ABSTRACT
BACKGROUND: Controlled ovarian stimulation (COS) with urinary gonadotrophins but not recombinant gonadotrophins, adversely affect the implantation process. In this study, we investigated the effects of urinary and recombinant gonadotrophins on gene expression profiles at implantation sites during the mouse peri-implantation period and the possible molecular mechanisms involved in the detrimental effects of urinary gonadotrophins using microarray technology. METHODS: Adult female CD1 mice were treated with (i) urinary human FSH (hFSH) and urinary HCG, (ii) recombinant hFSH and recombinant human LH or (iii) saline. Gene expression profiling with GeneChip mouse genome 430 2.0 arrays, containing 45 101 probe sets, was performed using implantation sites on embryonic day 5. Data were statistically analysed using Significance Analysis of Microarrays. Ten genes from the microarray analysis were selected for validation using quantitative RT-PCR (qRT-PCR). A parallel group of pregnant mice was allowed to give birth to study the effect of gonadotrophins on resorption. RESULTS: Urinary gonadotrophins differentially up-regulated the expression of 30 genes, increased resorption and reduced litter size, whereas recombinant gonadotrophins did not. Nine of the 10 genes were confirmed by qRT-PCR. CONCLUSIONS: Urinary gonadotrophins, but not recombinant gonadotrophins, exerted differential effects on gene expression during the murine peri-implantation period. These findings might contribute to improve protocols for COS, leading to higher successful pregnancy rates.
Subject(s)
Chorionic Gonadotropin/pharmacology , Chorionic Gonadotropin/urine , Embryo Implantation/physiology , Follicle Stimulating Hormone/pharmacology , Follicle Stimulating Hormone/urine , Gonadotropins/urine , Luteinizing Hormone/pharmacology , Recombinant Proteins/pharmacology , Animals , Female , Fetal Resorption/chemically induced , Gene Expression Profiling , Gene Expression Regulation, Developmental , Genes, MHC Class II/physiology , Humans , Mice , Oligonucleotide Array Sequence Analysis , Ovulation Induction , Pregnancy , Reverse Transcriptase Polymerase Chain Reaction , Up-RegulationABSTRACT
BACKGROUND: Ovarian stimulation in humans might affect the perinatal outcome and be considered as a stress factor in the implantation process. In this study we compared the effects of recombinant and urinary gonadotrophins during the mouse peri-implantation period. METHODS: Adult female CD1 mice were treated as follows (a) urinary hFSH and urinary hCG, (b) recombinant hFSH and recombinant hLH and (c) saline. The effects of the gonadotrophins on the expression of vascular endothelial growth factor120 (VEG120) and its receptors and the corticotrophin releasing hormone (CRH) system during the peri-implantation period were studied. The specific effects of the different gonadotrophins on the onset of implantation were also studied. RESULTS: Urinary gonadotrophin treatment caused lower levels of VEGF120, flt-1 and flk-1 mRNA levels, reduced the size of the embryo implantation site, delayed implantation and prolonged the gestational period. Both urinary hFSH and urinary hCG contributed to the adverse effects. Levels of CRH and CRHR1 expression were not influenced. Recombinant gonadotrophin treatment did not alter any of the parameters studied. CONCLUSIONS: Our results show that the VEGF system of the mouse uterus during the peri-implantation period is adversely affected by urinary gonadotrophins but not by recombinant gonadotrophins. The CRH system was not affected by the two types of gonadotrophins.
Subject(s)
Embryo Implantation , Gonadotropins/pharmacology , Gonadotropins/urine , Proteins/antagonists & inhibitors , Uterus/metabolism , Vascular Endothelial Growth Factor A/antagonists & inhibitors , Vascular Endothelial Growth Factor Receptor-2/antagonists & inhibitors , Animals , Chorionic Gonadotropin/pharmacology , Chorionic Gonadotropin/urine , Extracellular Matrix Proteins , Female , Follicle Stimulating Hormone/pharmacology , Follicle Stimulating Hormone/urine , Luteinizing Hormone/pharmacology , Mice , Mice, Inbred Strains , Myosin Heavy Chains , Nonmuscle Myosin Type IIB , Recombinant Proteins/pharmacology , Uterus/drug effects , Vascular Endothelial Growth Factor Receptor-1ABSTRACT
The aim of the present study was to establish whether estrogen and corticosteroids exert effects on vascular endothelial growth factor (VEGF)(164) expression in the hippocampus and nucleus paraventricularis of the hypothalamus by in situ hybridization. Female mice were ovariectomized and treated either with estradiol benzoate or vehicle and male mice were either adrenalectomized or sham-operated. Ovariectomy plus estrogen reduced VEGF(164) expression in the nucleus paraventricularis but not in the hippocampus. Adrenalectomy did not influence VEGF(164) mRNA levels in the hippocampus and nucleus paraventricularis. Our results show for the first time an inhibitory effect of estrogen on VEGF(164) expression in the nucleus paraventricularis and suggest a role for estrogen in the regulation of VEGF(164) expression and function in the central nervous system.
Subject(s)
Estradiol/analogs & derivatives , Estrogens/pharmacology , Paraventricular Hypothalamic Nucleus/drug effects , Receptors, Vascular Endothelial Growth Factor/metabolism , Adrenalectomy , Animals , Autoradiography , Corticosterone/metabolism , Estradiol/pharmacology , Female , Hippocampus/metabolism , In Situ Hybridization , Male , Mice , Mice, Inbred C57BL , Ovariectomy , Paraventricular Hypothalamic Nucleus/metabolism , RNA, Messenger/metabolism , Radioimmunoassay , Receptors, Vascular Endothelial Growth Factor/classification , Receptors, Vascular Endothelial Growth Factor/drug effects , Receptors, Vascular Endothelial Growth Factor/genetics , Tissue DistributionABSTRACT
BACKGROUND: Ovarian stimulation by gonadotrophin treatment exerts negative effects on implantation and embryonic development. We investigated whether gonadotrophin treatment affects VEGF(120) mRNA expression during the peri-implantation period. METHODS: Two groups of adult female CD1 mice were used: the hormone-treated group was injected i.p. with urinary human FSH (5 IU in 0.1 ml saline) and urinary HCG (5 IU in 0.1 ml saline). Spontaneously ovulating mice served as controls and received saline injections. The pregnant mice were killed on embryonic development (ED) days 0, 3, 4, 5 and 6 (day of vaginal plug detection is considered as ED0). The uteri with the implanted embryos were processed for in-situ hybridization for VEGF(120). A separate group of control and hormone-treated pregnant mice were allowed to give birth. Litter size, birthweight and length of gestational period were noted. RESULTS: Gonadotrophin treatment decreased VEGF(120) mRNA levels, delayed implantation, reduced the size of the embryo implantation site on ED5 and ED6 and prolonged the gestational period. CONCLUSIONS: Gonadotrophin treatment reduces VEGF(120) expression which may have serious consequences for normal embryonic development. The present data cannot establish whether this effect is a cause or consequence of delayed implantation.
