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Pulp cell cultures obtained with two different methods for in vitro cytotoxicity tests

  • O. CortésEmail author
  • C. Garcìa
  • L. Pérez
  • J. Boj
  • A. Alcaina
Article

Abstract

Aim: To describe two different protocols for obtaining primary pulp cell cultures, one derived from explants and the other following dissociation into single cell suspension by enzyme digestion. Methods: Human pulp tissue was obtained from three healthy premolars. The harvested pulp tissue was prepared for culture using physical methods (one of the premolars) and enzyme, type XI collagenase, (the two remaining premolars). Results: In the case of explant based culture, only limited growth was observed in some cases. However, by enzyme digestion, after two weeks cell growth was evident, and differences in cell type were observed according to the tooth involved. Conclusion: It has been possible to obtain abundant biological material using an enzyme digestion-based protocol for testing purposes, with low experimental variability, as all cells originated from the same individual.

Key words

Cell culture pulp fibroblasts explants enzyme digestion cell viability 

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References

  1. Ashkenazi M, Marouni M, Sarnat H.. In vitro viability, mitogenicity and clonogenic capacities of periodontal ligament fibroblasts after storage in four media supplemented with growth factors. Dent Traumatol 2001;17: 27–35.PubMedCrossRefGoogle Scholar
  2. Bencosme SA, Tsutsumi V..A fast method for processing biological bacterial for electron microscopy. Lab Invest 1970; 23: 447–51.PubMedGoogle Scholar
  3. Davis WL.Oral histology. Cell structure and function. WB Saunders Company, Philadelphia, 1986.Google Scholar
  4. Casas MJ, Kenny DJ, Johnston DH, Judd PL. Long-term outcomes of primary molar ferric sulfate pulpotomy and root canal therapy. Pediatr Dent 2004;26:44–8)PubMedGoogle Scholar
  5. Cortés O, Boj JR, Canalda C, Carreras M. Pulpal tissue reaction to formocresol versus ferric sulfate in pulpotomized rat teeth. J Clin Pediatr Dent 1997;21:247–54.Google Scholar
  6. Eisenmenger E, Zetner K..Veterinary dentistry. Society of Veterinary Dentistry.Lea and Febiger, Philadelphia, 1985.Google Scholar
  7. Feigal RJ, Yesilsoy C, Messer HH, Nelson J..Differential sensitivity of normal human pulp and transformed mouse fibroblasts to cytotoxic challenge. Arch Oral Biol 1985; 30: 609–613.PubMedCrossRefGoogle Scholar
  8. Freshney RI. Animal cell culture. A practical approach. 2nd edition. Oxford University Press, Oxford, 1992.Google Scholar
  9. Hashieh IA, Cosset A, Franquin JC, Camps J. In vitro cytotoxicity of one step dentin bonding systems. J Endodon 1999; 25: 89–92.CrossRefGoogle Scholar
  10. Hill SD, Berry CW, Seale NN, Kaga M.. Comparison of antimicrobial and citotoxic effects of gluataradhyde and formocresol. Oral Surg Oral Med Oral Pathol 1991; 71: 89–95.PubMedCrossRefGoogle Scholar
  11. Holan G, Eidelman E, Fuks AB. Long-term evaluation of pulpotomy in primary molars using mineral trioxide aggregate or formocresol. Pediatr Dent 2005;27:129–36.PubMedGoogle Scholar
  12. International Standards Organization.1992.10993–5. Biological evaluation of medical devices. Part 5. Tests for citotoxycity: in vitro methods.Google Scholar
  13. Jakoby WB, Pastan IH. Cell culture. Methods in enzymology. Academic Press, INC, California, 1979.Google Scholar
  14. Keiser K, Johnson CC, Tipton DA. Cytotoxicity of mineral trioxide aggregate using human periodontal ligament fibroblasts. J Endodon 2000; 26: 288–291.CrossRefGoogle Scholar
  15. Kuo MY, Lan WH, Lin SK, Tsai KS, Hahn LJ.Collagen gene expression in human dental pulp cell cultures. Arch Oral Biol 1992; 37: 945–952.PubMedCrossRefGoogle Scholar
  16. Langeland K, Guttuso J, Langeland L, Tobon G. Methods in the study of biologic responses to endodontic materials. Oral Surg Oral Med Oral Path 1969; 27: 522–542.CrossRefGoogle Scholar
  17. Lovschall H, Eiskjaer M, Arenhol-Bindslev D. Formaldehyde cytotoxicity in three human cell types assessed in three different assays. Toxicol in vitro 2002; 16: 63–69.PubMedCrossRefGoogle Scholar
  18. Mitchell PJ, Pitt TR, Torabinejad M, McDonald F.. Osteoblast biocompatibility of mineral trioxide aggregate. Biomaterials 1999:20;167–173.PubMedCrossRefGoogle Scholar
  19. Morgan SJ, Darling DC.. Animal cell culture. Bios Scientific Publishers Limited, Oxford, 1993.Google Scholar
  20. Nakashima M.. Establishment of primary cultures of pulp cells from bovine permanent incisors. Arch Oral Biol 1991; 36: 655–663.PubMedCrossRefGoogle Scholar
  21. Nakashima M. Bone morphogenetic proteins in dentin regeneration for potential use in endodontic therapy. Cytokine Growth Factor Rev 2005;16:369–76PubMedCrossRefGoogle Scholar
  22. Pascon E, Leonardo M, Safavi K, Langeland K. Tissue reaction to endodontic materials: methods, criteria, assessment, and observations. Oral Surg Oral Med Oral Pathol1991; 72: 222–237.PubMedCrossRefGoogle Scholar
  23. Salentijn M, Klyvert H. Dental and oral tissue. An introduction, 2nd edition. Lea and Febiger, Philadelphia, 1985.Google Scholar
  24. Stanford JW.. Recommmended standard practices for biological evaluation of dental materials. Int Dent J 1980; 30: 140–188.Google Scholar
  25. Tsukamoto Y, Fukutani S, Shin-Ike T, Kubota T, Sato S, Suzuki Y. Mineralized nodule formation by cultures of human dental derived fibroblasts. Arch Oral Biol 1992; 37:1045–1055.PubMedCrossRefGoogle Scholar
  26. van WyK CW, Olivier A, Maritz JS. Cultured pulp fibroblasts: are they suitable for in vitro cytotoxicity testing?. J Oral Pathol Med 2001; 30: 168–177.PubMedCrossRefGoogle Scholar
  27. Wen Sun H, Feigal R, Messer H. Cytotoxicity of glutaraldehyde and formaldehyde in relation to time of exposure and concentration. Pediatr Dent 1990; 12: 303–307.Google Scholar

Copyright information

© European Academy of Paediatric Dentistry 2006

Authors and Affiliations

  • O. Cortés
    • 1
    Email author
  • C. Garcìa
    • 1
  • L. Pérez
    • 1
  • J. Boj
    • 2
  • A. Alcaina
    • 1
  1. 1.School of Medicine and DentistryUniversity of MurciaMurciaSpain
  2. 2.Dental SchoolUniversity of BarcelonaBarcelonaSpain

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