**4. Regulation of RAS by associated microRNAs**

Given that there is further discussion of the biochemical functions of the RAS in other chapters, the discussion henceforth will focus on some of the most important components of the brain RAS and the miRNAs targeting them.

Since the vital function of the brain results in high physiological demands (i.e. requiring 20% of total cardiac output and a 10-fold higher oxygen and energy demand than other tissues), it requires strict coordination between blood flow and neuronal activity, a phenomena known as functional hyperaemia [77]. Cerebral blood flow is regulated by vasomotor, metabolic and neurogenic mechanisms, but can be modulated by vasoconstrictors such as Ang II and endothelin, vasodilators such as bradykinin, adenosine and other angiotensin ligands, while blood vessel capacity may be reduced or impeded by plaques of cholesterol, amyloid or fibrotic deposits.

Analysis by TargetScan [48], a software that predicts miRNA binding sites, suggests that 368 different miRNA families target RAS elements, the majority of which share transcripts. **Table 1** summarizes the total number of miRNAs and unique miRNAs with respect to RAS elements, as they have other targets outside the RAS. Angiotensin 4 receptor (also known as AT4R or IRAP) has 252 miRNA families associated with it, making it the highest amongst the RAS and approximately fivefold and threefold as many as that for arguably its better known receptors AT1R and AT2R. Notably, 88% of IRAP-associated miRNAs also regulate other RAS transcripts, suggesting its susceptibility to changes elsewhere in the RAS. In particular, IRAP has 28 miRNA families exclusively associated with it (also the most for RAS components), hinting at having high functional importance. Indeed, aminopeptidase B and dipeptidyl peptidase, necessary for Ang IV conversion, do not have exclusive miRNAs and thus may be subject to many regulatory effects.

A more in-depth examination of RAS-associated microRNAs, according to their functional impact in the RAS physiology is shown in **Table 2**. MiR-3163 targets the greatest number of RAS transcripts (N = 8) and may provide an over-arching level of regulation for the pathway


The total number of miRNAs represents miRNA families with binding sites at the 3′UTR region based on TargetScan [48]. Unique miRNAs are those considered solely with respect to other RAS elements.

**Table 1.** miRNA families targeting RAS elements.

miRNA transfer both propagates deleterious effects and helps recover cells from insults and prevent apoptosis. For example, miR-133 is increased in people with cardiovascular disease and is transferred through exosomes from multipotent mesenchymal stromal cells to astrocytes and neurons that promote recovery after stroke [60–63]. Furthermore, remote ischaemic conditioning, a technique of small cycles of ischaemia/reperfusion in distal extremities, was protective for cardiac and cerebrovascular effects in animal experiments and human clinical

Exosomal circulating miRNAs have many properties that arguably make them ideal biomarkers, including their presence in peripheral blood, detection in many biological fluids, their stability in RNase-rich body fluids and their tissue-specific expression patterns. These have been described in cardio-cerebrovascular disorders, diabetes, dyslipidemia and neurodegenerative disorders [1, 70–76]. Furthermore, human exosomes can be used therapeutically as a

Given that there is further discussion of the biochemical functions of the RAS in other chapters, the discussion henceforth will focus on some of the most important components of the

Since the vital function of the brain results in high physiological demands (i.e. requiring 20% of total cardiac output and a 10-fold higher oxygen and energy demand than other tissues), it requires strict coordination between blood flow and neuronal activity, a phenomena known as functional hyperaemia [77]. Cerebral blood flow is regulated by vasomotor, metabolic and neurogenic mechanisms, but can be modulated by vasoconstrictors such as Ang II and endothelin, vasodilators such as bradykinin, adenosine and other angiotensin ligands, while blood vessel capacity may be reduced or impeded by plaques of cholesterol, amyloid or fibrotic deposits.

Analysis by TargetScan [48], a software that predicts miRNA binding sites, suggests that 368 different miRNA families target RAS elements, the majority of which share transcripts. **Table 1** summarizes the total number of miRNAs and unique miRNAs with respect to RAS elements, as they have other targets outside the RAS. Angiotensin 4 receptor (also known as AT4R or IRAP) has 252 miRNA families associated with it, making it the highest amongst the RAS and approximately fivefold and threefold as many as that for arguably its better known receptors AT1R and AT2R. Notably, 88% of IRAP-associated miRNAs also regulate other RAS transcripts, suggesting its susceptibility to changes elsewhere in the RAS. In particular, IRAP has 28 miRNA families exclusively associated with it (also the most for RAS components), hinting at having high functional importance. Indeed, aminopeptidase B and dipeptidyl peptidase, necessary for Ang IV conversion, do not have exclusive miRNAs and thus may be subject to

A more in-depth examination of RAS-associated microRNAs, according to their functional impact in the RAS physiology is shown in **Table 2**. MiR-3163 targets the greatest number of RAS transcripts (N = 8) and may provide an over-arching level of regulation for the pathway

trials, with effects mediated by miRNAs such as miR-1 [64–69].

gene delivery vector to provide cells with heterologous miRNAs [53].

**4. Regulation of RAS by associated microRNAs**

brain RAS and the miRNAs targeting them.

178 Renin-Angiotensin System - Past, Present and Future

many regulatory effects.

as a whole, for example, in response to an external stimulus. miR-125-5p with five targets in common may function in a similar way, particularly since two of the targets are principal enzymes in RAS biochemistry. Yet, they make an ideal combination to block Ang II/AT1R and Ang IV/AT4R pathways and also shift the conversion of Ang I to Ang (1–7) via neprilysin and other peptidases to act on MasR.

In terms of RAS function, a group of microRNAs that can shift a predominant role of, for example, the Ang II/AT1R axis to opposing axes such as Ang(1–7)/MasR or Ang IV/AT4R could change cerebral blood flow, response to hypoxia and perhaps influence cognition and vice versa. Indeed, a panel of 17 miRNA families target aminopeptidase A and IRAP that could potentiate the formation of Ang IV (since aminopeptidase A converts Ang I and Ang II to Ang III, the Ang IV precursor for Ang IV). Thus, upregulation of those 17 miRNAs could modulate Ang III and IRAP to a greater extent than just one miRNA, such as miR-125. The net effect of reducing both ligand and receptor means that the function of the Ang IV/AT4R axis might be completely inhibited with likely deleterious effects on blood flow and cognitive performance. By contrast, downregulation of these miRNAs would increase the Ang IV/AT4R axis. The following section will discuss the effect of some specific miRNAs and their regulatory effects in RAS in the brain in health and in disease states.


From 164 combinations of overlapping targets and miRNAs in common, only 14 are included here, 12 which if increased would favour vasoconstriction and 2 would increase vasodilation. Others tend to influence multiple RAS pathways, an example, miR-3163, is given at the bottom of the table. miRNAs in bold are described further in the text.

**Table 2.** A summary of the subgroups of miRNAs according to their functional effect in the RAS.
