamonal J (2006) Matrix metalloproteinase-13 is highly expressed in destructive periodontal disease activity. J Periodontol 77(11):1863–1870. CrossRefPubMedGoogle Scholar
  39. 39.
    Matsui H, Yamasaki M, Nakata K, Amano K, Nakamura H (2011) Expression of MMP-8 and MMP-13 in the development of periradicular lesions. Int Endod J 44(8):739–745. CrossRefPubMedGoogle Scholar
  40. 40.
    Hong CY, Lin SK, Kok SH, Cheng SJ, Lee MS, Wang TM, Chen CS, Lin LD, Wang JS (2004) The role of lipopolysaccharide in infectious bone resorption of periapical lesion. J Oral Pathol Med 33(3):162–169CrossRefGoogle Scholar
  41. 41.
    Al-Rashed F, Kochumon S, Usmani S, Sindhu S, Ahmad R (2017) Pam3CSK4 induces MMP-9 expression in human monocytic THP-1 cells. Cell Physiol Biochem 41(5):1993–2003. CrossRefPubMedGoogle Scholar
  42. 42.
    Ahmad R, Shihab PK, Jasem S, Behbehani K (2014) FSL-1 induces MMP-9 production through TLR-2 and NF-kappaB /AP-1 signaling pathways in monocytic THP-1 cells. Cell Physiol Biochem 34(3):929–942. CrossRefPubMedGoogle Scholar
  43. 43.
    Lim R, Barker G, Lappas M (2014) The TLR2 ligand FSL-1 and the TLR5 ligand flagellin mediate pro-inflammatory and pro-labour response via MyD88/TRAF6/NF-kappaB-dependent signalling. Am J Reprod Immunol 71(5):401–417. CrossRefPubMedGoogle Scholar
  44. 44.
    Osorio C, Cavalla F, Paula-Lima A, Diaz-Araya G, Vernal R, Ahumada P, Gamonal J, Hernandez M (2015) H2 O2 activates matrix metalloproteinases through the nuclear factor kappa B pathway and Ca(2+) signals in human periodontal fibroblasts. J Periodontal Res 50(6):798–806. CrossRefPubMedGoogle Scholar
  45. 45.
    Sindhu S, Al-Roub A, Koshy M, Thomas R, Ahmad R (2016) Palmitate-induced MMP-9 expression in the human monocytic cells is mediated through the TLR4-MyD88 dependent mechanism. Cell Physiol Biochem 39(3):889–900. CrossRefPubMedGoogle Scholar
  46. 46.
    Nociti FH Jr, Casati MZ, Duarte PM (2015) Current perspective of the impact of smoking on the progression and treatment of periodontitis. Periodontol 67(1):187–210. CrossRefGoogle Scholar
  47. 47.
    Bergstrom J, Babcan J, Eliasson S (2004) Tobacco smoking and dental periapical condition. Eur J Oral Sci 112(2):115–120. CrossRefPubMedGoogle Scholar
  48. 48.
    Persic Bukmir R, Jurcevic Grgic M, Brumini G, Spalj S, Pezelj-Ribaric S, Brekalo Prso I (2016) Influence of tobacco smoking on dental periapical condition in a sample of Croatian adults. Wien Klin Wochenschr 128(7–8):260–265. CrossRefPubMedGoogle Scholar
  49. 49.
    Senturk RA, Sezgin Y, Bulut S, Ozdemir BH (2018) The effects of smoking on the expression of gelatinases in chronic periodontitis: a cross-sectional study. Braz Oral Res 32:e114. CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  • A. Fernández
    • 1
    • 2
  • A. M. Cárdenas
    • 1
    • 3
  • J. Astorga
    • 1
  • P. Veloso
    • 1
  • A. Alvarado
    • 2
  • P. Merino
    • 1
  • D. Pino
    • 2
  • D. Reyes-Court
    • 4
  • M. Hernández
    • 1
    • 5
    • 6
  1. 1.Laboratory of Periodontal Biology, Faculty of DentistryUniversidad de ChileSantiagoChile
  2. 2.Faculty of DentistryUniversidad Andres BelloSantiagoChile
  3. 3.Health sciences division, Faculty of DentistryUniversidad Santo TomásBucaramangaColombia
  4. 4.Department of Surgery, Faculty of DentistryUniversidad de ChileSantiagoChile
  5. 5.Department of Oral Pathology and Medicine, Faculty of DentistryUniversidad de ChileSantiagoChile
  6. 6.Dentistry Unit, Faculty of Health SciencesUniversidad Autónoma de ChileSantiagoChile

Personalised recommendations