**17. Side effects of our immune system**

relevant genes. miR-365 and miR-125b suppress the IL-6/STAT3 pathway in ALS microglia, by targeting IL-6 and STAT3, respectively, and they have a role in overall increase of TNFα mRNA levels. TNFα treatment increases miR-125b levels in microglia; thus it results in abnormal TNFα release. IL-6, a marker of activated microglia, is downregulated after symptom onset in ALS animal models and TNFα is upregulated in G93A mice and in ALS patients. miR-365 and miR-125b dysregulations might have a hand in pathological cytokine profile of ALS. People suggest that the pathogenesis of ALS involves astroglial dysfunction, with a dramatic loss of the excitatory amino acid transporter-2/glial transporter1 (EAAT2/GLT1) in both ALS patients and animal models [68–71]. The EAAT2/GLT1, is an important atroglial transporter, dynamically regulated by neurons, and it is involved in maintenance of extracel‐ lular glutamate concentration below neurotoxic level [72, 73]. The exosomes containing miR-124a and released from neurons can be directly internalized into astrocytes, which results in increase in miR-124a and GLT1 protein levels. miR-124a is downregulated in spinal cord of mutant SOD1 mouse models at end stage of disease, and in vivo injection of miR-124a oligonucleotides into spinal cord of ALS mice results in 30% increase in EAAT2/GLT1 expression. There is very little information about miRNA expression in ALS-induced muscle impairment. There is miR-23 upregulation in skeletal muscle biopsies, collected from ALS patients, and it is correlated with reduction in peroxisome proliferator-activated receptor coactivator-1 (PGC-1alpha) mRNA and protein, observed in both mouse and human muscle diseased samples. This coactivator is involved in muscle mitochondrial biogenesis and function. There is reduction in skeletal muscle mitochondrial function in ALS. It has been suggested that there might be a relation in miR-23a inhibition of PGC-1α and its downstream

It was reported that miR-206, a skeletal muscle-specific miRNA, strongly induced in ALS mouse model is in coincidence with the onset of neurological symptoms. It is a modifier of disease pathogenesis. In G93A-SOD1 mice, loss of miR-206 expression does not affect disease onset apparently, but accelerates disease progression, by skeletal muscle atrophy, kyphosis and paralysis, and reduces survival. In miR-206−/−/G93A-SOD1 mice, neuromuscular junctions (NMJs) are disorganized and reinnervation of denervated muscles by motor axons is post‐ poned in the absence of miR-206. Transcript derived from the *miR-206/133b* locus, which was originally identified as a synapse-associated non-coding RNA called 7H4, has a role in encoding components of the postsynaptic apparatus. The reported 7H4 sequence does not include miR-206; RT-PCR shows that miR-206 sequences are included in this synapse-enriched transcript. Histone deacetylase 4 (HDAC4) mRNA is one of the strongest computationally predicted targets of miR-206. HDAC4 protein expression is upregulated in skeletal muscle of miR-206−/− animals in comparison with wild-type controls after denervation event. *Hdac4* mRNA levels remain unchanged in miR-206−/− mice. It suggests that miR-206 acts in this case

signalling [74, 75].

152 Update on Amyotrophic Lateral Sclerosis

**16. Role of miR-206 in disease impediment**

by translational inhibition rather than by mRNA destabilization [21].

The immune system has a huge role in maintenance of physiological equilibrium within the CNS and in controlling neuronal cell death after adverse effects start. Butovsky et al. [65] performed a comparative analysis of miRNA expression profile of inflammatory monocytes and microglia from SOD1-G93A ALS mice to generate novel biomarkers and possible thera‐ peutics. Spleen monocytes, Ly6Chi, are recruited to the spinal cord where they proliferate during disease progression. They have pronounced proinflammatory profile in both miRNA and gene expression profile. This type of miRNA profile was found maintained in ALS patients and is apparently unique for ALS, when compared with other neuroinflammatory diseases like multiple sclerosis [65]. De Felice et al. [75, 76] did a miRNA expression profile of circulating leukocytes in a small sample of sALS patients and detected variation of eight distinct miRNAs. miR-338-3p expression is upregulated in brain tissue from ALS patients [31]. miRNAs can circulate in cell-free forms in body fluids like serum and plasma and act as signalling molecule between cells [77, 78]. Ever-increasing evidences support the idea that serum circulating secretory vesicles, including exosomes and shedding microvesicles, can function as intercel‐ lular shuttles of RNA and miRNA [60]. Muscle-enriched miR-206 is upregulated not only in SOD1-G93A mice muscles but also there is symptomatic increase in circulation in those mice. The whole transcriptome analysis of both mRNA and miRNA of sALS fibroblasts is done, in order to confirm whether dysregulated processes in the CNS might be reproduced in cells from peripheral tissues. The microprocessor complex gene DGCR8, the gene encoding Dicer enzyme and the RISC proteins AGO1/2 are downregulated in sALS fibroblasts with consequent decrease of multiple miRNAs. This creates a new horizon for generating new diagnostic pool for ALS [21].
