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VOLUME 22 , ISSUE 8 ( August, 2021 ) > List of Articles

ORIGINAL RESEARCH

Enhancement of Odontoblastic Differentiation of Stem Cells from Exfoliated Deciduous Tooth Using N-acetylcysteine—An In Vitro Study

Madhura Pawar, Vivek Pawar, Sanjay G Thete, Smita D Dutta, Pramod P Sadan, Rahul Maria, Dinraj Kulkarni

Keywords : DMP-1, DSPP, N-acetylcysteine, Odontogenic differentiation, Stem cells

Citation Information : Pawar M, Pawar V, Thete SG, Dutta SD, Sadan PP, Maria R, Kulkarni D. Enhancement of Odontoblastic Differentiation of Stem Cells from Exfoliated Deciduous Tooth Using N-acetylcysteine—An In Vitro Study. J Contemp Dent Pract 2021; 22 (8):882-889.

DOI: 10.5005/jp-journals-10024-3140

License: CC BY-NC 4.0

Published Online: 09-11-2021

Copyright Statement:  Copyright © 2021; Jaypee Brothers Medical Publishers (P) Ltd.


Abstract

Aim and objective: The study was conducted to evaluate the effects of N-acetylcysteine (NAC) on the propagation and differentiation of stem cells from human exfoliated deciduous teeth(SHED). Materials and methods: SHEDs were isolated by explant culture method and characterized for stem cell properties using flow cytometry method. MTT assay and Cell Counting Kit-8 (CCK-8) assay were used to examine the viability and proliferation of the SHEDs. The effects of NAC-induced osteo/odontoblastic differentiation of SHEDs were determined by functional staining for mineralization, and the gene expression of osteo/odontoblastic transcription factors and proteins was evaluated by real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analyses. Protein levels of collagen type 1 (COL1), dentin sialophosphoprotein (DSPP), and dentin matrix acidic phosphoprotein 1(DMP-1) were calculated by the Western blot method to assess the osteo/odontogenic differentiation. Results: SHEDs presented mesenchymal stem cell (MSC)-like characteristics on flow cytometric analysis. The cell viability and metabolic activity of SHEDs were increased with an increase in the concentrations of NAC from 0.5 to 10 nM. However, the concentrations of NAC from 0.5 to 2.5 mM did not affect cell proliferation. NAC incorporated at a concentration of 2.5 mM showed higher mineralization and considerably increased gene expression levels of runt-related transcription factor 2 (RUNX2), COL1A1, DSPP, and DMP-1. It significantly increased the protein expression of odontoblast-related matrix proteins like COL1, DSPP, and DMP-1. Conclusion: NAC regulates the healthy propagation of dental stem cells in vitro. Its effects on the differentiation of dental pulp SHEDs remain unidentified. This study explores that NAC can encourage the mineralization of SHEDs and differentiate them into the odontoblastic lineage. Clinical significance: The results propose that NAC could have a significant pharmacological role in activating and enhancing odontogenic differentiation of dental stem cells and possibly a prospect in regenerative dentistry.


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  1. Zhang N, Chen B, Wang W, et al. Isolation, characterization, and multi-lineage differentiation of stem cells from human exfoliated deciduous teeth. Mol Med Rep 2016;14(1):95–102. DOI: 10.3892/mmr.2016.5214.
  2. Zhu Q, Gao J, Tian G, et al. Adrenomedullin promotes the odontogenic differentiation of dental pulp stem cells through CREB/BMP2 signaling pathway. Acta Biochemic Biophys Sin 2017;49(7):609–616. DOI: 10.1093/abbs/gmx053.
  3. Garzon I, Martin-Piedra MA, Carriel V, et al. Bioactive injectable aggregates with nanofibrous microspheres and human dental pulp stem cells: a translational strategy in dental endodontics. J Tissue Eng Regen Med 2018;12(1):204–216. DOI: 10.1002/term.2397.
  4. Abuarqoub D, Awidi A, Abuharfeil N. Comparison of osteo/odontogenic differentiation of human adult dental pulp stem cells and stem cells from apical papilla in the presence of platelet lysate. Arch Oral Biol 2015;60(10):1545–1553. DOI: 10.1016/j.archoralbio.2015.07.007.
  5. Tangtrongsup S, Kisiday JD. Differential effects of the antioxidants N-acetylcysteine and pyrrolidine dithiocarbamate on mesenchymal stem cell chondrogenesis. Cell Mol Bioeng 2019;12(2):153–163. DOI: 10.1007/s12195-019-00566-3.
  6. Martacic J, Filipovic MK, Borozan S, et al. N-acetyl-l-cysteine protects dental tissue stem cells against oxidative stress in vitro. Clin Oral Invest 2018;22(8):2897–2903. DOI: 10.1007/s00784-018-2377-2.
  7. Zhang D, Li Q, Rao L, et al. Effect of 5-Aza-2′-deoxycytidine on odontogenic differentiation of human dental pulp cells. J Endod 2015;41(5):640–645. DOI: 10.1016/j.joen.2014.12.006.
  8. Zeng L, Zhao N, Han D, et al. DLX3 mutation negatively regulates odontogenic differentiation of human dental pulp cells. Arch Oral Biol 2017;77:12–17. DOI: 10.1016/j.archoralbio.2017.01.011.
  9. Zhou M, Liu NX, Shi SR, et al. Effect of tetrahedral DNA nanostructures on proliferation and osteo/odontogenic differentiation of dental pulp stem cells via activation of the notch signaling pathway. Nanomedicine 2018;14(4):1227–1236. DOI: 10.1016/j.nano.2018.02.004.
  10. Zou T, Dissanayaka WL, Jiang S, et al. Semaphorin 4D enhances angiogenic potential and suppresses osteo-/odontogenic differentiation of human dental pulp stem cells. J Endod 2017;43(2):297–305. DOI: 10.1016/j.joen.2016.10.019.
  11. Heng BC, Ye X, Liu Y, et al. Effects of recombinant overexpression of BCL2 on the proliferation, apoptosis, and osteogenic/odontogenic differentiation potential of dental pulp stem cells. J Endod 2016;42(4):575–583. DOI: 10.1016/j.joen.2016.01.013.
  12. Heng BC, Wang S, Gong T, et al. Ephrin B2 signalling enhances osteogenic/odontogenic differentiation of human dental pulp stem cells. Arch Oral Biol 2018;87:62–71. DOI: 10.1016/j.archoralbio.2017.12.014.
  13. Liu M, Zhao L, Hu J, et al. Endothelial cells, and endothelin-1 promote the odontogenic differentiation of dental pulp stem cells. Mol Med Rep 2018;18(1):893–901. DOI: 10.3892/mmr.2018.9033.
  14. Sonoda S, Mei YF, Atsuta I, et al. Exogenous nitric oxide stimulates the odontogenic differentiation of rat dental pulp stem cells. Sci Rep 2018;8(1):3419. DOI: 10.1038/s41598-018-21183-6.
  15. Ching HS, Luddin N, Rahman IA, et al. Expression of odontogenic and osteogenic markers in DPSCs and SHED: a review. Curr Stem Cell Res Ther 2017;12(1):71–79. DOI: 10.2174/1574888x11666160815095733.
  16. Rahimi S, Salarinasab S, Ghasemi N, et al. In vitro induction of odontogenic activity of human dental pulp stem cells by white Portland cement enriched with zirconium oxide and zinc oxide components. J Dent Res Dent Clin Dent Prospects 2019;13(1):3–10. DOI: 10.15171/joddd.2019.001.
  17. Teti G, Salvatore V, Focaroli S, et al. In vitro osteogenic and odontogenic differentiation of human dental pulp stem cells seeded on carboxymethyl cellulose-hydroxyapatite hybrid hydrogel. Front Physiol 2015;6:1–10. DOI: 10.3389/fphys.2015.00297.
  18. Lian M, Zhang Y, Shen Q, et al. JAB1 accelerates odontogenic differentiation of dental pulp stem cells. J Mol Histol 2016;47(3): 317–324. DOI: 10.1007/s10735-016-9672-5.
  19. Ngo VA, Jung JY, Koh JT, et al. Leptin induces odontogenic differentiation and angiogenesis in human dental pulp cells via activation of the mitogen-activated protein kinase signalling pathway. J Endod 2018;44(4):585–591. DOI: 10.1016/j.joen.2017.11.018.
  20. Qin W, Gao X, Ma T, et al. Metformin enhances the differentiation of dental pulp cells into odontoblasts by activating AMPK signalling. J Endod 2018;44(4):576–584. DOI: 10.1016/j.joen.2017.11.017.
  21. Hu X, Zhong Y, Kong Y, et al. Lineage-specific exosomes promote the odontogenic differentiation of human dental pulp stem cells (DPSCs) through TGFβ1/smads signalling pathway via transfer of microRNAs. Stem Cell Res Ther 2019;10(1):170. DOI: 10.1186/s13287-019-1278-x.
  22. Debeljak M J, Borozan S, Radovanovic A, et al. N-Acetyl-l-cysteine enhances ex-vivo amplification of deciduous teeth dental pulp stem cells. Arch Oral Biol 2016;70:32–38. DOI: 10.1016/j.archoralbio.2016.06.002.
  23. Tu M-G, Ho C-C, Hsu T-T, et al. Mineral trioxide aggregate with mussel-inspired surface nanolayers for stimulating odontogenic differentiation of dental pulp cells. J Endod 2018;44(6):963–970. DOI: 10.1016/j.joen.2018.02.018.
  24. Patil VR, Kharat AH, Kulkarni DG, et al. Long-term explant culture for harvesting homogeneous population of human dental pulp stem cells. Cell Biol Int 2018;42(12):1602–1610. DOI: 10.1002/cbin.11065.
  25. Onay EO, Yurtcu E, Terzi Y, et al. Odontogenic effects of two calcium silicate-based biomaterials in human dental pulp cells. Adv Clin Exp Med 2018;27(11):1541–1547. DOI: 10.17219/acem/74197.
  26. Wang S, Xia Y, Ma T, et al. Novel metformin-containing resin promotes odontogenic differentiation and mineral synthesis of dental pulp stem cells. Drug Deliv Transl Res 2019;9(1):85–96. DOI: 10.1007/s13346-018-00600-3.
  27. Atkuri KR, Mantovani JJ, Herzenberg LA, et al. N-acetylcysteine – a safe antidote for cysteine/glutathione deficiency. Curr Opin Pharmacol 2007;7(4):355–359. DOI: 10.1016/j.coph.2007.04.005.
  28. Berk M, Malhi GS, Gray LJ, et al. The promise of N-acetylcysteine in neuropsychiatry. Trends Pharmacol Sci 2013;34(3):167–177. DOI: 10.1016/j.tips.2013.01.001.
  29. Rushworth GF, Megson IL. Existing and potential therapeutic uses for N-acetylcysteine: the need for conversion to intracellular glutathione for antioxidant benefits. Pharmacol Ther 2014;142(2):150–159. DOI: 10.1016/j.pharmthera.2013.09.006.
  30. The ocharidou A, Bakopoulou A, Kontonasaki E, et al. Odontogenic differentiation and biomineralization potential of dental pulp stem cells inside Mg-based bioceramic scaffolds under low-level laser treatment. Lasers Med Sci 2017;32(1):201–210. DOI: 10.1007/s10103-016-2102-9.
  31. Paduano F, Marrelli M, White LJ, et al. Odontogenic differentiation of human dental pulp stem cells on hydrogel scaffolds derived from decellularized bone extracellular matrix and collagen type I. PLoS One 2016;11(2):148–225. DOI: 10.1371/journal.pone.0148225.
  32. Soares DG, Rosseto HL, Scheffel DS, et al. Odontogenic differentiation potential of human dental pulp cells cultured on a calcium-aluminate enriched chitosan-collagen scaffold. Clin Oral Investig 2017;21(9):2827–2839. DOI: 10.1007/s00784-017-2085-3.
  33. Kim Y, Park J-Y, Park H-J, et al. Pentraxin-3 modulates osteogenic/odontogenic differentiation and migration of human dental pulp stem cells. Int J Mol Sci 2019;20(22):57–78. DOI: 10.3390/ijms20225778.
  34. An S. The emerging role of extracellular Ca2+ in osteo/odontogenic differentiation and the involvement of intracellular Ca2+ signalling: from osteoblastic cells to dental pulp cells and odontoblasts. J Cell Physiol 2019;234(3):2169–2193. DOI: 10.1002/jcp.27068.
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