The Endometrial Proteome and Unexplained Infertility

In 25 to 35% of cases of primary infertility, doctors cannot reach a diagnosis that fully explains why a patient is infertile. This means that many women never learn why they cannot conceive and, as a result, fail to receive effective treatment. Manohar et al. (2015) attempt to set fertility research on the right track by examining the endometrial proteome, comparing protein abundancies in uterine biopsies from women presenting with primary infertility with those who have successfully carried a pregnancy.¹

The researchers chose to examine the endometrial proteome because the uterine lining is an important determinant of successful pregnancy. It is under cyclical flux, growing and diminishing with the hormones of the menstrual cycle. In this way, it can respond sensitively to enable implantation of the developing zygote post-conception. Without this nurturing and protective environment, placentation cannot take place and a pregnancy does not establish.

The research team examined uterine biopsies from women undergoing investigation for primary infertility (n = 12) who had been attempting unsuccessfully to become pregnant for over a year. The patients had normal hormonal profiles with ovulation, no uterine or anatomical abnormality, and a male partner with normal sperm counts showing no gross abnormalities. In other words, no cause for infertility could be determined. As a comparison, the researchers used uterine biopsies from fertile women, as proven by a previous successful pregnancy, who were presenting for surgical tubal ligation/re-connection (n = 12).

By measuring luteinizing hormone (LH) concentrations in morning urine samples, the research team established uterine biopsy collection on day LH+7. This meant researchers sampled each donor with the endometrium synchronized in its receptive phase for implantation. Using β3 integrin as a marker for infertility, the team confirmed reduced expression in the biopsies from infertile donors.

Following sample preparation, the team examined the proteins using 2-D polyacrylamide gel electrophoresis (PAGE), isoelectric focusing and sodium dodecyl (SDS)-PAGE. Examining band intensities, the team found that 12 proteins showed alterations in abundance between the infertile and fertile biopsy samples­­—8 proteins increased and 4 decreased with infertility.

They further identified these proteins of interest using liquid chromatography-mass spectrometry (LC-MS) following in-gel trypsin digestion, running samples through an Ultimate 3000 nano-high performance LC system coupled with electrospray ionization mass spectrometry.

Manohar et al. chose four proteins (IGK@protein, Apo-A1, HSPβ-1 and RPLPZ) for further investigation using SDS-PAGE followed by immunoblotting to confirm abundancies. This confirmed the results with increased abundancies for IGK@protein, Apo-A1 and HSPβ-1, and a reduction in RPLPZ with infertility. Using immunohistochemistry on tissue sections, the team investigated cellular location and protein expression differences between the biopsies from the fertile and infertile women donors. The team also cultured human endometrial stromal cells as an in vitro decidualization model for additional validation of the proteins detected.

In discussing the results obtained, Manohar et al. suggest that alterations in abundancies could relate to disruptions in hormonal signaling within the endometrium itself. Further research into these pathways could give answers to the role of endometrial function and its contribution in cases of infertility, perhaps opening areas for therapeutic discovery that would help women to conceive and maintain a pregnancy.

 

Reference

1. Manohar, M.M., et al. (2014) “Proteomic identification and analysis of human endometrial proteins associated with unexplained infertility,” Journal of Proteomics and Bioinformatics, 7(11). doi: 10.4172/jpb.1000340.