Sporadic Amyotrophic Lateral Sclerosis Skeletal Muscle Transcriptome Analysis: A Comprehensive Examination of Differentially Expressed Genes.

Amyotrophic lateral sclerosis (ALS) that comprises sporadic (sALS) and familial (fALS) cases, is a devastating neurodegenerative disorder characterized by progressive degeneration of motor neurons, leading to muscle atrophy and various clinical manifestations. However, the complex underlying mechanisms affecting this disease are not yet known. On the other hand, there is also no good prognosis of the disease due to the lack of biomarkers and therapeutic targets. Therefore, in this study, by means of bioinformatics analysis, sALS-affected muscle tissue was analyzed using the GEO GSE41414 dataset, identifying 397 differentially expressed genes (DEGs). Functional analysis revealed 320 up-regulated DEGs associated with muscle development and 77 down-regulated DEGs linked to energy metabolism. Protein-protein interaction network analysis identified 20 hub genes, including EIF4A1, HNRNPR and NDUFA4. Furthermore, miRNA target gene networks revealed 17 miRNAs linked to hub genes, with hsa-mir-206, hsa-mir-133b and hsa-mir-100-5p having been previously implicated in ALS. This study presents new potential biomarkers and therapeutic targets for ALS by correlating the information obtained with a comprehensive literature review, providing new potential targets to study their role in ALS.

Sex differences on constitutive long non-coding RNA expression: Modulatory effect of estradiol and testosterone in muscle cells.

INTRODUCTION: Despite the growing awareness of sexual dimorphism between males and females under pathological and physiological conditions, sex bias in biomedical research in animal models and patients is still present nowadays. The main objective of this work was to investigate sex differences in constitutive long non-coding RNA expression in spinal cord and skeletal muscle from wild-type mice. MATERIALS AND METHODS: To assess the influence of gender on long non-coding RNAs, we extracted RNA from tissues of male and female mice and analyzed the expression on nine long non-coding RNAs, selected for being among the most commonly studied or exerting an important role in muscle, at 50, 60, and 120 days of age. RESULTS AND DISCUSSION: We observed age- and tissue-dependent significant sex differences, being more prominent in skeletal muscle. We also studied the effect of sex steroid hormones on long non-coding RNA expression in vitro, noticing a modulation of long non-coding RNA levels upon estradiol and dihydrotestosterone treatment in muscle. CONCLUSIONS: Taken together, results obtained evidenced sex differences on constitutive long non-coding RNA expression and suggested an influence of steroid hormones complementary to other possible factors. These findings emphasize the importance of including both sexes in experimental design to minimize any potential sex bias.

New Insights into Endogenous Retrovirus-K Transcripts in Amyotrophic Lateral Sclerosis.

Retroviral reverse transcriptase activity and the increased expression of human endogenous retroviruses (HERVs) are associated with amyotrophic lateral sclerosis (ALS). We were interested in confirming HERVK overexpression in the ALS brain, its use as an accessory diagnostic marker for ALS, and its potential interplay with neuroinflammation. Using qPCR to analyze HERVK expression in peripheral blood mononuclear cells (PBMCs) and in postmortem brain samples from ALS patients, no significant differences were observed between patients and control subjects. By contrast, we report alterations in the expression patterns of specific HERVK copies, especially in the brainstem. Out of 27 HERVK copies sampled, the relative expression of 17 loci was >1.2-fold changed in samples from ALS patients. In particular, the relative expression of two HERVK copies (Chr3-3 and Chr3-5) was significantly different in brainstem samples from ALS patients compared with controls. Further qPCR analysis of inflammation markers in brain samples revealed a significant increase in NLRP3 levels, while TNFA, IL6, and GZMB showed slight decreases. We cannot confirm global HERVK overexpression in ALS, but we can report the ALS-specific overexpression of selected HERVK copies in the ALS brain. Our data are compatible with the requirement for better patient stratification and support the potential importance of particular HERVK copies in ALS.

Intermediate Repeat Expansion in the ATXN2 Gene as a Risk Factor in the ALS and FTD Spanish Population.

Intermediate CAG expansions in the gene ataxin-2 (ATXN2) are a known risk factor for ALS, but little is known about their role in FTD risk. Moreover, their contribution to the risk and phenotype of patients might vary in populations with different genetic backgrounds. The aim of this study was to assess the relationship of intermediate CAG expansions in ATXN2 with the risk and phenotype of ALS and FTD in the Spanish population. Repeat-primed PCR was performed in 620 ALS and 137 FTD patients in three referral centers in Spain to determine the exact number of CAG repeats. In our cohort, ≥27 CAG repeats in ATXN2 were associated with a higher risk of developing ALS (odds ratio [OR] = 2.666 [1.471-4.882]; p = 0.0013) but not FTD (odds ratio [OR] = 1.446 [0.558-3.574]; p = 0.44). Moreover, ALS patients with ≥27 CAG repeats in ATXN2 showed a shorter survival rate compared to those with <27 repeats (hazard ratio [HR] 1.74 [1.18, 2.56], p = 0.005), more frequent limb onset (odds ratio [OR] = 2.34 [1.093-4.936]; p = 0.028) and a family history of ALS (odds ratio [OR] = 2.538 [1.375-4.634]; p = 0.002). Intermediate CAG expansions of ≥27 repeats in ATXN2 are associated with ALS risk but not with FTD in the Spanish population. ALS patients carrying an intermediate expansion in ATXN2 show more frequent limb onset but a worse prognosis than those without expansions. In patients carrying C9orf72 expansions, the intermediate ATXN2 expansion might increase the penetrance and modify the phenotype.

Analysis of Plasma-Derived Exosomal MicroRNAs as Potential Biomarkers for Canine Idiopathic Epilepsy.

Epilepsy is one of the most prevalent complex neurological diseases in both the canine and human species, with the idiopathic form as its most common diagnosis. MicroRNAs (miRNAs) are small, noncoding RNA molecules that play a role in gene regulation processes and appear to be a promising biological target for convulsion control. These molecules have been reported as constituents of the internal content of exosomes, which are small extracellular vesicles released by cells. In this study, exosome samples were isolated from the plasma of 23 dogs, including 9 dogs with epilepsy responsive to treatment, 6 dogs with drug-resistant epilepsy, and 8 control dogs. Plasma exosomes were then characterized by electron transmission microscopy, nanoparticle tracking analysis, and dot blotting. Afterwards, the microRNA-enriched RNA content of exosomes was isolated, and miRNA quantification was performed by quantitative real-time PCR. Seven circulating miRNAs that have been previously described in the literature as potential diagnostic or prognostic biomarkers for epilepsy were evaluated. We observed significant differences in miR-16 (p < 0.001), miR-93-5p (p < 0.001), miR-142 (p < 0.001), miR-574 (p < 0.01), and miR-27 (p < 0.05) levels in dogs with refractory epilepsy compared to the control group. In drug-sensitive epileptic dogs, miR-142 (p < 0.01) showed significant differences compared to healthy dogs. Moreover, distinct levels of miR-16 (p < 0.05), miR-93-5p (p < 0.01), miR-132 (p < 0.05), and miR-574 (p < 0.05) were also found between drug-sensitive and drug-resistant epileptic dogs. Our results present plasma-circulating exosomes as an advantageous source of epileptic biomarkers, highlighting the potential of miRNAs as prognostic and diagnostic biomarkers of canine idiopathic epilepsy.

Repaglinide Induces ATF6 Processing and Neuroprotection in Transgenic SOD1G93A Mice.

The interaction of the activating transcription factor 6 (ATF6), a key effector of the unfolded protein response (UPR) in the endoplasmic reticulum, with the neuronal calcium sensor Downstream Regulatory Element Antagonist Modulator (DREAM) is a potential therapeutic target in neurodegeneration. Modulation of the ATF6-DREAM interaction with repaglinide (RP) induced neuroprotection in a model of Huntington's disease. Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder with no cure, characterized by the progressive loss of motoneurons resulting in muscle denervation, atrophy, paralysis, and death. The aim of this work was to investigate the potential therapeutic significance of DREAM as a target for intervention in ALS. We found that the expression of the DREAM protein was reduced in the spinal cord of SOD1G93A mice compared to wild-type littermates. RP treatment improved motor strength and reduced the expression of the ALS progression marker collagen type XIXα1 (Col19α1 mRNA) in the quadriceps muscle in SOD1G93A mice. Moreover, treated SOD1G93A mice showed reduced motoneuron loss and glial activation and increased ATF6 processing in the spinal cord. These results indicate that the modulation of the DREAM-ATF6 interaction ameliorates ALS symptoms in SOD1G93A mice.

Mitochondrial Aconitase Enzymatic Activity: A Potential Long-Term Survival Biomarker in the Blood of ALS Patients.

BACKGROUND: Amyotrophic lateral sclerosis (ALS) is a multisystemic, progressive, neurodegenerative disorder. Despite it being generally fatal within a period of 2-4 years, it is highly heterogeneous; as a result, survival periods may vary greatly among individual patients. Biomarkers can serve as tools for diagnosis, prognosis, indicators of therapeutic response, and future therapeutics. Free-radical-dependent mitochondrial damage is believed to play a crucial role in neurodegeneration in ALS. Mitochondrial aconitase, which is also known as aconitase 2 (Aco2), is a key Krebs cycle enzyme and is involved in the regulation of cellular metabolism and iron homeostasis. Aco2 is very sensitive to oxidative inactivation and can aggregate and accumulate in the mitochondrial matrix, causing mitochondrial dysfunction. Loss of Aco2 activity may therefore reflect increased levels of mitochondrial dysfunction due to oxidative damage and could be relevant to ALS pathogenesis. The aim of our study was to confirm changes in mitochondrial aconitase activity in peripheral blood and to determine whether such changes are dependent on, or independent of, the patient's condition and to propose the feasibility of using them as possible valid biomarkers to quantify the progression of the disease and as a predictor of individual prognosis in ALS. METHODS: We measured the Aco2 enzymatic activity in the platelets of blood samples taken from 22 controls and 26 ALS patients at different stages of disease development. We then correlated antioxidant activity with clinical and prognostic variables. RESULTS: Aco2 activity was significantly lower in the 26 ALS patients than in the 22 controls (p < 0.05). Patients with higher levels of Aco2 activity survived longer than those with lower levels (p < 0.05). Aco2 activity was also higher in patients with earlier onset (p < 0.05) and in those with predominantly upper motor neuron signs. CONCLUSIONS: Aco2 activity seems to be an independent factor that could be used in the long-term survival prognosis of ALS. Our findings suggest that blood Aco2 could be a leading candidate for use as a biomarker to improve prognosis. More studies are needed to confirm these results.

Hemizygous Granzyme A Mice Expressing the hSOD1G93A Transgene Show Slightly Extended Lifespan.

Granzyme A (gzmA), a serine protease involved in the modulation of the inflammatory immune response, is found at an elevated level in the serum from ALS patients. However, the influence of gzmA on the progression of ALS remains unclear. The aim of our work was to assess whether the absence of gzmA in an ALS murine model could help slow down the progression of the disease. Homozygous and hemizygous gzmA-deficient mice expressing the hSOD1G93A transgene were generated, and survival of these mice was monitored. Subsequently, gene and protein expression of inflammatory and oxidative stress markers was measured in the spinal cord and quadriceps of these mice. We observed the longest lifespan in gzmA+/- mice. GzmA gene and protein expression was downregulated in the spinal cord and serum from gmzA+/- mice, confirming that the increased survival of hemizygous mice is correlated with lower levels of gzmA. In addition, mRNA and protein levels of glutathione reductase (GSR), involved in oxidative stress, were found downregulated in the spinal cord and quadriceps of gmzA+/- mice, together with lower IL-1ß and IL-6 mRNA levels in hemyzigous mice. In summary, our findings indicate for the first time that reduced levels, but not the absence, of gzmA could slightly ameliorate the disease progression in this animal model.

Lessons to Learn from the Gut Microbiota: A Focus on Amyotrophic Lateral Sclerosis.

The gut microbiota is able to modulate the development and homeostasis of the central nervous system (CNS) through the immune, circulatory, and neuronal systems. In turn, the CNS influences the gut microbiota through stress responses and at the level of the endocrine system. This bidirectional communication forms the "gut microbiota-brain axis" and has been postulated to play a role in the etiopathology of several neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS). Numerous studies in animal models of ALS and in patients have highlighted the close communication between the immune system and the gut microbiota and, therefore, it is possible that alterations in the gut microbiota may have a direct impact on neuronal function and survival in ALS patients. Consequently, if the gut dysbiosis does indeed play a role in ALS-related neurodegeneration, nutritional immunomodulatory interventions based on probiotics, prebiotics, and/or postbiotics could emerge as innovative therapeutic strategies. This review aimed to shed light on the impact of the gut microbiota in ALS disease and on the use of potential nutritional interventions based on different types of biotics to ameliorate ALS symptoms.

Encapsulation of Large-Size Plasmids in PLGA Nanoparticles for Gene Editing: Comparison of Three Different Synthesis Methods.

The development of new gene-editing technologies has fostered the need for efficient and safe vectors capable of encapsulating large nucleic acids. In this work we evaluate the synthesis of large-size plasmid-loaded PLGA nanoparticles by double emulsion (considering batch ultrasound and microfluidics-assisted methodologies) and magnetic stirring-based nanoprecipitation synthesis methods. For this purpose, we characterized the nanoparticles and compared the results between the different synthesis processes in terms of encapsulation efficiency, morphology, particle size, polydispersity, zeta potential and structural integrity of loaded pDNA. Our results demonstrate particular sensibility of large pDNA for shear and mechanical stress degradation during double emulsion, the nanoprecipitation method being the only one that preserved plasmid integrity. However, plasmid-loaded PLGA nanoparticles synthesized by nanoprecipitation did not show cell expression in vitro, possibly due to the slow release profile observed in our experimental conditions. Strong electrostatic interactions between the large plasmid and the cationic PLGA used for this synthesis may underlie this release kinetics. Overall, none of the methods evaluated satisfied all the requirements for an efficient non-viral vector when applied to large-size plasmid encapsulation. Further optimization or alternative synthesis methods are thus in current need to adapt PLGA nanoparticles as delivery vectors for gene editing therapeutic technologies.

Therapeutic Assay with the Non-toxic C-Terminal Fragment of Tetanus Toxin (TTC) in Transgenic Murine Models of Prion Disease.

The non-toxic C-terminal fragment of the tetanus toxin (TTC) has been described as a neuroprotective molecule since it binds to Trk receptors and activates Trk-dependent signaling, activating neuronal survival pathways and inhibiting apoptosis. Previous in vivo studies have demonstrated the ability of this molecule to increase mice survival, inhibit apoptosis and regulate autophagy in murine models of neurodegenerative diseases such as amyotrophic lateral sclerosis and spinal muscular atrophy. Prion diseases are fatal neurodegenerative disorders in which the main pathogenic event is the conversion of the cellular prion protein (PrPC) into an abnormal and misfolded isoform known as PrPSc. These diseases share different pathological features with other neurodegenerative diseases, such as amyotrophic lateral sclerosis, Parkinson's disease or Alzheimer's disease. Hitherto, there are no effective therapies to treat prion diseases. Here, we present a pilot study to test the therapeutic potential of TTC to treat prion diseases. C57BL6 wild-type mice and the transgenic mice Tg338, which overexpress PrPC, were intracerebrally inoculated with scrapie prions and then subjected to a treatment consisting of repeated intramuscular injections of TTC. Our results indicate that TTC displays neuroprotective effects in the murine models of prion disease reducing apoptosis, regulating autophagy and therefore increasing neuronal survival, although TTC did not increase survival time in these models.

Inflammasome in ALS Skeletal Muscle: NLRP3 as a Potential Biomarker.

Since NLRP3 inflammasome plays a pivotal role in several neurodegenerative disorders, we hypothesized that levels of inflammasome components could help in diagnosis or prognosis of amyotrophic lateral sclerosis (ALS). Gene and protein expression was assayed by RT-PCR and Western blot. Spearman's correlation coefficient was used to determine the linear correlation of transcriptional expression levels with longevity throughout disease progression in mice models. Kaplan-Meier analysis was performed to evaluate MCC950 effects (NLRP3 inhibitor) on lifespan of SOD1G93A mice. The results showed significant alterations in NLRP3 inflammasome gene and protein levels in the skeletal muscle of SOD1G93A mice. Spearman's correlation coefficient revealed a positive association between Nlrp3 transcriptional levels in skeletal muscle and longevity of SOD1G93A mice (r = 0.506; p = 0.027). Accordingly, NLRP3 inactivation with MCC950 decreased the lifespan of mice. Furthermore, NLRP3 mRNA levels were significantly elevated in the blood of ALS patients compared to healthy controls (p = 0.03). In conclusion, NLRP3 could be involved in skeletal muscle pathogenesis of ALS, either through inflammasome or independently, and may play a dual role during disease progression. NLRP3 gene expression levels could be used as a biomarker to improve diagnosis and prognosis in skeletal muscle from animal models and also to support diagnosis in clinical practice with the blood of ALS patients.

What skeletal muscle has to say in amyotrophic lateral sclerosis: Implications for therapy.

Amyotrophic lateral sclerosis (ALS) is an adult onset disorder characterized by progressive neuromuscular junction (NMJ) dismantling and degeneration of motor neurons leading to atrophy and paralysis of voluntary muscles responsible for motion and breathing. Except for a minority of patients harbouring genetic mutations, the origin of most ALS cases remains elusive. Peripheral tissues, and particularly skeletal muscle, have lately demonstrated an active contribution to disease pathology attracting a growing interest for these tissues as therapeutic targets in ALS. In this sense, molecular mechanisms essential for cell and tissue homeostasis have been shown to be deregulated in the disease. These include muscle metabolism and mitochondrial activity, RNA processing, tissue-resident stem cell function responsible for muscle regeneration, and proteostasis that regulates muscle mass in adulthood. This review aims to compile scientific evidence that demonstrates the role of skeletal muscle in ALS pathology and serves as reference for development of novel therapeutic strategies targeting this tissue to delay disease onset and progression. LINKED ARTICLES: This article is part of a themed issue on Neurochemistry in Japan. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v178.6/issuetoc.

Competing Endogenous RNA Networks as Biomarkers in Neurodegenerative Diseases.

Protein aggregation is classically considered the main cause of neuronal death in neurodegenerative diseases (NDDs). However, increasing evidence suggests that alteration of RNA metabolism is a key factor in the etiopathogenesis of these complex disorders. Non-coding RNAs are the major contributor to the human transcriptome and are particularly abundant in the central nervous system, where they have been proposed to be involved in the onset and development of NDDs. Interestingly, some ncRNAs (such as lncRNAs, circRNAs and pseudogenes) share a common functionality in their ability to regulate gene expression by modulating miRNAs in a phenomenon known as the competing endogenous RNA mechanism. Moreover, ncRNAs are found in body fluids where their presence and concentration could serve as potential non-invasive biomarkers of NDDs. In this review, we summarize the ceRNA networks described in Alzheimer's disease, Parkinson's disease, multiple sclerosis, amyotrophic lateral sclerosis and spinocerebellar ataxia type 7, and discuss their potential as biomarkers of these NDDs. Although numerous studies have been carried out, further research is needed to validate these complex interactions between RNAs and the alterations in RNA editing that could provide specific ceRNET profiles for neurodegenerative disorders, paving the way to a better understanding of these diseases.

MicroRNA Alterations in a Tg501 Mouse Model of Prion Disease.

MicroRNAs (miRNAs) may contribute to the development and pathology of many neurodegenerative diseases, including prion diseases. They are also promising biomarker candidates due to their stability in body fluids. We investigated miRNA alterations in a Tg501 mouse model of prion diseases that expresses a transgene encoding the goat prion protein (PRNP). Tg501 mice intracranially inoculated with mouse-adapted goat scrapie were compared with age-matched, mock inoculated controls in preclinical and clinical stages. Small RNA sequencing from the cervical spinal cord indicated that miR-223-3p, miR-151-3p, and miR-144-5p were dysregulated in scrapie-inoculated animals before the onset of symptoms. In clinical-stage animals, 23 significant miRNA alterations were found. These miRNAs were predicted to modify the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways including prion disease, extracellular matrix interactions, glutaminergic synapse, axon guidance, and transforming growth factor-beta signaling. MicroRNAs miR-146a-5p (up in cervical spinal cord) and miR-342-3p (down in cervical spinal cord, cerebellum and plasma), both indicated in neurodegenerative diseases earlier, were verified by quantitative real-time polymerase chain reaction (qRT-PCR). Minimal changes observed before the disease onset suggests that most miRNA alterations observed here are driven by advanced prion-associated pathology, possibly limiting their use as diagnostic markers. However, the results encourage further mechanistic studies on miRNA-regulated pathways involved in these neurodegenerative conditions.

Neuroprotective Fragment C of Tetanus Toxin Modulates IL-6 in an ALS Mouse Model.

Neuroinflammation plays a significant role in amyotrophic lateral sclerosis (ALS) pathology, leading to the development of therapies targeting inflammation in recent years. Our group has studied the tetanus toxin C-terminal fragment (TTC) as a therapeutic molecule, showing neuroprotective properties in the SOD1G93A mouse model. However, it is unknown whether TTC could have some effect on inflammation. The objective of this study was to assess the effect of TTC on the regulation of inflammatory mediators to elucidate its potential role in modulating inflammation occurring in ALS. After TTC treatment in SOD1G93A mice, levels of eotaxin-1, interleukin (IL)-2, IL-6 and macrophage inflammatory protein (MIP)-1 alpha (α) and galectin-1 were analyzed by immunoassays in plasma samples, whilst protein expression of caspase-1, IL-1ß, IL-6 and NOD-, LRR- and pyrin domain-containing protein 3 (NLRP3) was measured in the spinal cord, extensor digitorum longus (EDL) muscle and soleus (SOL) muscle. The results showed reduced levels of IL-6 in spinal cord, EDL and SOL in treated SOD1G93A mice. In addition, TTC showed a different role in the modulation of NLRP3 and caspase-1 depending on the tissue analyzed. In conclusion, our results suggest that TTC could have a potential anti-inflammatory effect by reducing IL-6 levels in tissues drastically affected by the disease. However, further research is needed to study more in depth the anti-inflammatory effect of TTC in ALS.

Combined intramuscular and intraspinal transplant of bone marrow cells improves neuromuscular function in the SOD1G93A mice.

BACKGROUND: The simultaneous contribution of several etiopathogenic disturbances makes amyotrophic lateral sclerosis (ALS) a fatal and challenging disease. Here, we studied two different cell therapy protocols to protect both central and peripheral nervous system in a murine model of ALS. METHODS: Since ALS begins with a distal axonopathy, in a first assay, we performed injection of bone marrow cells into two hindlimb muscles of transgenic SOD1G93A mice. In a second study, we combined intramuscular and intraspinal injection of bone marrow cells. Fluorescence-activated cell sorting was used to assess the survival of the transplanted cells into the injected tissues. The mice were assessed from 8 to 16 weeks of age by means of locomotion and electrophysiological tests. After follow-up, the spinal cord was processed for analysis of motoneuron survival and glial cell reactivity. RESULTS: We found that, after intramuscular injection, bone marrow cells were able to engraft within the muscle. However, bone marrow cell intramuscular injection failed to promote a general therapeutic effect. In the second approach, we found that bone marrow cells had limited survival in the spinal cord, but this strategy significantly improved motor outcomes. Moreover, we also found that the dual cell therapy tended to preserve spinal motoneurons at late stages of the disease and to reduce microgliosis, although this did not prolong mice survival. CONCLUSION: Overall, our findings suggest that targeting more than one affected area of the motor system at once with bone marrow cell therapy results in a valuable therapeutic intervention for ALS.

Gene therapy for overexpressing Neuregulin 1 type I in skeletal muscles promotes functional improvement in the SOD1G93A ALS mice.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder affecting motoneurons (MNs), with no effective treatment currently available. The molecular mechanisms that are involved in MN death are complex and not fully understood, with partial contributions of surrounding glial cells and skeletal muscle to the disease. Neuregulin 1 (NRG1) is a trophic factor highly expressed in MNs and neuromuscular junctions. Recent studies have suggested a crucial role of the isoform I (NRG1-I) in the collateral reinnervation process in skeletal muscle, and NRG1-III in the preservation of MNs in the spinal cord, opening a window for developing novel therapies for neuromuscular diseases like ALS. In this study, we overexpressed NRG1-I widely in the skeletal muscles of the SOD1G93A transgenic mouse. The results show that NRG1 gene therapy activated the survival pathways in muscle and spinal cord, increasing the number of surviving MNs and neuromuscular junctions and reducing the astroglial reactivity in the spinal cord of the treated SOD1G93A mice. Furthermore, NRG1-I overexpression preserved motor function and delayed the onset of clinical disease. In summary, our data indicates that NRG1 plays an important role on MN survival and muscle innervation in ALS, and that viral-mediated overexpression of NRG1 isoforms may be considered as a promising approach for ALS treatment.

Impairment of autophagy in scrapie-infected transgenic mice at the clinical stage.

Autophagy appears to play a role in the etiology and progress of misfolded protein disorders. Although this process is dysregulated in prion diseases, it is unknown whether this impairment is a cause or a consequence of prion neuropathology. The study of autophagy during the progress of the disease could elucidate its role. For this purpose, we have investigated its regulation at different stages of the disease in Tg338 mice, a transgenic murine model that overexpresses the highly susceptible ovine VRQ prion protein allele. Mice were intracerebrally inoculated with mouse-adapted classical scrapie and euthanized at the preclinical and clinical stages of the disease. Regulation of autophagy was investigated analyzing the distribution of LC3-B and p62 proteins by immunohistochemistry. Moreover, the expression of genes involved in autophagy regulation was quantified by real-time PCR. LC3-B and p62 proteins were downregulated and upregulated, respectively, in the central nervous system of infected mice with clinical signs of scrapie. Accumulation of p62 correlated with scrapie-related lesions, suggesting an impairment of autophagy in highly prion-affected areas. In addition, Gas5 (growth arrest-specific 5), Atg5 (autophagy-related 5), and Fbxw7 (F-box and WD repeat domain containing 7) transcripts were downregulated in mesencephalon and cervical spinal cord of the same group of animals. The impairment of autophagic machinery seems to be part of the pathological process of scrapie, but only during the late stage of prion infection. Similarities between Tg338 mice and the natural ovine disease make them a reliable in vivo model to study prion infection and autophagy side by side.

Type XIX collagen: a promising biomarker from the basement membranes.

Among collagen members in the collagen superfamily, type XIX collagen has raised increasing interest in relation to its structural and biological roles. Type XIX collagen is a Fibril-Associated Collagen with Interrupted Triple helices member, one main subclass of collagens in this superfamily. This collagen contains a triple helix composed of three polypeptide segments aligned in parallel and it is associated with the basement membrane zone in different tissues. The molecular structure of type XIX collagen consists of five collagenous domains, COL1 to COL5, interrupted by six non-collagenous domains, NC1 to NC6. The most relevant domain by which this collagen exerts its biological roles is NC1 domain that can be cleavage enzymatically to release matricryptins, exerting anti-tumor and anti-angiogenic effect in murine and human models of cancer. Under physiological conditions, type XIX collagen expression decreases after birth in different tissues although it is necessary to keep its basal levels, mainly in skeletal muscle and hippocampal and telencephalic interneurons in brain. Notwithstanding, in amyotrophic lateral sclerosis, altered transcript expression levels show a novel biological effect of this collagen beyond its structural role in basement membranes and its anti-tumor and anti-angiogenic properties. Type XIX collagen can exert a compensatory effect to ameliorate the disease progression under neurodegenerative conditions specific to amyotrophic lateral sclerosis in transgenic SOD1G93A mice and amyotrophic lateral sclerosis patients. This novel biological role highlights its nature as prognostic biomarker of disease progression in and as promising therapeutic target, paving the way to a more precise prognosis of amyotrophic lateral sclerosis.