AUTHOR=Ducret Maxime , Fabre Hugo , Degoul Olivier , Atzeni Gianluigi , McGuckin Colin , Forraz Nico , Mallein-Gerin Frédéric , Perrier-Groult Emeline , Alliot-Licht Brigitte , Farges Jean-Christophe TITLE=Immunophenotyping Reveals the Diversity of Human Dental Pulp Mesenchymal Stromal Cells In vivo and Their Evolution upon In vitro Amplification JOURNAL=Frontiers in Physiology VOLUME=7 YEAR=2016 URL=https://www.frontiersin.org/journals/physiology/articles/10.3389/fphys.2016.00512 DOI=10.3389/fphys.2016.00512 ISSN=1664-042X ABSTRACT=

Mesenchymal stromal/stem cells (MSCs) from human dental pulp (DP) can be expanded in vitro for cell-based and regenerative dentistry therapeutic purposes. However, their heterogeneity may be a hurdle to the achievement of reproducible and predictable therapeutic outcomes. To get a better knowledge about this heterogeneity, we designed a flow cytometric strategy to analyze the phenotype of DP cells in vivo and upon in vitro expansion with stem cell markers. We focused on the CD31⁻ cell population to exclude endothelial and leukocytic cells. Results showed that the in vivo CD31⁻ DP cell population contained 1.4% of CD56⁺, 1.5% of CD146⁺, 2.4% of CD271⁺ and 6.3% of MSCA-1⁺ cells but very few Stro-1⁺ cells (≤ 1%). CD56⁺, CD146⁺, CD271⁺, and MSCA-1⁺ cell subpopulations expressed various levels of these markers. CD146⁺MSCA-1⁺, CD271⁺MSCA-1⁺, and CD146⁺CD271⁺ cells were the most abundant DP-MSC populations. Analysis of DP-MSCs expanded in vitro with a medicinal manufacturing approach showed that CD146 was expressed by about 50% of CD56⁺, CD271⁺, MSCA-1⁺, and Stro-1⁺ cells, and MSCA-1 by 15–30% of CD56⁺, CD146⁺, CD271⁺, and Stro-1⁺ cells. These ratios remained stable with passages. CD271 and Stro-1 were expressed by <1% of the expanded cell populations. Interestingly, the percentage of CD56⁺ cells strongly increased from P1 (25%) to P4 (80%) both in all sub-populations studied. CD146⁺CD56⁺, MSCA-1⁺CD56⁺, and CD146⁺MSCA-1⁺ cells were the most abundant DP-MSCs at the end of P4. These results established that DP-MSCs constitute a heterogeneous mixture of cells in pulp tissue in vivo and in culture, and that their phenotype is modified upon in vitro expansion. Further studies are needed to determine whether co-expression of specific MSC markers confers DP cells specific properties that could be used for the regeneration of human tissues, including the dental pulp, with standardized cell-based medicinal products.