The immune environment in human endometrium during the window of implantation

Abstract

Objective: Regulation of immune—related genes in endometrium during the window of implantation (WOI) is important for successful implantation and pregnancy maintenance. We have previously investigated global gene expression in human endometrium during the WOI compared to the proliferative phase, which revealed up— and down—regulation of several immune genes. Herein, we have validated these observations in human endometrium. Design: To compare specific immune gene expression in the WOI vs. proliferative endometrium, RT—PCR and Northern analysis were conducted. Materials/Methods: Endometrium in the late proliferative (cycle day 10—12) (n=3) and mid secretory phase (cycle day 20—24) (n=3) was obtained from normally cycling women (ages 18—39) after informed consent. Total RNA was prepared using Trizol (Invitrogen). RT—PCR was conducted using 1 g total RNA for cDNA synthesis followed by 30—35 cycles of PCR using gene—specific primers. For Northern analysis, 20 g total RNA was separated, transferred to Nytran membranes and probed with radiolabeled, gene—specific, cDNA probes. Results: Several immune—related genes were significantly up— regulated in secretory phase endometrium. RT—PCR confirmed the increase of the following genes in the secretory vs. proliferative phase: IL—15 (387bp), NKG5 (382bp), Natural killer associated transcript 2(NKAT2) (449bp), Decay accelerating factor (DAF) (399bp), Indoleamine 2, 3 dioxygenase (IDO) (462bp), Lymphotactin (304bp). Northern blot analysis of endometrial tissue confirmed expression and transcript size for each mRNA: IL—15(1.1kb), NKG5 (.745kb), NKAT2 (1.5kb), DAF (2.1kb), IDO (1.0 & 1.9kb), lymphotactin (1.5kb). Conclusions: This study unequivocally demonstrates up—regulation of several immune—related genes in human endometrium during the implantation window. These genes cover a wide variety of functions including, NK—cell proliferation (IL—15), chemokinesis (lymphotactin), antibacterial activity (NKG5), inhibition of T—cell proliferation (IDO), inhibition of the classical complement pathway ( DAF) and inhibition of NK—cell cytolytic activity (NKAT2). All these functions are important in the context of implantation and maternal tolerance of the fetus. Some patients with recurrent spontaneous abortion and unexplained infertility may have an underlying immune basis and the data presented herein serve as the basis for further investigation of their roles in implantation failure and repetitive miscarriage. Supported by: Supported by New Research Initiative of the Specialized Cooperative Centers Program in Reproduction Research (U54—31398) (LCG) and the NIH Office of Women’s Health (LCG).