title: Relaxin confers cytotrophoblast protection from hypoxia-reoxygenation injury through the phosphatidylinositol 3-kinase-Akt/protein kinase B cell survival pathway.
abstract: Preeclampsia is a hypertensive syndrome that manifests after 20 wk of gestation. Contemporary understanding of the maternal-fetal interface in preeclampsia suggests a major role for placental oxidative stress resulting from ischemia-reperfusion injury. We hypothesized that the pregnancy hormone relaxin would reduce cytotrophoblast apoptosis and necrosis (aponecrosis) and, hence, the export of placental debris into the maternal circulation. If so, then relaxin might be employed as a therapeutic intervention to diminish the activation of the maternal systemic inflammatory response central to the development of clinical disease. HTR-8/SVneo cells, a model for first trimester extravillous trophoblast, were subjected to serum deprivation and hypoxia or hypoxia-reoxygenation. The cells were treated with recombinant human relaxin or vehicle and apoptosis and/or necrosis evaluated by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL), CellEvent Caspase-3/7 and SYTOX AADvanced kit, and propidium iodide staining as determined by fluorescence microscopy or flow cytometry. To interrogate mechanisms of relaxin cytoprotection, HTR-8/SVneo cells were pretreated with pharmacological inhibitors of PI3-kinase LY294004, Akt/PKB MK-2206, or DMSO vehicle. HTR-8/SVneo cell identity was first confirmed by RT-PCR. The cells expressed placental alkaline phosphatase, aromatase, and human leukocyte antigen G. In addition, the cells expressed the relaxin receptor RXFP1 as well as H1 and H2 relaxins. Serum deprivation and hypoxia increased apoptotic cell death in HTR-8/SVneo cells, which was significantly ameliorated by concurrent treatment with relaxin. Serum deprivation and hypoxia-reoxygenation increased necrotic cell death in HTR-8/SVneo cells, which was also significantly rescued by concurrent treatment with relaxin. Pretreatment with LY294002 or MK-2206, to inhibit the phosphatidylinositol 3-kinase-Akt/protein kinase B cell survival pathway, significantly blunted the cytoprotective effect of relaxin. We demonstrated trophoblast cytoprotection by intervention with supraphysiological concentrations of relaxin, a process in part mediated through the PI3-kinase-Akt/PKB cell survival pathway. These results provide further rationale for clinical investigation of relaxin as a potential therapeutic in preeclampsia.
authors.affiliation: Department of Obstetrics and Gynecology, D. H. Barron Reproductive and Perinatal Biology Research Program, University of Florida College of Medicine, Gainesville, Florida; and.
authors.forename: Oluseyi
authors.initials: O
authors.lastname: Ogunleye
authors.affiliation: Department of Physiology and Functional Genomics, D. H. Barron Reproductive and Perinatal Biology Research Program, University of Florida College of Medicine, Gainesville, Florida.
authors.forename: Bertha
authors.initials: B
authors.lastname: Campo
authors.affiliation: Department of Physiology and Functional Genomics, D. H. Barron Reproductive and Perinatal Biology Research Program, University of Florida College of Medicine, Gainesville, Florida.
authors.forename: Diana
authors.initials: D
authors.lastname: Herrera
authors.affiliation: Department of Physiology and Functional Genomics, D. H. Barron Reproductive and Perinatal Biology Research Program, University of Florida College of Medicine, Gainesville, Florida.
authors.forename: Emiel D
authors.initials: ED
authors.lastname: Post Uiterweer
authors.affiliation: Department of Obstetrics and Gynecology, D. H. Barron Reproductive and Perinatal Biology Research Program, University of Florida College of Medicine, Gainesville, Florida; and kpconrad@ufl.edu.|Department of Physiology and Functional Genomics, D. H. Barron Reproductive and Perinatal Biology Research Program, University of Florida College of Medicine, Gainesville, Florida.
authors.forename: Kirk P
authors.initials: KP
authors.lastname: Conrad
mesh_terms: D001402:B-Lymphocytes / Q000187:drug effects* / Q000473:pathology; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects*; D019610:Cytoprotection / Q000187:drug effects; D006801:Humans; D058539:Phosphatidylinositol 3-Kinase / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D012065:Relaxin / Q000008:administration & dosage* / Q000378:metabolism; D015427:Reperfusion Injury / Q000378:metabolism* / Q000517:prevention & control; D015398:Signal Transduction / Q000187:drug effects; D014327:Trophoblasts / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology
chemical_list: D012065:Relaxin; D058539:Phosphatidylinositol 3-Kinase; D051057:Proto-Oncogene Proteins c-akt
keywords: H2 relaxin; HTR-8/SVneo cell; apoptosis; hypoxia; hypoxia-reoxygenation; necrosis; preeclampsia