Effect of Ultrasound on Human Umbilical Cord Perivascular-Stem Cell Expansion

Objective: In this study, we explored the effect of Low Intensity Pulsed Ultrasound (LIPUS) on the expansion, and state of differentiation, of HUCPVSC. Previous research has shown that low-intensity therapeutic ultrasound enhances bone and dental tissue formation and repair.

Methods: Different passages of HUCPVSC were expanded and were exposed to either LIPUS treatment (10 minutes per day for 1, 7 and 14 days) or sham treatment (Control). Baseline and end point (1,7 and 14 days) cell count, alkaline phosphatase, flow cytometry and nucleostemin [stem cell marker] mRNA gene expression were evaluated. Flow cytometry included (CD31 [endothelial cell marker], and CD90 [stem cell marker], CD 45 [differentiated hematopoietic cell marker], CD34 [hematopoietic cells and vascular endothelium marker], MHC I [Recognized during graft rejection and found on all nucleated cells], and MHC II [a marker for B-lymphocytes, macrophages and dendritic cells]. Assays were performed in triplicate (with an n=3 samples/assay). Statistical Analysis was performed using MANOVA tests to evaluate differences between LIPUS and control groups at different time points.

Results:

1. There were significant increases in cell count, ALP and DNA levels, being 1.5 times greater in the LIPUS treated groups compared with the control group at day 14 (P< 0.05).

2. There was a significantly increased CD90 expression in the LIPUS treated groups compared with the control group (P< 0.0001).

3. Neucleostemin gene expression was significantly greater in LIPUS treated groups compared with the control group (P< 0.05).

Conclusion: The results of this study suggest that LIPUS has the potential to enhance HUCPVSC expansion without differentiation. We continue to characterize the LIPUS-expanded cells for their application to craniofacial tissue engineering.

Acknowledgment: The Fund for Dentistry