Human relaxins (RLNH1, RLNH2), their receptor (RXFP1) and fetoplacental growth.

Relaxin, a systemic and placental hormone, has potential roles in fetoplacental growth. Human placenta expresses two RLN genes, RLNH1 and RLNH2 Maternal obesity is common and is associated with abnormal fetal growth. Our aims were to relate systemic and cord blood RLNH2, placental RLNs and their receptor (RXFP1) with fetoplacental growth in context of maternal body mass index, and associations with insulin-like growth factor 2 (IGF2) and vascular endothelial growth factor A (VEGFA) in the same placentas. Systemic, cord blood and placental samples were collected prior to term labor, divided by prepregnancy body mass index: underweight/normal (N = 25) and overweight/obese (N = 44). Blood RLNH2 was measured by ELISA; placental RLNH2, RLNH1, RXFP1, IGF2 and VEGFA were measured by quantitative immunohistochemistry and mRNAs were measured by quantitative reverse transcription PCR. Birthweight increased with systemic RLNH2 only in underweight/normal women (P = 0.036). Syncytiotrophoblast RLNH2 was increased in overweight/obese patients (P = 0.017) and was associated with placental weight in all subjects (P = 0.038). RLNH1 had no associations with birthweight or placental weight, but was associated with increased trophoblast and endothelial IGF2 and VEGFA, due to female fetal sex. Thus, while systemic RLNH2 may be involved in birthweight regulation in underweight/normal women, placental RLNH2 in all subjects may be involved in placental weight. A strong association of trophoblast IGF2 with birthweight and placental weight in overweight/obese women suggests its importance. However, an association of only RLNH1 with placental IGF2 and VEGFA was dependent upon female fetal sex. These results suggest that both systemic and placental RLNs may be associated with fetoplacental growth.

Relaxin, its receptor (RXFP1), and insulin-like peptide 4 expression through gestation and in placenta accreta.

This study was designed to show whether placental relaxin (RLN), its receptor (RXFP1), or insulin-like peptide 4 (INSL4) might have altered expression in patients with placenta accreta. The baseline expression of their genes through gestation (n = 34) was quantitated in the placental basal plate (BP) and villous trophoblast (TR), and compared to their expression in placenta accreta (n = 6). The proteins were also immunolocalized and quantitated in the accreta tissues. The messenger RNAs (mRNAs) of matrix metalloproteinase 9, -2, and tissue inhibitors of matrix metalloproteinase (TIMP)-1 were also measured. Results demonstrated that the BP and TR expressed low levels of RLN/RXFP1 and INSL4 through gestation. In accreta, increased RLN gene and protein in BP were associated with antepartum bleeding whereas INSL4 expression decreased throughout the TR. There were no changes in mRNAs for MMPs, but TIMP-1 was increased only in the invasive TR.

Identification of relaxin-responsive cells in the human choriodecidua at term.

The decidua of the human maternal-fetal interface is a local source of intrauterine relaxin, and the choriodecidua expresses its receptor (LGR7). Since these tissues consist of a variety of cells, we sought to identify the primary cell(s) responsible for LGR7 expression and relaxin responsiveness.

Relaxin and the human fetal membranes.

The human fetal membranes are complex tissues that perform many important functions during gestation. The extracellular matrix provides their strength to withstand the forces directed from the fetus and myometrium. Relaxin is a collagenolytic hormone that causes increased production of the matrix metalloproteinases. Its expression from the decidua is increased in patients with preterm premature rupture of the membranes, and its leucine-rich G receptor 7 is upregulated at preterm. The authors previously showed that relaxin is not involved in the infection-mediated cytokine response, but in the absence of infection, it causes increased secretion of both interleukin -6 and interleukin-8 from the membranes. In this article, the authors propose that relaxin is one of a number of sterile stimuli capable of causing increased proinflammatory cytokines, similar to but less robust than the effects of infection. These probably represent distinct inflammatory pathways involving different intracellular signaling events, which can result in either preterm premature rupture of the membranes or preterm labor. The current challenge is to fully understand these pathways and to clarify their similarities and differences.

Human decidual relaxin and preterm birth.

Relaxin in human pregnancy is both a systemic hormone from the corpus luteum and an autocrine/paracrine hormone at the maternal-fetal interface formed by the decidua/placenta and fetal membranes. We have focused our studies on the autocrine/paracrine roles of relaxin, especially in the preterm premature rupture of the fetal membranes, which causes 30-40% of preterm births. By using different techniques and different tissue collections, our laboratory has shown that expression of the relaxin genes and proteins in the decidua and placenta is increased in patients with preterm premature rupture of the fetal membranes. Relaxin binding and the expression of LGR7 are primarily in the chorion and decidua and are downregulated after spontaneous labor and delivery both at term and preterm. However, expression of LGR7 in the fetal membranes is significantly greater in all clinical situations at preterm than term, suggesting an important role for relaxin in these tissues at that time. The roles of the relaxin system in three potential causes of preterm birth are discussed: in the growth and proliferation of the membranes important for fetal membrane accommodation to fetal and placental growth, in acute infection, and in the inflammatory response leading to the initiation of labor.

Relaxin causes proliferation of human amniotic epithelium by stimulation of insulin-like growth factor-II.

OBJECTIVE: The study was conducted to determine whether relaxin has a proliferative effect on amniotic epithelial cells and to show that this effect is caused by its stimulation of the insulin-like growth factor-II (IGF-II) gene. STUDY DESIGN: Immunolocalization and Northern analysis were used to confirm the expression of IGF-II by the fetal cells in the membranes. Human amniotic epithelial (WISH) cells were treated with doses of IGF-II or human relaxin and their proliferative effects measured. The mechanism of the effect of relaxin on cellular proliferation was studied with the use of an IGF-II-blocking antibody and Northern analysis for IGF-II gene expression after treatment with relaxin. An in vivo correlate was sought by quantitation of relaxin gene expression in 10 fetal membranes from women with normally grown and large for gestational age infants. RESULTS: The amniotic epithelial and cytotrophoblast cells of the fetal membranes expressed IGF-II, as did the amniotic epithelial-like (WISH) cell line. Treatment of WISH cells with IGF-II or relaxin caused a significant (P <.03) and dose-related increase in WISH cell proliferation over 5 days. The concurrent treatment with a blocking antibody to IGF-II significantly decreased the proliferative response to IGF-II (P <.002) and relaxin (P <.002). Treatment with relaxin caused a significant increase (P <.003) in the transcription of IGF-II in 24 hours. In fetal membranes, the levels of relaxin gene expression correlated with fetal membrane surface area (r = 0.76) and was significantly greater (P <.008) in the membranes from macrosomic infants (4020-4729 g) compared with those normally grown (2855-3830 g). CONCLUSION: IGF-II and relaxin both caused the proliferation of WISH cells. Concurrent treatment with an IGF-II-blocking antibody abrogated the proliferative effects of both hormones. Relaxin increased the transcription of IGF-II, and its expression levels in the fetal membranes correlated with the membrane surface area as well as neonatal birth weight. These data suggest that relaxin is a growth factor for the fetal membranes.

Decidual relaxins: gene and protein up-regulation in preterm premature rupture of the membranes by complementary DNA arrays and quantitative immunocytochemistry.

OBJECTIVE: This study was undertaken to show both decidual relaxin gene and protein up-regulation in preterm premature rupture of the fetal membranes. STUDY DESIGN: Membranes after preterm premature rupture (n = 4) have been matched in pairs with preterm intact membranes (n = 4). These tissues were from patients without infection, labor, preeclampsia or intrauterine growth restriction, and none of the patients had a latency period of more than 8 hours. The messenger RNAs from these tissues were used on complementary DNA expression arrays; 488 genes were analyzed. Relaxin gene expression was quantitated from the arrays and in additional tissues by Northern analysis. The two relaxin proteins, H1 and H2, in the decidual cells were immunolocalized and quantitated by microdensitometry with the use of specific antisera that were raised to decapeptides over the region of least homologic features. The expression of five other genes that were selected from the arrays were quantitated by Northern analysis. RESULTS: Relaxin gene expression was up-regulated 3.4-fold on the complementary DNA arrays but was not confirmed on Northern analysis. On the other hand, protein analysis for relaxin H1 and H2 in the decidual cells showed them to be significantly up-regulated (P <.0001, for both proteins) in patients with preterm premature rupture of the membranes compared with control subjects. The 20 most highly expressed genes at preterm in tissues without rupture were determined. In addition, analysis of the genes that were up-regulated with preterm rupture of the membranes showed 30 differentially expressed genes. CONCLUSION: Relaxin gene expression in the decidua is up-regulated, and its protein expression is significantly increased with preterm rupture of the fetal membranes.