While ncRNAs have been implicated in regulating gene function within the cytoplasm due to the interaction of ncRNAs with ER-bound ribosomal polysomes, ncRNAs can also regulate RNA processing and translation, gene transcription, and cellular metabolic activity, indicating ncRNAs are functionally present in organelles such as the nucleus, ER, and mitochondria [40–44]. In one such example, Kim et al. explored the role of ncRNAs in the mitochondria and found lncND5 interacts with MRPP1 and controls mitochondrial gene expression [45]. Additionally, RMRP interacts with PNPASE and GRSF1, which regulates mitochondrial RNA processing, mtDNA replication, and ultimately mitochondrial-mediated oxidative phosphorylation capacity. These findings have significant consequences in relation to the development of therapies for metabolic disorders, such as cancer. For instance, investigators demonstrated that cytosolic lncRNAs and miRNAs are essential for controlling the expression of glucose transporters GLUT1-3 as well as glycolytic enzymes via HK2, which results in dysregulated glucose uptake in response to insulin and limits the conversion of glucose to glucose 6-phosphate thereby reducing the overall metabolic activity of mammalian cells [46, 47]. Xu et al. also found that lncMMPA alters the levels of ALDH1A3, affecting the rate of aerobic glycolysis in HCC, yet promotes M2 macrophage polarization [47]. Some of these metabolically linked-ncRNAs are also functionally active in the nucleus, and interact with DNA regulatory machinery to control cellular proliferation in response to glucose stimulation [48, 49]. In fact nuclear RNAi, and nuclear miRNA/lncRNA molecules have a crucial role in regulating the abundance of nascent mRNA transcripts post-transcription, and modulate co-transcriptional alternative splicing activity [50, 51]. Together, these findings indicate that in response to an environmental stimuli, ncRNAs function within every cellular compartment to regulate essential cellular processes associated with the transcription and translation of genetic information, chromatin remodeling [52], RNA processing, and DNA damage repair [53], which in turn, affects the maturation and function of mammalian cells.

Given the ever increasing complexity of RNA terminology, the RNA community recognized miRNAs, lncRNA, and circRNA as ncRNAs that function as endogenous genetic regulators of mRNA abundance as well as controlling the expression and activity of all other ncRNAs (i.e., tRNA, rRNA, snRNA). While a majority of this review will focus on miRNA biology as a leading ncRNA therapeutic approach, it should be appreciated that lncRNAs were one of the original ncRNAs identified to have the capacity to silence genetic information [54–57]. X-chromosome inactivation (XCI) mediated by XIST is an example of how a lncRNA can control an unbalanced dosage of genetic information within the developing female embryo [58]. Furthermore, since lncRNAs are found in low abundance it is thought lncRNAs mostly act “in cis” or near their respective sites of transcription and subsequently co-transcriptionally regulate the expression of neighboring genes. The ENCODE project has identified over 10,000 lncRNAs, many of which are differentially expressed between normal and cancerous cells [59]. Over the past decade, lncRNAs have been found to interact with chromatin, modulate gene expression, function as protein decoys, and govern cellular physiology [60]. The crucial role of lncRNA in eukaryotic cell biology and in normal cellular function indicate these ncRNAs may also be novel targets for therapeutic intervention. However, because the mechanisms of biogenesis and mRNA silencing by miRNAs are more well-established, this review primarily focuses on the utilization of miRNA in the identification and treatment of neurocognitive disorders that currently have little to no curative options.

miRNAs are small ncRNAs that modulate gene expression by binding to complementary sequences predominantly within the 3′UTR of target mRNAs [61, 62]. Similar to siRNA technology, the mechanism of action(s) linked to miRNAs indicates that miRNA/RNAi-based therapies could be used to silence, and in some cases repair, any misregulated genetic element within the human genome to correct a specific pathological deficit. miRNAs are, however, distinct from siRNAs in that they are endogenously expressed in the genome and transcribed in the nucleus by RNA polymerase II/III [63–65] (see Figure 1). The resulting primary miRNA (pri-miRNA) transcript is unique in that the pri-miRNA folds creating hairpin bulges similar to tRNA, which is recognized by the RNaseIII enzyme DROSHA [69–71]. Pri-miRNAs can originate from large intergenic regions of the genome, (i.e., areas with little protein coding activity), as well as from intronic regions, (i.e., from sequences processed and spliced out of newly transcribed mature mRNAs), and sometimes are polycistronic and contain multiple mature miRNA sequences [72]. Recently, a non-canonical pathway was identified that further categorized a subset of intronic miRNAs known as “miRtrons” (see Figure 1). These miRNA sequences mapped precisely to short hairpin introns and spanned each of the exon-intron junction borders [67, 73], meaning that when the spliceosome recognizes these junctions, a miRNA will be concomitantly produced with the exonic mature mRNA. Ultimately these findings indicate that an intronic sequence is no longer considered “junk DNA” and in some cases an entire intron codes for a miRNA that has significant downstream regulatory potential.

Canonical pri-miRNAs, as well as miRtrons, both contain sequential hairpin bulges, a terminal loop, are ∼85 nucleotides in length, and recognizable by the microprocessor complex containing DROSHA and cofactor DGCR8 (see Figure 1). In D. melanogaster, DGCR8 is called Pasha, yet performs similar functions [74–76]. The Drosha-DGCR8 heterodimer recognizes approximately one to two helical RNA turns of sequence from the pri-miRNA stem and splits the phosphodiester bond, releasing a short hairpin-shaped RNA ∼60 nucleotides in length [66]. These canonical pre-miRNAs and the pre-miRNAs generated from the miRtron pathway are exported to the cytoplasm via Exportin-5, and undergo further processing by the RNAse-III enzyme, Dicer [68, 77, 78]. Dicer cleaves the double-stranded pre-miRNA approximately two helical turns from the terminal loop [79], generating short double-stranded RNA fragments ∼22 nt in length known as the mature miRNA duplex. This duplex contains a guide and a star strand offset on the 5′ ends, allowing for strand selection to occur via Argonaute and the RNA-induced silencing complex (RISC) [80–82].

The focus on the miRNA biogenesis pathway is an important area of research, as many second and third generation siRNA therapeutics utilize the miRNA biogenesis pathway to induce a particular outcome of gene repression, activation, or editing [83–85]. Duplexed siRNAs will also undergo strand selection, and can be engineered with sequence bias to favor a particular guide siRNA. Both the guide miRNA or synthetic siRNA is incorporated into RISC which contains, AGO/TRBP and binds to complementary mRNAs within the 3′ UTR resulting in stalling of polysomal translational machinery, the degradation of the mRNA through slicing activity of AGO2, the storage or degradation of mRNA in P-bodies, or the transport of mRNA to exosomes or other cellular regions, such as dendrites (see Figure 1) [86–89].

While much attention has been given to the role of miRNAs in posttranscriptional gene silencing, recent studies have highlighted the broader functions of miRNAs in regulating nuclear chromatin dynamics. Catalanotto et al. highlight the nuclear function(s) of miRNAs, including their involvement in transcriptional regulation and their ability to recognize gene targets at the promoter level [90]. Politz et al. investigated the presence and localization of miRNAs within the nucleolus, and observed that certain nucleolar miRNAs contribute to ribosome biosynthesis and associate with the granular component, where ribosome assembly occurs [91]. Interestingly, in Alzheimer’s patients, reduction in nucleolar size is notable, concomitant with reduced rRNA transcription [92]. In Parkinson’s, nucleolar integrity is maintained by ɑ-synuclein co-localization with nucleolin, which is regulated by the presence of miR-7 and miRNA-153 [93, 94]. Therefore, miRNAs may regulate the pathophysiology of many neurological disorders, absent of the canonical gene repression mechanisms mentioned above. In fact a number of synthetic RNAi-based therapies have utilized this approach to generate oligonucleotide based therapies such as AMT-130, and VY-HTT01 to correct the HTT gene defect in individuals with Huntington’s [95], as well as antisense mediated regulation of ATXN and CAG repeats in individuals with spinocerebellar ataxia [96].

Neuroinflammation is a complex homeostatic process that is essential in mitigating exogenous pathogens from entering the central nervous system (CNS) [97]. Normally, pro-inflammatory responses allow for the acute recruitment of macrophages, natural killer cells, and T-cells to attack these pathogens in a robust manner [98]. Once the pathogen is removed, inflammatory cytokine signaling decreases, and the presence of inflammatory cells diminishes. A key factor in the inflammatory process is the activation of microglia, which safeguards the brain from neuronal damage through the release of neuroprotective cytokines and other inflammatory factors [99–101]. Chronic neuroinflammation is the continued presence of inflammatory cytokine signaling absent of a pathogen, which results in a persistent stress response within the microglia of the CNS, causing disruptions in normal neuronal communication, damage to the neuronal circuitry, and neurodegeneration [102]. The etiology of various neurodegenerative diseases such as Alzheimer’s, Parkinson’s, amyotrophic lateral sclerosis (ALS), and certain forms of epilepsy, are all thought to involve a neuro-inflammatory component.

Recently, the heterogeneic classification of the microglia into M0, M1, M2a, M2b, and M2c cell types indicate that specific microglial cells are responsible for initiating differential cellular responses during neuronal perturbations [103]. For instance, M2 cells tend to secrete anti-inflammatory cytokines and neurotrophic factors, as well as support oligodendrogenesis and remyelination [104]. By contrast, M1 cells respond to bacterial byproducts such as LPS and IFN-y, causing a release of TNF-α and IL-6 inducing oxidative stress and neuronal dysfunction [105]. Given the genetic basis for classifying microglial cell types has been established, researchers can now elucidate how M1 cells remain activated post infection, or how M2 cells are prevented from secreting anti-inflammatory cytokines that promote neurogenesis. One of these regulatory processes involves ncRNA, since miRNA activity and expression are repeatedly involved in cell-fate decisions during cellular differentiation [106–108]. Elucidating the mechanisms of M0 to M1 or M2 cell type transition would be an important area of study so as to further develop candidate therapeutics to delay or abate the progression of neurodegenerative disease.

Independent of the bifurcation of microglial cell-fate, miRNAs also play a significant role in the activation or suppression of neuroinflammation by controlling the abundance of central mediators of NLRP3 priming and/or activation of the inflammasome [109] (see Figure 2). For instance, miR-155 regulates the activity of NF-kB, a major protein complex that controls DNA transcription and cytokine production, supports maturation of dendritic cells, promotes recruitment of neutrophils, stimulates M1 polarization of macrophage and microglial cells, and activates T- and B-cells [122]. Interestingly, the miR-155 host gene is found within the BIC gene cluster, a region that contains transcription factor binding sites for NF-kB, SMAD4, and other interferon regulatory factors [123, 124]. As expected, miR-155 is expressed in a coordinated feedback loop with a series of transcription factors that contain miRNA bindings sites for miR-155 [125]. This regulatory loop ensures robust gene expression of pro-inflammatory cytokines is rapidly expressed during an acute infection, while simultaneously activating a regulatory fail safe to dampen unabated inflammatory signaling. Guil et al. identified a similar feedback mechanism in which RNA-binding protein hnRNP A1 modulates the processing of miR-18a [126]. Specifically, hnRNP A1 binds exclusively to pri-miR-18a, facilitating DROSHA processing, in turn promoting a rapid depletion of hnRNP A1 which results in diminished miR-18a processing and reduced pre-miR-18a levels.

Since these initial discoveries, hundreds of miRNAs have been identified to either stimulate or abrogate anti-inflammatory responses (see Figure 2) [127]. For instance, miR-223, is highly expressed in the hematopoietic system and regulates granulopoiesis, a process essential in differentiating bone-marrow derived myeloblasts so as to re-generate the depleted neutrophil reservoir during and after an acute infection [128]. However, miR-223 keeps neutrophils and monocytes in an inactive state, absent of inflammatory stimuli. Another anti-inflammatory miRNA, miR-124, controls the levels of TNF-a in the microglia, dampens the abundance of reactive oxygen species, and promotes M2 type polarization [129, 130]. Identifying these anti-inflammatory miRNAs is the first step in the process of developing oligonucleotide based therapeutics to maintain the activity of healthy microglia and promote neuronal differentiation. In support of this approach, imbalances in miRNA levels, such as miR-155, miR-18a, and miR-124 results in heightened inflammation in vivo. In zebrafish, levels of miR-18a spike one to 5 days post retinal injury [131]. In miR-18a^mi502 altered animals, photoreceptor regeneration was delayed, and was fully restored once the continued presence of active inflammation was abated by dexamethasone treatment. In mice, the loss of miR-223 results in a protective phenotype using a MOG induced EAE model [101, 132] These mice exhibit decreased spinal cord inflammation and reduced demyelination. This seems counterintuitive, yet using a stable miR-223 knockout model, developmentally, these mice most likely have a depleted neutrophil reservoir. Reducing the concentration of miR-223 also results in the decline of inflammatory responses and the presence of increased autophagy within the microglia. miR-155 has been extensively studied using a variety of knockout and overexpression models. In mice it is clear that miR-155 modulates the polarization of the microglia and can promote neuroinflammation by inhibiting inflammasome signaling mediators such as SIRT1 and SOCS1, and by promoting IL-6 production [110, 133]. Specifically, in APPtg and 3xTg Alzheimer mouse models, the persistently elevated levels of miR-155 was associated with the proliferation of dendritic cells, as well as T regulatory cells that support neuroinflammation.

While proper miRNA production leads to healthy cell function, regulation of gene expression, and maintenance of normal intracellular pathways, the disruption of miRNA biogenesis can also contribute to a variety of illnesses (see Figure 2). Tang et al., identified miR-709 as a nuclear miRNA involved in the regulation of miR-15/16 biogenesis [134]. Specifically, miR-709 binds to pri-miR-15a/16-1, ∼0.8 kB away from the pre-miRNA hairpin bulge, and recruits chromatin remodeling complexes that prevents the processing of pri-miR-15a/16-1 by DROSHSA. Interestingly, both miR-15/16 and miR-709 have important roles in controlling neuroinflammation in vivo. miR-709 controls glutamatergic signaling and endosomal trafficking in cortical neurons, resulting in balanced sleep-wake cycles [135]. The deletion of mir-15/16 in T-regulatory cells results in animals with extensive lymphoproliferative disease and systemic tissue inflammation [136]. Some of the phenotypes could be linked to the abundance and activity of Toll-like receptors (TLRs), which are expressed in dendritic cells, and macrophages, as well as epithelial cells. TLRs are the first line of defense against pathogens by recognizing pathogen-associated molecular patterns (PAMPs) [137]. When activated, TLRs trigger signaling pathways that involve various molecules and transcription factors, leading to the release of inflammatory cytokines. In the central nervous system, TLRs are present mainly in microglia and astroglial cells, but also in some neurons and other glial cells. While TLR activation is a defense mechanism against invaders and tissue damage, excessive activation can disrupt the balance of immune responses, resulting in the production of persistent pro-inflammatory molecules [138]. This can lead to neuroinflammation and damage in the nervous system, contributing to various neurodegenerative diseases. miR-15a, has been shown to target TLR4-associated pathways in bone marrow derived macrophages affecting the mortality of mice using a model of sepsis [139] (see Figure 2). Furthermore, miR-155 activity is induced by TLR signaling, while miR-146a was reported to control cytokine and TLR signaling through a negative feedback regulatory loop with IRAK1 and TRAF6 [140, 141]. Taken together, these studies indicate, miRNAs can intersect with TLR signaling and modulate the process of an inflammatory response. Specifically, when TLR4 is activated by substances that are harmful to the nervous system, such as LPS, a pattern of expression that includes numerous miRNA clusters, such as miR-15, miR-182, and miR-200, result in a coordinated effort to stimulate acute inflammatory response to remove the pathogen. However, dysregulated miRNA signaling can result in persistent inflammatory cues that promote a diseased physiological state by promoting neuroinflammation. Here we discuss relevant neurological disorders that are the result of mis-regulated miRNA expression and activity, which could serve as the basis for therapeutic intervention using oligonucleotide therapeutics to abate or slow the onset of these diseases.

Epilepsy is a group of disorders characterized by recurrent seizures [142], and is one of the most common neurological conditions that affects at least 50 million people worldwide [143, 144]. Epilepsy follows a bimodal age distribution pattern, meaning that infants and elderly are at the highest risk, with the highest incidence rate being in individuals over 70 years of age [145]. Onset rates in young children have decreased with an increase in perinatal care and sanitation, leading to a decrease in infectious disease. However, epilepsy rates in the elderly have increased due to increased life expectancy, resulting in higher rates of other neurological disorders and cancers that may contribute to the etiology of epilepsy [146]. The International League Against Epilepsy (ILAE) defines the disorder as a patient having 1) two seizures, separated by more than 24 h, occurring without a clear cause, 2) one seizure without a clear cause, and the chance of having more seizures in the next 10 years being over 60%, or 3) diagnosis of an epilepsy syndrome [147]. Although the definition of epilepsy suggests that it is a single disorder, it is more accurate to describe epilepsy as a group of disorders with diverse etiologies and outcomes. Different epilepsy syndromes are characterized by seizure type(s), age of onset, developmental status, comorbid features, and etiology [142]. The ILAE has defined six etiologic categories; Structural, Genetic, Infectious, Metabolic, Immune, and Unknown [148]. Understating the exact genetic underpinnings that manifest these specific etiologies is crucial in understanding how to better treat individuals with epileptic syndromes.

ALS is a fatal neurodegenerative disorder affecting motor neurons in the spinal cord, brainstem, and cerebral cortex [164, 165]. Currently, there are no effective therapeutic options available that delay or slow the onset of ALS. The disease primarily strikes in mid-to-late adulthood, progressively leading to muscle weakness, atrophy, and ultimately respiratory muscle dysfunction, resulting in death [166]. Most ALS cases are sporadic, while only 5%–10% present with a familial history [165]. The median survival is around 3 years, with only 10% surviving beyond 8 years, emphasizing the need for improved diagnostic biomarkers and therapeutic interventions [165]. Repeat expansions in the C9ORF72 gene and mutations in superoxide dismutase 1 (SOD1) are common genetic causes of ALS, while TAR DNA-binding protein (TDP-43) and fused in sarcoma/translated in liposarcoma (FUS/TLS) mutations represent a smaller fraction [165]. Importantly, aberrant miRNA expression profiles are observed in both ALS mouse models and patients, highlighting the potential role of miRNAs in disease progression [165].

Microglia-mediated neuroinflammation plays a complex role in ALS progression. Inflammatory responses are observed in both SOD-1-transgenic mice and ALS patients, influencing disease progression [138]. ALS is gradually being appreciated as an inflammatory disease, since dysfunction of normal microglia and astrocytes contribute to the phenotypes of ALS [138]. For instance, microglia, the immune cells of the CNS, normally promote synaptic maturation and remodeling, control of excessive neuronal activity, and promote oligodendrocyte progenitor cell development. In ALS, the microglia attempt to clear pathological aggregates, at the expense of enhanced proinflammatory cytokine signaling, ROS and nitric oxide production, excessive synaptic pruning, resulting in a neurotoxic state that generates reactive C3⁺ astrocytes. Interestingly, targeting peripheral Ly6Chi monocytes with anti-Ly6C mAb reduces microglial infiltration and modifies the disease course in SOD1 mouse models indicating the monocytes and resident microglia in SOD1 mice influence ALS disease progression [167].

Astrocyte-mediated neuroinflammation also plays a complex role in ALS progression. Astrocytes are instrumental in regulating neuronal synapse release, facilitate neurotransmitter clearance, support glutamate reuptake, and promote antioxidant production. In ALS, these astrocytes are not able to maintain these functions and excessive pro-inflammatory cytokines result in the secretion of neurotoxic factors, increased ROS production, and reduced neurotrophic factors resulting in neuronal degeneration [167]. Many of the phenotypic consequences of astrocyte-mediated neuroinflammation are still being investigated; however, it was recently appreciated that motor neuron cell death is triggered by astrocyte mediated activation of a certain form of programmed cell death called necroptosis. This form of cell death is described as the loss of plasma membrane integrity due to activation of RIPK1 and MLKL. Some investigators are determining if regulators of these kinases could be used as therapeutics to abate the progression of ALS. Here, we argue that miRNAs that target these kinases such as miR-421, miR-381, and miR-425 could serve as effective treatments for ALS.

The greatest supporting evidence for the involvement of miRNAs in ALS etiology is the genetic link to TDP-43. TDP-43 is a member of TAR-binding proteins that are responsible for proper RNA processing. It is known that the generation of a subset of pre-miRNAs requires TDP-43 interactions with the nuclear Drosha complex for the proper cleaving of the cognate parental pri-miRNA. Additionally, cytoplasmic TDP-43 associates with the Dicer complex by binding to the terminal loops of the pre-miRNA facilitating Dicer-mediated cleavage of these pre-miRNAs. Studies by Kawahara et al., clearly show that TDP-43 is indispensable for neuronal outgrowth in cell culture models. Taken together, it is unquestionable that the non-coding RNA regulatory pathway is emerging as a central contributor to ALS pathogenesis.

Mutations in a number of RNA binding genes, such as C9ORF72, SOD1, TDP-43, and FUS result in altered RNA metabolism and correlate with an increased abundance of the protein aggregates similarly found in ALS patients [164]. Specifically, miRNA profiling reveals a series of dysregulated miRNAs in ALS (see Table 1). miR-218 is one miRNA of interest, given the expression of miR-218 is enriched in motor neurons, and was found in the exosomes released from cultured neurons post injury. In cultured primary astrocytes, it was found that miR-218 targets EAAT2, resulting in the modulation of glutamate re-uptake. EAAT2 expression is downregulated in post-mortem brain tissues from ALS patients, and overexpression of EAAT2 in the SOD1 model mentioned above protected animals from L-glutamate induced cytotoxicity. Finally, intracerebroventricular injections of miR-218 inhibitor restored normal astrocyte function as well as GFAP and Cx43 expression. These findings indicate that key miRNAs are vitally important to ALS pathogenesis, and furthermore, that differential expression of a particular miRNA alone is insufficient to predict their involvement with respect to disease progression. Exemplified in the case above, the persistent expression of miR-218 in motor neurons resulted in the chronic excitability of glutamatergic neurons, which was only corrected with the introduction of miR-218 inhibitors.

miR-494-3p is another miRNA that was found to be dysregulated in C9ORF72-ALS derived astrocytes. These astrocytes have a reduction in secreted antioxidant proteins, resulting in a state of oxidative stress and reduced cell survival for co-cultured motor neurons. Restoring the levels of miR-494-3p using miRNA mimics in these C9ORF72-ALS derived astrocytes resulted in healthy co-cultures of motor neurons. The genetic targets and the mechanisms of action of miR-494-3p are still unclear, but some studies point to the regulation of pro-inflammatory mediators such TRAF3, and IKKb/NF-kB resulting in the attenuation of M1 macrophage activation. miR-124 is another miRNA associated with ALS progression, given miR-124 has been linked with motor neuron dysfunction and degeneration in the SOD1 mouse model [168]. Specifically, the overexpression of miR-124 resulted in the early onset apoptosis of wild type motor neurons, while miR-124 inhibitor restored neurite outgrowth mitochondrial dynamics, synaptic signaling, and axonal transport of glutamate in SOD1 motor neurons.

As the field moves towards utilizing miRNAs as biomarkers for ALS onset and progression, some have identified additional mechanistic actions for miRNA-mediated progression of ALS. For instance, the accumulation of glycogen in the lumbar spinal cord of SOD1 ALS mice indicates that certain metabolic abnormalities may be an underlying cause of the neuroinflammatory pathways that support ALS progression [169] (see Figure 2). In support of this notion, GYS1^Nestin-KO mice showed significant deficits in learning capacity, as well as altered neurogenesis. Furthermore, normal astrocytes tend to accumulate glycogen similar to muscle cells for utilization during times of high energy demand. Since astrocytes have low levels of glucose-6-phosphate, astrocytes tend to release lactate instead of glucose, which is consumed by the surrounding neurons. Unfortunately, persistent levels of lactate secretion due to overabundant glycogen storage in ALS-astrocytes will also result in the aberrant activation of microglia that in turn secrete pro-inflammatory cytokines TNF-α, IL-6, an IL-1. In support of this hypothesis, miR-338-3p was shown to inhibit glycogenolysis in ALS mice resulting in glycogen accumulation within the spinal cord, while the use of miR-338-3p inhibitors restored some functionality in ALS mice. Interestingly, highly expressed circulating miRNAs such as miR-338-3p, as well as miR-206, miR-183, miR-451, and miR-125a/b are candidate biomarkers for ALS [170–172].

As ALS remains a rare, yet complex disorder with multiple facets of pathogenesis, the potential to cure the disease lies in the continued elucidation of the ncRNA transcriptome which involves the understanding of both miRNA dysregulation and the function of hnRNPs, such as TDP-43. By investigating these pleiotropic regulators in a cell contextual manner, new genetic players that control protein folding, neuroinflammation, glycogen storage, and necroptosis within astrocytes, microglia, and motor neurons will hold the key in determining the next series of therapeutic targets that will slow the progression of ALS. Interestingly, researchers just reported that intravenous application of a subset of anti-inflammatory T-cells attenuates some of the pro-neuroinflammatory phenotypes that hasten ALS progression.

Parkinson’s Disease is a neurodegenerative disorder characterized by bradykinesia, rest tremors, and rigidity [173]. While clinical signs will result in a diagnosis of probable Parkinson’s, a definitive diagnosis requires pathological testing to identify the presence of Lewy bodies, which is an abnormal aggregation of α-synuclein protein. The underlying biological mechanisms of Parkinson’s involve the loss of dopaminergic neurons, particularly in the substantia nigra within the basal ganglia. There is no definitive cause of Parkinson’s, but evidence suggests that genetic and environmental factors play a role in its development [174]. One of the contributing factors of Parkinson’s development is microglia cell dysfunction. Microglia are non-neural cells of the CNS that regulate inflammation, regeneration, and metabolism of neurons. Microglia and astrocytes both secrete pro-inflammatory cytokines, when an infection is present, yet the chronic secretion of pro-inflammatory cytokines result in neuroinflammation associated with the development of Parkinson’s [120]. RNA metabolism may also play a role in the pathogenesis of Parkinson’s. Specifically, genes associated with inherited Parkinson’s have been shown to interact with the RNA translation initiation process and RNA translation components [175]. Although there is no cure for Parkinson’s, treatment options focus on providing symptomatic relief, such as drugs that increase dopamine levels in the striatum or operate on dopamine receptors within the brain [176].

Neuroinflammation is believed to contribute to the development of Parkinson’s. In particular, evidence suggests that neuroinflammation perpetuates the death of dopaminergic neurons by activating the microglia and promoting the infiltration of T-cells at sites of neuronal injury [177]. This pro-inflammatory response specific only to dopaminergic neurons implicates a level of cell context specificity analogous to the regulatory functions of miRNA. In fact, dysregulation of microRNA biogenesis appears to disrupt the balance of inflammatory processes in the brain, and results in the heightening of neuroinflammatory cues specifically associated with the Parkinsonian phenotype. The loss of Dicer in striatal dopaminergic neurons using a conditional knockout mouse model resulted in animals having shortened lifespan, an expansion of the resident astrocyte population, yet no neuronal degeneration [178]. However, the loss of Dicer throughout the entire midbrain of these animals resulted in progressive dopaminergic neuronal loss. Furthermore, patients with chromosomal 22q11.2 deletion syndrome, which includes the genetic loss of DGCR8, have a higher occurrence of Parkinson’s as confirmed by pronounced Lewy body formation in post-mortem brain specimens [178]. Additionally, specific miRNAs such as miR-433, miR-16, and miR-7 regulate the genetic pathways associated with α-synuclein abundance, which is a precursor for α-synuclein protein aggregation and Lewy body formation. miR138 and miR-184 have also been reported to control the LRRK2 and E2F1 genes respectively resulting in aberrant lysosomal activity of astrocytes. Understanding the connection between microRNAs and neuroinflammatory processes, such as the generation of reactive astrocytes, may provide valuable insights into the cellular mechanisms of Parkinson’s and lead to the discovery of novel therapeutics to slow disease progression [178] (see Table 1).

Alzheimer’s Disease (AD) is the most common neurodegenerative disease (NDD) and the most common form of dementia. The clinical signs of Alzheimer’s include progressive cognitive decline and memory loss, along with difficulties with language processing and problem solving. The cause of Alzheimer’s is unknown, but evidence suggests that a combination of genetics, environment, and lifestyle factors play a role in its development. There is currently no cure for Alzheimer’s, and current therapeutics treat the symptoms of the disease. Alzheimer’s is characterized by the accumulation of amyloid beta plaques and tau protein tangles in the brain, which leads to the loss of synaptic connections and neurons, which in turn leads to neuroinflammation, oxidative stress, and synaptic dysfunction. Specifically, there is increasing evidence that neuroinflammation plays a critical role in the progression of the disease. Increased concentration of proinflammatory cytokines have been found in patients with Alzheimer’s, and PET scans have shown increased microglial inflammation in the brains of patients with Alzheimer’s. Microglia use microRNAs to rapidly respond to inflammation [121].

In the brains of individuals with Alzheimer’s, there are elevated levels of specific microRNAs, including miRNA-9, miRNA-34a, miRNA-125b, miRNA-146a, and miRNA-155. These miRNAs increase in activation when they are around proinflammatory cytokines and amyloid beta, both of which are associated with Alzheimer’s inflammation. These miRNAs are influenced by the protein NFkB, which is elevated in the brains of Alzheimer’s patients. Alexandrov et al. investigated the levels of these miRNAs in the cerebrospinal fluid (CSF) of Alzheimer’s patients as compared to individuals without Alzheimer’s [186]. The results showed that the miRNAs listed above were significantly higher in the CSF of patients with Alzheimer’s than without, indicating their potential role as biomarkers.

Huntington’s disease is a rare neurodegenerative disorder caused by an autosomal dominantly-inherited CAG-trinucleotide repeat-expansion encoding for glutamine in the huntingtin (HTT) gene, leading to protein misfolding and aggregation of aberrant proteins [153, 155, 197]. It is characterized by cognitive, motor, and psychiatric disturbance. At the cellular level, mutant HTT results in neuronal dysfunction and death through a number of mechanisms, including disruption of proteostasis, transcription and mitochondrial function, and direct toxicity of the mutant protein [197].

Transcriptional dysregulation is one of the major pathogenic processes in HD. Transcriptional alterations are present in Huntington’s mice before the onset of disease symptoms and in asymptomatic HD-mutation humans, suggesting that perturbed gene expression is one of the causes of pathogenesis. Zsindely et al. [153] investigated miRNAs that are misregulated in a Drosophila model of Huntington’s. miRNAs function as post-transcriptional regulators of gene expression. Therefore, miRNA dysregulation introduces a second layer of altered gene regulatory mechanisms besides the transcriptional misregulation of mRNAs in HD and can pinpoint those molecular processes that are part of the response to mHtt-induced proteopathic stress [153].

They determined the functional effects of three down-regulated (dme-mir-137, dme-mir-219, and dme-mir-1010) and two up-regulated (dme-mir-10 and dme-mir-305) miRNAs on mHtt-induced pathology. Overexpression of both dme-mir-137 and dme-mir-1010 ameliorated mHtt-induced phenotypes, indicating that the down-regulation of these miRNAs is pathological. Overexpression of dme-mir-219, on the other hand, was detrimental both in the Huntington’s model and in control flies, suggesting that in this case, the observed phenotypic outcome is due to a toxic effect of dme-mir-219 overexpression that is independent of Huntington’s pathology. Overexpression of dme-miR-10 also exacerbated all analyzed phenotypes of Huntington’s flies, and it also had a mildly negative effect on the eclosion rate of control flies. This suggests that flies are sensitive to the level of dme-mir-10-5p, and increased expression of this miRNA might be a part of the molecular basis of pathology in the HD model. Overexpression of dme-mir-305 alleviated all analyzed phenotypes of Huntington’s flies and had significant detrimental effects on the viability, longevity, and motor activity of control flies [153].

Fukuoka et al. [154] investigated the use of miR-132 as a potential treatment for Huntington’s disease (HD). A recombinant adeno-associated virus (rAAV) miRNA expression system was used to deliver miR-132 to the R6/2 mice. Findings demonstrated that giving Huntington’s mice access to miR-132 enhanced motor function and lengthened their lives. The therapy had no impact on the expression of disease-causing mutant HTT genes and their byproducts. Supplementing with miR-132 may be a treatment strategy for easing Huntington’s symptoms and delaying the onset of the disease without specifically addressing the underlying genetic etiology. Although it might not offer a permanent cure for Huntington’s, it exhibits promise as a prospective therapeutic approach [154].

Degeneration of the medium spiny neurons (MSNs) in the striatum of the basal ganglia is the key pathological feature of Huntington’s [198]. Using a high-throughput miRNA interaction reporter assay (HiTmIR) to study the interactions between miR-34a-5p and HD associated genes, Hart et al. (2023) identified NDUFA9, TAF4B, NRF1, POLR2J2, DNALI1, HIP1, TGM2, and POLR2G as direct miR-34a-5p target genes. Down-regulation of miR-34a-5p in MSNs of R6/2 mice and human HD brain samples might contribute to an elevated intracellular Ca²⁺ level, increased UPR and ROS levels resulting in a pronounced activation of apoptosis. These results lay the ground for future therapeutic interventions using miRNA-34 [155].

Although Huntington’s mainly affects the CNS, Diez-Planelles et al. [199] hypothesized that ubiquitous expression of mutant HTT contributes to disturbances in circulating miRNA profile in symptomatic Huntington’s patients, reflecting changes in both the central nervous system (CNS) and peripheral tissues. To test their hypothesis, they analyzed plasma samples from symptomatic patients with 40–45 CAG repeats in the HTT gene, the most common range among Huntington’s patients, screening for changes in the Huntington’s circulating miRNA profile of healthy matched controls. The Unified Huntington’s Disease Rating Scale and Total Functional Capacity, two disease severity indicators, were found to be linked with the expression levels of specific circulating miRNAs. Additionally, the study found that Huntington’s patients had an overexpression of circulating miRNAs that regulate metabolism. According to these results, circulating miRNA profiles may be used as possible biomarkers for tracking the development of the disease and developing customized treatments for Huntington’s patients [199].

The expression of miRNAs in the prefrontal cortex (BA9) of Huntington’s patients and normal subjects was examined by Hoss et al. [156] Next-generation miRNA sequence analysis was performed on tissue samples from 26 Huntington’s, 2 Huntington’s gene-positive, and 36 control brains. Neuropathological information available for all Huntington’s brains included age at disease onset, CAG-repeat size, Vonsattel grade, and Hadzi-Vonsattel striatal and cortical scores, a continuous measure of the extent of neurodegeneration. Linear models were performed to examine the relationship of miRNA expression to these clinical features, and messenger RNA targets of associated miRNAs were tested for gene ontology term enrichment.

75 differentially-expressed miRNA’s were identified in the tissue samples. Nine were significantly associated with Vonsattel grade of neuropathological involvement and three of these, miR-10b-5p, miR-10b-3p, and miR-302a-3p, significantly related to the Hadzi-Vonsattel striatal score (a continuous measure of striatal involvement) after adjustment for CAG length. Five miRNAs (miR-10b-5p, miR-196a-5p, miR-196b-5p, miR-10b-3p, and miR-106a-5p) were identified as having a significant relationship to CAG length-adjusted age of onset including miR-10b-5p, the mostly strongly overexpressed miRNA in Huntington’s cases. These results provide insight into striatal involvement and support a role for these miRNAs, particularly miR-10b-5p, in HD pathogenicity. The miRNAs identified in our studies of postmortem brain tissue may be detectable in peripheral fluids and thus warrant consideration as accessible biomarkers for disease stage, rate of progression, and other important clinical characteristics of Huntington’s [156].

Chang et al. [157] examined 13 miRNA’s (miR-1, mirR-9, miR-9*, miR-10b, miR-29a, miR-29b, miR-124a, miR-132, miR-155, miR-196a, miR-196b, miR-330, and miR-615) that previously demonstrated alterations in the brains of Huntington’s patients or that are potential regulators of differentially-expressed genes in brains of Huntington’s patients, to examine their expression in the peripheral leukocytes of Huntington’s patients healthy controls. Expression levels of miR-9* were much lower in Huntington’s patients compared with those in healthy controls, suggesting that miR-9* may be involved in the pathogenesis of Huntington’s. miR-9 is highly conserved across vertebrate species and shows brain-specific expression. The complementary strand miR-9* is also crucial for brain development. These findings suggest that miR-9*-mediated gene regulation is important for synaptic plasticity and memory, both of which play an important role in the generation of abnormal movements and psychiatric abnormalities in Huntington’s [157].