Efecto de dos soluciones irrigadoras de quitosán sobre la liberación de proteínas bioactivas de la dentina radicular

ilustraciones, gráficas, tablas

Autores:
Quijano Guauque, Sara Beatriz
Tipo de recurso:
Fecha de publicación:
2016
Institución:
Universidad Nacional de Colombia
Repositorio:
Universidad Nacional de Colombia
Idioma:
spa
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oai:repositorio.unal.edu.co:unal/81276
Acceso en línea:
https://repositorio.unal.edu.co/handle/unal/81276
https://repositorio.unal.edu.co/
Palabra clave:
610 - Medicina y salud::613 - Salud y seguridad personal
Dentina
Endodoncia Regenerativa
Quitosano
Dentin
Regenerative Endodontics
Chitosan
Regeneración
Endodoncia
Quitosano
Péptidos y Proteínas de Señalización Intercelular
Dentina.
Regeneration
Endodontics
Chitosan
Intercellular Signaling Peptides and Proteins
Dentin
Rights
openAccess
License
Atribución-NoComercial-SinDerivadas 4.0 Internacional
id UNACIONAL2_16a1f15f39a30d3be59364d1b4dd9b44
oai_identifier_str oai:repositorio.unal.edu.co:unal/81276
network_acronym_str UNACIONAL2
network_name_str Universidad Nacional de Colombia
repository_id_str
dc.title.spa.fl_str_mv Efecto de dos soluciones irrigadoras de quitosán sobre la liberación de proteínas bioactivas de la dentina radicular
dc.title.translated.eng.fl_str_mv Effect of two irrigating solutions of chitosan on the release of bioactive proteins from root dentin
title Efecto de dos soluciones irrigadoras de quitosán sobre la liberación de proteínas bioactivas de la dentina radicular
spellingShingle Efecto de dos soluciones irrigadoras de quitosán sobre la liberación de proteínas bioactivas de la dentina radicular
610 - Medicina y salud::613 - Salud y seguridad personal
Dentina
Endodoncia Regenerativa
Quitosano
Dentin
Regenerative Endodontics
Chitosan
Regeneración
Endodoncia
Quitosano
Péptidos y Proteínas de Señalización Intercelular
Dentina.
Regeneration
Endodontics
Chitosan
Intercellular Signaling Peptides and Proteins
Dentin
title_short Efecto de dos soluciones irrigadoras de quitosán sobre la liberación de proteínas bioactivas de la dentina radicular
title_full Efecto de dos soluciones irrigadoras de quitosán sobre la liberación de proteínas bioactivas de la dentina radicular
title_fullStr Efecto de dos soluciones irrigadoras de quitosán sobre la liberación de proteínas bioactivas de la dentina radicular
title_full_unstemmed Efecto de dos soluciones irrigadoras de quitosán sobre la liberación de proteínas bioactivas de la dentina radicular
title_sort Efecto de dos soluciones irrigadoras de quitosán sobre la liberación de proteínas bioactivas de la dentina radicular
dc.creator.fl_str_mv Quijano Guauque, Sara Beatriz
dc.contributor.advisor.none.fl_str_mv Castellanos Parra, Jaime Eduardo
García Guerrero, Claudia Carmiña
dc.contributor.author.none.fl_str_mv Quijano Guauque, Sara Beatriz
dc.contributor.researchgroup.spa.fl_str_mv Invendo
dc.subject.ddc.spa.fl_str_mv 610 - Medicina y salud::613 - Salud y seguridad personal
topic 610 - Medicina y salud::613 - Salud y seguridad personal
Dentina
Endodoncia Regenerativa
Quitosano
Dentin
Regenerative Endodontics
Chitosan
Regeneración
Endodoncia
Quitosano
Péptidos y Proteínas de Señalización Intercelular
Dentina.
Regeneration
Endodontics
Chitosan
Intercellular Signaling Peptides and Proteins
Dentin
dc.subject.decs.spa.fl_str_mv Dentina
Endodoncia Regenerativa
Quitosano
dc.subject.decs.eng.fl_str_mv Dentin
Regenerative Endodontics
Chitosan
dc.subject.proposal.spa.fl_str_mv Regeneración
Endodoncia
Quitosano
Péptidos y Proteínas de Señalización Intercelular
Dentina.
dc.subject.proposal.eng.fl_str_mv Regeneration
Endodontics
Chitosan
Intercellular Signaling Peptides and Proteins
Dentin
description ilustraciones, gráficas, tablas
publishDate 2016
dc.date.issued.none.fl_str_mv 2016
dc.date.accessioned.none.fl_str_mv 2022-03-17T19:12:17Z
dc.date.available.none.fl_str_mv 2022-03-17T19:12:17Z
dc.type.spa.fl_str_mv Trabajo de grado - Maestría
dc.type.driver.spa.fl_str_mv info:eu-repo/semantics/masterThesis
dc.type.version.spa.fl_str_mv info:eu-repo/semantics/acceptedVersion
dc.type.content.spa.fl_str_mv Other
dc.type.redcol.spa.fl_str_mv http://purl.org/redcol/resource_type/TM
status_str acceptedVersion
dc.identifier.uri.none.fl_str_mv https://repositorio.unal.edu.co/handle/unal/81276
dc.identifier.instname.spa.fl_str_mv Universidad Nacional de Colombia
dc.identifier.reponame.spa.fl_str_mv Repositorio Institucional Universidad Nacional de Colombia
dc.identifier.repourl.spa.fl_str_mv https://repositorio.unal.edu.co/
url https://repositorio.unal.edu.co/handle/unal/81276
https://repositorio.unal.edu.co/
identifier_str_mv Universidad Nacional de Colombia
Repositorio Institucional Universidad Nacional de Colombia
dc.language.iso.spa.fl_str_mv spa
language spa
dc.relation.references.spa.fl_str_mv 1. Xie Z, Shen Z, Zhan P, Yang J, Huang Q, Huang S, Chen L, Lin Z. Functional Dental Pulp Regeneration: Basic Research and Clinical Translation. Int J Mol Sci. 2021 Aug 20;22(16):8991
2. Smith JG, Smith AJ, Shelton RM, Cooper PR. Dental Pulp Cell Behavior in Biomimetic Environments. J Dent Res. 2015 Nov;94(11):1552-9.
3. Huang X, Li Z, Liu A, Liu X, Guo H, Wu M, Yang X, Han B, Xuan K. Microenvironment Influences Odontogenic Mesenchymal Stem Cells Mediated Dental Pulp Regeneration. Front Physiol. 2021 Apr 22;12:656588.
4. Jeeruphan T, Jantarat J, Yanpiset K, Suwannapan L, Khewsawai P, Hargreaves KM. Mahidol study 1: comparison of radiographic and survival outcomes of immature teeth treated with either regenerative endodontic or apexification methods: a retrospective study. J Endod. 2012 Oct;38(10):1330-6.
5. Jung C, Kim S, Sun T, Cho YB, Song M. Pulp-dentin regeneration: current approaches and challenges. J Tissue Eng. 2019; 10:2041731418819263.
6. Zhang X, Li H, Sun J, Luo X, Yang H, Xie L, Yang B, Guo W, Tian W. Cell-derived micro-environment helps dental pulp stem cells promote dental pulp regeneration. Cell Prolif. 2017 Oct;50(5):e12361.
7. Dung SZ, Gregory RL, Li Y, Stookey GK. Effect of lactic acid and proteolytic enzymes on the release of organic matrix components from human root dentin. Caries Res. 1995;29(6):483-9.
8. Widbiller M, Schweikl H, Bruckmann A, Rosendahl A, Hochmuth E, Lindner SR, Buchalla W, Galler KM. Shotgun Proteomics of Human Dentin with Different Prefractionation Methods. Sci Rep. 2019 Mar 14;9(1):4457.
9. Kim JY, Xin X, Moioli EK, Chung J, Lee CH, Chen M, Fu SY, Koch PD, Mao JJ. Regeneration of dental-pulp-like tissue by chemotaxis-induced cell homing. Tissue Eng Part A. 2010 Oct;16(10):3023-31.
10. Srisuwan T, Wattanapakkavong K. Direct effect of transforming growth factor-beta 1 (TGF-β1) on human apical papilla cell proliferation and mineralisation. Aust Endod J. 2021 Oct 1. doi: 10.1111/aej.12572. Epub ahead of print.
11. Zhao S, Sloan AJ,Murray PE, Lumley PJ, Smith AJ. Ultrastructural localisation of TGF-beta exposure in dentine by chemical treatment. Histochem J. 2000 Aug;32(8):489-94.
12. Zhang M, Jiang F, Zhang X, Wang S, Jin Y, Zhang W, Jiang X. The Effects of Platelet-Derived Growth Factor-BB on Human Dental Pulp Stem Cells Mediated Dentin-Pulp Complex Regeneration. Stem Cells Transl Med. 2017 Dec;6(12):2126-2134.
13. Matsushita K, Motani R, Sakuta T, Yamaguchi N, Koga T, Matsuo K, Nagaoka S, Abeyama K, Maruyama I, Torii M. The role of vascular endothelial growth factor in human dental pulp cells: induction of chemotaxis, proliferation, and differentiation and activation of the AP-1-dependent signaling pathway. J Dent Res. 2000 Aug;79(8):1596-603.
14. Yuan GH, Yang GB, Wu LA, Chen Z, Chen S. Potential Role of Dentin Sialoprotein by Inducing Dental Pulp Mesenchymal Stem Cell Differentiation and Mineralization for Dental Tissue Repair. Dent Hypotheses. 2010 Jan 1;1(2):69-75.
15. Galler KM, Buchalla W, Hiller KA, Federlin M, Eidt A, Schiefersteiner M, Schmalz G. Influence of root canal disinfectants on growth factor release from dentin. J Endod. 2015 Mar;41(3):363-8.
16. Aksel H, Albanyan H, Bosaid F, Azim AA. Dentin Conditioning Protocol for Regenerative Endodontic Procedures. J Endod. 2020 Aug;46(8):1099-1104.
17. Fong D, Duceppe N, Hoemann CD. Mesenchymal stem cell detachment with trace trypsin is superior to EDTA for in vitro chemotaxis and adhesion assays. Biochem Biophys Res Commun. 2017 Mar 11;484(3):656-661.
18. Widbiller M, Schmalz G. Endodontic regeneration: hard shell, soft core. Odontology. 2021 Apr;109(2):303-312.
19. Del Carpio-Perochena A, Bramante CM, Duarte MA, de Moura MR, Aouada FA, Kishen A. Chelating and antibacterial properties of chitosan nanoparticles on dentin. Restor Dent Endod. 2015 Aug;40(3):195-201.
20. Silva PV, Guedes DF, Nakadi FV, Pécora JD, Cruz-Filho AM. Chitosan: a new solution for removal of smear layer after root canal instrumentation. Int Endod J. 2013 Apr;46(4):332-8.
21. Shrestha A, Kishen A. Antibacterial Nanoparticles in Endodontics: A Review. J Endod. 2016 Oct;42(10):1417-26.
22. (ISO) ISO/TS 11405:2015(en) Dentistry — Testing of adhesion to tooth structure. ISO/TC 106/SC 12015.
23. de Moura MR, Aouada FA, Avena-Bustillos RJ, McHugh TH, Krochta JM, Mattoso LHC. Improved barrier and mechanical properties of novel hydroxypropyl methylcellulose edible films with chitosan/tripolyphosphate nanoparticles. J Food Eng 2009; (92) 448–53
24. Kassaee M, Hosseini S, Elahi SH. A New Nano-Chitosan Irrigant with Superior Smear Layer Removal and Penetration. Nanochem Res 2016; 1: 150-56. doi 10.7508/NCR.2016.02.002.
25. Martinho FC, Leite FRM, Arruda-Vasconcelos R, Louzada LM, Darveau RP, Gomes BPFA. Influence of Bacterial Profiles in Cytokine and Clinical Features of Endodontic Disease. J Endod. 2021 Aug;47(8):1265-1271
26. Morgan E, Varro R, Sepulveda H, Ember JA, Apgar J, Wilson J, Lowe L, Chen R, Shivraj L, Agadir A, Campos R, Ernst D, Gaur A. Cytometric bead array: a multiplexed assay platform with applications in various areas of biology. Clin Immunol. 2004 Mar;110(3):252-66.
27. Zeng Q, Nguyen S, Zhang H, Chebrolu HP, Alzebdeh D, Badi MA, Kim JR, Ling J, Yang M. Release of Growth Factors into Root Canal by Irrigations in Regenerative Endodontics. J Endod. 2016 Dec;42(12):1760-1766.
28. Evanko SP, Raines EW, Ross R, Gold LI, Wight TN. Proteoglycan distribution in lesions of atherosclerosis depends on lesion severity, structural characteristics, and the proximity of platelet-derived growth factor and transforming growth factor-beta. Am J Pathol. 1998 Feb;152(2):533-46.
29. Goldberg M, Kulkarni AB, Young M, Boskey A. Dentin: structure, composition and mineralization. Front Biosci (Elite Ed). 2011 Jan 1;3:711-35.
30. Diao S, Lin X, Wang L, Dong R, Du J, Yang D, Fan Z. Analysis of gene expression profiles between apical papilla tissues, stem cells from apical papilla and cell sheet to identify the key modulators in MSCs niche. Cell Prolif. 2017 Jun;50(3):e12337.
31. Sadaghiani L, Gleeson HB, Youde S, Waddington RJ, Lynch CD, Sloan AJ. Growth Factor Liberation and DPSC Response Following Dentine Conditioning. J Dent Res. 2016 Oct;95(11):1298-307.
32. Ivica A, Zehnder M, Mateos JM, Ghayor C, Weber FE. Biomimetic Conditioning of Human Dentin Using Citric Acid. J Endod. 2019 Jan;45(1):45-50.
33. Ferreira LN, Puppin-Rontani RM, Pascon FM. Effect of Intracanal Medicaments and Irrigants on the Release of Transforming Growth Factor Beta 1 and Vascular Endothelial Growth Factor from Cervical Root Dentin. J Endod. 2020 Nov;46(11):1616-1622
34. Tavares S, Pintor A, Mourão CFAB, Magno M, Montemezzi P, Sacco R, Alves G, Scelza MZ. Effect of Different Root Canal Irrigant Solutions on the Release of Dentin-Growth Factors: A Systematic Review and Meta-Analysis. Materials (Basel). 2021 Oct 5;14(19):5829
35. Baker SM, Sugars RV, Wendel M, Smith AJ, Waddington RJ, Cooper PR, Sloan AJ. TGF-beta/extracellular matrix interactions in dentin matrix: a role in regulating sequestration and protection of bioactivity. Calcif Tissue Int. 2009 Jul;85(1):66-74.
36. He H, Yu J, Liu Y, et al. Effects of FGF2 and TGFbeta1 on the differentiation of human dental pulp stem cells in vitro. Cell Biol Int 2008;32:827
37. Khan JA, Hasan A, Dossa S, Ali B. Effect of Natural and Artificial Dentin Conditioners on the Release of Vascular Endothelial Growth Factor. J Endod. 2021 May;47(5):800-805
38. Usuelli M, Meyer T, Mezzenga R, Mitsi M. VEGF and VEGFR2 bind to similar pH-sensitive sites on fibronectin, exposed by heparin-mediated conformational changes. J Biol Chem. 2021 Jan-Jun;296:100584.
39. Mochizuki M, Güç E, Park AJ, Julier Z, Briquez PS, Kuhn GA, Müller R, Swartz MA, Hubbell JA, Martino MM. Growth factors with enhanced syndecan binding generate tonic signalling and promote tissue healing. Nat Biomed Eng. 2020 Apr;4(4):463-475.
40. Yoshiba K, Yoshiba N, Nakamura H, Iwaku M, Ozawa H. Immunolocalization of fibronectin during reparative dentinogenesis in human teeth after pulp capping with calcium hydroxide. J Dent Res. 1996 Aug;75(8):1590-7. doi: 10.1177/00220345960750081101.
41. Suzuki S, Sreenath T, Haruyama N, Honeycutt C, Terse A, Cho A, Kohler T, Müller R, Goldberg M, Kulkarni AB. Dentin sialoprotein and dentin phosphoprotein have distinct roles in dentin mineralization. Matrix Biol. 2009 May;28(4):221-9.
42. Ritchie H. The functional significance of dentin sialoprotein-phosphophoryn and dentin sialoprotein. Int J Oral Sci. 2018 Nov 5;10(4):31.
43. Mohammadi Z, Shalavi S, Jafarzadeh H. Ethylenediaminetetraacetic acid in endodontics. Eur J Dent. 2013 Sep;7(Suppl 1):S135-S142.
44. Sayin TC, Serper A, Cehreli ZC, Kalayci S. Calcium loss from root canal dentin following EDTA, EGTA, EDTAC, and tetracycline-HCl treatment with or without subsequent NaOCl irrigation. J Endod. 2007 May;33(5):581-4.
45. Vold IMN, Vårum KM, Guibal E, Smidsrød O. Binding of ions to chitosan—selectivity studies. Carbohydr Polym. 2003;54(4):471–7
46. Webster A, Halling MD, Grant DM. Metal complexation of chitosan and its glutaraldehyde cross-linked derivative. Carbohydr Res. 2007 Jul 2;342(9):1189-201.
47. Rotstein I, Dankner E, Goldman A, Heling I, Stabholz A, Zalkind M. Histochemical analysis of dental hard tissues following bleaching. J Endod 1996;22:23– 6.
48. Aquino-Martínez R, Monroe DG, Ventura F. Calcium mimics the chemotactic effect of conditioned media and is an effective inducer of bone regeneration. PLoS One. 2019 Jan 4;14(1):e0210301.
49. Kishen A, Shi Z, Shrestha A, Neoh KG. An investigation on the antibacterial and antibiofilm efficacy of cationic nanoparticulates for root canal disinfection. J Endod. 2008;34(12):1515-20.
dc.rights.spa.fl_str_mv Derechos reservados al autor, 2016
dc.rights.coar.fl_str_mv http://purl.org/coar/access_right/c_abf2
dc.rights.license.spa.fl_str_mv Atribución-NoComercial-SinDerivadas 4.0 Internacional
dc.rights.uri.spa.fl_str_mv http://creativecommons.org/licenses/by-nc-nd/4.0/
dc.rights.accessrights.spa.fl_str_mv info:eu-repo/semantics/openAccess
rights_invalid_str_mv Atribución-NoComercial-SinDerivadas 4.0 Internacional
Derechos reservados al autor, 2016
http://creativecommons.org/licenses/by-nc-nd/4.0/
http://purl.org/coar/access_right/c_abf2
eu_rights_str_mv openAccess
dc.format.extent.spa.fl_str_mv 23 páginas
dc.format.mimetype.spa.fl_str_mv application/pdf
dc.publisher.spa.fl_str_mv Universidad Nacional de Colombia
dc.publisher.program.spa.fl_str_mv Bogotá - Odontología - Maestría en Odontología
dc.publisher.department.spa.fl_str_mv Departamento de Ciencias Básicas y Medicina Oral
dc.publisher.faculty.spa.fl_str_mv Facultad de Odontología
dc.publisher.place.spa.fl_str_mv Bogotá, Colombia
dc.publisher.branch.spa.fl_str_mv Universidad Nacional de Colombia - Sede Bogotá
institution Universidad Nacional de Colombia
bitstream.url.fl_str_mv https://repositorio.unal.edu.co/bitstream/unal/81276/7/1032389456.2022.pdf
https://repositorio.unal.edu.co/bitstream/unal/81276/6/license.txt
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repository.name.fl_str_mv Repositorio Institucional Universidad Nacional de Colombia
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spelling Atribución-NoComercial-SinDerivadas 4.0 InternacionalDerechos reservados al autor, 2016http://creativecommons.org/licenses/by-nc-nd/4.0/info:eu-repo/semantics/openAccesshttp://purl.org/coar/access_right/c_abf2Castellanos Parra, Jaime Eduardod005422b5147ff1af3f046ffaf4328e6García Guerrero, Claudia Carmiña73644d50f4fcb71b6604b6cd2f0a89c3Quijano Guauque, Sara Beatrizf970ba20d9998c1f63f628df2f517ee2Invendo2022-03-17T19:12:17Z2022-03-17T19:12:17Z2016https://repositorio.unal.edu.co/handle/unal/81276Universidad Nacional de ColombiaRepositorio Institucional Universidad Nacional de Colombiahttps://repositorio.unal.edu.co/ilustraciones, gráficas, tablasLa regeneración pulpo-dentinal, requiere desinfección y acondicionamiento intraconducto para cumplir los objetivos terapéuticos. Los quelantes modifican la superficie y liberan moléculas bioactivas que estimulan las celúlas madre de la papila apical (SCAPs). El ácido etilendiaminotetraacético (EDTA), quelante capaz de liberar proteínas inmersas en la matriz, ejerce un efecto residual sobre las SCAPs, y altera negativamente la superficie al utilizarse con hipoclorito de sodio. Quitosán (QS) en solución o nano-particulado (QSnp), polímero natural, antimicrobiano y quelante, induce estabilidad biológica, ampliando las alternativas para el acondicionamiento dentinal. Por tal motivo el objetivo de este trabajo fue determinar la capacidad de dos preparaciones de quitosán para liberar proteínas de la matriz dentinal radicular y describir los cambios en la composición química, posterior al efecto quelante. Materiales y Métodos: Estudio experimental cuantitativo in-vitro para cuantificar sialoproteína dentinal (DSP), factor de Crecimiento Transformante Beta (TGFß1), factor de crecimiento del Endotelio Vascular (VEFG) y el factor de crecimiento derivado de plaquetas Isoforma BB (PDGF-BB) liberadas con agentes quelantes. 24 hemiraices (n=6) fueron distribuidas en 4 grupos durante 5 minutos en EDTA, QS y QSnp, Posterior incubación a 37°C durante 7 días, se recogieron sobrenadantes y se congelaron hasta su procesamiento. Un ensayo ELISA cuantificó DSP. Para TGFß1, VEFG y PDGF-BB, un panel de citoquinas fue realizado. Kruskal-Wallis con Wilcoxon identificó diferencias entre los grupos. Espectroscopia Raman permitió el análisis no destructivo de los cambios químicos superficiales. Resultados: La liberación de TGF-β1, VEGF, y DSP dentinal, fue identificable en todos los grupos. La liberación de TGF- β1, ocurrió con todos los quelantes y mayor la mayor liberación de DSP ocurrió con QSnp. PDGF-BB no superó los límites de detección. Conclusión: La utilización de agentes quelantes permitió la solubilización de TGF-β1, VEGF, y DSP inmersos en la matriz dentinal radicular siendo QSnp quien promovió una liberación significativa de DSP. La espectroscopia Raman identificó la capacidad de quelación para QS y QSnp y su interacción con otros componentes de la matriz. (Texto tomado de la fuente)Introduction: Dentin- Pulp regeneration requires disinfection and intracanal conditioning to achieve therapeutic objectives. Chelators modify the surface and release bioactive molecules that stimulate stem cells of the apical papilla (SCAPs). Ethylenediaminetetraacetic acid (EDTA), a chelating agent capable of releasing proteins embedded in the matrix, exerts a residual effect on SCAPs, and negatively alters the surface when is used with sodium hypochlorite. Chitosan solution (QS) or nanoparticulate (QSnp), is a natural, antimicrobial, and chelating polymer, that induces biological stability, broadening the alternatives for dentin conditioning. For this reason, the objective of this work was to determine the capacity of two chitosan preparations to release proteins from the root dentin matrix and to describe the changes in the chemical composition, after the chelating effect. Materials and Methods: Quantitative in-vitro experimental study to quantify dentinal sialoprotein (DSP), Transforming Growth Factor Beta (TGFß1), Vascular Endothelial Growth Factor (VEFG) and plateletderived growth factor Isoform BB (PDGF-BB) released with chelating agents. 24 Hemi-roots (n=6) were distributed in 4 groups and immersed for 5 minutes in the chelating solution. Subsequent incubation at 37°C for 7 days, supernatants were collected and frozen until processing. An ELISA assay quantified DSP. For TGFß1, VEFG, and PDGF-BB, a cytokine bead array was performed. Kruskal-Wallis with Wilcoxon identified differences between groups. Raman spectroscopy allowed a non-destructive analysis of the chemical changes in the dentinal surface. Results: The release of TGF-β1, VEGF, and dentinal DSP was identifiable in all groups. The release of TGF-β1 occurred with all the chelators and the greatest release of DSP occurred with QSnp. PDGF-BB did not exceed detection limits. Conclusion: The use of chelating agents allowed the solubilization of TGF-β1, VEGF, and DSP immersed in the radicular dentinal matrix. QSnp promotes a significant release of DSP. Raman spectroscopy identified the chelating ability for QS and QSnp and their interaction with other matrix components.Convocatoria para el fortalecimiento de alianzas interdisciplinarias de investigación y creación artística de la Sede Bogotá de la Universidad Nacional De Colombia 2018- Código 41999MaestríaMagíster en OdontologíaEstudio experimental cuantitativo in-vitroMicroambiente seguro para endodoncia regenerativa23 páginasapplication/pdfspaUniversidad Nacional de ColombiaBogotá - Odontología - Maestría en OdontologíaDepartamento de Ciencias Básicas y Medicina OralFacultad de OdontologíaBogotá, ColombiaUniversidad Nacional de Colombia - Sede Bogotá610 - Medicina y salud::613 - Salud y seguridad personalDentinaEndodoncia RegenerativaQuitosanoDentinRegenerative EndodonticsChitosanRegeneraciónEndodonciaQuitosanoPéptidos y Proteínas de Señalización IntercelularDentina.RegenerationEndodonticsChitosanIntercellular Signaling Peptides and ProteinsDentinEfecto de dos soluciones irrigadoras de quitosán sobre la liberación de proteínas bioactivas de la dentina radicularEffect of two irrigating solutions of chitosan on the release of bioactive proteins from root dentinTrabajo de grado - Maestríainfo:eu-repo/semantics/masterThesisinfo:eu-repo/semantics/acceptedVersionOtherhttp://purl.org/redcol/resource_type/TM1. Xie Z, Shen Z, Zhan P, Yang J, Huang Q, Huang S, Chen L, Lin Z. Functional Dental Pulp Regeneration: Basic Research and Clinical Translation. Int J Mol Sci. 2021 Aug 20;22(16):89912. Smith JG, Smith AJ, Shelton RM, Cooper PR. Dental Pulp Cell Behavior in Biomimetic Environments. J Dent Res. 2015 Nov;94(11):1552-9.3. Huang X, Li Z, Liu A, Liu X, Guo H, Wu M, Yang X, Han B, Xuan K. Microenvironment Influences Odontogenic Mesenchymal Stem Cells Mediated Dental Pulp Regeneration. Front Physiol. 2021 Apr 22;12:656588.4. Jeeruphan T, Jantarat J, Yanpiset K, Suwannapan L, Khewsawai P, Hargreaves KM. Mahidol study 1: comparison of radiographic and survival outcomes of immature teeth treated with either regenerative endodontic or apexification methods: a retrospective study. J Endod. 2012 Oct;38(10):1330-6.5. Jung C, Kim S, Sun T, Cho YB, Song M. Pulp-dentin regeneration: current approaches and challenges. J Tissue Eng. 2019; 10:2041731418819263.6. Zhang X, Li H, Sun J, Luo X, Yang H, Xie L, Yang B, Guo W, Tian W. Cell-derived micro-environment helps dental pulp stem cells promote dental pulp regeneration. Cell Prolif. 2017 Oct;50(5):e12361.7. Dung SZ, Gregory RL, Li Y, Stookey GK. Effect of lactic acid and proteolytic enzymes on the release of organic matrix components from human root dentin. Caries Res. 1995;29(6):483-9.8. Widbiller M, Schweikl H, Bruckmann A, Rosendahl A, Hochmuth E, Lindner SR, Buchalla W, Galler KM. Shotgun Proteomics of Human Dentin with Different Prefractionation Methods. Sci Rep. 2019 Mar 14;9(1):4457.9. Kim JY, Xin X, Moioli EK, Chung J, Lee CH, Chen M, Fu SY, Koch PD, Mao JJ. Regeneration of dental-pulp-like tissue by chemotaxis-induced cell homing. Tissue Eng Part A. 2010 Oct;16(10):3023-31.10. Srisuwan T, Wattanapakkavong K. Direct effect of transforming growth factor-beta 1 (TGF-β1) on human apical papilla cell proliferation and mineralisation. Aust Endod J. 2021 Oct 1. doi: 10.1111/aej.12572. Epub ahead of print.11. Zhao S, Sloan AJ,Murray PE, Lumley PJ, Smith AJ. Ultrastructural localisation of TGF-beta exposure in dentine by chemical treatment. Histochem J. 2000 Aug;32(8):489-94.12. Zhang M, Jiang F, Zhang X, Wang S, Jin Y, Zhang W, Jiang X. The Effects of Platelet-Derived Growth Factor-BB on Human Dental Pulp Stem Cells Mediated Dentin-Pulp Complex Regeneration. Stem Cells Transl Med. 2017 Dec;6(12):2126-2134.13. Matsushita K, Motani R, Sakuta T, Yamaguchi N, Koga T, Matsuo K, Nagaoka S, Abeyama K, Maruyama I, Torii M. The role of vascular endothelial growth factor in human dental pulp cells: induction of chemotaxis, proliferation, and differentiation and activation of the AP-1-dependent signaling pathway. J Dent Res. 2000 Aug;79(8):1596-603.14. Yuan GH, Yang GB, Wu LA, Chen Z, Chen S. Potential Role of Dentin Sialoprotein by Inducing Dental Pulp Mesenchymal Stem Cell Differentiation and Mineralization for Dental Tissue Repair. Dent Hypotheses. 2010 Jan 1;1(2):69-75.15. Galler KM, Buchalla W, Hiller KA, Federlin M, Eidt A, Schiefersteiner M, Schmalz G. Influence of root canal disinfectants on growth factor release from dentin. J Endod. 2015 Mar;41(3):363-8.16. Aksel H, Albanyan H, Bosaid F, Azim AA. Dentin Conditioning Protocol for Regenerative Endodontic Procedures. J Endod. 2020 Aug;46(8):1099-1104.17. Fong D, Duceppe N, Hoemann CD. Mesenchymal stem cell detachment with trace trypsin is superior to EDTA for in vitro chemotaxis and adhesion assays. Biochem Biophys Res Commun. 2017 Mar 11;484(3):656-661.18. Widbiller M, Schmalz G. Endodontic regeneration: hard shell, soft core. Odontology. 2021 Apr;109(2):303-312.19. Del Carpio-Perochena A, Bramante CM, Duarte MA, de Moura MR, Aouada FA, Kishen A. Chelating and antibacterial properties of chitosan nanoparticles on dentin. Restor Dent Endod. 2015 Aug;40(3):195-201.20. Silva PV, Guedes DF, Nakadi FV, Pécora JD, Cruz-Filho AM. Chitosan: a new solution for removal of smear layer after root canal instrumentation. Int Endod J. 2013 Apr;46(4):332-8.21. Shrestha A, Kishen A. Antibacterial Nanoparticles in Endodontics: A Review. J Endod. 2016 Oct;42(10):1417-26.22. (ISO) ISO/TS 11405:2015(en) Dentistry — Testing of adhesion to tooth structure. ISO/TC 106/SC 12015.23. de Moura MR, Aouada FA, Avena-Bustillos RJ, McHugh TH, Krochta JM, Mattoso LHC. Improved barrier and mechanical properties of novel hydroxypropyl methylcellulose edible films with chitosan/tripolyphosphate nanoparticles. J Food Eng 2009; (92) 448–5324. Kassaee M, Hosseini S, Elahi SH. A New Nano-Chitosan Irrigant with Superior Smear Layer Removal and Penetration. Nanochem Res 2016; 1: 150-56. doi 10.7508/NCR.2016.02.002.25. Martinho FC, Leite FRM, Arruda-Vasconcelos R, Louzada LM, Darveau RP, Gomes BPFA. Influence of Bacterial Profiles in Cytokine and Clinical Features of Endodontic Disease. J Endod. 2021 Aug;47(8):1265-127126. Morgan E, Varro R, Sepulveda H, Ember JA, Apgar J, Wilson J, Lowe L, Chen R, Shivraj L, Agadir A, Campos R, Ernst D, Gaur A. Cytometric bead array: a multiplexed assay platform with applications in various areas of biology. Clin Immunol. 2004 Mar;110(3):252-66.27. Zeng Q, Nguyen S, Zhang H, Chebrolu HP, Alzebdeh D, Badi MA, Kim JR, Ling J, Yang M. Release of Growth Factors into Root Canal by Irrigations in Regenerative Endodontics. J Endod. 2016 Dec;42(12):1760-1766.28. Evanko SP, Raines EW, Ross R, Gold LI, Wight TN. Proteoglycan distribution in lesions of atherosclerosis depends on lesion severity, structural characteristics, and the proximity of platelet-derived growth factor and transforming growth factor-beta. Am J Pathol. 1998 Feb;152(2):533-46.29. Goldberg M, Kulkarni AB, Young M, Boskey A. Dentin: structure, composition and mineralization. Front Biosci (Elite Ed). 2011 Jan 1;3:711-35.30. Diao S, Lin X, Wang L, Dong R, Du J, Yang D, Fan Z. Analysis of gene expression profiles between apical papilla tissues, stem cells from apical papilla and cell sheet to identify the key modulators in MSCs niche. Cell Prolif. 2017 Jun;50(3):e12337.31. Sadaghiani L, Gleeson HB, Youde S, Waddington RJ, Lynch CD, Sloan AJ. Growth Factor Liberation and DPSC Response Following Dentine Conditioning. J Dent Res. 2016 Oct;95(11):1298-307.32. Ivica A, Zehnder M, Mateos JM, Ghayor C, Weber FE. Biomimetic Conditioning of Human Dentin Using Citric Acid. J Endod. 2019 Jan;45(1):45-50.33. Ferreira LN, Puppin-Rontani RM, Pascon FM. Effect of Intracanal Medicaments and Irrigants on the Release of Transforming Growth Factor Beta 1 and Vascular Endothelial Growth Factor from Cervical Root Dentin. J Endod. 2020 Nov;46(11):1616-162234. Tavares S, Pintor A, Mourão CFAB, Magno M, Montemezzi P, Sacco R, Alves G, Scelza MZ. Effect of Different Root Canal Irrigant Solutions on the Release of Dentin-Growth Factors: A Systematic Review and Meta-Analysis. Materials (Basel). 2021 Oct 5;14(19):582935. Baker SM, Sugars RV, Wendel M, Smith AJ, Waddington RJ, Cooper PR, Sloan AJ. TGF-beta/extracellular matrix interactions in dentin matrix: a role in regulating sequestration and protection of bioactivity. Calcif Tissue Int. 2009 Jul;85(1):66-74.36. He H, Yu J, Liu Y, et al. Effects of FGF2 and TGFbeta1 on the differentiation of human dental pulp stem cells in vitro. Cell Biol Int 2008;32:82737. Khan JA, Hasan A, Dossa S, Ali B. Effect of Natural and Artificial Dentin Conditioners on the Release of Vascular Endothelial Growth Factor. J Endod. 2021 May;47(5):800-80538. Usuelli M, Meyer T, Mezzenga R, Mitsi M. VEGF and VEGFR2 bind to similar pH-sensitive sites on fibronectin, exposed by heparin-mediated conformational changes. J Biol Chem. 2021 Jan-Jun;296:100584.39. Mochizuki M, Güç E, Park AJ, Julier Z, Briquez PS, Kuhn GA, Müller R, Swartz MA, Hubbell JA, Martino MM. Growth factors with enhanced syndecan binding generate tonic signalling and promote tissue healing. Nat Biomed Eng. 2020 Apr;4(4):463-475.40. Yoshiba K, Yoshiba N, Nakamura H, Iwaku M, Ozawa H. Immunolocalization of fibronectin during reparative dentinogenesis in human teeth after pulp capping with calcium hydroxide. J Dent Res. 1996 Aug;75(8):1590-7. doi: 10.1177/00220345960750081101.41. Suzuki S, Sreenath T, Haruyama N, Honeycutt C, Terse A, Cho A, Kohler T, Müller R, Goldberg M, Kulkarni AB. Dentin sialoprotein and dentin phosphoprotein have distinct roles in dentin mineralization. Matrix Biol. 2009 May;28(4):221-9.42. Ritchie H. The functional significance of dentin sialoprotein-phosphophoryn and dentin sialoprotein. Int J Oral Sci. 2018 Nov 5;10(4):31.43. Mohammadi Z, Shalavi S, Jafarzadeh H. Ethylenediaminetetraacetic acid in endodontics. Eur J Dent. 2013 Sep;7(Suppl 1):S135-S142.44. Sayin TC, Serper A, Cehreli ZC, Kalayci S. Calcium loss from root canal dentin following EDTA, EGTA, EDTAC, and tetracycline-HCl treatment with or without subsequent NaOCl irrigation. J Endod. 2007 May;33(5):581-4.45. Vold IMN, Vårum KM, Guibal E, Smidsrød O. Binding of ions to chitosan—selectivity studies. Carbohydr Polym. 2003;54(4):471–746. Webster A, Halling MD, Grant DM. Metal complexation of chitosan and its glutaraldehyde cross-linked derivative. Carbohydr Res. 2007 Jul 2;342(9):1189-201.47. Rotstein I, Dankner E, Goldman A, Heling I, Stabholz A, Zalkind M. Histochemical analysis of dental hard tissues following bleaching. J Endod 1996;22:23– 6.48. Aquino-Martínez R, Monroe DG, Ventura F. Calcium mimics the chemotactic effect of conditioned media and is an effective inducer of bone regeneration. PLoS One. 2019 Jan 4;14(1):e0210301.49. Kishen A, Shi Z, Shrestha A, Neoh KG. An investigation on the antibacterial and antibiofilm efficacy of cationic nanoparticulates for root canal disinfection. J Endod. 2008;34(12):1515-20.Sistema Nanoestructurado para Irrigación en Endodoncia - Convocatoria para el fortalecimiento de alianzas interdisciplinarias de investigación y creación artística de la Sede Bogotá de la Universidad Nacional De Colombia 2018Universidad Nacional de ColombiaEstudiantesInvestigadoresORIGINAL1032389456.2022.pdf1032389456.2022.pdfTrabajo de Grado de Maestría en Odontologíaapplication/pdf515250https://repositorio.unal.edu.co/bitstream/unal/81276/7/1032389456.2022.pdfcd9cb16edfac6de20e62c228b9816b21MD57LICENSElicense.txtlicense.txttext/plain; charset=utf-84074https://repositorio.unal.edu.co/bitstream/unal/81276/6/license.txt8153f7789df02f0a4c9e079953658ab2MD56THUMBNAIL1032389456.2022.pdf.jpg1032389456.2022.pdf.jpgGenerated Thumbnailimage/jpeg5091https://repositorio.unal.edu.co/bitstream/unal/81276/8/1032389456.2022.pdf.jpg2c20734cb42d7b9756200a9d3d212c96MD58unal/81276oai:repositorio.unal.edu.co:unal/812762023-08-02 23:04:18.157Repositorio Institucional 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EVESURBIFBPUiBMQSBTRUNSRVRBUsONQSBHRU5FUkFMLiAqTEEgVEVTSVMgQSBQVUJMSUNBUiBERUJFIFNFUiBMQSBWRVJTScOTTiBGSU5BTCBBUFJPQkFEQS4gCgpBbCBoYWNlciBjbGljIGVuIGVsIHNpZ3VpZW50ZSBib3TDs24sIHVzdGVkIGluZGljYSBxdWUgZXN0w6EgZGUgYWN1ZXJkbyBjb24gZXN0b3MgdMOpcm1pbm9zLiBTaSB0aWVuZSBhbGd1bmEgZHVkYSBzb2JyZSBsYSBsaWNlbmNpYSwgcG9yIGZhdm9yLCBjb250YWN0ZSBjb24gZWwgYWRtaW5pc3RyYWRvciBkZWwgc2lzdGVtYS4KClVOSVZFUlNJREFEIE5BQ0lPTkFMIERFIENPTE9NQklBIC0gw5psdGltYSBtb2RpZmljYWNpw7NuIDE5LzEwLzIwMjEK

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