By integrated analysis of three recently published DNA microarray studies with human tissue, thirty-eight genes were found to be differentially expressed in the same direction in fibroid compared to adjacent uterine myometrium by at least two research groups. Among these genes, twelve with rat orthologs were identified as estrogen-regulated from our array study investigating uterine expression in ovariectomized rats treated with estrogen. Functional and pathway analyses of the twelve genes suggested multiple molecular mechanisms for estrogen-dependent cell survival and tumor growth. Firstly, estrogen increased expression of the anti-apoptotic PCP4 gene and suppressed the expression of growth inhibitory receptors PTGER3 and TGFBR2. Secondly, estrogen may antagonize PPARγ signaling, thought to inhibit fibroid growth and survival, at two points in the PPAR pathway: 1) through increased ANXA1 gene expression which can inhibit phospholipase A2 activity and in turn decrease arachidonic acid synthesis, and 2) by decreasing L-PGDS expression which would reduce synthesis of PGJ2, an endogenous ligand for PPARγ. Lastly, estrogen affects retinoic acid (RA) synthesis and mobilization by regulating expression of CRABP2 and ALDH1A1. RA has been shown to play a significant role in the development of uterine fibroids in an animal model. |
Functional genomics involves the analysis of large datasets of information derived from
various biological experiments. One such type of large-scale experiment involves monitoring
the expression levels of thousands of genes simultaneously under a particular condition,
called gene expression analysis. Microarray technology makes this possible and the quantity
of data generated from each experiment is enormous, dwarfi ng the amount of data generated
by genome sequencing project. Microarray technology has become one of the indispensable tools that many biologists use to monitor genome wide expression levels of genes in a given organism
Genomics refers to the comprehensive study of genes and their function. Recent advances in bioinformatics and high-throughput technologies such as microarray analysis are bringing about a revolution in our understanding of the molecular mechanisms underlying normal and dysfunctional biological processes. Microarray studies and other genomic techniques are also stimulating the discovery of new targets for the treatment of disease which is aiding drug development, immunotherapeutics and gene therapy. In this site, we have compiled an extensive list of resources to assist reseachers interested in establishing a microarray platform and performing expression profiling experiments.
Gene expression profiling or microarray analysis has enabled the measurement of thousands of genes in a single RNA sample. There are a variety of microarray platforms that have been developed to accomplish this and the basic idea for each is simple: a glass slide or membrane is spotted or "arrayed" with DNA fragments or oligonucleotides that represent specific gene coding regions. Purified RNA is then fluorescently- or radioactively labeled and hybridized to the slide/membrane. In some cases, hybridization is done simultaneously with reference RNA to facilitate comparison of data across multiple experiments. After thorough washing, the raw data is obtained by laser scanning or autoradiographic imaging . At this point, the data may then be entered into a database and analyzed by a number of statistical methods.
A number of issues must be addressed before establishing a microarray platform and beginning expression profiling studies, in particular, the overall cost. For a cDNA microarray platform, one must purchase a clone set, robot, printing pins and the reagents needed for DNA amplification and purification. The cost of these materials can vary significantly, but one can expect to need at least $100,000 to establish such a platform. However, once the process of printing and hybridizing microarrays has been optimized, the cost per experiment will fall dramatically. Thus, one must decide if the number of planned experiments is enough to warrant the time and cost of establishing a microarray platform. If not, it may be more prudent to seek the services of an academic microarray core facility or a commercial entity. |
