Determining the LIF-sensitive period for implantation using a LIF-receptor antagonist

Abstract

Uteri of Lif null mice do not support embryo implantation. Since deletion of some genes often prevents the survival of null mice to adulthood, we have used a proven inhibitor of leukaemia inhibitory factor (LIF) signalling to identify the precise window of time during which LIF is required in vivo, and assessed the cellular expression of several LIF-associated targets. On day 4 of pregnancy, mice were injected with hLIF-05 (inhibitor) into the uterine lumen, with corresponding volumes of PBS (vehicle) injected into the contralateral horn. On days 5 and 6, the number of implantation sites was recorded and the uteri processed for immunohistochemistry. Blockade of LIF on day 4 reduced embryo implantation by 50% (P%0.0001) and was effective maximally between 0930 and 1230 h. Antagonism of LIF signalling was evidenced by a lack of phosphorylated STAT3 in the luminal epithelium (LE). Amphiregulin was absent from the LE on day 4 evening and H-type-1 antigen expression was retained in the LE on day 5 in inhibited uteri. Interleukin-1a and oncostatin M expression were reduced in the stroma on day 6, following LIF inhibition. Unexpectedly, PTGS2 expression in stroma was unaffected by LIF inhibition in vivo, in contrast to Lif null mice. In summary, this suggests that LIF signalling is effective for implantation during a discrete time window on day 4 and antagonism of LIF signalling recapitulates many features exhibited in Lif null uteri. The data presented validates the use of antagonists to investigate tissue specific and temporal cytokine signalling in reproductive function.

Physiological and molecular determinants of embryo implantation

“…Lif -deficient female mice exhibit implantation failure and supplementation with LIF rescues this defect (Chen et al, 2000; Stewart et al, 1992). Moreover, pharmological blocking LIF action in the uterus significantly reduces the phosphorylation of the downstream signaling molecule signal transducer and activator of transcription (STAT) 3 in the uterus, resulting in implantation failure (Menkhorst et al, 2011; Mohamet et al, 2009; White et al, 2007). The importance of LIF signaling in implantation is further evidenced by the observations that inactivation of gp130 and STAT3 also causes implantation failure (Catalano et al, 2005; Cheng et al, 2001; Daikoku et al, 2011; Ernst et al, 2001).…”

Leukemia Inhibitory Factor: Roles in Embryo Implantation and in Nonhormonal Contraception

“…In LIF-deficient female mice, EGF-like growth factors such as amphiregulin (Ar), heparin binding epidermal growth factor (HB-EGF), and epiregulin (Ereg) were not expressed at the site of blastocyst apposition [36], although expressions of EGF receptors were not affected [36]. The dependency of Ar on LIF was evident from lack of expression of this growth factor in uterine luminal epithelia following administration of inhibitor to LIF (hLIF-05) [37]. LIF was also required to induce expression of implantation genes including Msx-1 and Wnt-4 [38].…”

Leukemia Inhibitory Factor Enhances Endometrial Stromal Cell Decidualization in Humans and Mice

“…Unlike in women, in mice decidualization of ESC occurs post-implantation. LIF −/− female mice do not undergo artificial decidualization [14] and intraluminal administration of a short-acting LIF inhibitor during the peri-implantation period results in less extensive desmin filaments (decidual marker) than in the control mice [15]. Further, intraluminal injections of LIF into Fox2a null females partially rescues the formation of a deciduoma during artificial decidualization [16].…”