The primary goal of this study was to examine the effects of human dental pulp stem cell-derived exosomes on the carrageenan-induced acute inflammation in mice. Exosomes were purified by differential ultracentrifugation from the supernatants of stem cells derived from the dental pulp of human exfoliated deciduous teeth (SHEDs) cultivated in serum-free medium. At 1 h post-carrageenan injection, exosomes derived from supernatants of 2 × 106 SHEDs were administered by intraplantar injection to BALB/c mice; 30 mg/kg of prednisolone and phosphate-buffered saline (PBS) were used as positive and negative controls, respectively. Edema was measured at 6, 24, and 48 h after carrageenan injection. For the in vivo imaging experiments, AngioSPARK750, Cat B 750 FAST, and MMPSense 750 FAST were administered into the mouse tail vein 2 h post-carrageenan injection. Fluorescence images were acquired at 6, 24, and 48 h after edema induction by IVIS Spectrum in vivo imaging system. Exosomes significantly reduced the carrageenan-induced edema at all the time points studied (by 39.5, 41.6, and 25.6 % at 6, 24, and 48 h after injection, respectively), to similar levels seen with the positive control (prednisolone). In vivo imaging experiments revealed that, both exosomes and prednisolone suppress activities of cathepsin B and matrix metalloproteinases (MMPs) at the site of carrageenan-induced acute inflammation, showing more prominent effects of prednisolone at the early stages, while exosomes exerted their suppressive effects gradually and at later time points. Our study demonstrates for the first time that exosomes derived from human dental pulp stem cells suppress carrageenan-induced acute inflammation in mice.
SHED exosomes carrageenan acute inflammation
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This research was funded by the European Social Fund under the Global Grant measure (No. VP1-3.1-ŠMM-07-K-03-016).
Compliance with Ethical Standards
Human material was collected under the approval of the Lithuanian Bioethics Committee.
All procedures with animals were carried out in accordance with the guidelines of the European Union and were approved by the Lithuanian Ethics Committee on the use of the laboratory animals under State Veterinary Service.
Conflicts of Interest
The authors declare that there are no conflicts of interest.
Supplemental table 1Immunophenotypic characterization of SHEDs. SHEDs grown under serum-free medium MSC NutriStem XF medium (Biological Industries) were characterized by FACS for the expression of negative (CD45, CD34) and positive (CD90, CD73, CD105 and CD146) markers of MSC-like cells. (JPEG 260 kb) (PDF 75 kb)
Vesicle size distribution in exosomal preparations. Particle size range was estimated in PBS using a dynamic light scattering device (Brookhaven Instruments Corp., NY, USA). Measurements were performed in triplicate at a temperature of 25 °C (PDF 75 kb) (JPEG 260 kb)
White, E.S., and A.R. Mantovani. 2013. Inflammation, wound repair, and fibrosis: reassessing the spectrum of tissue injury and resolution. The Journal of Pathology 229: 141–144.PubMedCentralCrossRefPubMedGoogle Scholar
McCarberg, B.H., and Cryer, B. 2014. Evolving therapeutic strategies to improve nonsteroidal anti-inflammatory drug safety. American Journal of Therapeutics.Google Scholar
Santiago, T., and J.A.P. da Silva. 2014. Safety of low- to medium-dose glucocorticoid treatment in rheumatoid arthritis: myths and reality over the years. Annals of the New York Academy of Sciences 1318: 41–49.CrossRefPubMedGoogle Scholar
Bianco, P., X. Cao, P. Frenette, J. Mao, P. Robey, P. Simmons, et al. 2013. The meaning, the sense and the significance: translating the science of mesenchymal stem cells into medicine. Nature Medicine 19: 35–42.PubMedCentralCrossRefPubMedGoogle Scholar
Ratajczak, J., K. Miekus, M. Kucia, J. Zhang, R. Reca, P. Dvorak, et al. 2006. Embryonic stem cell-derived microvesicles reprogram hematopoietic progenitors: evidence for horizontal transfer of mRNA and protein delivery. Leukemia 20: 847–56.CrossRefPubMedGoogle Scholar
Zhang, B., Yin, Y., Lai, R.C., and Lim, S.K. 2014. Immunotherapeutic potential of extracellular vesicles. Frontiers in Immunology 5.Google Scholar
Zitvogel, L., A. Regnault, A. Lozier, J. Wolfers, C. Flament, D. Tenza, et al. 1998. Eradication of established murine tumors using a novel cell-free vaccine: dendritic cell-derived exosomes. Nature Medicine 4: 594–600.CrossRefPubMedGoogle Scholar
Admyre, C., S.M. Johansson, K.R. Qazi, J.J. Filen, R. Lahesmaa, M. Norman, et al. 2007. Exosomes with immune modulatory features are present in human breast milk. Journal of Immunology 179: 1969–78.CrossRefGoogle Scholar
Kordelas, L., V. Rebmann, A.K.K. Ludwig, S. Radtke, J. Ruesing, T.R. Doeppner, et al. 2014. MSC-derived exosomes: a novel tool to treat therapy-refractory graft-versus-host disease. Leukemia 28: 970–973.PubMedGoogle Scholar
Zhang, B., Y. Yin, R.C. Lai, S.S. Tan, A.B... Choo, and S.K. Lim. 2014. Mesenchymal stem cells secrete immunologically active exosomes. Stem Cells and Development 23: 1233–1244.Google Scholar
Pivoriuunas, A., A. Surovas, V. Borutinskaite, D. Matuzeviccius, G. Treigyte, J. Savickiene, et al. 2010. Proteomic analysis of stromal cells derived from the dental pulp of human exfoliated deciduous teeth. Stem Cells and Development 19: 1081–93.CrossRefPubMedGoogle Scholar
Thery, C., Amigorena, S., Raposo, G., Clayton, A. 2006. Isolation and characterization of exosomes from cell culture supernatants and biological fluids. Current Protocols in Cell Biology; Chapter 3:Unit 3 22.Google Scholar
Buono, C., J.J. Anzinger, M. Amar, and H.S. Kruth. 2009. Fluorescent pegylated nanoparticles demonstrate fluid-phase pinocytosis by macrophages in mouse atherosclerotic lesions. Journal of Clinical Investigation 119: 1373–81.PubMedCentralCrossRefPubMedGoogle Scholar
Barber, P.A., D. Rushforth, S. Agrawal, and U.I. Tuor. 2012. Infrared optical imaging of matrix metalloproteinases (MMPs) up regulation following ischemia reperfusion is ameliorated by hypothermia. BMC Neuroscience 13: 76.PubMedCentralCrossRefPubMedGoogle Scholar
Perretti, M., and A. Ahluwalia. 2000. The microcirculation and inflammation: site of action for glucocorticoids. Microcirculation 7: 147–61.CrossRefPubMedGoogle Scholar
Perretti, M., and F. D’Acquisto. 2009. Annexin A1 and glucocorticoids as effectors of the resolution of inflammation. Nature Reviews Immunology 9: 62–70.CrossRefPubMedGoogle Scholar
Aalberts, M., F.M. van Dissel-Emiliani, N.P. van Adrichem, M. van Wijnen, M.H. Wauben, T.A. Stout, et al. 2012. Identification of distinct populations of prostasomes that differentially express prostate stem cell antigen, annexin A1, and GLIPR2 in humans. Biology of Reproduction 86: 82.CrossRefPubMedGoogle Scholar
Ronquist, G.K., A. Larsson, A. Stavreus-Evers, and G. Ronquist. 2012. Prostasomes are heterogeneous regarding size and appearance but affiliated to one DNA-containing exosome family. Prostate 72: 1736–45.CrossRefPubMedGoogle Scholar
Subra, C., D. Grand, K. Laulagnier, A. Stella, G. Lambeau, M. Paillasse, et al. 2010. Exosomes account for vesicle-mediated transcellular transport of activatable phospholipases and prostaglandins. Journal of Lipid Research 51: 2105–20.PubMedCentralCrossRefPubMedGoogle Scholar
Napimoga, M.H., C.A. da Silva, V. Carregaro, T.S. Farnesi-de-Assuncao, P.M. Duarte, N.F. de Melo, et al. 2012. Exogenous administration of 15d-PGJ2-loaded nanocapsules inhibits bone resorption in a mouse periodontitis model. Journal of Immunology 189: 1043–52.CrossRefGoogle Scholar
Andersson, S.E., L. Kallstrom, M. Malm, A. Miller-Larsson, and B. Axelsson. 1995. Inhibition of nitric oxide synthase reduces Sephadex-induced oedema formation in the rat lung: dependence on intact adrenal function. Inflammation Research 44: 418–22.CrossRefPubMedGoogle Scholar