Stro-1 Positive And Stro-1 Negtive Human Mesenchymal Stem Cells Express Different Levels Of Immunosuppression

Mesenchymal stem cells (MSCs) have been shown to elicit immunosuppressive effect on allongeneic lymphocyte response. However, MSCs are heterogneous and data on the inhibitory ablities of different MSC subsets are lacking. We previously reported that the functions of MSCs depend on their Stro-1 phenotype: Stro-1+ cells home in higher number than Stro-1" cells in the majortity of mouse tissue, While Stro-1" cells support better engraftment of haematopoietic progenitors. According to this study, we hypothesized that Stro-1+ cells might display more signifcant immunosuppressive properties which would explain better crossing of the xenogeneic barrier and more widespread homing to target organs. To explore this hypothesis, we isolated Stro-1+cells from human MSCs and add 1,000~30, 000 Stro-l+cells or Stro-1'cells to mixed lymphocyte reaction (MLR) and to mitogen response assays with PHA plus IL-2. With both Stro-1+ cells or Stro-1 "cells, we observed a dose-dependent reduction of lymphocyte proliferation, but, importantly, this suppressive effect was significantly more powerful with Stro-l+cells as compared to Stro-1" cells (P<0.05). As an example, as few as 1,000 Stro-1+ MSCs inhibit lymphocyte proliferation more effectively than 10,000 Stro-1" cells. To investigate whether the difference of suppressive effect we observed between Stro-1+cells and Stro-1 "cells still exist when MSC subsets are separated physically from PBL, we performed MLR in the upper chamber of a transwell and we seeded the lower chamber either with Stro-1+ cells or Stro-1" cells. Our results showed that Stro-1+ cells still displayed more effectiv immnosuppression than that observed in Stro-1" cells ((P<0.05) in such condition.However, actual physical contact between MSC and peripheral blood cell (PBL) could increase such suppression, which indicates that effective concentration of soluble factor(s) could not be excluded from the immunosuppression of MSCs. We infer that such factor(s) might be significantly produced to a high level by Stro-l+cells than by Stro-1" cells. Furthermore, to detect which inhibitory factor(s) involves in the different suppression of MSC subsets, we evaluated cytokine and chemokine genes expression in both Stro-1+ cells and Stro-1" cells by relative quantitative real-time RT-PCR. No significant difference in TGF-beta1, IL-10 (both are well-known inhibitors of T-cell activation and proliferation) and SDF-1 (a chemokine, which together with its receptor CXCR4 induces stem cell homing) levels was found between the two subsets. This finding suggests that the candidate T-cell inhibitory factors are not responsible for the more profound inhibition of immunoreactivity by Stro-1+ cells than by Stro-1" cells; other factor(s) might be identified in further investigation. In conclusion, our finding clearly has possible therapeutic implication: a pre-selection before ex vivo expansion of MSCs for in vivo infusion may be relevant to exert more profound inhibitory effect only with fewer amounts of expanded cells, especially in those situation where the incidence of GVHD is high, such as unrelated donor heamatopoietic stem cell transplation, and haploid stem cell transplantation. Besides this series of experiments, we also estalished a new MSC culture method: use Platelet-Rich Plasma (PRP) in place of FBS to provide a safer and more effective culture condition to expand MSCs for clinical purpose.