Increasing the levels of a tiny RNA molecule known as microRNA-483 enhanced the function of endothelial cells and eased the symptoms of pulmonary arterial hypertension (PAH) in rat models of the disease, a new study shows.
The results support the potential of microRNA-483 as a potential therapy for PAH.
The study, “MicroRNA-483 amelioration of experimental pulmonary hypertension,” was published in the journal EMBO Molecular Medicine.
The improper functioning of endothelial cells — those lining the inner wall of blood vessels – is characteristic of PAH progression. Compromised endothelial cells produce higher-than-normal levels of pro-inflammatory molecules called cytokines, and of molecules that constrict blood vessels (vasoconstrictors), while lowering those that open (dilate) blood vessels, called vasodilators. This results in increased blood pressure in the lung’s vascular system.
There are medicines that can target impaired endothelial cells, but their efficacy is limited.
MicroRNAs are a special class of short RNA molecules capable of regulating gene activity. They have been increasingly recognized as potential therapeutic targets.
One such short RNA molecule is MicroRNA-483 (miR-483), which contains two strands, called miR-483-3p and miR- 483-5p. In a previous study, researchers at the Xi’an Jiaotong University Health Science Center, in China, observed that miR-483 enhanced the function of endothelial cells by promoting an anti-scarring (fibrotic) effect.
Now, the team investigated whether miR-483 plays a role in PAH.
The scientists first assessed whether the levels of miR-483 in the blood differed between animals with PAH and healthy ones (control group). Compared with the controls, those with PAH had significantly lower levels of the two miR strands that compose miR-483 — especially those with more severe disease.
Levels of miR-483-3p below 27% were able to identify PAH with a sensitivity of 88.4% and a specificity of 56.8%, the results suggested. Meanwhile, levels of miR-483-5p below 26% did the same with a sensitivity of 82.1% and a specificity of 48.9%. Of note, a test’s sensitivity is its ability to correctly identify those with a given disease. Specificity refers to correctly identifying those without the disease.
After dividing the PAH animals according to their mortality risk, the researchers then saw that those with an intermediate or high risk had lower miR-483-3p levels than those with a low mortality risk.
To assess if these lower miR483 levels were in fact associated with endothelial cells, the team isolated extracellular vesicles, called EVs, from the blood of both those with PAH and healthy controls. To identify EVs originating from endothelial cells they used a marker called CD144.
The results showed that EVs from the PAH animals had lower levels of miR-483 compared with healthy controls. Of note, EVs are an important mode of intercellular communication, and are known for their capacity to carry proteins and DNA and RNA molecules, including microRNAs. They are increasingly recognized as potential biomarkers for diagnosis.
“The lower levels of miR‐483 found in circulation and CD144‐enriched EVs of IPAH patients suggested that the expression of miR‐483‐3p/‐5p might affect genes and pathways involved in PAH,” the researchers wrote.
To better understand the role of miR-483 in endothelial cells, the researchers then increased the levels of this microRNA in human pulmonary arterial endothelial cells grown in the lab.
Those results showed that cells altered their behavior, with changes in several pathways, including a lower activity of processes related to cell adhesion, programmed cell death (apoptosis), metabolism, inflammation, migration, and proliferation.
A bioinformatic analysis identified several of the molecular players targeted by miR-483-3p and miR-483-5p. All of them played a role in the altered pathways that had been identified.
Next, the researchers developed a genetic rat model, named EC-miR-483-Tg, in which endothelial cells had stable levels of miR-483. The compound monocrotaline (MCT) was used to induced PAH in this rat model and also in control animals.
After MCT administration, the levels of PAH-related genes were higher in control animals than in EC-miR-483-Tg engineered rats, the results showed.
At a physiological level, the mean pulmonary artery pressure (mPAP) and Fulton index (a measure of heart enlargement) were alleviated in the engineered group of rats treated with MCT. Moreover, the rats had less thickened blood vessels and less permeable endothelium compared with control rats.
Another animal model was then tested, in which Sugen 5416 (SU5416, a blocker of endothelial growth factor receptor) and chronic hypoxia were used to induce PAH. Compared with controls, this rat model also had a reduced increase of mPAP and Fulton index, the researchers found.
The researchers then tested whether the administration of miR-483 could ease the symptoms of PAH. They delivered miR-483 using harmless fluorescent viruses administered directly into the trachea. Because the viruses were fluorescent, the researchers were able to confirm that the miR was successfully delivered to the lungs.
PAH induction using MCT in these rats showed that this miR-483 therapy suppressed PAH-related genes, reducing the effect of the disease, compared with controls.
“Overexpression of miR‐483 in endothelial cells (EC s) inhibits inflammatory and fibrogenic responses and leads to ameliorated pulmonary hypertension phenotypes in rats,” the researchers wrote.
In addition, miR-483 reduced heart enlargement and the thickness and blockage of blood vessels. It also improved the survival of MCT-treated rats.
Taken together, the findings suggested that miR-483 constitutes both an useful biomarker and a therapeutic target for PAH.
“Our findings herein demonstrate the potential use of … administered miR-483 or agents that elevate endogenous [internal] miR-483 to maintain functional endothelium, at least during the early stages of PAH,” the team concluded.
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