Identification of novel endometrial targets for contraception using microarray technology

S Mirkin, S Oehninger

The Jones Institute for Reproductive Medicine, Norfolk, VA, United States

Introduction A series of temporally and spatially coordinated morphological and molecular changes are necessary for the preparation of the receptive endometrium. We propose that altering events associated with endometrial changes essential for implantation would provide an opportunity for blocking implantation and hence could be used as a strategy for developing contraception methods.

Objectives The objective of this study was to identify candidate genes that could potentially be of high functional significance during embryo implantation in the human endometrium and could be used as a contraceptive targets.

Materials and methods We compare gene expression profiles of the mid luteal phase (receptive endometrium) versus the early luteal phase (pre-receptive endometrium) in healthy fertile volunteers. Endometrial biopsies were obtained on day LH + 3 (day 16, early luteal phase) and on day LH+ 8 (day 21, receptive endometrium). Total RNA was extracted for gene expression analysis using DNA microarrays. cDNA synthesis and in vitro transcription were carried out to create biotinylated cRNA's using the Affymetrix GeneChip Fluidics Station 400, and chemically fragmented cRNA samples were hibrydized on Affymetrix Hu 95A microarray containing 12,000 full length previously characterized genes. Following optical scanning data were analyzed using the GeneChip Analysis Suite. Gene expression profiles were compared and concordant genes were analyzed using SAM software.

Results With a false discovery rate of 0, the analysis revealed that 107 genes were significantly and differently expressed (> 2-fold) between early luteal phase and the receptive endometrium (mid luteal phase). Of these, 49 genes were up regulated whereas 58 were down regulated. More than + 4 fold changes were demonstrated for 27 genes (msh homeo box homolog 1 ¯7.45 fold; thymidylate synthetase ¯ 5.90 fold; ZW10 interactor ¯ 5.10 fold; KIAA0146 ­ 32.00 fold; interleukin 15 ­ 7.50 fold). Some of those genes were not previously reported as being involved in the human embryo implantation. Random genes were selected and their expression were validated using real time PCR.

Conclusions We have identified new candidate genes that involved in the transition of early luteal phase to the endometrial receptivity phase. We reported 107 genes that are potential candidates targets for contraception. Ongoing functional studies will provide proof of the principle that these genes are indeed contraceptive targets.