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CC BY 4.0 · Eur J Dent 2023; 17(01): 173-182
DOI: 10.1055/s-0042-1744370
Original Article

Analysis of Thrombin-Activated Platelet-Derived Exosome (T-aPDE) Potential for Dental Pulp Regeneration: In-Vitro Study

Dini Asrianti Bagio 
1   Department of Conservative Dentistry, Faculty of Dentistry, Universitas Indonesia, Jakarta, Indonesia
,
Indah Julianto 
2   Department of Dermato Venereology, Faculty of Medicine, Universitas Sebelas Maret, Solo Surakarta, Indonesia
,
Anggraini Margono 
1   Department of Conservative Dentistry, Faculty of Dentistry, Universitas Indonesia, Jakarta, Indonesia
,
Endang Suprastiwi 
1   Department of Conservative Dentistry, Faculty of Dentistry, Universitas Indonesia, Jakarta, Indonesia
› Institutsangaben Funding This study was financially supported by HIBAH PUBLIKASI WCU UI Nomor: NKB-494/UN2.RST/HKP.05.00/2021, Universitas Indonesia.
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Abstract

Objective This study analyzed the potential of various concentrations of the thrombin-activated platelet-derived exosome (T-aPDE) to regenerate the dental pulp by performing an in-vitro analysis of the cell viability, migration activity, and vascular endothelial growth factor A (VEGF-A) expression of human dental pulp stem cells (hDPSCs).

Material and Methods The hDPSCs were collected from nine third molar teeth of nine healthy donors and were isolated and cultured using the explant method. They were harvested between the third and fourth passages and starved, after which they were seeded in the following treatments: Dulbecco's Modified Eagle Medium and 10% platelet-rich plasma-thrombin as the control groups, and 0.5, 1, and 5% T-aPDE as the experimental groups. All groups had three biological triplicates (Triplo) and two number of experiments. The T-aPDE was analyzed using transmission electron microscopy testing, particle size analyzer, and CD63 +  and CD81 +  specific immune phenotyping flow cytometry tests for plasma exosomes. The cell viability was evaluated using the colorimetric assay of activity cellular enzymes (MTT assay); the migration activity, using scratch assay; and the VEGF-A expression, using enzyme-linked immunosorbent assay.

Results The highest viability absorbance value of hDPSCs after 24, 48, 72 hours of observation was in the 5% T-aPDE group (p<0.05). Whereas, the closest distance result of migratory activation hDPSCs was also in the same group (p<0.05). However the highest VEGF-A expression of hDSPCs was noted in the same group at 72 hours observation (p<0.05).

Statistical Analysis The data were analyzed using one-way analysis of variance and the Kruskal–Wallis test. The statistical power was set at p <0.05

Conclusion The 5% T-aPDE had a higher potential to induce dental pulp regeneration than the other groups.

Keywords

platelet - exosome - thrombin - dental pulp - stem cells


Publikationsverlauf

Artikel online veröffentlicht:
21. Juni 2022

© 2022. The Author(s). This is an open access article published by Thieme under the terms of the Creative Commons Attribution License, permitting unrestricted use, distribution, and reproduction so long as the original work is properly cited. (https://creativecommons.org/licenses/by/4.0/)

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  • References

  • 1 Žižka R, Šedý J. Paradigm shift from stem cells to cell free regenerative endodontic procedures: a critical review. Stem Cells Dev 2017; 26 (03) 147-153
    CrossrefPubMedGoogle Scholar
  • 2 Kim SG, Malek M, Sigurdsson A, Lin LM, Kahler B. Regenerative endodontics: a comprehensive review. Int Endod J 2018; 51 (12) 1367-1388
    CrossrefPubMedGoogle Scholar
  • 3 American Association of Endodontists. Scope of endodontics: regenerative endodontics. Accessed November 2, 2020 at: https://www.aae.org/specialty/wp-content/uploads/sites/2/2017/06/scopeofendo_regendo.pdf
    Google Scholar
  • 4 He L, Zhong J, Gong Q. et al. Regenerative endodontics by cell homing. Dent Clin North Am 2017; 61 (01) 143-159
    CrossrefPubMedGoogle Scholar
  • 5 Martínez CE, Smith PC, Alvarado VAP. The influence of platelet-derived products on angiogenesis and tissue repair: a concise update. Front Physiol 2015; 6: 290
    CrossrefPubMedGoogle Scholar
  • 6 Li X, Hou J, Wu B, Chen T, Luo A. Effects of platelet-rich plasma and cell coculture on angiogenesis in human dental pulp stem cells and endothelial progenitor cells. J Endod 2014; 40 (11) 1810-1814
    CrossrefPubMedGoogle Scholar
  • 7 Kobayashi E, Flückiger L, Fujioka-Kobayashi M. et al. Comparative release of growth factors from PRP, PRF, and advanced-PRF. Clin Oral Investig 2016; 20 (09) 2353-2360
    CrossrefPubMedGoogle Scholar
  • 8 Jin R, Song G, Chai J, Gou X, Yuan G, Chen Z. Effects of concentrated growth factor on proliferation, migration, and differentiation of human dental pulp stem cells in vitro. J Tissue Eng 2018; 9: 2041731418817505
    CrossrefPubMedGoogle Scholar
  • 9 Saeed MA, El-Rahman MA, Helal ME, Zaher AR, Grawish ME. Efficacy of human platelet rich fibrin exudate vs fetal bovine serum on proliferation and differentiation of dental pulp stem cells. Int J Stem Cells 2017; 10 (01) 38-47
    CrossrefPubMedGoogle Scholar
  • 10 Illmilda MD, Bagio DA. Advanced platelet rich fibrin (A-PRF) supplemented conditioned media for human dental pulp stem cell proliferation. J Int Dent Med Res 2019; 12 (02) 396-400
    Google Scholar
  • 11 Bagio DA, Julianto I, Suprastiwi E, Margono A. Ideal concentration of advanced-platelet rich fibrin (A-PRF) conditioned media for human dental pulp stem cells differentiation. Pesqui Bras Odontopediatria Clin Integr 2019; 19 (01) 1-9
    CrossrefGoogle Scholar
  • 12 Masuki H, Okudera T, Watanebe T. et al. Growth factor and pro-inflammatory cytokine contents in platelet-rich plasma (PRP), plasma rich in growth factors (PRGF), advanced platelet-rich fibrin (A-PRF), and concentrated growth factors (CGF). Int J Implant Dent 2016; 2 (01) 19
    CrossrefPubMedGoogle Scholar
  • 13 Fujioka-Kobayashi M, Miron RJ, Hernandez M, Kandalam U, Zhang Y, Choukroun J. Optimized platelet rich fibrin with the low speed concept: growth factor release, biocompatibility and cellular response. J Periodontol 2017; 88 (01) 112-121
    CrossrefPubMedGoogle Scholar
  • 14 Straum OK. The optimal platelet concentration in platelet-rich plasma for proliferation of human cells in vitro-diversity, biases, and possible basic experimental principles for further research in the field: a review. PeerJ 2020; 8: e10303
    CrossrefPubMedGoogle Scholar
  • 15 Jha N, Ryu JJ, Choi EH, Kaushik NK. Generation and role of reactive oxygen and nitrogen species induced by plasma, lasers, chemical agents, and other systems in dentistry. Oxid Med Cell Longev 2017; 2017: 7542540
    PubMedGoogle Scholar
  • 16 Saghiri MA, Asatourian A, Sorenson CM, Sheibani N. Role of angiogenesis in endodontics: contributions of stem cells and proangiogenic and antiangiogenic factors to dental pulp regeneration. J Endod 2015; 41 (06) 797-803
    CrossrefPubMedGoogle Scholar
  • 17 Bronckaers A, Hilkens P, Fanton Y. et al. Angiogenic properties of human dental pulp stem cells. PLoS One 2013; 8 (08) e71104
    CrossrefPubMedGoogle Scholar
  • 18 Zhang M, Jiang F, Zhang X. et al. The effects of platelet-derived growth factor-BB on human dental pulp stem cells mediated dentin-pulp complex regeneration. Stem Cells Transl Med 2017; 6 (12) 2126-2134
    CrossrefPubMedGoogle Scholar
  • 19 Cocucci E, Meldolesi J. Ectosomes and exosomes: shedding the confusion between extracellular vesicles. Trends Cell Biol 2015; 25 (06) 364-372
    CrossrefPubMedGoogle Scholar
  • 20 Vlassov AV, Magdaleno S, Setterquist R, Conrad R. Exosomes: current knowledge of their composition, biological functions, and diagnostic and therapeutic potentials. Biochim Biophys Acta 2012; 1820 (07) 940-948
    CrossrefPubMedGoogle Scholar
  • 21 Kumeda N, Ogawa Y, Akimoto Y, Kawakami H, Tsujimoto M, Yanoshita R. Characterization of membrane integrity and morphological stability of human salivary exosomes. Biol Pharm Bull 2017; 40 (08) 1183-1191
    CrossrefPubMedGoogle Scholar
  • 22 Boukouris S, Mathivanan S. Exosomes in bodily fluids are a highly stable resource of disease biomarkers. Proteomics Clin Appl 2015; 9 (3-4): 358-367
    CrossrefPubMedGoogle Scholar
  • 23 Inal JM, Kosgodage U, Azam S, Stratton D, Antwi-Baffour S, Lange S. Blood/plasma secretome and microvesicles. Biochim Biophys Acta 2013; 1834 (11) 2317-2325
    CrossrefPubMedGoogle Scholar
  • 24 Torreggiani E, Perut F, Roncuzzi L, Zini N, Baglìo SR, Baldini N. Exosomes: novel effectors of human platelet lysate activity. Eur Cell Mater 2014; 28: 137-151 , discussion 151
    CrossrefPubMedGoogle Scholar
  • 25 Liu X, Wang L, Ma C, Wang G, Zhang Y, Sun S. Exosomes derived from platelet-rich plasma present a novel potential in alleviating knee osteoarthritis by promoting proliferation and inhibiting apoptosis of chondrocyte via Wnt/β-catenin signaling pathway. J Orthop Surg Res 2019; 14 (01) 470
    CrossrefPubMedGoogle Scholar
  • 26 Guo SC, Tao SC, Yin WJ, Qi X, Yuan T, Zhang CQ. Exosomes derived from platelet-rich plasma promote the re-epithelization of chronic cutaneous wounds via activation of YAP in a diabetic rat model. Theranostics 2017; 7 (01) 81-96
    CrossrefPubMedGoogle Scholar
  • 27 Stanko P, Altanerova U, Jakubechova J, Repiska V, Altaner C. Dental mesenchymal stem/stromal cells and their exosomes. Stem Cells Int 2018; 2018: 8973613
    CrossrefPubMedGoogle Scholar
  • 28 Hu X, Zhong Y, Kong Y, Chen Y, Feng J, Zheng J. Lineage-specific exosomes promote the odontogenic differentiation of human dental pulp stem cells (DPSCs) through TGFβ1/smads signaling pathway via transfer of microRNAs. Stem Cell Res Ther 2019; 10 (01) 170
    CrossrefPubMedGoogle Scholar
  • 29 Huang CC, Narayanan R, Alapati S, Ravindran S. Exosomes as biomimetic tools for stem cell differentiation: applications in dental pulp tissue regeneration. Biomaterials 2016; 111: 103-115
    CrossrefPubMedGoogle Scholar
  • 30 Ivica A, Ghayor C, Zehnder M, Valdec S, Weber FE. Pulp-derived exosomes in a fibrin-based regenerative root filling material. J Clin Med 2020; 9 (02) 1-12
    CrossrefPubMedGoogle Scholar
  • 31 Naz S, Khan FR, Zohra RR. et al. Isolation and culture of dental pulp stem cells from permanent and deciduous teeth. Pak J Med Sci 2019; 35 (04) 997-1002
    PubMedGoogle Scholar
  • 32 Franco D, Franco T, Schettino AM, Filho JM, Vendramin FS. Protocol for obtaining platelet-rich plasma (PRP), platelet-poor plasma (PPP), and thrombin for autologous use. Aesthetic Plast Surg 2012; 36 (05) 1254-1259
    CrossrefPubMedGoogle Scholar
  • 33 Martins TS, Catita J, Rosa IM, da Cruz E Silva OAB, Henriques AG. Exosome isolation from distinct biofluids using precipitation and column-based approaches. PLoS One 2018; 13 (06) e0198820
    CrossrefPubMedGoogle Scholar
  • 34 Lötvall J, Hill AF, Hochberg F. et al. Minimal experimental requirements for definition of extracellular vesicles and their functions: a position statement from the International Society for Extracellular Vesicles. J Extracell Vesicles 2014; 3: 26913
    CrossrefPubMedGoogle Scholar
  • 35 Martin-Piedra MA, Garzon I, Oliveira AC. et al. Cell viability and proliferation capability of long-term human dental pulp stem cell cultures. Cytotherapy 2014; 16 (02) 266-277
    CrossrefPubMedGoogle Scholar
  • 36 Grada A, Otero-Vinas M, Prieto-Castrillo F, Obagi Z, Falanga V. Research techniques made simple: analysis of collective cell migration using the wound healing assay. J Invest Dermatol 2017; 137 (02) e11-e16
    CrossrefPubMedGoogle Scholar
  • 37 Hilkens P, Gervois P, Fanton Y. et al. Effect of isolation methodology on stem cell properties and multilineage differentiation potential of human dental pulp stem cells. Cell Tissue Res 2013; 353 (01) 65-78
    CrossrefPubMedGoogle Scholar
  • 38 Helwa I, Cai J, Drewry MD. et al. A comparative study of serum exosome isolation using differential ultracentrifugation and three commercial reagents. PLoS One 2017; 12 (01) e0170628
    CrossrefPubMedGoogle Scholar
  • 39 Caradec J, Kharmate G, Hosseini-Beheshti E, Adomat H, Gleave M, Guns E. Reproducibility and efficiency of serum-derived exosome extraction methods. Clin Biochem 2014; 47 (13-14): 1286-1292
    CrossrefPubMedGoogle Scholar
  • 40 Kosaka N, Yoshioka Y, Hagiwara K, Tominaga N, Ochiya T. Functional analysis of exosomal microRNA in cell-cell communication research. Methods Mol Biol 2013; 1024: 1-10
    CrossrefPubMedGoogle Scholar
  • 41 Caruana A, Savina D, Macedo JP, Soares SC. From platelet-rich plasma to advanced platelet-rich fibrin: biological achievements and clinical advances in modern surgery. Eur J Dent 2019; 13 (02) 280-286
    Artikel in Thieme ConnectPubMedGoogle Scholar