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CD146 Expression Influences Periapical Cyst Mesenchymal Stem Cell Properties

Abstract

Recent studies have identified a new human dental derived progenitor cell population with multi-lineage differentiation potential referred to as human periapical cyst mesenchymal stem cells (hPCy-MSCs). In the present study, we compared two subpopulations of hPCy-MSCs characterised by the low or high expression of CD146 to establish whether this expression can regulate their stem cell properties. Using flow cytometry, we evaluated the stem cell marker profile of hPCy-MSCs during passaging. Furthermore, CD146Low and CD146High cells were sorted by magnetic beads and subsequently both cell populations were evaluated for differences in their proliferation, self-renewal, stem cell surface markers, stemness genes expression and osteogenic differentiation potential.

We found that hPCy-MSCs possessed a stable expression of several mesenchymal stem cell surface markers, whereas CD146 expression declined during passaging.

In addition, sorted CD146Low cells proliferated significantly faster, displayed higher colony-forming unit-fibroblast capacity and showed higher expression of Klf4 when compared to the CD146High subset. Significantly, the osteogenic potential of hPCy-MSCs was greater in the CD146Low than in CD146High population. These results demonstrate that CD146 is spontaneously downregulated with passaging at both mRNA and protein levels and that the high expression of CD146 reduces the proliferative, self-renewal and osteogenic differentiation potential of hPCy-MSCs. In conclusion, our study demonstrates that changes in the expression of CD146 can influence the stem cell properties of hPCy-MSCs.

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References

  1. 1.

    Egusa, H., Sonoyama, W., Nishimura, M., Atsuta, I., & Akiyama, K. (2012). Stem cells in dentistry--part I: stem cell sources. Journal of Prosthodontic Research, 56, 151–165.

    Article  PubMed  Google Scholar 

  2. 2.

    Tatullo, M., Marrelli, M., & Paduano, F. (2015). The regenerative medicine in oral and maxillofacial surgery: the most important innovations in the clinical application of mesenchymal stem cells. International Journal of Medical Sciences, 12, 72–77.

    Article  PubMed  PubMed Central  Google Scholar 

  3. 3.

    Gronthos, S., Mankani, M., Brahim, J., Robey, P. G., & Shi, S. (2000). Postnatal human dental pulp stem cells (DPSCs) in vitro and in vivo. Proceedings of the National Academy of Sciences of the United States of America, 97, 13625–13630.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  4. 4.

    Seo, B. M., Miura, M., Gronthos, S., Bartold, P. M., Batouli, S., Brahim, J., et al. (2004). Investigation of multipotent postnatal stem cells from human periodontal ligament. Lancet (London, England), 364, 149–155.

    CAS  Article  PubMed  Google Scholar 

  5. 5.

    Miura, M., Gronthos, S., Zhao, M., Lu, B., Fisher, L. W., Robey, P. G., et al. (2003). SHED: stem cells from human exfoliated deciduous teeth. Proceedings of the National Academy of Sciences of the United States of America, 100, 5807–5812.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  6. 6.

    Morsczeck, C., Gotz, W., Schierholz, J., Zeilhofer, F., Kuhn, U., Mohl, C., et al. (2005). Isolation of precursor cells (PCs) from human dental follicle of wisdom teeth. Matrix Biology , 24, 155–165.journal of the International Society for Matrix Biology

    CAS  Article  PubMed  Google Scholar 

  7. 7.

    Sonoyama, W., Liu, Y., Fang, D., Yamaza, T., Seo, B. M., Zhang, C., et al. (2006). Mesenchymal stem cell-mediated functional tooth regeneration in swine. PloS One, 1, e79.

    Article  PubMed  PubMed Central  Google Scholar 

  8. 8.

    Huang, G. T., Gronthos, S., & Shi, S. (2009). Mesenchymal stem cells derived from dental tissues vs. those from other sources: their biology and role in regenerative medicine. Journal of Dental Research, 88, 792–806.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  9. 9.

    Kabir, R., Gupta, M., Aggarwal, A., Sharma, D., Sarin, A., & Kola, M. Z. (2014). Imperative role of dental pulp stem cells in regenerative therapies: a systematic review. Nigerian journal of surgery , 20, 1–8.official publication of the Nigerian Surgical Research Society

    PubMed  PubMed Central  Google Scholar 

  10. 10.

    Rizk, A., & Rabie, A. B. (2013). Human dental pulp stem cells expressing transforming growth factor beta3 transgene for cartilage-like tissue engineering. Cytotherapy, 15, 712–725.

    CAS  Article  PubMed  Google Scholar 

  11. 11.

    Xin, L. Z., Govindasamy, V., Musa, S., & Abu Kasim, N. H. (2013). Dental stem cells as an alternative source for cardiac regeneration. Medical Hypotheses, 81, 704–706.

    CAS  Article  PubMed  Google Scholar 

  12. 12.

    Graziano, A., d'Aquino, R., Laino, G., & Papaccio, G. (2008). Dental pulp stem cells: a promising tool for bone regeneration. Stem Cell Reviews, 4, 21–26.

    Article  PubMed  Google Scholar 

  13. 13.

    Ishkitiev, N., Yaegaki, K., Imai, T., Tanaka, T., Fushimi, N., Mitev, V., et al. (2015). Novel management of acute or secondary biliary liver conditions using hepatically differentiated human dental pulp cells. Tissue Engineering Part A, 21, 586–593.

    CAS  Article  PubMed  Google Scholar 

  14. 14.

    Yang, R., Chen, M., Lee, C. H., Yoon, R., Lal, S., & Mao, J. J. (2010). Clones of ectopic stem cells in the regeneration of muscle defects in vivo. PloS One, 5, e13547.

    Article  PubMed  PubMed Central  Google Scholar 

  15. 15.

    Bianco, J., De Berdt, P., Deumens, R., & des Rieux, A. (2016). Taking a bite out of spinal cord injury: Do dental stem cells have the teeth for it? Cellular and Molecular Life Sciences , 73, 1413–1437.CMLS

    CAS  Article  PubMed  Google Scholar 

  16. 16.

    Guimaraes, E. T., Cruz Gda, S., Almeida, T. F., Souza, B. S., Kaneto, C. M., Vasconcelos, J. F., et al. (2013). Transplantation of stem cells obtained from murine dental pulp improves pancreatic damage, renal function, and painful diabetic neuropathy in diabetic type 1 mouse model. Cell Transplantation, 22, 2345–2354.

    Article  PubMed  Google Scholar 

  17. 17.

    Arthur, A., Rychkov, G., Shi, S., Koblar, S. A., & Gronthos, S. (2008). Adult human dental pulp stem cells differentiate toward functionally active neurons under appropriate environmental cues. Stem Cells (Dayton, Ohio), 26, 1787–1795.

    CAS  Article  PubMed  Google Scholar 

  18. 18.

    Marrelli, M., Paduano, F., & Tatullo, M. (2013). Cells isolated from human periapical cysts express mesenchymal stem cell-like properties. International Journal of Biological Sciences, 9, 1070–1078.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  19. 19.

    Tatullo M, Falisi G, Amantea M, Rastelli C, Paduano F, Marrelli M. Dental pulp stem cells and human periapical cyst mesenchymal stem cells in bone tissue regeneration: comparison of basal and osteogenic differentiated gene expression of a newly discovered mesenchymal stem cell lineage. Journal of Biological Regulators and Homeostatic Agents 2015; 29:713–718.

  20. 20.

    Marrelli, M., Paduano, F., & Tatullo, M. (2015). Human periapical cyst-mesenchymal stem cells differentiate into neuronal cells. Journal of Dental Research, 94, 843–852.

    CAS  Article  PubMed  Google Scholar 

  21. 21.

    Kouroupis, D., Churchman, S. M., McGonagle, D., & Jones, E. A. (2014). The assessment of CD146-based cell sorting and telomere length analysis for establishing the identity of mesenchymal stem cells in human umbilical cord. F1000Research, 3, 126.

    PubMed  PubMed Central  Google Scholar 

  22. 22.

    Ulrich, C., Abruzzese, T., Maerz, J. K., Ruh, M., Amend, B., Benz, K., et al. (2015). Human Placenta-Derived CD146-Positive Mesenchymal Stromal Cells Display a Distinct Osteogenic Differentiation Potential. Stem Cells and Development, 24, 1558–1569.

    CAS  Article  PubMed  Google Scholar 

  23. 23.

    Hagmann, S., Frank, S., Gotterbarm, T., Dreher, T., Eckstein, V., & Moradi, B. (2014). Fluorescence activated enrichment of CD146+ cells during expansion of human bone-marrow derived mesenchymal stromal cells augments proliferation and GAG/DNA content in chondrogenic media. BMC Musculoskeletal Disorders, 15, 322.

    Article  PubMed  PubMed Central  Google Scholar 

  24. 24.

    Zhu, W., Tan, Y., Qiu, Q., Li, X., Huang, Z., Fu, Y., et al. (2013). Comparison of the properties of human CD146+ and CD146- periodontal ligament cells in response to stimulation with tumour necrosis factor alpha. Archives of Oral Biology, 58, 1791–1803.

    CAS  Article  PubMed  Google Scholar 

  25. 25.

    Luo, Y., Zheng, C., Zhang, J., Lu, D., Zhuang, J., Xing, S., et al. (2012). Recognition of CD146 as an ERM-binding protein offers novel mechanisms for melanoma cell migration. Oncogene, 31, 306–321.

    CAS  Article  PubMed  Google Scholar 

  26. 26.

    Shi, S., & Gronthos, S. (2003). Perivascular niche of postnatal mesenchymal stem cells in human bone marrow and dental pulp. Journal of Bone and Mineral Research , 18, 696–704.the official journal of the American Society for Bone and Mineral Research

    Article  PubMed  Google Scholar 

  27. 27.

    Sivasankar, V., & Ranganathan, K. (2015). Growth characteristics and expression of CD73 and CD146 in cells cultured from dental pulp. Journal of Investigative and Clinical Dentistry. doi:10.1111/jicd.12155.

    PubMed  Google Scholar 

  28. 28.

    Shi, S., Bartold, P. M., Miura, M., Seo, B. M., Robey, P. G., & Gronthos, S. (2005). The efficacy of mesenchymal stem cells to regenerate and repair dental structures. Orthodontics & craniofacial research, 8, 191–199.

    CAS  Article  Google Scholar 

  29. 29.

    Covas, D. T., Panepucci, R. A., Fontes, A. M., Silva Jr., W. A., Orellana, M. D., Freitas, M. C., et al. (2008). Multipotent mesenchymal stromal cells obtained from diverse human tissues share functional properties and gene-expression profile with CD146+ perivascular cells and fibroblasts. Experimental Hematology, 36, 642–654.

    CAS  Article  PubMed  Google Scholar 

  30. 30.

    Paduano, F., Marrelli, M., White, L. J., Shakesheff, K. M., & Tatullo, M. (2016). Odontogenic differentiation of human dental pulp stem cells on hydrogel scaffolds derived from Decellularized bone extracellular matrix and Collagen Type I. PloS One, 11, e0148225.

    Article  PubMed  PubMed Central  Google Scholar 

  31. 31.

    Huang, C. E., FW, H., CH, Y., Tsai, L. L., Lee, T. H., Chou, M. Y., et al. (2014). Concurrent expression of Oct4 and Nanog maintains mesenchymal stem-like property of human dental pulp cells. International Journal of Molecular Sciences, 15, 18623–18639.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  32. 32.

    Imbert, A. M., Garulli, C., Choquet, E., Koubi, M., & Aurrand-Lions, M. (2012). Chabannon C. CD146 expression in human breast cancer cell lines induces phenotypic and functional changes observed in Epithelial to Mesenchymal Transition. PloS One, 7, e43752.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  33. 33.

    Wagner, W., Bork, S., Horn, P., Krunic, D., Walenda, T., Diehlmann, A., et al. (2009). Aging and replicative senescence have related effects on human stem and progenitor cells. PloS One, 4, e5846.

    Article  PubMed  PubMed Central  Google Scholar 

  34. 34.

    Mitchell, J. B., McIntosh, K., Zvonic, S., Garrett, S., Floyd, Z. E., Kloster, A., et al. (2006). Immunophenotype of human adipose-derived cells: temporal changes in stromal-associated and stem cell-associated markers. Stem Cells (Dayton, Ohio), 24, 376–385.

    Article  PubMed  Google Scholar 

  35. 35.

    Halfon, S., Abramov, N., Grinblat, B., & Ginis, I. (2011). Markers distinguishing mesenchymal stem cells from fibroblasts are downregulated with passaging. Stem Cells and Development, 20, 53–66.

    CAS  Article  PubMed  Google Scholar 

  36. 36.

    Gharibi, B., & Hughes, F. J. (2012). Effects of medium supplements on proliferation, differentiation potential, and in vitro expansion of mesenchymal stem cells. Stem cells translational medicine, 1, 771–782.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  37. 37.

    Russell, K. C., Tucker, H. A., Bunnell, B. A., Andreeff, M., Schober, W., Gaynor, A. S., et al. (2013). Cell-surface expression of neuron-glial antigen 2 (NG2) and melanoma cell adhesion molecule (CD146) in heterogeneous cultures of marrow-derived mesenchymal stem cells. Tissue Engineering Part A, 19, 2253–2266.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  38. 38.

    Tormin, A., Li, O., Brune, J. C., Walsh, S., Schutz, B., Ehinger, M., et al. (2011). CD146 expression on primary nonhematopoietic bone marrow stem cells is correlated with in situ localization. Blood, 117, 5067–5077.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  39. 39.

    Sharma, M. B., Limaye, L. S., & Kale, V. P. (2012). Mimicking the functional hematopoietic stem cell niche in vitro: recapitulation of marrow physiology by hydrogel-based three-dimensional cultures of mesenchymal stromal cells. Haematologica, 97, 651–660.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  40. 40.

    Russell, K. C., Phinney, D. G., Lacey, M. R., Barrilleaux, B. L., Meyertholen, K. E., & O'Connor, K. C. (2010). In vitro high-capacity assay to quantify the clonal heterogeneity in trilineage potential of mesenchymal stem cells reveals a complex hierarchy of lineage commitment. Stem Cells (Dayton, Ohio), 28, 788–798.

    CAS  Article  PubMed  Google Scholar 

  41. 41.

    Sorrentino, A., Ferracin, M., Castelli, G., Biffoni, M., Tomaselli, G., Baiocchi, M., et al. (2008). Isolation and characterization of CD146+ multipotent mesenchymal stromal cells. Experimental Hematology, 36, 1035–1046.

    CAS  Article  PubMed  Google Scholar 

  42. 42.

    Baksh, D., Yao, R., & Tuan, R. S. (2007). Comparison of proliferative and multilineage differentiation potential of human mesenchymal stem cells derived from umbilical cord and bone marrow. Stem Cells (Dayton, Ohio), 25, 1384–1392.

    CAS  Article  PubMed  Google Scholar 

  43. 43.

    Harkness L, Zaher W, Ditzel N, Isa A, Kassem M. CD146/MCAM defines functionality of human bone marrow stromal stem cell populations. Stem cell research & therapy 2016; 7:4.

  44. 44.

    Espagnolle, N., Guilloton, F., Deschaseaux, F., Gadelorge, M., Sensebe, L., & Bourin, P. (2014). CD146 expression on mesenchymal stem cells is associated with their vascular smooth muscle commitment. Journal of Cellular and Molecular Medicine, 18, 104–114.

    CAS  Article  PubMed  Google Scholar 

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Acknowledgments

The present study was supported by “ICARE Project - Infrastruttura Calabrese per la medicina Rigenerativa: “Generazione di biobanche per la criopreservazione di cellule staminali umane e di tessuto osseo per uso clinico e design e sviluppo di bioscaffold innovativi”. PON03PE_00009_2.2. The authors are grateful to Dr. Raghavendra Vasudeva Murthy and Dr. Pasquale Marrazzo for editing the manuscript.

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Correspondence to Marco Tatullo.

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Francesco Paduano; Massimo Marrelli and Marco Tatullo equally contributed to this work.

Electronic Supplementary Material

Supplementary Fig. 1
figure6

MSC surface marker expression on dental stem cells (DSCs) from various passages. Surface marker expression levels of CD146, CD13, CD29, CD44, CD73, CD90, CD105 and CD45 in a dental pulp stem cells (DPSCs), b dental follicle stem cells (DFPCs) and c periodontal ligament stem cells (PDLSCs) during various passages (p2, p4, p6, p8). (GIF 48 kb)

Supplementary Fig. 2
figure7

Growth kinetics and the osteogenic potential of hPCy-MSCs during expansion in vitro. a Long term growth curves of hPCy-MSCs, each obtained from an individual donor (n = 8). b Osteogenic differentiation of hPCy-MSCs at passage 0 and passage 10. Cells were exposed to osteogenic induction medium for 3 weeks, followed by staining with Alizarin Red. (GIF 176 kb)

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Appendix

Appendix

Table 1 Primer sequences used for real-time PCR

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Paduano, F., Marrelli, M., Palmieri, F. et al. CD146 Expression Influences Periapical Cyst Mesenchymal Stem Cell Properties. Stem Cell Rev and Rep 12, 592–603 (2016). https://doi.org/10.1007/s12015-016-9674-4

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Keywords

  • Dental stem cells (DSCs)
  • Human periapical cyst mesenchymal stem cells (hPCy-MSCs)
  • CD146
  • MSC surface markers
  • Magnetic cell sorting
  • Stemness genes
  • Osteogenic differentiation