**4.2 Alzheimer's disease**

Alzheimer's disease (AD) is an age-related neurodegenerative disorder characterized by the progressive loss of cognition and memory due to severe neuronal cell loss. Hallmarks of the disease include the formation of extracellular amyloid plaques and intracellular neurofibrillary tangles of hyperphosphorylated tau. Given the high degree of cell death and chronic inflammation in the CNS, it is unsurprising that a large number of miRNAs are differentially expressed in AD [82–85], however, a number of miRNAs have also been shown to affect the pathogenic mechanisms of the disease.

Alternative splicing of amyloid precursor protein (APP), the parent molecule of pathogenic Aβ, is regulated by miR-124. Indeed, miR-124 is downregulated in the AD brain and its expression was shown to inhibit polypyrimidine tract binding protein 1 (PTBP1) resulting in increased APP with exon 7 and 8 inclusion [86]. Alterations of this kind have been associated with increased Aβ production [87]. Furthermore, miR-98 reduces the expression of insulin-like growth factor 1 (IGF-1) which is involved in the processing of APP. Overexpression of miR-98 downregulates IGF-1 resulting in increased Aβ production and tau phosphorylation [88]. The expression of tau is affected by the levels of miR-34a and miR-26b [89, 90]. Overexpression of miR-26b also leads to aberrant cell cycle entry that involves the nuclear export and activation of cyclin-dependent kinase 5 (CDK5), a major kinase involved in tau phosphorylation [89]. Finally, pro-inflammatory NFκB-associated miRNAs such as miR-7, miR-9, miR-34a, miR-125b, miR-46a and miR-155 are all upregulated in AD [85]. Presenilin 2 (PS2) mutations have been implicated in the development of autosomal dominant AD, and microglial knockout of PS2 reduces miR-146 expression and results in an increased pro-inflammatory response [91]. The level of inflammation in the CNS is a strong determining factor for disease progression in AD [92].

Limited work has been carried out to date on the involvement of tsRNA in AD; however, similar to PD, mutation of tsRNA-generating enzyme Ang has been identified. In an Italian cohort of AD patients nonsense mutations in *ANG* were identified with 0.2% frequency resulting in a 51 amino acid shortening in the protein [93].

#### **4.3 Stroke**

Stroke remains one of the leading causes of death and disability worldwide, conferring a high morbidity, disability, and mortality. Cerebral ischaemia triggers a complex cascade of physiological, biochemical and gene expression changes primarily resulting from impaired cellular energetics and the collapse of ion gradients. In particular, ischaemia-mediated glutamate elevation and subsequent over-activation of glutamate N-methyl-D-aspartate (NMDA) receptors is central to excitotoxic neuronal injury and cell death during ischaemic stroke [94, 95].

miR-107 has been shown to play a key role in the regulation of excitotoxicity in ischaemic neuronal injury, associated with increased glutamate accumulation both *in vivo* and in ischaemic stroke patients [96]. Increased miR-107 following ischaemic stroke inhibits GLT-1 expression, an abundant glutamate transporter, resulting in the accumulation of glutamate. Hypoxamir miR-210 has been widely reported as a miRNA ubiquitously expressed in ischaemic cells and tissues, with a central role in adaptation to low-oxygen environments such as tumourigenesis and ischaemia [97]. Robust induction of miR-210-3p following ischaemic stroke *in vivo* has been associated with modulation of PI3K-p70S6K signaling in response to AMPK activation and NMDA receptor-mediated glutamate excitotoxicity [98]. A number of other miRNAs have also been reported to play roles in the regulation of glutamate neurotransmission and excitotoxicity in ischaemic stroke, including miR-223, miR-181, miR-125a, miR-125b, miR-1000, miR-132 and miR-124a [99].

miR-223 has been shown to regulate the functional expression of glutamate receptor AMPAR subunit GluR2 and NMDAR subunit NR2B, which control neuronal excitability in response to glutamate, reducing neuronal excitability and cell death by inhibition of NMDA-induced calcium influx in hippocampal neurons [100]. One of the most abundantly expressed neuronal miRNAs, dysregulation of miR-124 has been implicated in many CNS disorders and has been shown to be downregulated following ischaemic stroke [101]. Downregulation of miR-124 *in vivo* following ischaemic stroke has been associated with upregulation of deathassociated protein kinase 1 (DAPK1), identified as a direct target of miR-124, caspase-3, and cleaved caspase-3, while over-expression of miR-124 was shown to significantly decrease DAPK1, caspase-3, cleaved caspase-3 levels and reduce

*Emerging Roles of Non-Coding RNA in Neuronal Function and Dysfunction DOI: http://dx.doi.org/10.5772/intechopen.101327*

NMDA- and oxygen-glucose deprivation (OGD)-induced neuronal death *in vivo* [102]. Moreover, the neuroprotective role of miR-124 has been associated with decreased expression of pro-apoptotic protein Bax and increased expression of anti-apoptotic Bcl-2 and Bcl-xl [103].

In the context of inflammation associated with cerebral ischaemia, miR-181c has been shown to inhibit prominent pro-inflammatory cytokine TNF-α in response to OGD, reducing microglial activation and neuronal cell death [104]. Furthermore, miR-216a, miR-3437b and miR-126-3p or -5p have also been associated with regulation of TNF following cerebral ischaemia.

Recent studies have shown tiRNAs to be upregulated following ischaemia in models of OGD *in vitro* and following ischaemic-reperfusion injury *in vivo.* Rapid and response-specific increases in tiRNA levels have been shown to correlate with degree of tissue damage, highlighting the potential role of tiRNA detection as a stress biomarker of injury [55, 56, 105, 106]. Furthermore, upregulation of 5′tiRNA fragments has been shown to inhibit endothelial angiogenesis following ischaemic stroke, indicating a role in modulating cerebral responses to ischaemic injury [106].

#### **4.4 Amyolateral sclerosis**

Amyolateral sclerosis (ALS) is the third most common neurodegenerative disease and is characterized by the rapid degeneration of cortical and spinal motor neurons leading to paralysis and death within 3–5 years of diagnosis [107, 108]. Approximately 90% of cases are sporadic, however a number of genetic mutations have been identified that account for 11% of sporadic and 70% of familial ALS [107, 109]. Mutations involving superoxide dismutase (SOD1), fused in sarcoma (FUS), TAR DNA-binding protein 43 (TDP43) and a hexanucleotide repeat expansion on chromosome 9 in open reading frame 72 (C9ORF72) have all been associated with ALS pathology [109].

Deregulation of miR-142-3p has been identified in both SOD1 and TDP-43 mutant mice, as well as in serum from ALS patients. Subsequent bioinformatic analysis identified TDP-43 and C9orf72 as targets of miR-142-3p, further implicating this miRNA in ALS pathology [110]. The skeletal muscle-specific miRNA, miR-206, regulates myogenesis, promotes the formation of neuromuscular junctions and is upregulated in ALS [111, 112]. This protective response occurs early in disease progression and plateaus [111], and higher levels of miR-206 are found in spinal ALS which is associated with lower atrophy rates [113]. Upregulation of miR-155 has been identified in both sporadic and familial ALS, and its inhibition in SOD1 mutant mice resulted in increased survival [114]. Finally, a number of miRNA associated with regulation of oxidative stress are altered in ALS. X-linked inhibitor of apoptosis (XIAP) and the Nrf2-ARE pathway have been closely associated with neuronal dysfunction in ALS and are regulated by miR-34a and miR-27a [115, 116].

As seen with other neurodegenerative diseases, mutations to *ANG* have been identified in ALS and repeatedly validated in independent cohorts [79, 117–119]. Characterization of these mutations determined a reduction in ribonuclease activity and nuclear translocation of Ang [117]. Interestingly, the Ang-generated tiRNA 5′ValCAC is increased in SOD1G93A mice at symptom onset and correlate with Ang expression and slower disease progression. Furthermore, increased 5′ValCAC in ALS patient serum samples is correlated with slower disease progression [120].

#### **4.5 Epilepsy**

Epilepsy is a heterogeneous group of disorders characterized by spontaneous and recurrent seizures that affects approximately 50 million people worldwide [121].

In the majority of instances, seizures can be controlled, however approximately 30% of cases are treatment resistant. Seizures arise from abnormal synchronous activity in hyperexcitable neuronal networks and while this can be attributed to altered electrophysiological properties of ion channels and neurotransmitter systems, converging lines of research have also indicated a central role for the regulation of protein translation [122, 123].

As described in Section 2.2.1, miRNAs play a key role in neuronal excitability and connectivity making them prime targets in epilepsy research. The growth, spine density and arborization of dendrites are directly regulated by miR-132, miR-134 and miR-9 [20–24, 124, 125]. miR-132 is significantly increased in the hippocampus of experimental mice undergoing seizure and its inhibition has been shown to increase neuronal survival and reduce seizure frequency [126, 127]. Upregulation of miR-134 has been identified in resected hippocampal and neocortical tissue of patients with treatment-resistant temporal-lobe epilepsy [128]. This was also observed in a number of animal models where inhibition of miR-134 was shown to reduce seizure occurrence and increase spine volume in hippocampal neurons [128–130]. Neuronal potassium channel expression is regulated by miR-92a and miR-324. miR-92a has been shown to be increased in temporal lobe epilepsy patients, and in animal models of epilepsy inhibition of miR-324 delays the onset of spontaneous seizures [131, 132]. Finally, the Ca2+ extruding pump ATP2B4 and the sodium-potassium-chloride transporter NKCC1 are regulated by miR-129 and miR-101a respectively. miR-129 is increased in temporal lobe epilepsy patients and inhibition of miR-1219 and miR-101a have been shown to reduce hyperexcitability in animal models of epilepsy [133, 134].

Recently, serum from two independent cohorts of temporal-lobe epilepsy patients have revealed increased levels of three 5′tRFS, 5′AlaTGC, 5′GluCTC and 5′GlyGCC. These tRFs were detected in resected hippocampal and cortical tissue and were not associated with any disease related lesions. Furthermore, these fragments were detected in primary mouse hippocampal neurons and their expression was shown to be activity-related [135].
