RESUMEN
Mutations in the LINC-HELLP non-coding RNA (HELLPAR) have been associated with familial forms of the pregnancy-specific HELLP syndrome. These mutations negatively affect extravillous trophoblast (EVT) differentiation from a proliferative to an invasive state and disturb the binding of RNA splicing complex proteins PCBP1, PCBP2, and YBX1 to LINC-HELLP. In this study, by using both in vitro and ex vivo experiments, we investigate if these proteins are involved in the regulation of EVT invasion during placentation. Additionally, we study if this regulation is due to alternative mRNA splicing. HTR-8/SVneo extravillous trophoblasts and human first trimester placental explants were used to investigate the effect of siRNA-mediated downregulation of PCBP1, PCBP2, and YBX1 genes on the differentiation of EVTs. Transwell invasion assays and proliferation assays indicated that upon knockdown of PCBP2 and, to a lesser extent, YBX1 and PCBP1, EVTs fail to differentiate toward an invasive phenotype. The same pattern was observed in placental explants where PCBP2 knockdown led to approximately 80% reduction in the number of explants showing any EVT outgrowth. Of the ones that still did show EVT outgrowth, the percentage of proliferating EVTs was significantly higher compared to explants transfected with non-targeting control siRNAs. To further investigate this effect of PCBP2 silencing on EVTs, we performed whole transcriptome sequencing (RNA-seq) on HTR-8/SVneo cells after PCBP2 knockdown. PCBP2 knockdown was found to have minimal effect on mRNA expression levels. In contrast, PCBP2 silencing led to a switch in splicing for a large number of genes with predominant functions in cellular assembly and organization, cellular function and maintenance, and cellular growth and proliferation and the cell cycle. EVTs, upon differentiation, alter their function to be able to invade the decidua of the mother by changing their cellular assembly and their proliferative activity by exiting the cell cycle. PCBP2 appears to be a paramount regulator of these differentiation mechanisms, where its disturbed binding to LINC-HELLP could contribute to dysfunctional placental development as seen in the HELLP syndrome.
RESUMEN
Human C2orf69 is an evolutionarily conserved gene whose function is unknown. Here, we report eight unrelated families from which 20 children presented with a fatal syndrome consisting of severe autoinflammation and progredient leukoencephalopathy with recurrent seizures; 12 of these subjects, whose DNA was available, segregated homozygous loss-of-function C2orf69 variants. C2ORF69 bears homology to esterase enzymes, and orthologs can be found in most eukaryotic genomes, including that of unicellular phytoplankton. We found that endogenous C2ORF69 (1) is loosely bound to mitochondria, (2) affects mitochondrial membrane potential and oxidative respiration in cultured neurons, and (3) controls the levels of the glycogen branching enzyme 1 (GBE1) consistent with a glycogen-storage-associated mitochondriopathy. We show that CRISPR-Cas9-mediated inactivation of zebrafish C2orf69 results in lethality by 8 months of age due to spontaneous epileptic seizures, which is preceded by persistent brain inflammation. Collectively, our results delineate an autoinflammatory Mendelian disorder of C2orf69 deficiency that disrupts the development/homeostasis of the immune and central nervous systems.
Asunto(s)
Encefalitis/genética , Enfermedades Mitocondriales/genética , Animales , Evolución Biológica , Sistemas CRISPR-Cas , Línea Celular , Encefalitis/mortalidad , Femenino , Genes Recesivos , Glucógeno/metabolismo , Humanos , Inflamación/genética , Masculino , Proteínas de la Membrana/genética , Enfermedades Mitocondriales/mortalidad , Linaje , Convulsiones/genética , Convulsiones/mortalidad , Pez Cebra/genéticaRESUMEN
Preeclampsia is a frequent gestational hypertensive disorder with equivocal pathophysiology. Knockout of peptide hormone ELABELA (ELA) has been shown to cause preeclampsia-like symptoms in mice. However, the role of ELA in human placentation and whether ELA is involved in the development of preeclampsia in humans is not yet known. In this study, we show that exogenous administration of ELA peptide is able to increase invasiveness of extravillous trophoblasts in vitro, is able to change outgrowth morphology and reduce trophoblast proliferation ex vivo, and that these effects are, at least in part, independent of signaling through the Apelin Receptor (APLNR). Moreover, we show that circulating levels of ELA are highly variable between women, correlate with BMI, but are significantly reduced in first trimester plasma of women with a healthy BMI later developing preeclampsia. We conclude that the large variability and BMI dependence of ELA levels in circulation make this peptide an unlikely candidate to function as a first trimester preeclampsia screening biomarker, while in the future administering ELA or a derivative might be considered as a potential preeclampsia treatment option as ELA is able to drive extravillous trophoblast differentiation.
Asunto(s)
Diferenciación Celular , Hormonas Peptídicas/sangre , Placenta/metabolismo , Trofoblastos/citología , Adulto , Apelina/sangre , Índice de Masa Corporal , Línea Celular , Proliferación Celular , Estudios de Cohortes , Femenino , Humanos , Preeclampsia/sangre , Embarazo , Primer Trimestre del Embarazo/sangre , GemelosAsunto(s)
Placentación , Preeclampsia , Biomarcadores , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , EmbarazoRESUMEN
The apelinergic-axis (Apelin, Elabela and their receptor APJ) is involved in many physiological and pathological processes. Both Elabela/APJ and Apelin/APJ are implicated in the pathophysiology of preeclampsia in rodents. However, the findings regarding the apelinergic axis in human preeclamptic placental development have been rather conflicting. In this systematic review we present an overview of the current evidence regarding the pathophysiological role of Apelin, Elabela and their receptor in human preeclamptic pregnancies. The databases used for this systematic review were Pubmed, Scopus, Google Scholar and ClinicalTrials.gov. The reference lists of the selected studies were also screened for any additional studies. The last search was performed on the 25th of March 2019. Thirteen studies were included and subjected to quality assessment so that only high quality datasets were finally selected and included in this systematic evaluation. In total, 410 women that developed preeclampsia or IUGR and 409 healthy control pregnancies were included. The findings of this review suggest that circulating Apelin levels are increased in early onset/severe preeclamptic patients while Apelin levels in severe preeclamptic placenta tissues appear to show the opposite. Circulating Elabela levels in early-onset preeclamptic women do not differ from controls, while its levels in late-onset preeclampsia remain inconclusive. The studies on Elabela and APJ expression in placental tissues require larger sample sizes with defined preeclampsia subtypes to draw any definite conclusions. Large cohort studies with affected and control groups matched for Body Mass Index and gestational age at sampling are essential in order to substantiate other current findings.