MicroRNA dysregulation in lung injury: the role of the miR-26a/EphA2 axis in regulation of endothelial permeability

Abstract

MicroRNAs (miRNAs) are noncoding RNAs that regulate gene expression in many diseases, although the contribution of miRNAs to the pathophysiology of lung injury remains obscure. We hypothesized that dysregulation of miRNA expression drives the changes in key genes implicated in the development of lung injury. To test our hypothesis, we utilized a model of lung injury induced early after administration of intratracheal bleomycin (0.1 U). Wild-type mice were treated with bleomycin or PBS, and lungs were collected at 4 or 7 days. A profile of lung miRNA was determined by miRNA array and confirmed by quantitative PCR and flow cytometry. Lung miR-26a was significantly decreased 7 days after bleomycin injury, and, on the basis of enrichment of predicted gene targets, it was identified as a putative regulator of cell adhesion, including the gene targets EphA2, KDR, and ROCK1, important in altered barrier function. Lung EphA2 mRNA, and protein increased in the bleomycin-injured lung. We further explored the miR-26a/EphA2 axis in vitro using human lung microvascular endothelial cells (HMVEC-L). Cells were transfected with miR-26a mimic and inhibitor, and expression of gene targets and permeability was measured. miR-26a regulated expression of EphA2 but not KDR or ROCK1. Additionally, miR-26a inhibition increased HMVEC-L permeability, and the disrupted barrier integrity due to miR-26a was blocked by EphA2 knockdown, shown by VE-cadherin staining. Our data suggest that miR-26a is an important epigenetic regulator of EphA2 expression in the pulmonary endothelium. As such, miR-26a may represent a novel therapeutic target in lung injury by mitigating EphA2-mediated changes in permeability.

Ephs and Ephrins in Adult Endothelial Biology

“…In a study of Miao et al, it was shown that VEGF activates the intracellular PI3K/Akt and Erk1/2 signaling pathways resulting in the increased expression of EphA2 which then, in turn, contributes to an increase in paracellular permeability [38]. The role of (ephrinA1-induced) EphA2 forward signaling in reducing the endothelial barrier function has also been shown in several other in vitro and in vivo studies, where the increased expression of EphA2 increases vascular permeability, and reduced expression decreases (ephrinA1-induced) vascular permeability [28,37,39,75]. Opposite to the increase in vascular permeability upon EphA2 forward signaling, the exposure of endothelial cells to EphA4 recombinant protein could protect the endothelial barrier against TNFα-induced vascular leakage.…”

“…The regulation of ephrins by miRNAs has been described for the ephrinA3 ligand, where increased levels of miR-210 repressed the protein expression of ephrinA3, and decreased levels of miR-210 increased ephrinA3 expression [16][17][18]. Expression of the EphA2 receptor was shown to be regulated by miR-26a, where a mimic of miR-26a decreased EphA2 expression, and the use of a miR-26a inhibitor increased EphA2 expression [28]. In addition, ephrinB2 and EphB4 are described to be potentially regulated by miR-20b [27], EphB2 and EphB4 by miR-520h [30] and EphA7 is described as a direct target of miR-137 [29].…”

Extracellular vesicles released from p18 overexpressing pulmonary endothelial cells are barrier protective – potential implications for acute respiratory distress syndrome

“…We next sought to understand the potential mechanism through which EDEVs isolated from LMVEC overexpressing p18 wt could protect the pulmonary endothelium. A range of studies have demonstrated the key role of miRNAs in regulating endothelial barrier function 27 – 29 and the potential for EDEVs to act as carriers for these miRNA. 9 Our next experiments therefore studied the profile of miRNA present within EDEVs isolated from cells overexpressing p18 wt or GFP control.…”

Platelet activation in experimental murine neonatal pulmonary hypertension

“…Interstitial platelets were defined for positive staining for CD41‐APC and CD42b‐FITC but negative staining for CD41‐BV421 (CD41‐APC Hi , CD42b‐FITC Hi , CD41‐BV421 Lo ). Quantification was performed using 123eCount beads (Thermofisher) as previously described (Good et al, 2018).…”