Neuroinflammation and Epilepsy: From Pathophysiology to Therapies Based on Repurposing Drugs.

Neuroinflammation and epilepsy are different pathologies, but, in some cases, they are so closely related that the activation of one of the pathologies leads to the development of the other. In this work, we discuss the three main cell types involved in neuroinflammation, namely (i) reactive astrocytes, (ii) activated microglia, and infiltration of (iii) peripheral immune cells in the central nervous system. Then, we discuss how neuroinflammation and epilepsy are interconnected and describe the use of different repurposing drugs with anti-inflammatory properties that have been shown to have a beneficial effect in different epilepsy models. This review reinforces the idea that compounds designed to alleviate seizures need to target not only the neuroinflammation caused by reactive astrocytes and microglia but also the interaction of these cells with infiltrated peripheral immune cells.

Exploring miRNA-target gene pair detection in disease with coRmiT.

A wide range of approaches can be used to detect micro RNA (miRNA)-target gene pairs (mTPs) from expression data, differing in the ways the gene and miRNA expression profiles are calculated, combined and correlated. However, there is no clear consensus on which is the best approach across all datasets. Here, we have implemented multiple strategies and applied them to three distinct rare disease datasets that comprise smallRNA-Seq and RNA-Seq data obtained from the same samples, obtaining mTPs related to the disease pathology. All datasets were preprocessed using a standardized, freely available computational workflow, DEG_workflow. This workflow includes coRmiT, a method to compare multiple strategies for mTP detection. We used it to investigate the overlap of the detected mTPs with predicted and validated mTPs from 11 different databases. Results show that there is no clear best strategy for mTP detection applicable to all situations. We therefore propose the integration of the results of the different strategies by selecting the one with the highest odds ratio for each miRNA, as the optimal way to integrate the results. We applied this selection-integration method to the datasets and showed it to be robust to changes in the predicted and validated mTP databases. Our findings have important implications for miRNA analysis. coRmiT is implemented as part of the ExpHunterSuite Bioconductor package available from https://bioconductor.org/packages/ExpHunterSuite.

Autophagy drives the conversion of developmental neural stem cells to the adult quiescent state.

Neurogenesis in the adult mammalian brain relies on the lifelong persistence of quiescent neural stem cell (NSC) reservoirs. Little is known about the mechanisms that lead to the initial establishment of quiescence, the main hallmark of adult stem cells, during development. Here we show that protein aggregates and autophagy machinery components accumulate in developmental radial glia-like NSCs as they enter quiescence and that pharmacological or genetic blockade of autophagy disrupts quiescence acquisition and maintenance. Conversely, increasing autophagy through AMPK/ULK1 activation instructs the acquisition of the quiescent state without affecting BMP signaling, a gatekeeper of NSC quiescence during adulthood. Selective ablation of Atg7, a critical gene for autophagosome formation, in radial glia-like NSCs at early and late postnatal stages compromises the initial acquisition and maintenance of quiescence during the formation of the hippocampal dentate gyrus NSC niche. Therefore, we demonstrate that autophagy is cell-intrinsically required to establish NSC quiescence during hippocampal development. Our results uncover an important role of autophagy in the transition of developmental NSCs into their dormant adult form, paving the way for studies directed at further understanding the mechanisms of stem cell niche formation and maintenance in the mammalian brain.

Beneficial Effect of Fingolimod in a Lafora Disease Mouse Model by Preventing Reactive Astrogliosis-Derived Neuroinflammation and Brain Infiltration of T-lymphocytes.

Lafora disease (LD; OMIM#254780) is a rare, devastating, and fatal form of progressive myoclonus epilepsy that affects young adolescents and has no treatment yet. One of the hallmarks of the disease is the accumulation of aberrant poorly branched forms of glycogen (polyglucosans, PGs) in the brain and peripheral tissues. The current hypothesis is that this accumulation is causative of the pathophysiology of the disease. Another hallmark of LD is the presence of neuroinflammation. We have recently reported the presence of reactive glia-derived neuroinflammation in LD mouse models and defined the main inflammatory pathways that operate in these mice, mainly TNF and IL-6 signaling pathways. In addition, we described the presence of infiltration of peripheral immune cells in the brain parenchyma, which could cooperate and aggravate the neuroinflammatory landscape of LD. In this work, we have checked the beneficial effect of two compounds with the capacity to ameliorate neuroinflammation and reduce leukocyte infiltration into the brain, namely fingolimod and dimethyl fumarate. Our results indicate a beneficial effect of fingolimod in reducing reactive astrogliosis-derived neuroinflammation and T-lymphocyte infiltration, which correlated with the improved behavioral performance of the treated Epm2b-/- mice. On the contrary, dimethyl fumarate, although it was able to reduce reactive astrogliosis, was less effective in preventing neuroinflammation and T-lymphocyte infiltration and in modifying behavioral tests.

Treatment of metastatic squamous cell carcinoma arising in sacrococcygeal pilonidal sinus: a case report series.

Background: Squamous cell carcinoma (SCC) arising in a sacrococcygeal pilonidal sinus is rare, with cases of metastatic disease being even rarer. Among published cases, almost none have reported on systemic treatment. Objective: This disease has a poorer prognosis than other forms of cutaneous SCC; therefore, our objective is to shed some light on the treatment of metastatic disease. Methods: We present a series of nine cases treated at a single center, four of whom received systemic treatment. Additionally, other previously reported cases of metastatic disease are included in an attempt to draw stronger conclusions. Results: Four patients were treated under several treatment regimens, with a median progression-free survival of only 2 months and two instances of partial response (18%). The best result was achieved with cemiplimab. Across all the cases, there was a trend toward a benefit of the use of systemic treatment (HR 0.41, 95% CI 0.15-1.12, p = 0.083; median overall survival 13 vs. 8 months). Limitations: Limitations include the significant lack of information on previously published cases and the extremely heterogeneous nature of the existing information. Conclusion: The initial systemic treatment should be an anti-PD-1, as with other SCCs. After progression on anti-PD-1, there is no strong evidence to support the recommendation of a specific treatment or sequence: options include cetuximab and/or chemotherapy (platinum, paclitaxel, 5-fluorouracyl).

Deciphering the Polyglucosan Accumulation Present in Lafora Disease Using an Astrocytic Cellular Model.

Lafora disease (LD) is a neurological disorder characterized by progressive myoclonus epilepsy. The hallmark of the disease is the presence of insoluble forms of glycogen (polyglucosan bodies, or PGBs) in the brain. The accumulation of PGBs is causative of the pathophysiological features of LD. However, despite the efforts made by different groups, the question of why PGBs accumulate in the brain is still unanswered. We have recently demonstrated that, in vivo, astrocytes accumulate most of the PGBs present in the brain, and this could lead to astrocyte dysfunction. To develop a deeper understanding of the defects present in LD astrocytes that lead to LD pathophysiology, we obtained pure primary cultures of astrocytes from LD mice from the postnatal stage under conditions that accumulate PGBs, the hallmark of LD. These cells serve as novel in vitro models for studying PGBs accumulation and related LD dysfunctions. In this sense, the metabolomics of LD astrocytes indicate that they accumulate metabolic intermediates of the upper part of the glycolytic pathway, probably as a consequence of enhanced glucose uptake. In addition, we also demonstrate the feasibility of using the model in the identification of different compounds that may reduce the accumulation of polyglucosan inclusions.

Development and validation of a clinical score for identifying patients with high risk of latent autoimmune adult diabetes (LADA): The LADA primary care-protocol study.

BACKGROUND: Latent autoimmune diabetes in adults (LADA) is a type of diabetes mellitus showing overlapping characteristics between type 1 Diabetes Mellitus and type 2 Diabetes Mellitus (T2DM), and autoimmunity against insulin-producing pancreatic cells. For its diagnosis, at least one type of anti-pancreatic islet antibody (GADAb is the most common) is required. Many authors recommend performing this measure in all newly diagnosed patients with DM, but it is not possible in Primary Health Care (PHC) due to its high cost. Currently, a relevant proportion of patients diagnosed as T2DM could be LADA. Confusing LADA with T2DM has clinical and safety implications, given its different therapeutic approach. The main objective of the study is to develop and validate a clinical score for identifying adult patients with DM at high risk of LADA in PHC. METHODS: This is an observational, descriptive, cross-sectional study carried out in Primary Care Health Centers with a centralized laboratory. All people over 30 years of age diagnosed with diabetes within a minimum of 6 months and a maximum of 4 years before the start of the study will be recruited. Individuals will be recruited by consecutive sampling. The study variables will be obtained through clinical interviews, physical examinations, and electronic medical records. The following variables will be recorded: those related to Diabetes Mellitus, sociodemographic, anthropometric, lifestyle habits, laboratory parameters, presence of comorbidities, additional treatments, personal or family autoimmune disorders, self-perceived health status, Fourlanos criteria, and LADA diagnosis (as main variable) according to current criteria. DISCUSSION: The study will provide an effective method for identifying patients at increased risk of LADA and, therefore, candidates for antibody testing. However, a slight participation bias is to be expected. Differences between participants and non-participants will be studied to quantify this potential bias.

Age-Related microRNA Overexpression in Lafora Disease Male Mice Provides Links between Neuroinflammation and Oxidative Stress.

Lafora disease is a rare, fatal form of progressive myoclonus epilepsy characterized by continuous neurodegeneration with epileptic seizures, characterized by the intracellular accumulation of aberrant polyglucosan granules called Lafora bodies. Several works have provided numerous evidence of molecular and cellular alterations in neural tissue from experimental mouse models deficient in either laforin or malin, two proteins related to the disease. Oxidative stress, alterations in proteostasis, and deregulation of inflammatory signals are some of the molecular alterations underlying this condition in both KO animal models. Lafora bodies appear early in the animal's life, but many of the aforementioned molecular aberrant processes and the consequent neurological symptoms ensue only as animals age. Here, using small RNA-seq and quantitative PCR on brain extracts from laforin and malin KO male mice of different ages, we show that two different microRNA species, miR-155 and miR-146a, are overexpressed in an age-dependent manner. We also observed altered expression of putative target genes for each of the microRNAs studied in brain extracts. These results open the path for a detailed dissection of the molecular consequences of laforin and malin deficiency in brain tissue, as well as the potential role of miR-155 and miR-146a as specific biomarkers of disease progression in LD.

TNF and IL6/Jak2 signaling pathways are the main contributors of the glia-derived neuroinflammation present in Lafora disease, a fatal form of progressive myoclonus epilepsy.

Lafora disease (LD; OMIM#254780) is a rare form of progressive myoclonus epilepsy (prevalence <1:1,000,000) characterized by the accumulation of insoluble deposits of aberrant glycogen (polyglucosans), named Lafora bodies, in the brain but also in peripheral tissues. LD is the most severe form of the group of progressive myoclonus epilepsies, since patients present a rapid deterioration and dementia with amplification of seizures, leading to death after a decade from the onset of the first symptoms. We have recently described that reactive glia-derived neuroinflammation should be considered a novel hallmark of LD since we observed a florid upregulation of differentially expressed genes in both LD mouse lines, which were mainly related to mediators of inflammatory response. In this work, we define an upregulation of the expression of mediators of the TNF and IL6/JAK2 signaling pathways in LD. In addition, we describe the activation of the non-canonical form of the inflammasome. Furthermore, we describe the infiltration of peripheral immune cells in the brain parenchyma, which could aggravate glia-derived neuroinflammation. Finally, we describe CXCL10 and S100b as blood biomarkers of the disease, which will allow the study of the progression of the disease using serum blood samples. We consider that the identification of these initial inflammatory changes in LD will be very important to implement possible anti-inflammatory therapeutic strategies to prevent the development of the disease.

Papel de la ecocardiografía speckle-tracking (rastreo de marcas) y la resonancia magnética cardíaca-feature tracking en la circulación de Fontan. Diferencias entre los ventrículos derecho e izquierdo / Speckle-tracking echocardiography and MCR-feature tracking in Fontan circulation. Differences between right and left ventricles

Resumen Introducción: En los pacientes con DCPT, la disfunción ventricular es inevitable, y más temprana en VU derechos. La deformación miocárdica por STE y RMC-FT parece promisoria. Objetivo: Analizar la función ventricular mediante STE y RMC-FT en pacientes con DCPT, en comparación con RMC convencional según la morfología del VU y la posible implicación en su diagnóstico temprano. Método: Se recogieron medidas del strain longitudinal y circunferencial por STE y RMC-FT, volúmenes ventriculares y FE por RMC en 64 pacientes con DCPT. Resultados: La morfología ventricular no se relacionó con disfunción por RMC. Los VU derechos tuvieron valores por STE y RMC-FT disminuidos respecto de los VU izquierdos, con FE similares. Existe correlación entre STE y RMC-FT, no equivalentes, con buena factibilidad y reproducibilidad. Conclusiones: La RMC-FT y el STE son técnicas útiles en el diagnóstico temprano y la vigilancia de la función ventricular en VU derechos con FE preservada.

Abstract Introduction: In patients with TCPC, the development of ventricular dysfunction is inevitable and is more precocious in SRVs. Myocardial deformation by STE and CMR-FT is promising. Objective: To analize ventricular function in patients with TCPC using STE and CMR-FT compared with conventional cMRI, depending on SV morphology, to determine their role in early diagnosis of ventricular dysfunction. Method: Sixty-four patients with TCPC were included. Longitudinal and circumferential strain by STE and CMR-FT and ventricular volume and EF were obtained. Results: Dysfunction analyzed by cMRI showed no association with ventricular morphology. SRVs had lower values in STE and CMR-FT compared with SLVs, with similar EF. While not equivalent, correlation was observed between the STE and the CMR-FT values, demonstrating good feasibility and reproducibility. Conclusion: The strain data in CMR-FT and STE could be useful for diagnosis and monitoring of ventricular function and as markers of early SRV dysfunction with preserved EF.

Pharmacological Modulation of Glutamatergic and Neuroinflammatory Pathways in a Lafora Disease Mouse Model.

Lafora disease (LD) is a fatal rare neurodegenerative disorder that affects young adolescents and has no treatment yet. The hallmark of LD is the presence of polyglucosan inclusions (PGs), called Lafora bodies (LBs), in the brain and peripheral tissues. LD is caused by mutations in either EPM2A or EPM2B genes, which, respectively, encode laforin, a glucan phosphatase, and malin, an E3-ubiquitin ligase, with identical clinical features. LD knockout mouse models (Epm2a - / - and Epm2b - / -) recapitulate PG body accumulation, as in the human pathology, and display alterations in glutamatergic transmission and neuroinflammatory pathways in the brain. In this work, we show the results of four pre-clinical trials based on the modulation of glutamatergic transmission (riluzole and memantine) and anti-neuroinflammatory interventions (resveratrol and minocycline) as therapeutical strategies in an Epm2b - / - mouse model. Drugs were administered in mice from 3 to 5 months of age, corresponding to early stage of the disease, and we evaluated the beneficial effect of the drugs by in vivo behavioral phenotyping and ex vivo histopathological brain analyses. The behavioral assessment was based on a battery of anxiety, cognitive, and neurodegenerative tests and the histopathological analyses included a panel of markers regarding PG accumulation, astrogliosis, and microgliosis. Overall, the outcome of ameliorating the excessive glutamatergic neurotransmission present in Epm2b - / - mice by memantine displayed therapeutic effectiveness at the behavioral levels. Modulation of neuroinflammation by resveratrol and minocycline also showed beneficial effects at the behavioral level. Therefore, our study suggests that both therapeutical strategies could be beneficial for the treatment of LD patients. A mouse model of Lafora disease (Epm2b-/-) was used to check the putative beneficial effect of different drugs aimed to ameliorate the alterations in glutamatergic transmission and/or neuroinflammation present in the model. Drugs in blue gave a more positive outcome than the rest.

Solar Cell Parameter Extraction Method from Illumination and Dark I-V Characteristics.

A novel method to extract the seven parameters of the double-diode model of solar cells using the current-voltage (I-V) characteristics under illumination and in the dark is presented. The algorithm consists of two subroutines which are alternatively run to adjust all the parameters of the cell in an iterative process. Curve fitting of the light I-V characteristics ensures accuracy in the prediction of the maximum power point, whereas simultaneously fitting the dark I-V characteristics results in a set of physically meaningful parameters that provide information about the physical performance of the photovoltaic devices. Experimental I-V curves of in-house solar cells are used to validate the proposed parameter extraction method, which can be furthermore applied to other types of p-n junction-based photovoltaic devices.

Neuroprotective Effect of IND1316, an Indole-Based AMPK Activator, in Animal Models of Huntington Disease.

Aggregation of mutant huntingtin, because of an expanded polyglutamine track, underlies the cause of neurodegeneration in Huntington disease (HD). However, it remains unclear how some alterations at the cellular level lead to specific structural changes in HD brains. In this context, the neuroprotective effect of the activation of AMP-activated protein kinase (AMPK) appears to be a determinant factor in several neurodegenerative diseases, including HD. In the present work, we describe a series of indole-derived compounds able to activate AMPK at the cellular level. By using animal models of HD (both worms and mice), we demonstrate the in vivo efficacy of one of these compounds (IND1316), confirming that it can reduce the neuropathological symptoms of this disease. Taken together, in vivo results and in silico studies of druggability, allow us to suggest that IND1316 could be considered as a promising new lead compound for the treatment of HD and other central nervous system diseases in which the activation of AMPK results in neuroprotection.

An empirical pipeline for personalized diagnosis of Lafora disease mutations.

Lafora disease (LD) is a fatal childhood dementia characterized by progressive myoclonic epilepsy manifesting in the teenage years, rapid neurological decline, and death typically within ten years of onset. Mutations in either EPM2A, encoding the glycogen phosphatase laforin, or EPM2B, encoding the E3 ligase malin, cause LD. Whole exome sequencing has revealed many EPM2A variants associated with late-onset or slower disease progression. We established an empirical pipeline for characterizing the functional consequences of laforin missense mutations in vitro using complementary biochemical approaches. Analysis of 26 mutations revealed distinct functional classes associated with different outcomes that were supported by clinical cases. For example, F321C and G279C mutations have attenuated functional defects and are associated with slow progression. This pipeline enabled rapid characterization and classification of newly identified EPM2A mutations, providing clinicians and researchers genetic information to guide treatment of LD patients.

Gene expression analysis method integration and co-expression module detection applied to rare glucide metabolism disorders using ExpHunterSuite.

High-throughput gene expression analysis is widely used. However, analysis is not straightforward. Multiple approaches should be applied and methods to combine their results implemented and investigated. We present methodology for the comprehensive analysis of expression data, including co-expression module detection and result integration via data-fusion, threshold based methods, and a Naïve Bayes classifier trained on simulated data. Application to rare-disease model datasets confirms existing knowledge related to immune cell infiltration and suggest novel hypotheses including the role of calcium channels. Application to simulated and spike-in experiments shows that combining multiple methods using consensus and classifiers leads to optimal results. ExpHunter Suite is implemented as an R/Bioconductor package available from https://bioconductor.org/packages/ExpHunterSuite . It can be applied to model and non-model organisms and can be run modularly in R; it can also be run from the command line, allowing scalability with large datasets. Code and reports for the studies are available from https://github.com/fmjabato/ExpHunterSuiteExamples .

Beneficial Effects of Metformin on the Central Nervous System, with a Focus on Epilepsy and Lafora Disease.

Metformin is a drug in the family of biguanide compounds that is widely used in the treatment of type 2 diabetes (T2D). Interestingly, the therapeutic potential of metformin expands its prescribed use as an anti-diabetic drug. In this sense, it has been described that metformin administration has beneficial effects on different neurological conditions. In this work, we review the beneficial effects of this drug as a neuroprotective agent in different neurological diseases, with a special focus on epileptic disorders and Lafora disease, a particular type of progressive myoclonus epilepsy. In addition, we review the different proposed mechanisms of action of metformin to understand its function at the neurological level.

TRIM32 and Malin in Neurological and Neuromuscular Rare Diseases.

Tripartite motif (TRIM) proteins are RING E3 ubiquitin ligases defined by a shared domain structure. Several of them are implicated in rare genetic diseases, and mutations in TRIM32 and TRIM-like malin are associated with Limb-Girdle Muscular Dystrophy R8 and Lafora disease, respectively. These two proteins are evolutionary related, share a common ancestor, and both display NHL repeats at their C-terminus. Here, we revmniew the function of these two related E3 ubiquitin ligases discussing their intrinsic and possible common pathophysiological pathways.

Modulators of Neuroinflammation Have a Beneficial Effect in a Lafora Disease Mouse Model.

Lafora disease (LD; OMIM#274780) is a fatal rare neurodegenerative disorder characterized by generalized epileptic seizures and the presence of polyglucosan inclusions (PGs), called Lafora bodies (LBs), typically in the brain. LD is caused by mutations in two genes EPM2A or EPM2B, which encode respectively laforin, a glucan phosphatase, and malin, an E3-ubiquitin ligase. Much remains unknown about the molecular bases of LD and, unfortunately, appropriate treatment is still missing; therefore patients die within 10 years from the onset of the disease. Recently, we have identified neuroinflammation as one of the initial determinants in LD. In this work, we have investigated anti-inflammatory treatments as potential therapies in LD. With this aim, we have performed a preclinical study in an Epm2b-/- mouse model with propranolol, a ß-adrenergic antagonist, and epigallocatechin gallate (EGCG), an antioxidant from green tea extract, both of which displaying additional anti-inflammatory properties. In vivo motor and cognitive behavioral tests and ex vivo histopathological brain analyses were used as parameters to assess the therapeutic potential of propranolol and EGCG. After 2 months of treatment, we observed an improvement not only in attention defects but also in neuronal disorganization, astrogliosis, and microgliosis present in the hippocampus of Epm2b-/- mice. In general, propranolol intervention was more effective than EGCG in preventing the appearance of astrocyte and microglia reactivity. In summary, our results confirm the potential therapeutic effectiveness of the modulators of inflammation as novel treatments in Lafora disease.

Endocytosis of the glutamate transporter 1 is regulated by laforin and malin: Implications in Lafora disease.

Lafora disease (LD) is a fatal rare type of progressive myoclonus epilepsy that appears during early adolescence. The disease is caused by mutations in EPM2A or EPM2B genes, which encode laforin, a glucan phosphatase, and malin, an E3-ubiquitin ligase, respectively. Although the exact roles of laforin and malin are still not well understood, it is known that they work as a complex in which laforin recruits targets that will be ubiquitinated by malin. Recently, we suggested that the type of epilepsy that accompanies LD could be due to deficiencies in the function of the astrocytic glutamate transporter GLT-1. We described that astrocytes from LD mouse models presented decreased levels of GLT-1 at the plasma membrane, leading to increased levels of glutamate in the brain parenchyma. In this work, we present evidence indicating that in the absence of a functional laforin/malin complex (as in LD cellular models) there is an alteration in the ubiquitination of GLT-1, which could be the cause of the reduction in the levels of GLT-1 at the plasma membrane. On the contrary, overexpression of the laforin/malin complex promotes the retention of GLT-1 at the plasma membrane. This retention may be due to the direct ubiquitination of GLT-1 and/or to an opposite effect of this complex on the dynamics of the Nedd4.2-mediated endocytosis of the transporter. This work, therefore, presents new pieces of evidence on the regulation of GLT-1 by the laforin/malin complex, highlighting its value as a therapeutic target for the amelioration of the type of epilepsy that accompanies LD.

Neuroinflammation and progressive myoclonus epilepsies: from basic science to therapeutic opportunities.

Progressive myoclonus epilepsies (PMEs) are a group of genetic neurological disorders characterised by the occurrence of epileptic seizures, myoclonus and progressive neurological deterioration including cerebellar involvement and dementia. The primary cause of PMEs is variable and alterations in the corresponding mutated genes determine the progression and severity of the disease. In most cases, they lead to the death of the patient after a period of prolonged disability. PMEs also share poor information on the pathophysiological bases and the lack of a specific treatment. Recent reports suggest that neuroinflammation is a common trait under all these conditions. Here, we review similarities and differences in neuroinflammatory response in several PMEs and discuss the window of opportunity of using anti-inflammatory drugs in the treatment of several of these conditions.