S-Space College of Medicine/School of Medicine (의과대학/대학원) Dept. of Medicine (의학과) Theses (Master's Degree_의학과)
In vivo bioluminescence reporter gene imaging for the activation of neuronal differentiation induced by neuronal activator neurogenin 1 (Ngn1) in neuronal precursor cells
- 의과대학 의학과
- Issue Date
- 서울대학교 대학원
- 학위논문 (석사)-- 서울대학교 대학원 : 의학과 핵의학 전공, 2013. 2. 정재민.
- Purpose: The facilitation capability of neuronal lineages derived from transplanted stem cells is essential to improve the low efficacy of neuronal differentiation in stem cell therapy in vivo. Neurogenin 1 (Ngn1), a basic helix-loop-helix factor, has been used as an activator of neuronal differentiation. In this study, we monitored the in vivo activation of neuronal differentiation by Ngn1 in neuronal precursor cells using neuron-specific promoter-based optical reporters.
Methods: The NeuroD promoter coupled with the firefly luciferase reporter system (pNeuroD-Fluc) was used to monitor differentiation in F11 neuronal precursor cells. In vitro luciferase activity was measured and normalized by protein content. The in vivo-jetPEITM system was used for in vivo transgene delivery. The IVIS 100 imaging system was used to monitor in vivo luciferase activity.
Results: The Ngn1-induced neuronal differentiation of F11 cells generated neurite outgrowth within 2 days of Ngn1 induction. Immunofluorescence staining demonstrated that early and late neuronal marker expression (βIII-tubulin, NeuroD, MAP2, NF-M, and NeuN) was significantly increased at 3 days after treatment with Ngn1. When Ngn1 and the pNeuroD-Fluc vector were co-transfected into F11 cells, we observed an approximately 11-fold increase in the luciferase signal. An in vivo study showed that bioluminescence signals were gradually increased in Ngn1-treated F11 cells until 3 days.
Conclusions: In this study, we examined the in vivo tracking of neuronal differentiation induced by Ngn1 using an optical reporter system. This reporter system could be used effectively to monitor the activation efficiency of neuronal differentiation in grafted stem cells treated with Ngn1 for stem cell therapy.