Objectives: Calcium hydroxide, a root canal temporary sealer, has long been used and it has anti-bacterial and anti-inflammatory activity. To investigate the properties of a newly developed calcium hydroxide paste comprising silicon-substituted hydroxyapatite (Si-HA), we examined the anti-inflammatory activity of the new calcium hydroxide paste in RAW 264.7 macrophages stimulated by lipopolysaccharide (LPS), which causes infection of the root canal. Methods: The test materials, including Calcipex II as control group and the newly developed TRC paste, were extracted from cell culture media and then diluted for experiment. In LPS-stimulated RAW 264.7 cells, the cytotoxicity and nitric oxide (NO) production of test materials were measured by MTT assay and Griess reagents, respectively. Also, the expression of the inducible nitric oxide synthase (iNOS) was assessed by western blotting. Results: The IC50 values of Calcipex II and TRC paste were 17.6 ㎎/㎖ and 13.5 ㎎/㎖, respectively. The level of NO, increased by LPS, was dose-dependently inhibited more by TRC paste than Calcipex II treatment. In addition, iNOS expression was decreased by 71% and 92% at concentrations of 2 ㎎/㎖ and 20 ㎎/㎖ of TRC paste, respectively. Conclusions: We demonstrated that the Si-HA calcium hydroxide paste has a slightly improved anti-inflammatory property and further studies are needed before clinical recommendations are proposed.
1. Byström A, Claesson R, Sundqvist G. The antibacterial effect of camphorated paramonochlorophenol, camphorated phenol and calcium hydroxide in the treatment of infected root canals. Endod Dent Traumatol 1985;1:170-5.
2. Athanassiadis B, Abbott PV, Walsh LJ. The use of calcium hydroxide, antibiotics and biocides as antimicrobial medicaments in endodontics. Aust Dent J 2007;52:S64-S82. https://doi.org/10.1111/j.1834-7819.2007.tb00527.x
3. Narita H, Itoh S, Imazato S, Yoshitake F, Ebisu S. An explanation of the mineralization mechanism in osteoblasts induced by calcium hydroxide. Acta Biomater 2010;6:586-90. https://doi.org/10.1016/j.actbio.2009.08.005
4. Han B, Wang X, Liu J, Liang F, Qu X, Yang Z, Gao X. The biological performance of calcium hydroxide–loaded microcapsules. J Endod 2013;39:1030-4. https://doi.org/10.1016/j.joen.2013.04.014
5. Silva LAB, Nelson-Filho P, Leonardo MR, Rossi MA, Pansani CA. Effect of calcium hydroxide on bacterial endotoxin in vivo . J Endod 2002;28:94-8.
6. Balamurugan A, Rebelo AH, Lemos AF, Rocha JH, Ventura JM, Ferreira JM. Suitability evaluation of sol–gel derived Si-substituted hydroxyapatite for dental and maxillofacial applications through in vitro osteoblasts response. Dent Mater 2008;24:1374-80. https://doi.org/10.1016/j.dental.2008.02.017
7. Torres N, Oh S, Appleford M, Dean DD, Jorgensen JH, Ong JL, et al. Stability of antibacterial self-assembled monolayers on hydroxyapatite. Acta Biomater 2010;6:3242-55. https://doi.org/10.1016/j.actbio.2010.02.032
8. Stanić V, Jana ć kovi ć D, Dimitrijevi ć S, Tanaskovi ć S.B, Mitri ć M, Pavlovi ć MS, et al. Synthesis of antimicrobial monophase silver-doped hydroxyapatite nanopowders for bone tissue engineering. Appl Surf Sci 2011;257:4510-8. https://doi.org/10.1016/j.apsusc.2010.12.113
9. Schonfeld SE, Greening AB, Glick DH, Frank AL, Simon JH, Herles SM. Endotoxic activity in periapical lesions. Oral Surg Oral Med Oral Pathol 1982;53:82-7.
10. Kakehashi S, Stanley HR, Fitzgerald RJ. The effects of surgical exposures of dental pulps in germ-free and conventional laboratory rats. Oral Surg Oral Med Oral Pathol 1965;20:340-9. https://doi.org/10.1016/0030-4220(65)90166-0
11. Jiang J, Zuo J, Chen SH, Holliday LS. Calcium hydroxide reduces lipopolysaccharide-stimulated osteoclast formation. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2003;95:348-54. https://doi.org/10.1067/moe.2003.18
12. Kilbourn RG, Griffith OW. Overproduction of nitric oxide in cytokine-mediated and septic shock. J Natl Cancer Inst 1992;84:827-31.
13. Bogdan C, Rollinghoff M, Diefenbach A. The role of nitric oxide in innate immunity. Immunol rev 2000;173:17-26.
14. da Silva RAB, Leonardo MR, da Silva LAB, Faccioli LH, Medeiros AI. Effect of a calcium hydroxide–based paste associated to chlorhexidine on RAW 264.7 macrophage cell line culture. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2008;106:e44-e51. https://doi.org/10.1016/j.tripleo.2008.06.027
15. Petel R, Moskovitz M, Tickotsky N, Halabi A, Goldstein J, Houri-Haddad Y. Cytotoxicity and proliferative effects of Iodoform-containing root canal-filling material on RAW 264.7 macrophage and RKO epithelial cell lines. Arch Oral Biol 2013;58:75-81. https://doi.org/10.1016/j.archoralbio.2012.06.014
16. Mahoney JR Jr, Beutler BA, Le Trang N, Vine W, Ikeda Y, Kawakami M, et al. Lipopolysaccharide-treated RAW 264.7 cells produce a mediator that inhibits lipoprotein lipase in 3T3-L1 cells. J Immunol 1985;134:1673-75.
17. KIM MS, Ha SM, Choi YM. Porous composite comprising silicon-substituted hydroxyapatite and beta-tricalcium phosphate, and process for preparing the same. Google Patents; 2010.
18. Roh J, Kim JY, Choi YM, Ha SM, Kim KN, Kim KM. Bone regeneration using a mixture of silicon-substituted coral HA and β-TCP in a rat calvarial bone defect model. Materials 2016;9:E97. https://doi.org/10.3390/ma9020097
19. Green LC, Wagner DA, Glogowski J, Skipper PL, Wishnok JS, Tannenbaum SR. Analysis of nitrate, nitrite, and [15N]nitrate in biological fluids. Anal Biochem 1982;126:131-8.
20. Kim D, Kim E. Antimicrobial effect of calcium hydroxide as an intracanal medicament in root canal treatment: a literature review - Part I. In vitro studies. Restor Dent Endod 2014;39:241-52. https://doi.org/10.5395/rde.2014.39.4.241
21. Desai S, Chandler N. Calcium hydroxide-based root canal sealers: a review. J Endod 2009;35: 475-80. https://doi.org/10.1016/j.joen.2008.11.026
22. Kim JW, Cho KM, Park SH, Song SG, Park MS, Jung HR, et al. Overfilling of calcium hydroxide-based paste Calcipex II produced a foreign body granuloma without acute inflammatory reaction. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2009;107:e73-6. https://doi.org/10.1016/j.tripleo.2008.10.019
23. Olsen JJ, Thorn JJ, Korsgaard N, Pinholt EM. Nerve lesions following apical extrusion of non-setting calcium hydroxide: A systematic case review and report of two cases. J Craniomaxillofac Surg 2014;42:757-62. https://doi.org/10.1016/j.jcms.2013.11.007
24. Sutha S, Kavitha K, Karunakaran G, Rajendran V. In-vitro bioactivity, biocorrosion and antibacterial activity of silicon integrated hydroxyapatite/chitosan composite coating on 316 L stainless steel implants. Mater Sci Eng C Mater Biol Appl 2013;33:4046-54. https://doi.org/10.1016/j.msec.2013.05.047
25. Thian ES, Huang J, Best SM, Barber ZH, Brooks RA, Rushton N, et al. The response of osteoblasts to nanocrystalline silicon-substituted hydroxyapatite thin films. Biomaterials 2006;27:2692-8. https://doi.org/10.1016/j.biomaterials.2005.12.019
26. Patel N, Best SM, Bonfield W, Gibson IR, Hing KA, Damien E, et al. A comparative study on the in vivo behavior of hydroxyapatite and silicon substituted hydroxyapatite granules. J Mater Sci Mater Med 2002;13:1199-206. https://doi.org/10.1023/A:1021114710076
27. Fava LRG, Saunders WP. Calcium hydroxide pastes: classification and clinical indications. Int Endod J 1999;32:257-82. https://doi.org/10.1046/j.1365-2591.1999.00232.x
28. Kleinert H, Pautz A, Linker K, Schwarz PM. Regulation of the expression of inducible nitric oxide synthase. Eur J Pharmacol 2004;500:255-66. https://doi.org/10.1016/j.niox.2010.04.007