Serum miR-34a as Indicator of Impaired Fibrinolytic Capacity in Pediatric Thrombosis Through Inadequate Regulation of the ACE/PAI-1 Axis

Pediatric thrombosis (PT) represents a rare condition that can manifest from neonatal life to adolescence, encompassing life-threatening complications. Its pathogenesis is attributed to immature hemostasis in conjunction with environmental and genetic factors, predominantly including those resulting in increased levels of plasminogen activator inhibitor 1 (PAI-1), the principal inhibitor of fibrinolysis, which is subject to upstream regulation by angiotensin-converting enzyme (ACE). Although the implication of microRNAs (miRNAs), epigenetic modulators of gene expression, has been demonstrated in adult thrombosis, evidence is lacking in the pediatric setting. Here, we investigated the involvement of two miRNA regulators of PAI-1 (<i>SERPINE1</i> gene) in PT, in relation to clinical and genetic parameters that induce PAI-1 fluctuations. Following bioinformatic target-prediction, miRNA expression was assessed by quantitative real-time PCR in serum-samples of 19 pediatric patients with thrombosis (1-18 months post-incident), and 19 healthy controls. Patients were genotyped for the <i>SERPINE1</i>-4G/5G and <i>ACE</i>-I/D polymorphisms by PCR-based assays. Genotypic and thrombosis-related clinical data were analyzed in relation to miRNA-expression. Two miRNAs (miR-145-5p, miR-34a-5p) were identified to target <i>SERPINE1</i> mRNA, with miR-34a additionally targeting the mRNA of <i>ACE</i>. The expression of miR-34a was significantly decreased in patients compared to controls (<i>p</i> = 0.029), while no difference was observed in miR-145 expression. Within patients, miR-34a expression demonstrated a peak 1-3 months post-thrombosis and was diminished upon treatment completion (<i>p</i> = 0.031), followed by a slight long-term increase. MiR-34a levels differed significantly by thrombosis site (<i>p</i> = 0.019), while a significant synergistic interaction between site and onset type (provoked/unprovoked) was detected (<i>p</i> = 0.016). Finally, an epistatic modification was observed in cerebral cases, since double homozygosity (4G/4G + D/D) led to a miR-34 decrease, with D/D carriership reversing the 4G/4G-induced upregulation of miR-34a (<i>p</i> = 0.006). Our findings suggest that in pediatric thrombosis, downregulation of miR-34a is indicative of impaired fibrinolytic capacity, attributed to deficient regulation of the inhibitory ACE/PAI-1 axis.