Establishing a murine model of experimental apical periodontitis induced by Fusobacterium nucleatum

doi:10.12307/2022.030

Abstract

Abstract: BACKGROUND: Apical periodontitis is a common oral inflammatory disease. Research on animal models for this disease has been extensively carried out, but there is yet no unified method for establishing such models.
OBJECTIVE: To establish a murine model of experimental apical periodontitis model induced by Fusobacterium nucleatum.
METHODS: Twenty-five BALB/c mice were divided into normal control group (n=5) and experimental group (n=20). Mice in the normal control group were sacrificed at 0 day to collect the lower jaws. In the experimental group, the root canal of the lower first molar was exposed followed by injection of Fusobacterium nucleatum suspension, and then the tooth crown was sealed with temporary restorative material. In the experimental group, five mice were selected at each observational time points (1, 3, 5, 7 days postoperatively), and samples of the mandible were collected accordingly. Micro-CT was performed to analyze alveolar bone resorption. The periapical tissue changes were observed by hematoxylin-eosin staining. Tartrate resistant acid phosphatase staining was applied to detect the expression of osteoclasts.
RESULTS AND CONCLUSION: Micro-CT scan showed that the apical periodontal space of the lower first molars began to widen at 3 days postoperatively and significant alveolar bone resorption was developed at 7 days. Three-dimensional reconstruction of the apical region showed decreased bone volume fraction and increased trabecular separation in the apical region at 7 days postoperatively (P < 0.05). Hematoxylin-eosin staining showed that inflammatory cell infiltration was obviously visible in the apical region at 3 days postoperatively, and then the infiltration gradually expanded. Tartrate resistant acid phosphatase staining showed a sustained increase in the number of osteoclasts in the apical region at 3, 5, and 7 days postoperatively. To conclude, the murine model of experimental apical periodontitis can be successfully established by inoculating Fusobacterium nucleatum into the root canals, based on which, we can further explore the etiology, pathology and treatment of apical periodontitis.

Key words: apical periodontitis, Fusobacterium nucleatum, Micro-CT, hematoxylin-eosin staining, osteoclast, animal model, mouse

CLC Number:

Wang Jiajia, Liu Jie, Wang Min. Establishing a murine model of experimental apical periodontitis induced by Fusobacterium nucleatum[J]. Chinese Journal of Tissue Engineering Research, 2022, 26(2): 176-181.

Figures/Tables 5

References

[1] NAIR PN. Apical periodontitis: a dynamic encounter between root canal infection and host response. Periodontol 2000. 1997;13:121-148.
[2] COLIĆ M, GAZIVODA D, VUCEVIĆ D, et al. Proinflammatory and immunoregulatory mechanisms in periapical lesions. Mol Immunol. 2009;47(1):101-113.
[3] MÁRTON IJ, KISS C. Overlapping protective and destructive regulatory pathways in apical periodontitis. J Endod. 2014;40(2):155-163.
[4] SHAH A, LEE D, SONG M, et al. Clastic cells are absent around the root surface in pulp-exposed periapical periodontitis lesions in mice. Oral Dis. 2018;24(1-2):57-62.
[5] 梅陵宣,刘正,张濒.实验性鼠根尖周炎组织学动态观察[J].实用口腔医学杂志,2002,18(6):500-503.
[6] DE ROSSI A, ROCHA LB, ROSSI MA. Interferon-gamma, interleukin-10, Intercellular adhesion molecule-1, and chemokine receptor 5, but not interleukin-4, attenuate the development of periapical lesions. J Endod. 2008;34(1):31-38.
[7] 陈兴兴,谷苗,吕海鹏,等.不同方法建立实验性狗根尖周炎模型的比较[J].口腔医学研究,2009,25(4):416-419.
[8] SILVA L, NELSON-FILHO P, LEONARDO MR, et al. Effect of calcium hydroxide on bacterial endotoxin in vivo. J Endod. 2002;28(2):94-98.
[9] 王青山,高艳,荣丽,等.内毒素诱导大鼠根尖周炎的X线片及组织病理学改变[J].上海口腔医学,2008,17(4):416-419.
[10] BRENNAN CA, GARRETT WS. Fusobacterium nucleatum - symbiont, opportunist and oncobacterium. Nat Rev Microbiol. 2019;17(3):156-166.
[11] 谢晓婷,王以玲,肖水清.具核梭杆菌的致病性与其可引起的口腔疾病[J].中国医学创新,2017,14(10):145-148.
[12] BOUILLAGUET S, MANOIL D, GIRARD M, et al. Root Microbiota in Primary and Secondary Apical Periodontitis. Front Microbiol. 2018;9: 2374.
[13] ZANDI H, KRISTOFFERSEN AK, ØRSTAVIK D, et al. Microbial Analysis of Endodontic Infections in Root-filled Teeth with Apical Periodontitis before and after Irrigation Using Pyrosequencing. J Endod. 2018;44(3): 372-378.
[14] 郭惠杰,王嘉德.慢性根尖周炎患牙根尖外表面的细菌生物膜[J].北京大学学报(医学版),2009,41(5):571-574.
[15] BULLMAN S, PEDAMALLU CS, SICINSKA E, et al. Analysis of Fusobacterium persistence and antibiotic response in colorectal cancer. Science. 2017;358(6369):1443-1448.
[16] 罗玉龙,葛全兴,聂玉强,等.具核梭杆菌感染与结直肠癌的关系[J].广东医学,2015,36(14):2161-2164.
[17] HAN YW, REDLINE RW, LI M, et al. Fusobacterium nucleatum induces premature and term stillbirths in pregnant mice: implication of oral bacteria in preterm birth. Infect Immun. 2004;72(4):2272-2279.
[18] SWIDSINSKI A, DÖRFFEL Y, LOENING-BAUCKE V, et al. Acute appendicitis is characterised by local invasion with Fusobacterium nucleatum/necrophorum. Gut. 2011;60(1):34-40.
[19] PETERSEN J, GLAßL EM, NASSERI P, et al. The association of chronic apical periodontitis and endodontic therapy with atherosclerosis. Clin Oral Investig. 2014;18(7):1813-1823.
[20] PAULA-SILVA FWG, RIBEIRO-SANTOS FR, PETEAN IBF, et al. Root canal contamination or exposure to lipopolysaccharide differentially modulate prostaglandin E 2 and leukotriene B 4 signaling in apical periodontitis. J Appl Oral Sci. 2020;28:e20190699.
[21] 凌均棨,韦曦,刘红艳.难治性根尖周炎的病因及防治策略[J].中华口腔医学杂志,2010,45(1):52-57.
[22] SIQUEIRA JF JR, RÔÇAS IN. Exploiting molecular methods to explore endodontic infections: Part 1--current molecular technologies for microbiological diagnosis. J Endod. 2005;31(6):411-423.
[23] XIONG H, WEI L, PENG B. The Presence and involvement of interleukin-17 in apical periodontitis. Int Endod J. 2019;52(8): 1128-1137.
[24] NAIR PN. On the causes of persistent apical periodontitis: a review. Int Endod J. 2006;39(4):249-281.
[25] SEGATA N, HAAKE SK, MANNON P, et al. Composition of the adult digestive tract bacterial microbiome based on seven mouth surfaces, tonsils, throat and stool samples. Genome Biol. 2012;13(6):R42.
[26] RICKARD AH, GILBERT P, HIGH NJ, et al. Bacterial coaggregation: an integral process in the development of multi-species biofilms. Trends Microbiol. 2003;11(2):94-100.
[27] BRADSHAW DJ, MARSH PD, WATSON GK, et al. Role of Fusobacterium nucleatum and coaggregation in anaerobe survival in planktonic and biofilm oral microbial communities during aeration. Infect Immun. 1998;66(10):4729-4732.
[28] 郭杨,张玉杰,肖水清.具核梭杆菌生物学特性及检测手段的研究进展[J].国际口腔医学杂志,2012,39(6):770-774.
[29] BACHRACH G, IANCULOVICI C, NAOR R, et al. Fluorescence based measurements of Fusobacterium nucleatum coaggregation and of fusobacterial attachment to mammalian cells. FEMS Microbiol Lett. 2005;248(2):235-240.
[30] 张竹砚.慢性根尖周炎动物模型的研究进展[J].口腔材料器械杂志, 2020,29(2):47-51.
[31] LUNGOVÁ V, RADLANSKI RJ, TUCKER AS, et al. Tooth-bone morphogenesis during postnatal stages of mouse first molar development. J Anat. 2011;218(6):699-716.
[32] 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;20:340-349.
[33] 周维君,车英林,杨瑞琨,等.根尖周炎模型小鼠DKK1,SFRP1及骨形成相关分子Runx2,Osteocalcin的表达变化[J].临床口腔医学杂志, 2017,33(12):715-719.
[34] ANDRADA AC, AZUMA MM, FURUSHO H, et al. Immunomodulation Mediated by Azithromycin in Experimental Periapical Inflammation. J Endod. 2020 Aug 5:S0099-2399(20)30574-4. doi: 10.1016/j.joen.2020.07.028.
[35] 王艳青.Cyclophilin A在根尖周炎中的表达及作用研究[D].武汉:武汉大学,2019.
[36] 张琛,侯本祥.根尖周炎动物模型中CD14、TLR4的表达[J].中华口腔医学杂志,2007,42(3):148-149.
[37] PAULA-SILVA FWG, ARNEZ MFM, PETEAN IBF, et al. Effects of 5-lipoxygenase gene disruption on inflammation, osteoclastogenesis and bone resorption in polymicrobial apical periodontitis. Arch Oral Biol. 2020;112:104670.
[38] WANG L, ZHANG H, DONG M, et al. Role of the Btk-PLCγ2 Signaling Pathway in the Bone Destruction of Apical Periodontitis. Mediators Inflamm. 2019;2019:8767529.
[39] TAMMA R, ZALLONE A. Osteoblast and osteoclast crosstalks: from OAF to Ephrin. Inflamm Allergy Drug Targets. 2012;11(3):196-200.
[40] 李福军,彭彬.大鼠实验性根尖周病的酶组织化学研究[J].口腔医学研究,2003,19(2):109-111.
[41] ZUBERY Y, DUNSTAN CR, STORY BM, et al. Bone resorption caused by three periodontal pathogens in vivo in mice is mediated in part by prostaglandin. Infect Immun. 1998;66(9):4158-4162.
[42] JOHNSON L, ALMEIDA-DA-SILVA CLC, TAKIYA CM, et al. Oral infection of mice with Fusobacterium nucleatum results in macrophage recruitment to the dental pulp and bone resorption. Biomed J. 2018;41(3):184-193.