Interestingly, the exhibited ratios suggest that the processed allograft stimulates the viability of undifferentiated MSCs, while this effect is not as obvious for differentiated MSCs. mean seeding efficiency than control MSCs. Dynamic seeding resulted in BAY 1000394 (Roniciclib) a standard distribution of cells among the surface of the nerve allograft. No cells were located inside the nerve allograft after seeding. Conclusion Differentiated MSCs can be dynamically seeded on the surface of a processed nerve allograft, in a similar fashion as undifferentiated MSCs. Schwann-like differentiated MSCs have a significantly higher seeding efficiency after 12 hours of dynamic seeding. We conclude that differentiation of MSCs into Schwan-like cells may improve the seeding strategy and the ability of nerve allografts to support axon regeneration. 0.05 was considered statistically significant. RESULTS Mesenchymal stem cell collection Flow cytometric analysis showed that this cultured rat MSCs were positive for mesenchymal stem cell markers CD29 (88.2%) and CD90 (88.3%) and unfavorable for the hematopoetic cell markers CD34 (91.1%) and CD45 (86.0%), demonstrating that MSCs were definitively the cell lineage utilized in this study. Mesenchymal stem cell differentiation into Schwann-like cells The morphology of differentiated MSCs changed in vitro in a more spindle-like shape, common for Schwann cells. In contrast to undifferentiated MSCs, Schwann cells and differentiated MSCs showed positive immunofluorescence for Schwann cell surface markers S100 Calcium Binding Protein B (S100B), Glial Fibrillary Acidic Protein (GFAP) and BAY 1000394 (Roniciclib) Nerve Growth Factor Receptor (NGFR/P75NTR), which verified successful differentiation into Schwann-like cells.18 Cell viability The cell viability of undifferentiated and differentiated MSCs were equal and remained constant during the first three time points, after which the viability of both cell types slightly decreased. No decrease in cell viability was observed upon co-culture of MSCs with the processed nerve allograft. The viability of differentiated MSCs in the vicinity of an allograft methods the viability of differentiated MSCs alone over time (p=0.270), while the viability of undifferentiated MSCs with the allograft increased over time compared to undifferentiated MSCs alone; the increased viability ratio between 2 and 3 days of culture was statistically significant (p=0.025). The differences between both cell-group ratios after 3 and 7 days of culture were statistically significant as well (p=0.009 and p=0.026). These results imply that chemical processing BAY 1000394 (Roniciclib) of nerve allografts does not generate a surface that decreases cell-viability. Interestingly, the exhibited ratios suggest that the processed allograft stimulates the viability of undifferentiated MSCs, while this effect is not as obvious for differentiated MSCs. The absorbance ratios, and thus the viability ratios of the different cell cultures over time is shown in Physique 1. Open in a separate window Physique 1. Metabolic activity of undifferentiated and differentiated MSCs after a processed nerve allograft was launched to their well. The activity is usually expressed as a ratio of the metabolic activity of MSCs without a processed nerve allograft. Undifferentiated MSCs experienced a significantly higher metabolic ratio after 3 (p=0.009) and 7 (p=0.026) days of culture. Error bars = SEM. *=statistical significance, p 0.05 Cell adhesion After six hours of seeding, only 24.38% (6.34) of undifferentiated MSCs and 43.33% (3.02) of differentiated MSCs were attached to the processed nerve allografts. Subsequently, these percentages increased to 80.00% (1.73) (undifferentiated MSCs) and 94.54% (1.50) (differentiated MSCs) after 12 hours of dynamic seeding and changed to 82.00% (5.92) (undifferentiated MSCs) and 77.50% (6.67) (differentiated MSCs) after 24 hours. Between group analyses did not show any significant differences. Within group analyses showed a significant conversation between seeding efficiency and seeding period Rabbit Polyclonal to GATA4 for undifferentiated MSCs (p=0.021) and differentiated MSCs (p=0.007). Pairwise comparisons revealed that this increase between 6 and 12 hours of seeding (p=0.029) undifferentiated MSCs was statistically significant, but not between 12 and 24 hours (p=0.486). The increased seeding efficiency.
