The Role of TBX3 in Human ES Cell Differentiation
试验速览
- 阶段
- 不适用
- 状态
- 撤回
- 主要终点
- The cell morphology and biochemical changes will be examined for the effects of TBX3 knockdown on hES cell differentiation, including testing human ß-CG estradiol and progesterone.
研究概览
简要总结
Stem cells can develop into every cell, every tissue and every organ in the human body, e.g., they can make any kind of cells in the human body. Stem cells reproduce themselves many times over and over. Their almost limitless potential has made stem cells a significant focus of medical research. But before scientists can use stem cells for medical purposes, they must first learn how to harness their power. They cannot treat disease until they learn how to manipulate stem cells to get them to develop into specific tissues or organs.
We know that turning genes on and off is crucial to the process differentiation, so we can add some factor into the culture dish and observe stem cells to differentiate into specific types of cells. But some sort of signal is needed to actually trigger the stem cells to differentiate. We are still searching for that signal. If we can ultimately learn how to direct stem cells to differentiate into one type of tissue or another, then we can use them to treat patients. In this proposal, we will first examine this step.
We propose a novel approach to understanding differentiation of human embryo stem (hES) cells, by studying TBX3, a protein called a transcription factor that controls the expression of other genes. In humans, the loss of function of TBX3 causes Ulnar-Mammary Syndrome, a genetic disorder that can pass from one generation to the next. Furthermore, our preliminary results show that TBX3 is downstream mediator of another protein, BMP4. BMP4 is a known key regulator for hES cell differentiation. Thus, TBX3 is an attractive candidate as a downstream mediator of BMP4 in hES cell differentiation. We will test TBX3 effects on hES cell differentiation if down-regulate TBX3 in hES cells with a technology called siRNA knockdown. We will identify the genes controlled by TBX3 with a recently invented powerful technology called CHIP-GLAS. This technique allows us to examine thousands of genes on a small chip in a single experiment. We expect that the innovative experiments proposed here will open a new avenue to understanding the signal of hES cell differentiation.
详细描述
PURPOSE OF STUDY The purpose of this study is to understand what triggers stem cells to differentiate into one type of tissue or another. The study of TBX3, a protein called a transcription factor that controls the expression of other genes, shows that it is a downstream mediator of the protein BMP4, a known key regulator for hES cell differentiation.
Hypothesis and Aims
To understand the role of TBX3 in hES cell differentiation, we hypothesize that TBX3 is a downstream mediator of BMP4 and play an important role in hES cell differentiation. We propose the following specific aims:
- To examine TBX3 function in hES cell differentiation. NIH approved hES cells will be cultured in a conditioned medium with BMP4 to induce differentiation and TBX3 expression. TBX3 will be knocked down with siRNA technology. The cell morphology and biochemical changes will be examined for the effects of TBX3 knockdown on hES cell differentiation, including testing human ß-CG estradiol and progesterone. The gene expression profile will also be examined with Affymetrix gene expression chip and verified with real-time RT-PCR.
- To identify the genome-wide downstream targets directly bound by TBX3 in hESC with Chromatin immunoprecipitation-guided ligation and selection (CHIP-GLAS). To identify the targets of TBX3 in BMP4-induced hES cells, we will perform ChIP-GLAS analysis. CHIP-GLAS is a novel high-throughput technique with combination of chromatin precipitation and oligonucleotide-based microarray. This technology will allow us to identify the genome-wide downstream target genes directly bound by TBX3 in hES cells.
We expect that the proposed experiments will identify a key regulator for hES cell differentiation and open a new avenue to understanding the function of TBX3 in hES cells. The CHIP-GLAS promoter array provides us a cutting edge tool and allows us to examine the broad function of TBX3. We anticipate that the study will significantly enhance our understanding of TBX3 function in hES cell differentiation.
研究设计
- 研究类型
- Observational
- 观察模型
- Case Only
- 时间视角
- Other
入排标准
- 性别
- All
- 接受健康志愿者
- 否
入选标准
- 未提供
排除标准
- 未提供
结局指标
主要结局
The cell morphology and biochemical changes will be examined for the effects of TBX3 knockdown on hES cell differentiation, including testing human ß-CG estradiol and progesterone.
时间窗: 3 years
To identify the targets of TBX3 in BMP4-induced hES cells, we will perform ChIP-GLAS analysis. CHIP-GLAS will allow us to identify the genome-wide downstream target genes directly bound by TBX3 in hES cells.
时间窗: 3 years
次要结局
未报告次要终点