Errors during chromosome separation have a major impact on human health, contributing to tumor formation in somatic cells and producing gametes with the wrong number of chromosomes in the germline. The rate of chromosome malsegregation during meiosis is unusually high in humans compared with other mammals and frequently leads to aneuploid conception, embryo implantation failure, miscarriage or the birth of children affected by severe developmental syndromes. Most meiotic errors occur during oogenesis and their frequency increases dramatically as women age. This leads to significant reproductive health problems for women of advanced reproductive age, an issue of growing importance in Western countries, where the proportion of women delaying reproduction is increasing. The success rates of assisted reproduction procedures also decline rapidly as women age. Consequently, some couples require multiple treatment cycles to achieve a pregnancy and endure significant emotional, physical and financial demands, as a result. Data from screening of preimplantation embryos suggests that increased chromosome abnormality is the primary cause of reduced assisted reproduction success rates in this patient group. The cause of the age-related increase in chromosome malsegregation is currently unknown. In most cells, proper chromosome separation is ensured by surveillance systems (checkpoints) that regulate the progression of the cell cycle. One such checkpoint, the spindle assembly checkpoint, modulates the timing of anaphase initiation in response to the improper alignment of chromosomes at the metaphase plate. This checkpoint has been shown to be essential for accurate chromosome segregation in a variety of cultured cells. If the integrity of such surveillance systems deteriorates in aging oocytes, this could explain the age related increase in aneuploidy. Our preliminary data suggest that the abundance of mRNA transcripts from critical checkpoint genes is reduced in the oocytes of older women, potentially impairing checkpoint function and leading to age-related aneuploidy. We will explore this possibility by using RNA interference to examine the functionality of the spindle checkpoint during oogenesis. Additionally, we will employ real-time fluorescent RT-PCR to quantify the expression levels of checkpoint genes in oocytes from women of varying ages. This will expose any age related differences. Finally, we will employ comparative genomic hybridization of polar bodies to categorize oocytes as normal or aneuploid. The number of transcripts in oocytes from each category will then be deduced using RTPCR, revealing any relationship between checkpoint gene expression and aneuploidy.
