Effects of Rac1 Inhibition on Hepatic Differentiation of Mesenchymal Stromal Cells

• Main Authors: Nan-Yuan Teng, 滕南淵
• Other Authors: Oscar K. Lee
• Format: Others
• Language: en_US
• Published: 2012
• Online Access: http://ndltd.ncl.edu.tw/handle/96623134971732384424

碩士 === 國立陽明大學 === 臨床醫學研究所 === 100 === One of the most important components in regenerative medicine is cell, and in vitro differentiation of specific cell lineages from stem cells provides a stable cell supply for regenerative medicine. Understanding the mechanisms underlying differentiation process may be beneficial for finding more efficient and effective ways to derive lineage-specific cells. Previously, we successfully developed a two-step protocol to differentiate human bone marrow-derived mesenchymal stromal cells into hepatocyte-like cells in vitro. The protocol is also applicable to mouse mesenchymal stromal cells. During the hepatic commitment process, an obvious cell morphological change was observed, from fibroblast-like morphology to polygonal epithelial morphology. Besides, cell motility was decreased along the progress of differentiation. These phenomena suggested that hepatic differentiation of MSCs was a process associated with mesenchymal-epithelial transition. Thus, we hypothesized that acceleration of mesenchymal-epithelial transition can promote hepatic differentiation of mesenchymal stromal cells. We planned to accelerate mesenchymal-epithelial transition during hepatic differentiation of mouse mesenchymal stromal cells by using the Rac1 inhibitor, NSC23766, to inhibit the activity of Rac1, which is reported to be required for growth factor-induced mesenchymal phenotype. Quantitative real time PCR and functional assays were used to evaluate the effects of Rac1 inhibition on hepatic differentiation. According to our data, Rac1 inhibition made differentiating cells more epithelial and compact within 7 days. Immunofluorescent staining revealed that the epithelial morphology caused by Rac1 inhibitor was not related to E-cadherin. Furthermore, Rac1 inhibition upregulated the expression level of hepatic marker genes. Rac1-inhibited group also performed better in PAS stain as well as CYP3A4 activity assay. Consistent with findings in mouse mesenchymal stromal cells, Rac1 inhibition also led to similar results in hepatic differentiation of human mesenchymal stromal cells. Taken together, our data suggest that Rac1 inhibition might promote hepatic differentiation of mesenchymal stromal cells partly, if not wholly, through accelerating mesenchymal-epithelial transition.