Interleukin 1 beta is induced by interleukin 11 during decidualization of human endometrial stromal cells, but is not released in a bioactive form

Abstract

Blastocyst implantation is dependent on the differentiation of endometrial stromal cells (ESC) into decidual cells. Decidualization of human ESC in vitro is enhanced by interleukin 11 (IL11), with associated changes in gene expression. Genes downstream of IL11 may provide targets for the treatment of implantation failure or the development of non-hormonal contraceptives. This study aimed to examine the effect of IL11 on interleukin 1 beta (IL1B) mRNA and protein expression during in vitro decidualization of ESC. Cells were decidualized with 17β-estradiol and medroxyprogesterone acetate in the presence or absence of exogenous IL11, and IL1B mRNA was quantified by real-time RT–PCR. Inactive proIL1B and bioactive IL1B in cell lysates and conditioned media were measured using specific immunoassays. Secretion of bioactive IL1B from decidualizing ESC was investigated by in vitro stimulation of decidualizing cells with lipopolysaccharide, interferon gamma or human chorionic gonadotropin. Immunohistochemistry was carried out on cycling and pregnant decidua using an antibody specific for bioactive IL1B. Exogenous IL11 increased by 28-fold the abundance of IL1B mRNA in decidualizing ESC, and total immunoreactive IL1B was also increased. However, this was not reflected in bioactive IL1B secretion from these cells, and none of the tested stimuli were able to induce its release. Bioactive IL1B was detected in vivo at very low levels and at discrete foci in late secretory phase and first trimester decidua. This regulation of latent and bioactive IL1B at the fetal–maternal interface may prime decidual cells to respond rapidly to immunological challenge or to signals from the blastocyst during implantation.

Introduction

Decidualization of endometrial stromal cells is essential for blastocyst implantation and establishment of pregnancy. Proliferation and differentiation of stromal cells into decidual cells during the late secretory phase of the menstrual cycle is mediated by a complex network of hormones, growth factors and cytokines, including interleukin 11 (IL11) (Tanaka et al., 2001, Dimitriadis et al., 2002, Dimitriadis et al., 2005, Karpovich et al., 2005). During in vitro decidualization of human endometrial stromal cells (ESC) with either 8-bromo-cAMP or progesterone, exogenous IL11 enhances the morphological and molecular markers of decidualization (Tanaka et al., 2001, Dimitriadis et al., 2002). We have previously demonstrated using cDNA microarray analysis that IL11 alters ESC gene expression during progesterone-induced decidualization, with the most upregulated gene being interleukin 1 beta (IL1B) (White et al., 2005).

IL1B is a pro-inflammatory cytokine with known roles in acute and chronic inflammation, fever and tissue remodelling (reviewed in Dinarello (1997)). The IL1 system includes two ligands (IL1A and IL1B), two cell surface receptors (IL1R1 and IL1R2), a receptor accessory protein (IL1RAP) and a naturally occurring antagonist (IL1RA). IL1A and IL1B are encoded by different genes, but have identical biological activities.

Production of bioactive IL1B requires transcription, translation, activation and secretion, each of which is independently regulated (reviewed in Dinarello (1997)). Human IL1B is first translated as a 31 kDa latent precursor (proIL1B) with no IL1 bioactivity. ProIL1B is found within intracellular vesicles (endolysosomes) or associated with microtubules, and has no known membrane-bound form. Within endolysosomes, proIL1B is cleaved by the IL1B-converting enzyme (ICE, caspase 1) to form the 17 kDa bioactive IL1B and a propiece (Cerretti et al., 1992, Thornberry et al., 1992). Following proIL1B activation, the endolysosomes fuse with the plasma membrane and release a combination of bioactive IL1B, proIL1B and propiece. This form of secretion is dependent on both energy and temperature, and can be specifically inhibited or stimulated by a variety of factors.

Extracellular proIL1B can be cleaved by a number of enzymes (trypsin, elastase, chymotrypsin, mast cell chymase, granzyme A and cathepsin G) to form the bioactive protein (reviewed in Dinarello (1997) and Fantuzzi et al. (1997)). Importantly, the matrix metalloproteinases MMP2, 3 and 9 (Schonbeck et al., 1998), and meprin B (Herzog et al., 2005), are also capable of processing proIL1B into its bioactive form. Once secreted, the propiece also has biological activity as a chemoattractant for fibroblasts (Higgins et al., 1993). While membrane-bound IL1A activity has been demonstrated in human monocytes (Bailly et al., 1990), bioactive IL1B is thought to be predominantly extracellular.

During the human menstrual cycle, IL1B mRNA is maximally expressed in the endometrium during the late secretory phase (Kauma et al., 1990, Talbi et al., 2005). At this time, immunoreactive IL1B protein is detectable in endothelial cells and unidentified cells within the stromal compartment (Simon et al., 1993). Proliferative phase endometrium shows weak IL1B immunostaining in the stroma and epithelium (Tabibzadeh and Sun, 1992). Expression of immunoreactive IL1B by decidual cells during the menstrual cycle has not previously been examined.

In first trimester human implantation sites, IL1B protein is present in the villous cytotrophoblast, syncytiotrophoblast and decidua, with activated macrophages also staining positively at each of these sites (Simon et al., 1994a). Both IL1B mRNA and protein are expressed in much greater abundance in first trimester decidua compared to non-pregnant endometrium (Kauma et al., 1990).

Using genetically deficient mouse models, it has been demonstrated that neither IL1B (Zheng et al., 1995), IL1R1 (Abbondanzo et al., 1996) nor ICE (Kuida et al., 1995, Li et al., 1995) are essential for ovulation, fertilization, implantation or parturition, but functional compensation by the structurally related cytokine IL18 has not been excluded. Simon et al. (1994b) have reported that IL1 signalling is essential for mouse implantation, but the phenotype may be strain-specific (Abbondanzo et al., 1996). There is evidence that IL1 is detrimental to pregnancy in some strains of mice (Arck et al., 1997), but mice deficient in IL1RA have reduced fertility (Hirsch, 2000, Irikura et al., 2002). Thus, the question of whether IL1B inhibits or enhances decidualization and implantation in rodents is still unclear. Exogenous recombinant human IL1B inhibits both progesterone- and cAMP-induced human decidualization in vitro (Inoue et al., 1994, Vicovac et al., 1994, Frank et al., 1995, Mizuno et al., 1999), as does IL1A (Kariya et al., 1991).

As IL1B is the most upregulated gene during IL11-enhanced human decidualization (White et al., 2005) and, given the evidence that the IL1 system participates in fetal–maternal communication during implantation (Librach et al., 1994, Simon et al., 1994a, Simon et al., 1994b), the interaction between IL11 and IL1B may have functional significance in the establishment of pregnancy. We examined therefore the effect of IL11 on IL1B mRNA and protein expression by decidualizing ESC, using real-time quantitative RT–PCR, immunostaining and quantitative immunoassays for proIL1B and bioactive IL1B.