| Summary: | 碩士 === 國防醫學院 === 海底醫學研究所 === 100 === Stem cell therapy is to inject non-pharmacological stem cells into the human body and utilize the strong regenerative and repairing capacity of stem cells to renew and regenerate the tissue structures of damaged or aged organs, thus to repair the unhealthy body and normalize the tissues of each organ. Currently, stem cell therapy has been applied clinically to cure cancers, stroke, diabetes, etc.. As for hyperoxia, many diseases and symptoms must be cured by hyperoxia therapy, i.e., carbon monoxide poisoning and would healing, etc.. Hyperoxia therapy increases the oxygen concentration in the human body, but there has not been conclusive research result about the effect of high oxygen levels on the physiological behaviors of adult stem cells. To date many researches have indicated that the hypoxic environment in each organ and tissue of our human bodies is indispensible for preventing differentiation of adult stem cells as well as maintaining their quiescence and self-renewal. Under hyperoxic environment, previous studies have indicated that the high level of ROS (reactive oxygen species) increases stem cell apoptosis, and that hyperoxia can promote stem cell migration. In addition to the well-known bone marrow stem cells, in recent years it was found that neural crest stem cells isolated from adult hair follicles and dental follicles are almost as pluripotent as embryonic stem cells. However, currently it is not clear yet how the levels of oxygen affect the behaviors of neural crest stem cells. Therefore, my thesis research is aimed to investigate whether oxygen concentrations affect the proliferation, migration and apoptosis of neural crest stem cells isolated from adult hair follicle.
Our experiments used 20% oxygen as the control environment, and could be divided into three parts. In the first part, we confirmed that culturing under different oxygen concentrations (1%、20%、40% and 80% O2) led to different total cell numbers of neural crest stem cells. In the second part, we performed EdU and TUNEL staining to identify the proliferating and apoptotic neural crest stem cells, respectively. In the third part, we performed immunostaining to analyze expression of proteins involved in cell migration, including actin, cortactin and CXCR4.
We found that the total cell numbers of neural crest stem cells were significantly decreased under high oxygen concentrations (40% and 80% O2). The EdU staining showed that the proliferation rates of neural crest stem cells were comparable between hypoxic and normoxic culturing environments, whereas the proliferation rate was significantly lower under hyperoxia compared with that under normoxia. The TUNEL staining showed that the apoptosis rates of neural crest stem cells were comparable between hypoxic and normoxic culturing environments, whereas the apoptosis rate was significantly higher under hyperoxia compared with that under normoxia. As for the immunostaining with migration markers, we did not observe significant differences among hypoxic, normoxic and hyperoxic environments.
In summary, while culturing under hypoxia, the total number of neural crest stem cells has a small decrease without significant changes in both the proliferation and apoptosis rates. While culturing under hyperoxia, the total number and proliferation rate of neural crest stem cells both significantly decrease, while their apoptosis rate significantly increases. On the other hand, alteration of oxygen concentrations has no significant effect on neural crest stem cell migration.
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