Abstract
BackgroundExtensive uterine adaptations, including angiogenesis, occur prior to implantation in early pregnancy and are potentially regulated by the peptide hormone relaxin. This was investigated in two studies. First, we took a microarray approach using human endometrial stromal (HES) cells treated with relaxin in vitro to screen for target genes. Then we aimed to investigate whether or not relaxin deficiency in mice affected uterine expression of representative genes associated with angiogenesis and uterine remodeling, and also blood vessel proliferation in the pre-implantation mouse endometrium.MethodsNormal HES cells were isolated and treated with recombinant human relaxin (10 ng/ml) for 24 h before microarray analysis. Reverse transcriptase PCR was used to analyze gene expression of relaxin and its receptor (Rxfp1) in ovaries and uteri; quantitative PCR was used to analyze steroid receptor, angiogenesis and extracellular matrix remodeling genes in the uteri of wild type (Rln+/+) and Rln-/- mice on days 1-4 of pregnancy. Immunohistochemistry localized endometrial endothelial cell proliferation and mass spectrometry measured steroid hormones in the plasma.ResultsMicroarray analysis identified 63 well-characterized genes that were differentially regulated in HES cells after relaxin treatment. Expression of some of these genes was increased in the uterus of Rln+/+ mice by day 4 of pregnancy. There was significantly higher vascular endothelial growth factor A (VegfA), estrogen receptor 1 (Esr1), progesterone receptor (Pgr), Rxfp1, egl-9 family hypoxia-inducible factor 1 (Egln1), hypoxia inducible factor 1 alpha (Hif1α), matrix metalloproteinase 14 (Mmp14) and ankryn repeat domain 37 (Ankrd37) in Rln-/- compared to Rln+/+ mice on day 1. Progesterone receptor expression and plasma progesterone levels were higher in Rln-/- mice compared to Rln+/+ mice. However, endometrial angiogenesis was not advanced as pre-implantation endothelial cell proliferation did not differ between genotypes.ConclusionsRelaxin treatment modulates expression of a variety of angiogenesis-related genes in HES cells. However, despite accelerated uterine gene expression of steroid receptor, progesterone and angiogenesis and extracellular matrix remodeling genes in Rln-/- mice, there was no impact on angiogenesis. We conclude that although relaxin deficiency results in phenotypic changes in the pre-implantation uterus, endogenous relaxin does not play a major role in pre-implantation angiogenesis in the mouse uterus.Electronic supplementary materialThe online version of this article (doi:10.1186/s12958-016-0148-y) contains supplementary material, which is available to authorized users.
Highlights
Extensive uterine adaptations, including angiogenesis, occur prior to implantation in early pregnancy and are potentially regulated by the peptide hormone relaxin
Despite accelerated uterine gene expression of steroid receptor, progesterone and angiogenesis and extracellular matrix remodeling genes in Rln-/- mice, there was no impact on angiogenesis
We conclude that relaxin deficiency results in phenotypic changes in the pre-implantation uterus, endogenous relaxin does not play a major role in pre-implantation angiogenesis in the mouse uterus
Summary
Extensive uterine adaptations, including angiogenesis, occur prior to implantation in early pregnancy and are potentially regulated by the peptide hormone relaxin. Maturation and remodeling of the endometrium and its blood vessels are primarily regulated by the ovarian steroid hormones estrogen and progesterone [5, 6]. Relaxin treatment of ovariectomized rats and rhesus monkeys primed with estrogen increases uterine weight relative to controls, and causes increased arteriole number per unit area, dilated blood vessels on the endometrial luminal surface and pronounced endothelial cell proliferation in arterioles and capillaries of the endometrium [9,10,11,12]. In the context of human physiology, women in a clinical trial for the treatment of scleroderma reported heavier or irregular menstrual bleeding if they were treated with relaxin, possibly due to increased endometrial vascularization [15, 16]
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