Evolutionary forward genomics reveals novel insights into the genes and pathways dysregulated in recurrent early pregnancy loss.

study questionAre the genes that gained novel expression in the endometria of Eutherian (placental) mammals more likely to be dysregulated in patients with endometrial-associated recurrent early pregnancy loss (REPL)? SUMMARY ANSWER: There was a significant enrichment of genes dysregulated in REPL patients among the Eutherian-specific endometrial genes. WHAT IS KNOWN ALREADY: Pregnancy loss is the most common complication of human pregnancy. REPL has multiple etiologies, including dysregulation of endometrial function, leading to 'suboptimal' implantation. Although the implantation process is tightly regulated in Eutherian (placental) mammals, the molecular factors contributing to dysregulated endometrial gene expression patterns in women with REPL are largely unknown. STUDY DESIGN, SIZE, DURATION: Endometrial biopsies were obtained from 32 REPL patients during the mid-luteal phase, and evaluated for glandular development arrest based on elevated nuclear cyclin E levels in gland cells, and for out-of-phase endometrial development based on histology. Gene expression levels were measured using Illumina Human HT-12v4 BeadChip arrays. PARTICIPANTS/MATERIALS, SETTING,

methodsDifferentially expressed genes were identified between patients with (i) out-of-phase (n = 10) versus normal (n = 22) histological dating and (ii) abnormally elevated (n = 9) versus normal (n = 23) cyclin E levels in the nuclei of endometrial glands, using a likelihood ratio test. Enrichment of dysregulated genes in REPL endometria among Eutherian-specific genes was tested by permutation. Gene ontology and pathway enrichment analyses were carried out for the dysregulated genes. MAIN RESULTS AND THE ROLE OF CHANCE: Fifty-eight and eighty-one genes were identified as differentially expressed at P < 0.001 in women with out-of-phase histological dating and abnormally elevated glandular cyclin E levels, respectively. Genes that were recruited into endometrial expression during the evolution of pregnancy in Eutherian mammals were significantly enriched for dysregulated genes (P = 0.002 for histology, P = 0.021 for cyclin E), as well as for genes involved in immune response and signaling pathways with essential roles in implantation and endometrial biology. LIMITATIONS, REASONS FOR CAUTION: Small sample size limits the statistical power to detect dysregulated genes, and the lack of non-REPL control women does not allow us to test for the contribution of these genes to overall risk of REPL. WIDER IMPLICATIONS OF THE

findingsEnrichment of functional gene categories, as well as genes gained expression in the Eutherian endometria, help to identify molecular etiologies that contribute to normal functioning of the endometrium. These pathways are also strong candidates for successful pregnancy outcomes. Using the evolutionary history of mammalian gene expression in the endometrial tissue may be a promising approach to discover genes involved in female reproductive disorders. STUDY FUNDING/COMPETING INTERESTS: This work is supported by National Institutes of Health (NIH) grant R01 HD21244 to C.O. Authors declare no competing interests.