Evaluation of growth, stemness, and angiogenic properties of dental pulp stem cells cultured in cGMP xeno-/serum-free medium

Abstract

This study was aimed to investigate the effects of cGMP xeno-/serum-free medium (XSF, Irvine Scientific) on the properties of human dental pulp stem cells (DPSCs). DPSCs, from passage 2, were cultured in XSF or fetal bovine serum (FBS)-supplemented medium, and sub-cultured up to passage 8. Cumulative population doublings (PDs) and the number of colony-forming-units (CFUs) were determined. qRT-PCR, ELISA, and in vitro assays were used to assess angiogenic capacity. Flow cytometry was used to measure CD73, CD90, and CD105 expression. Differentiation into osteo-, adipo-, and chondrogenic cell lineages was performed. DPSCs showed more elongated morphology, a reduced rate of proliferation at later passages, and lower CFU counts in XSF compared with FBS. Expression of angiogenic factors at the gene and protein levels varied in the two media and with passage number, but cells grown in XSF had more in vitro angiogenic activity. The majority of early and late passage DPSCs cultured in XSF expressed CD73 and CD90. In contrast, the percentage of CD105 positive DPSCs in XSF medium was significantly lower with increased passage whereas the majority of cells cultured in FBS were CD105 positive. Switching XSF-cultured DPSCs to medium supplemented with human serum restored the expression of CD105. The tri-lineage differentiation of DPSCs cultured under XSF and FBS conditions was similar. We showed that despite reduced CD105 expression levels, DPSCs expanded in XSF medium maintained a functional MSC phenotype. Furthermore, restoration of CD105 expression is likely to occur upon in vivo transplantation, when cells are exposed to human serum.

Hollow-fiber bioreactor production of extracellular vesicles from human bone marrow mesenchymal stromal cells yields nanovesicles that mirrors the immuno-modulatory antigenic signature of the producer cell

“…When post-bioreactor MSCs were seeded back into their normal culture medium in a 2D adherent state (flask-based), cells were shown to completely restore their CD105 levels up to a degree matching the expression level of the pre-bioreactor processed cells. Akin to our results for the plasticity of the CD105 expression level, Qu et al observed similar results where MSCs re-expressed CD105 after the cells grown under serum-free conditions were seeded back into serumcontaining medium [44]. Noteworthy, in contrast to the cells, analysis of the EV immunophenotyping surface marker profile did show the presence of CD105 expression level on EVs produced throughout the 25-day production scheme, suggesting that the MSCs still produced CD105 when inoculated into the bioreactor, but regulated its expression towards the EVs instead of the cellular membrane.…”

Impact of Type 2 Diabetes Mellitus on Human Bone Marrow Stromal Cell Number and Phenotypic Characteristics

“…Interestingly, late passage cultures demonstrated small but apparent differences in this characteristic expression pattern including a reduced CD90 population and increased negative marker population (Supplementary Figure S2A). Though changes to MSC surface marker profiles have been noted in late passage MSCs, the loss of CD90 which is considered a robust marker of MSC identity has not been previously recorded to our knowledge [20][21][22][23][24][25][26]. The presence of T2DM in the host did not affect this characteristic expression pattern in early passages (Figure 2A) or the altered expression at late passages (Supplementary Figure S2B,C).…”

Maxillofacial-Derived Mesenchymal Stem Cells: Characteristics and Progress in Tissue Regeneration

“…According to in vitro research, both fresh and cryopreserved dental MSCs isolated from humans and animals successfully meet mesenchymal stem cell criteria. Maxillofacialderived MSCs express MSC and neural crest-related genes and markers including STRO-1, CD13, CD29, CD44, CD59, CD73, CD90, CD105, CD106, and CD146 and were negative for hematopoietic lineage markers including CD14, CD31, CD34, and CD45 [3,7,8,[30][31][32][33][34][35][36][37][38] (Table 1).…”