Quantification and Characterization of the Nucleolar Channel System of Human Endometrium

Abstract

Nucleolar channel systems (NCSs) are membranous structures discovered in 1960 with electron microscopy that are unique to the nuclei of endometrial epithelial cells (EECs). NCSs are now detected with indirect immunofluorescence using antibodies to nuclear pore complex (NPC) proteins enriched in NCSs. NCSs are present in ∼50% of all EECs during the human implantation window: cycle days 19-24 of an idealized 28-day menstrual cycle. NCSs are unique to healthy human endometrium: they are not found in other hormonally sensitive tissue such as breast, human endometrial stromal cells, endometrial carcinoma tissue, or even in baboon endometrium.;Currently, there is no in vitro system to induce NCSs and the only way to study NCSs is with endometrial tissue specimens. Immunofluorescence has previously revealed NCSs contain a subset each of NPC, inner nuclear membrane and endoplasmic reticulum proteins. We now add the nuclear transport factors Ran and karyopherins alpha and beta1. Additionally, the centrosomal component gamma-tubulin is the first component of NCS cores.;To determine the prevalence of NCSs in endometrial tissue, a semi-quantitative grading scheme (absent/low vs. normal) was employed. With this, we confirmed the midluteal association of NCSs and demonstrated no association between NCSs and overall fertility status or unexplained infertility. Building on this, a method to quantify NCSs absolutely was developed and validated. Based upon studies examining the regional preference for implantation, we hypothesized that NCS presence varies by uterine region and quantitatively determined this by uterine region. NCSs were found preferentially in the upper five regions of the endometrium (16.9%, IQR=12.7-23.4) versus the lower uterine segment (6.1%, IQR=3.0-9.9). In another study, we quantified NCSs in specimens from women having undergone ovarian hyperstimulation (OH). OH is a hormonal regimen used during in vitro fertilization and known to affect the endometrium. During control natural cycles, NCSs were absent on day 16 (0.0%, IQR=0.0-0.0), while after OH NCSs were robust on day 16 (40.4%, IQR=22.6-53.3) suggesting that the NCS window is widened by OH.;Together, this data suggests a physiological role for NCSs in the human endometrium during the implantation window. Moreover, our NCS quantification method is a benchmark for future studies.