Potential epigenetic markers of clinical diagnostics/therapeutic targets in preeclampsia

Preeclampsia (PE) is characterized by new-onset hypertension after the 20th week of gestation. This condition can be accompanied by new-onset proteinuria end endothelial dysfunction, leading to widespread end-organ injury [1]. Among hypertensive disorders of pregnancy, PE occurs in about 8% of pregnancies and is the most significant responsible for maternal and fetal deaths in the world [2].

The placenta is the central organ for pregnancy and the development of PE, and trophoblasts perform its main functions. While its pathophysiology is complex, abnormal placental development and the absence of an invasive endothelial phenotype of cytotrophoblasts lead to primary factors in PE, such as poor trophoblastic invasion and impaired vascular remodeling [3].

There has been an increasing search for effective predictive biomarkers for PE, facilitating early diagnosis, constant monitoring of high-risk cases and scheduled delivery. Prediction before the 16th week of gestation in women at high risk of developing PE has clinical utility in preventing early-onset PE (EOPE) with premature birth associated with perinatal morbidity [4]. Prophylactic administration of acetylsalicylic acid can reduce PE and premature birth onset [4]. The use of low doses of acetylsalicylic acid is recommended in gestation for the prevention of PE in high-risk pregnant women. Aspirin interacts with different pathways, which modulates the epigenetic landscape in placenta stem cells derived from PE [5]. Further prediction of women at risk of developing late-onset PE (LOPE) allows for greater surveillance and scheduled birth before the worst outcome of PE, thereby reducing maternal and fetal deaths [6].

Epigenetic mechanisms include DNA methylation (DNAm), histone modifications, and noncoding RNAs, which regulate gene expression during development and in differentiated tissues [7] and play essential roles in placenta development. Notably, changes in patterns of epigenetic markers may be causes or consequences of abnormal placental development and several other environmental factors, with implications for maternal morbidity, fetal development and future diseases in the mother and fetus [3].

Several studies have shown the roles of epigenetic mechanisms in placental development, as reviewed elsewhere [8]. DNAm is an incorporation of CH₃ at carbon 5 of cytosine [9] and is the most well-known mechanism in placental development and PE. Differences in global DNAm levels in placentas of PE have been reported, mainly in EOPE and preterm PE [10]. Moreover, 365 differentially methylated CpG loci were identified in the placenta. In comparison, maternal peripheral blood showed only 71 loci, which demonstrates an inconsistency between the DNAm profiles in peripheral blood and placenta, even in pathological cases [11]. However, DNAm analyses in white blood cells isolated from maternal peripheral blood collected in the first trimester of pregnancy have 75% concordance with the results of analyses in the placental chorionic tissue [12].

The search for biomarkers is based on the analysis of the target tissue and can be through candidate genes [11,12] or global analysis [10]. However, since accessing the placenta during pregnancy is invasive, this change must be detected at a peripheral level. Cell-free DNAm showed differences between control and pregnancies with PE at around 12 weeks of gestation and predicted 72% of patients with EOPE at 80% specificity [13].

Peripheral blood, serum, and urine from pregnant women with PE showed an increase in fetal and total cell-free DNA in any type of PE, and this is associated with the severity of the disease and an adverse maternal-fetal outcome [14]. Former studies have the limitation of evaluating fetal cell-free DNA based only on genomic sequences of male DNA; this limitation has already been overcome with new methodologies, which can identify total, fetal, and mitochondrial cell-free DNA. The data indicate the participation of cell-free DNA in the pathophysiology of PE. However, the exact mechanisms, and whether this increase in cell-free DNA, is a cause or consequence of the mechanisms that trigger PE are unknown. Furthermore, data heterogeneity highlights the importance of methodological and longitudinal standardization for these analyses [14].

The TIMP3 gene promoter showed hypomethylation, with an increased TIMP-3 expression in corresponding placentas from different types of PE compared with controls, including between EOPE and LOPE [15]. These findings indicate the role of DNAm at the TIMP3 promoter as an epigenetic mechanism in PE. The invasion of trophoblastic cells and, consequently, the placentation process requires a balance between the levels of MMPs and their inhibitory TIMPs. Additionally, TIMP3 acts as an antiangiogenic factor by blocking VEGF, thus participating in early and late PE development [15].

Interestingly, TIMP-3 concentrations in plasma were increased in pregnant with PE compared with control, and these concentrations were positively correlated with circulating concentrations of MMP-2 and TIMP-1 in PE [16]. Indeed, the deregulation in placental expression of MMPs and TIMPs and their circulating levels was demonstrated in hypertensive disorders of pregnancy [16,17]. These data demonstrate that TIMP3 is a promising gene candidate for longitudinal clinical studies in cohorts of pregnant women at risk of developing PE.

MicroRNAs (miRNAs) are stable in plasma and have been explored as disease biomarkers. However, identifying biomarkers that are a good predictor for identifying women at high risk of developing PE was initially challenged by issues regarding sample handling and study design, which have hampered robust analysis of miRNAs as biomarkers for predicting PE [18]. It was claimed that standard methodological approaches needed to be established to make progress toward the clinical application of miRNAs. Systematic review and meta-analysis concluded that circulating miRNAs can be a non-invasive biomarker with high potential and accuracy for diagnosing and predicting PE [19]. However, prospective and longitudinal studies must be conducted to understand these data better.

Seven circulating miRNAs were suggested as potential biomarkers of pregnancy complications, including PE [20]. These miRNAs were increased in the first trimester of gestation and participate in the pathophysiology of PE in different ways, such as, for example, the miR-125b affects the trophoblastic invasion, reduces the migration and function of endothelial cells and membrane markers related to cell adhesion, proliferation and growth is the target of this miRNA [20]. Expression profile of circulating miRNAs in plasma obtained between 20 and 25th week of gestation of pregnant women who then developed PE showed 23 miRNAs upregulated; the miR-204-5p interact with genes MMP-9, TGFBR2 and SIRT1 that participates in the regulation of endothelial function, arterial remodeling, cell migration and vascular aging [21].

Exosomes are extracellular vesicles that may contain genetic content, proteins, lipids, and metabolites [22]. An extensive review of exosomes and their use as biomarkers and therapeutics showed that both strategies have limitations, such as the difficulty in measuring due to their low quantity and determining the cell type that generated the exosome. Placenta-derived exosomes are involved in cellular communication, and their content is related to the physiological condition of the cell that produces it [22]. Furthermore, it can be detected in peripheral blood from the sixth week of gestation, increases throughout pregnancy, and increases even further before and during pathological conditions of pregnancy, hence the interest in its use as a potential biomarker for early diagnosis [23].

The exosomes demonstrated a potential for diagnosis and differentiation of EOPE and LOPE. The groups have unique exomiRNA associated with biological pathways that characterize each group [23]. These exosomes can express immunoregulatory markers, which, for example, block the T cell response, helping the appropriate recognition of the fetus and thus regulating inflammatory pathways. In the last trimester of pregnancy, these are already involved in metabolic pathways. exomiRNA has been identified in pregnant women with PE, such as miR-153 and miR-325-3p, which are related to endothelial cell dysfunction [23].

Clinical guidelines, first-trimester combined algorithms, sFlt1:PIGF ratio, and PIGF alone testing are used as PE prediction tests. These are not applied to all pregnant women; they have varying sensitivity and predictive value depending on the type of PE and generally have a good negative predictive value but a low positive predictive value [24]. Using biomarkers based on epigenetic changes related to target organs or key processes of PE in a multi-omics approach combined with existing biomarkers at the appropriate time of pregnancy can increase the sensitivity and predictive value of the tests. Thus, reducing hospitalizations and comorbidities related to the mother and fetus.

In conclusion, epigenetic mechanisms play a role in the physiopathology of PE (Supplementary Figure S1). Most of these epigenetic mechanisms are already used as biomarkers or therapies in different types of cancer, which is a pathophysiological process with similar biological pathways due to characteristics such as cell proliferation, migration, and invasion. Performing predictive and non-invasive analyses at different gestational ages is difficult in PE. Several studies have analyzed these mechanisms in the past decades, mainly DNAm in the placenta, umbilical cord blood and other tissues; however, these data are inconsistent.

An ideal longitudinal study design could include the collection of peripheral blood during the first or second trimester of gestation, which can facilitate early detection and interventions in pregnant women with risk criteria for developing PE. Next, the results should undergo thorough statistical analyses to identify sites or genes with potential clinical utility as biomarkers with high accuracy and specificity, and longitudinal studies for validation can be carried out in different cohorts so that they can be translated into clinical practices.

Supplemental material

Supplemental data for this article can be accessed at https://doi.org/10.1080/17501911.2024.2383558

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The authors have no financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.

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