| Summary: | 博士 === 國立陽明大學 === 臨床醫學研究所 === 106 === Human adult mesenchymal stem cells (hMSCs) possess self-renewal and multi-lineage differentiation potential. These cells function not only to maintain the homeostasis of tissues and organs but also play an important role in the repair and replenishment of damaged tissues and organs. hMSCs has the potential to differentiate into osteoblasts and can be induced to differentiate in-vitro. Understanding the factors regulating hMSCs during osteogenic differentiation and its mechanism may further provide insights to improve the production and quality of hMSCs for medical and industrial applications. The purpose of this study is to explore the role of both biophysical and biochemical cues during osteogenesis. Specifically, the effect of stereotopography substrate and intermittent administration of the parathyroid hormone (PTH) during osteogenic differentiation was investigated.
Stereo-topographical silicon nanowires (SiNWs) that were precisely produced by controlled methods to consistently yield various dimensions and growth orientation were used in this study. hMSCs were cultured on stereo SiNWs of different lengths in the absence of biochemical osteogenic induction cues displayed a spherical and less-elongated morphology and showed an approximately 10% loss of cell viability compared to those grown on two-dimensional (2–D) flat silicon wafer. Moreover, osteogenic gene expression of COL1a1 and RUNX2 and F-actin, phosphorylated focal adhesion kinase (pFAK), vinculin and alpha 2 integrin in hMSCs cultured on the shortest SiNWs was significantly higher than those grown on the longer SiNWs and 2–D flat silicon wafer. Stereo-topographical cues provided by SiNWs are able to regulate osteogenic differentiation of hMSCs via cytoskeleton remodeling and this is correlated with the differential expression of alpha 2/beta 1 integrin heterodimers and the focal adhesion molecules pFAK and vinculin.
The recombinant N-terminal (1–34 amino acids) fragment of the parathyroid hormone [PTH (1–34)] was identified to promote osteogenesis, however the mode of PTH treatment can affect different signaling pathways and fate of cells. This study examined and compared the effect of intermittent and continuous PTH (1–34) administration on hMSCs during osteogenic induction. Relative gene expression of osteoblast-specific genes demonstrated significant upregulation of RUNX2, type I Collagen, ALP, and Osterix and increased alkaline phosphatase (ALP) activity in the presence of PTH (1–34). Between the two modes of treatment, Intermittent PTH (1–34) administration increased PKC activity at day 7 of osteogenic differentiation, whereas inhibition of PKC activity attenuated these effects. In addition, the specific isoform PKCδ was activated upon treatment. These findings demonstrate that intermittent PTH (1–34) treatment enhances the osteogenesis of hMSCs by upregulating osteoblast-specific genes via PKCδ activation.
In conclusion, this dissertation investigated the effects of the biophysical property of stereo-topographical SiNWs and biochemical property of PTH (1-34) on hMSCs lineage commitment and differentiation. This study also proposed mechanistic explanation on mechanical sensor mediators and the signal transduction involved during osteogenic differentiation of hMSCs. Importantly, this study presented the relationship between stereo-topographical structures and osteogenic differentiation and provided insights into the design of scaffolds to enhance the biocompatibility of bionic composites for tissue engineering. Also, the identified role of PKC signaling during PTH-induced osteogenesis may provide insights on drug development and therapeutic interventions for osteoporosis.
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