MicroRNA roles in brain development and stroke implications

MicroRNAs (miRNAs) are formed by a group of nonprotein coding genes that are present in all living organisms. MicroRNAs are involved in multiple biological reactions such as cell growth, cell death, tissue differentiation, and embryonic development.45 Variations in miRNA sequences affect miRNA regulation and have been associated with many human disorders.46 It has also been found that miRNAs work as gene regulators in cerebrovascular diseases.47 Due to their unique structures and functions, it has been proposed that miRNAs might be used as biomarkers in certain human diseases.48 For instance, miRNAs play a key role in regulatory processes of leukocyte gene expression in AIS.49 It has been shown that miRNA alterations following central nervous system injury stimulate neuronal cell death mediated by inflammation and oxidative stress.50 Specific miRNAs have been shown in experimental studies to correlate with specific findings. For example, miRNA-17-92 was found to have a role in regulating the genes for T-box protein 2, phosphatase homolog and tensin homolog. These genes are known to play a role in radial glial cells.51 It was found that miRNA-17-92 was upregulated in mice after stroke, which boosted the proliferation of neuronal progenitor cells.52 In addition, the miRNA-124 cluster has been shown to have a role in the differentiation of neuronal progenitor cells. For instance, knockout mice of miRNA-124 caused reduction in brain size and anatomical abnormalities.53 Hence, following an AIS, high levels of miRNA-124 were attributed to different cellular processes, such as inflammation, edema, cell death, and neurogenesis.52, 54, 55 Lalwani et al. found out that miRNA-142-3p represses vascular endothelial cahedrins in zebrafish and as a result mediates vascular integrity. Increased levels of miRNA-142-3p were correlated with vascular hemorrhage, whereas low levels caused abnormal vascular remodeling.56 Furthermore, Liu et al. found that miRNA-142-3p was elevated in rodents following ischemic stroke, which indicates its role in secular remodeling.52 Similarly, miRNA-126 targets vascular endothelial growth factor (VEGF). Thus, it mediates vascular development.57-59 In rodents, miRNA-126 inhibits VEGF and affects retinal neovascularization post-ischemic stroke.60, 61 Moreover, in humans, elevated levels of miRNA-126 were established as a biomarker for AIS.62

MicroRNA involvement in stroke formation and miRNAs as diagnostic tools for AIS

Atherosclerosis, diabetes, and hypertension are comorbidities mostly associated with stroke.63, 64 It has been reported that hypertension is the number one risk factor for stroke formation due to its effects on vessel elasticity, making them easy to rupture, which leads to hemorrhagic stroke.65 In one of the studies carried out on rats with hypertension, it was shown the levels of miRNA-155 were reduced in these rats.66 In addition, it was observed that miRNA-155 plays a role in vessel relaxation as it targets nitric oxide synthase and angiotensin II receptors.54, 67 Hence, miRNA-155 has a crucial role in stroke formation as it modulates blood pressure. Moreover, miRNA-22 has been found to target chromogranin A, leading to a boost in catestatin, which regulates blood pressure.68-70 In animal studies, rats that were treated with miRNA-22 antagonist had a decrease in blood pressure.71 In a comparison of hypertensive and normotensive rats, a sequence of 24 miRNAs has been shown to be expressed in the brainstem of hypertensive rats.72 Similarly, a sequence of 30 miRNAs were found to be upregulated in human endothelium of vasculature that was thought to play a role in hypertension.73

Alterations in miRNAs were found to occur in atherosclerotic vessel walls and serum, implying their involvement in atherosclerosis formation and progression.74 For instance, miRNA-155 was found to play a role in the inflammatory processes that accompany atherogenesis by targeting pro-inflammatory transcription factors such as Ets1 and AT1R, thus being atheroprotective.75 In mouse studies, knockdown of miRNA-155 led to Fas apoptosis protein downregulation, along with TNF-α downregulation.76, 77 In addition, antagonism of miRNA-155 enhanced lipid uptake and inflammation.78 Similarly, downregulation of miRNA-320a, which is involved in VEGF signaling pathways, as well as miRNA-92a, which regulates shear stress genes, had atheroprotective effects.79-81 Moreover, type II diabetic patients who had ischemic stroke were found to have low levels of miRNA-223 and high levels of miRNA-144. It was found that miRNA-144 levels were low in the serum of diabetic patients following stroke.82 In diabetic mice, it was established that the levels of miRNA-200a and miRNA-466a were downregulated in neural stem cells.83

In 2009, the first study to compare the expression of miRNAs in healthy people and patients with acute ischemia, as per WHO clinical criteria, was published by Tan et al. They found out that 157 miRNAs were expressed in stroke patients. Among those, 138 miRNAs were highly expressed; among them, 17 were upregulated (miR-25, 181a, 513a5p, 550, 602, 665, 891a, 923, 933, 939, 1184, 1246, 1261, 1275, 1285, 1290, and let-7e). Hence, it was concluded that these miRNAs could be used in the diagnosis of AIS as well as in the differentiation of large and small artery stroke.84 In 2013, the Tan et al. research team found that 21 miRNAs were similarly expressed in all AIS patients (hsa-miR-1258, 125a5p, 1260, 1273, 149, 220b, 23a, 25, 26b, 29b1, 302e, 34b, 4835p, 488, 4903p, 498, 506, 659, 890, 920, and 934). Among these 21 miRNAs, four were downregulated (miR-25, 34b, 4835p, and 498).85

Levels of miRNAs in blood were compared between patients who had strokes and controls, and it was found that miR-122, 148a, 19a, 320, and 4429 were low and miR-363 and 487b were elevated in patients with AIS.86 Similarly, comparison of miR-210 levels in blood in ischemic stroke patients and healthy controls showed that miR-122 levels were downregulated in stroke patients, especially during the 7–14-day period following stroke onset, which suggested that the miR-122 level could be useful in stroke diagnosis a few days after stroke onset.87

Researchers in one study found that miR-16 was higher in ischemic stroke patients compared to hemorrhagic stroke patients with an odds ratio of 9.75, which indicates that miRNA is not only a stroke diagnostic biomarker, but also could be used in distinguishing between ischemic and hemorrhagic stroke.88 In contrast, miR-21 was reported previously to have cardioprotective effects in ischemia reperfusion-induced cardiocyte cell death. It was identified that miR-21 is significantly downregulated in AIS patients within the first 24 h, suggesting its potential as a diagnostic tool at the early stage of cerebral ischemia.89 Furthermore, Let-7 is a type of miRNA, with a family of 12 members in humans, that plays an important role in central nervous system gene expression regulation.90 It has been proposed that Let-7 promotes neurodegeneration by the activation of RNA-sensing Toll-like receptors.91 Serum Let-7e was found to be elevated in ischemic stroke patients within the first 24 h of stroke onset with a specificity and sensitivity of 73.4% and 82.8%, respectively, for the diagnosis of ischemic stroke.92 In contrast, another member of the Let family, Let-7c-5p, was reduced in the plasma of ischemic stroke patients within the first 24 h.93