Using Oct4,HRE dual promoter driven reporter gene to identify cancer stem cells via multi-modalities molecular image technique

• Main Authors: Hsiang-Lin Yu, 游祥霖
• Other Authors: C. Allen Chang
• Format: Others
• Language: zh-TW
• Published: 2012
• Online Access: http://ndltd.ncl.edu.tw/handle/26554945002919127053

碩士 === 國立陽明大學 === 生物醫學影像暨放射科學系 === 100 === Cancer/tumor stem cell, also known as tumor initiating cell, is a small subpopulation of cancer cell with stem cell property, process self-renewal and differentiation capacity. Several evidences suggest that cancer stem cells are involved in chemo-resistance, radio-resistances, tumor relapse and metastasis. Thus the issue of how to identify cancer stem cells in a tumor and target them is a emergent topic of cancer research. Oct4, also known as POU5f1, is a member of POU transcription factor family, which localizes in human 6th chromosome (6p21.31). Oct4 transcriptional factor only expressed in undifferentiated embryonic stem cells, therefore it is regarded as a unique gene in stem cell. Interestingly, it was found in many cancer researches that Oct4 is also expressed in many kinds of cancer, for example, brain cancer, breast cancer, and thyroid cancer. Therefore, the over-expression of Oct4 in cancer cells is also regarded as a specific gene expression in tumor stem cells. Tumor microenvironment profoundly regulates the gene expression of tumor cell. Hypoxia is one of the characteristics in tumor microenvironment. Generally, hypoxia affects a variety of tumor cell properties such as radio-resistance, chemo-resistance, neovascularization and metastasis. With up-regulating HIF signal pathway, it makes cancer cells behave different gene signature in hypoxia region. The previous research has demonstrated that the hypoxia induced factor-2 (HIF-2) could up-regulate the transcription factor –Oct4. Therefore, the hypoxia region may be the niche of cancer stem cells. We constructed dual promoters reporter gene system in the research, which includes: (1) Red fluorescent protein (mcherry) and luciferase gene driven by Oct4 promoter; (2) Enhance green fluorescent protein( EGFP) driven by hypoxia response element(HRE) promoter. After transfecting the dual promoter reporter gene system into human non-small cell lung cancer cell line (H1299) and establishing the stable clone, we wondered if the cells in the hypoxia area have higher Oct4 expression level in the S.C. xenograft tumor model. Then separated the mcherry+EGFP+cells which are putative cancer stem cells with flow cytometry and analyzed the difference of gene expression between cancer cells and cancer stem cells.