matured MII or IVM-MII oocytes were collected and processed rapidly. RNA was extracted and amplified using a 2-round in vitro transcription procedure, then labeled and hybridized to Applied Biosystems Human Genome Survey Microarrays. This array interrogates 29,098 genes. Up-or down-regulated genes were classified according to the molecular function, biological process, or pathway in which they participate. Statistical analysis was employed to compare each class of genes between samples.RESULTS: The in vitro transcription protocol generated an average of 12 mg of RNA per sample. GV, in vivo matured-MII and IVM-MII oocytes expressed 14,188, 16,402 and 18,299 genes, respectively. There was extensive overlap in the genes expressed by each group, but also some significant differences. Samples showing the least similarity in expression profile were GV and in vivo MII oocytes. IVM-MIIs also displayed significant differences in expression when compared to in vivo MII oocytes, indicating that the process of in vitro maturation does not perfectly mimic that achieved in vivo. A disproportionate number of genes for storage proteins displayed down-regulation in IVM-MII oocytes (relative to in vivo MII). A reduction in expression was also seen for genes involved in processes of homeostasis. Significant up-regulation of gene expression for the ubiquitin proteasome pathway was observed in both IVM-MII and GV samples.CONCLUSIONS: This study provided a detailed insight into gene expression of human oocytes, assessing virtually every gene in the genome. GV oocytes were highly active for processes related to transcription, RNA processing and early stages of protein synthesis, whereas MII oocytes were active for pathways involved in processing of mature proteins. The results suggest that although IVM-MII oocytes closely resemble in vivo-MII oocytes for pathways related to nuclear maturity, a number of those associated with cytoplasmic functions continue to be expressed in a GV-like manner. Additionally, IVM-MII oocytes have significant differences in the expression of genes related to cellular storage and homeostasis. The pathways abnormally expressed during IVM highlight specific deficiencies in culture media and point the way to further optimization of IVM protocols.

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In two separate prospectively randomized trials, intracytoplasmic sperm injection (ICSI) cycles were studied in a controlled manner to monitor the effects of either bovine oviductal epithelial cell co-culture (n = 119) or assisted hatching by zona drilling (n = 100). In the first study, immediately following ICSI, all eggs were placed directly either onto partial monolayers of bovine oviductal cells or into regular culture medium. Although the embryo developmental rate was apparently compromised in part by the presence of the co-culture cells, ultimately there were no significant differences in either the viable pregnancy rate (31.6% co-culture versus 29.0% control) or the embryonic implantation rate (11.4% co-culture versus 13.6% control). Assisted hatching also had no significant impact on ICSI cycle outcome in terms of either the viable pregnancy rate (30.0% assisted hatching versus 32.0% control) or the embryonic implantation rate (8.5% assisted hatching versus 13.5% control). However, in female patients aged > or = 35 years, assisted hatching appeared to convey a marginally significant benefit in terms of both the viable pregnancy rate (35.5% assisted hatching versus 11.1% control) and the embryonic implantation rate (10.3% assisted hatching versus 3.1% control). It seems that the overall improvement of ICSI cycle outcome cannot be achieved by the general application of either co-culture or assisted hatching. Nevertheless, it is possible that there remain specific patient groups that might benefit from selected use of either of these modalities.

Endometriosis affects 10-15% of women of reproductive age and is a common cause of infertility and pelvic pain. Although endometriosis is characterized by abnormal growth or turn-over of cells, the genetic changes involved remain unclear. We employed a multi-color fluorescence in situ hybridization (FISH) strategy to determine the incidence of somatic chromosomal numeric alterations in severe/late stage endometriosis. Using alpha-satellite sequence-specific DNA probes for chromosomes 7, 8, 11, 12, 16, 17, and 18, simultaneous two- and three-color FISH were performed to evaluate the frequency of monosomic, disomic, and trisomic cells in normal control and endometriotic tissue specimens. In one of four endometriosis samples studied, a significantly higher frequency of monosomy for chromosome 17 (14.8%, chi 2(4) = 53.3, P < 0.0001) and 16 (8.8%, chi 2(4) = 11.4, P < 0.05) was observed. An increased number of cells with chromosome 11 trisomy (14.8%, chi 2(4) = 96.2, P < 0.0001) were detected in a second case. In a third case, a distinct colony of nuclei with chromosome 16 monosomy (14.1%, chi 2(4) = 21.39, P < 0.005) was detected. Acquired chromosome-specific aneuploidy may be involved in endometriosis, reflecting clonal expansion of chromosomally abnormal cells. That candidate tumor suppressor genes and oncogenes have been mapped to chromosomes 11, 16, and 17 suggests that chromosomal loss or gain plays a role in the development and/or progression of endometriosis.

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