Expression and localization of CRAMP in rat tooth germ and during reparative dentin formation
- 265 Downloads
Cathelicidin-related antimicrobial peptide (CRAMP) is an antimicrobial peptide in mice and rats homologous to LL-37 in humans. In addition to its antibacterial activity, CRAMP has various physiological functions by binding to formyl peptide receptor 2 (FPR2). However, the role of these peptides in teeth is unknown. Therefore, we investigated the role of CRAMP and FPR2 in tooth development, reparative dentin formation, and defense response.
Material and methods
First, we examined the localization of CRAMP and FPR2 during tooth development by immunohistochemical analysis. Next, we investigated the localization of CRAMP, FPR2, and CD68-positive macrophages by immunohistochemical analysis during pulp inflammation and reparative dentin formation after cavity preparation. Finally, we analyzed the effect of lipopolysaccharide (LPS) on the expression of CRAMP and FPR2 in dental pulp cells by real-time reverse transcription PCR.
At the late bell stage in tooth development, CRAMP was detected in odontoblasts, and FPR2 was observed in the sub-odontoblastic layer. In mature teeth, CRAMP was not detected, but FPR2 continued to be localized in the sub-odontoblastic layer. After cavity preparation, CRAMP-positive cells and macrophages were found in dental pulp tissues below the cavity at an early stage of repair. At subsequent stages of reparative dentin formation, CRAMP was observed in odontoblast-like cells that contacted reparative dentin. FPR2 immunoreactivity was also detected in odontoblast-like cells and neighboring cells. LPS stimulated the expression of CRAMP mRNA in dental pulp cells in vitro.
Localization of CRAMP and its receptor FPR2-positive cells were observed during physiological and reparative dentin formation.
CRAMP/LL-37 has a possibility that induce reparative dentin formation.
KeywordsAntimicrobial peptide Tooth development Cavity preparation Odontoblast Reparative dentin
We express our sincere thanks to Dr. Takahashi of the Institute for Oral Science, Matsumoto Dental University for helpful discussions and encouragement.
This work was supported by Japan Society for the Promotion of Science (JSPS) KAKENHI Grant Number JP26893301.
Compliance with ethical standards
Conflict of interest
The authors have no conflicts of interest to declare.
All experiments were conducted in accordance with the guidelines for studies with laboratory animals of the Matsumoto Dental University Experimental Animal Committee.
For this type of study, formal consent is not required.
- 1.Soehnlein O, Wantha S, Simsekyilmaz S, Döring Y, Megens RT, Mause SF, Drechsler M, Smeets R, Weinandy S, Schreiber F, Gries T, Jockenhoevel S, Möller M, Vijayan S, van Zandvoort MA, Agerberth B, Pham CT, Gallo RL, Hackeng TM, Liehn EA, Zernecke A, Klee D, Weber C (2011) Neutrophil-derived cathelicidin protects from neointimal hyperplasia. Sci Transl Med 3:103ra98CrossRefPubMedPubMedCentralGoogle Scholar
- 3.Chromek M, Slamová Z, Bergman P, Kovács L, Podracká L, Ehrén I, Hökfelt T, Gudmundsson GH, Gallo RL, Agerberth B, Brauner A (2006) The antimicrobial peptide cathelicidin protects the urinary tract against invasive bacterial infection. Nat Med 12(6):636–641. https://doi.org/10.1038/nm1407 CrossRefPubMedGoogle Scholar
- 4.Kovach MA, Ballinger MN, Newstead MW, Zeng X, Bhan U, Yu FS, Moore BB, Gallo RL, Standiford TJ (2012) Cathelicidin-related antimicrobial peptide is required for effective lung mucosal immunity in Gram-negative bacterial pneumonia. J Immunol 189(1):304–311. https://doi.org/10.4049/jimmunol.1103196 CrossRefPubMedPubMedCentralGoogle Scholar
- 7.Mookherjee N, Brown KL, Bowdish DM, Doria S, Falsafi R, Hokamp K, Roche FM, Mu R, Doho GH, Pistolic J, Powers JP, Bryan J, Brinkman FS, Hancock RE (2006) Modulation of the TLRmediated inflammatory response by the endogenous human host defense peptide LL-37. J Immunol 176(4):2455–2464. https://doi.org/10.4049/jimmunol.176.4.2455 CrossRefPubMedGoogle Scholar
- 8.Yang D, Chen Q, Schmidt AP, Anderson GM, Wang JM, Wooters J, Oppenheim JJ, Chertov O (2000) LL-37, the neutrophil granule- and epithelial cell-derived cathelicidin, utilizes formyl peptide receptor-like 1 (FPRL1) as a receptor to chemoattract human peripheral blood neutrophils, monocytes, and T cells. J Exp Med 192(7):1069–1074. https://doi.org/10.1084/jem.192.7.1069 CrossRefPubMedPubMedCentralGoogle Scholar
- 9.Kurosaka K, Chen Q, Yarovinsky F, Oppenheim JJ, Yang D (2005) Mouse cathelin-related antimicrobial peptide chemoattracts leukocytes using formyl peptide receptor-like 1/mouse formyl peptide receptor-like 2 as the receptor and acts as an immune adjuvant. J Immunol 174(10):6257–6265. https://doi.org/10.4049/jimmunol.174.10.6257 CrossRefPubMedGoogle Scholar
- 10.Koczulla R, von Degenfeld G, Kupatt C, Krötz F, Zahler S, Gloe T, Issbrücker K, Unterberger P, Zaiou M, Lebherz C, Karl A, Raake P, Pfosser A, Boekstegers P, Welsch U, Hiemstra PS, Vogelmeier C, Gallo RL, Clauss M, Bals R (2003) An angiogenic role for the human peptide antibiotic LL-37/hCAP-18. J Clin Invest 111(11):1665–1672. https://doi.org/10.1172/JCI17545 CrossRefPubMedPubMedCentralGoogle Scholar
- 17.Tani-Ishii N, Wang CY, Stashenko P (1995) Immunolocalization of bone-resorptive cytokines in rat pulp and periapical lesions following surgical pulp exposure. Oral Microbiol Immunol 10(4):213–219. https://doi.org/10.1111/j.1399-302X.1995.tb00145.x CrossRefPubMedGoogle Scholar
- 26.Zhao H, Feng J, Seidel K, Shi S, Klein O, Sharpe P, Chai Y (2014) Secretion of Shh by a neurovascular bundle niche supports mesenchymal stem cell homeostasis in the adult mouse incisor. Cell Stem Cell 14(2):160–173. https://doi.org/10.1016/j.stem.2013.12.013 CrossRefPubMedPubMedCentralGoogle Scholar
- 27.Sarmiento BF, Aminoshariae A, Bakkar M, Bonfield T, Ghosh S, Montagnese TA, Mickel AK (2016) The expression of the human cathelicidin LL-37 in the human dental pulp: an in vivo study. Int J Pharm 1:5Google Scholar
- 31.Maddox JF, Hachicha M, Takano T, Petasis NA, Fokin VV, Serhan CN (1997) Lipoxin A4 stable analogs are potent mimetics that stimulate human monocytes and THP-1 cells via a G-protein-linked lipoxin A4 receptor. J Biol Chem 272(11):6972–6978. https://doi.org/10.1074/jbc.272.11.6972 CrossRefPubMedGoogle Scholar
- 32.Krishnamoorthy S, Recchiuti A, Chiang N, Yacoubian S, Lee CH, Yang R, Petasis NA, Serhan CN (2010) Resolvin D1 binds human phagocytes with evidence for proresolving receptors. Proc Natl Acad Sci U S A 107(4):1660–1665. https://doi.org/10.1073/pnas.0907342107 CrossRefPubMedPubMedCentralGoogle Scholar
- 34.Coffelt SB, Marini FC, Watson K, Zwezdaryk KJ, Dembinski JL, LaMarca HL, Tomchuck SL, Honer ZU, Bentrup K, Danka ES, Henkle SL, Scandurro AB (2009) The pro-inflammatory peptide LL-37 promotes ovarian tumor progression through recruitment of multipotent mesenchymal stromal cells. Proc Natl Acad Sci U S A 106(10):3806–3811. https://doi.org/10.1073/pnas.0900244106 CrossRefPubMedPubMedCentralGoogle Scholar
- 35.Kittaka M, Shiba H, Kajiya M, Fujita T, Iwata T, Rathvisal K, Ouhara K, Takeda K, Fujita T, Komatsuzawa H, Kurihara H (2013) The antimicrobial peptide LL37 promotes bone regeneration in a ratcalvarial bone defect. Peptides 46:136–142. https://doi.org/10.1016/j.peptides.2013.06.001 CrossRefPubMedGoogle Scholar