Immunophenotyping Reveals the Diversity of Human Dental Pulp Mesenchymal Stromal Cells In vivo and Their Evolution upon In vitro Amplification

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.

Different Concentrations of C5a Affect Human Dental Pulp Mesenchymal Stem Cells Differentiation

“…In the present study, Cyagen Human Mesenchymal Stem Cell Growth Medium kits were used to culture the primary dental pulp cells and subsequently, stem cell-related marker expression was detected by ow cytometry. MSCA-1 was expressed by 5.6% of DPSCs, which was consistent with a previous report (21). General high glucose DMEM with fetal bovine serum was used to culture DPSCs, which induced MSCA-1 expression in 1.1% of DPSCs, as determined by ow cytometry.…”

Evaluation of bone marrow microenvironment could change how myelodysplastic syndromes are diagnosed and treated

“…Ducret et al found an important diminution of CD271 + and STRO-1 + cells (<1%) in the culture expanded MSC populations compared with fresh human dental pulp (37). In addition, the percentage of CD56 + cells strongly increased from P1 (25%) to P4 (80%) in amplified MSC subpopulations from human dental pulp (38). Likewise, in in vitro cultures of normal BM samples changes were noted in the proportion of STRO-1 + and CD73 + MSCs compared with the initial seeded sample.…”

“…However, MSC amplification in cultures produces changes in their phenotypic and genomic imprinting profiles and distorts their real distribution in comparison with in vivo models (36,37). In addition, the percentage of CD56 + cells strongly increased from P1 (25%) to P4 (80%) in amplified MSC subpopulations from human dental pulp (38). In addition, the percentage of CD56 + cells strongly increased from P1 (25%) to P4 (80%) in amplified MSC subpopulations from human dental pulp (38).…”

Dental Mesenchymal Stem Cell-Based Translational Regenerative Dentistry: From Artificial to Biological Replacement

“…CD56 is a cell adhesion molecule which belongs to the superfamily of immunoglobulin receptors. It is abundantly expressed in the central nervous systems and hence it mediates several neuronal functions by controlling intercellular adhesion, neurite outgrowth, and cell migration, proliferation, survival and differentiation; all of which are characteristics of cells migrating from the cephalic neural crest (Ducret et al, 2016 ).…”

“…They allow for the expression of stem/progenitor cell markers and preserve amplification kinetics without inducing karyotype abnormality while maintaining the differentiation potential of DPSCs into osteoblast/odontoblast cells. Although there are many successful advanced therapies with medicinal products and good regulations from the FDA “21 CFR Part 1271” and the European Medicines Agency (European Directive 1394/2007), further studies are required to determine more specific properties that could be used for the regeneration of human tissues, with standardized cell-based medicinal products (Ducret et al, 2015b , 2016 ).…”