Neddylation orchestrates the complex transcriptional and posttranscriptional program that drives Schwann cell myelination.

Myelination is essential for neuronal function and health. In peripheral nerves, >100 causative mutations have been identified that cause Charcot-Marie-Tooth disease, a disorder that can affect myelin sheaths. Among these, a number of mutations are related to essential targets of the posttranslational modification neddylation, although how these lead to myelin defects is unclear. Here, we demonstrate that inhibiting neddylation leads to a notable absence of peripheral myelin and axonal loss both in developing and regenerating mouse nerves. Our data indicate that neddylation exerts a global influence on the complex transcriptional and posttranscriptional program by simultaneously regulating the expression and function of multiple essential myelination signals, including the master transcription factor EGR2 and the negative regulators c-Jun and Sox2, and inducing global secondary changes in downstream pathways, including the mTOR and YAP/TAZ signaling pathways. This places neddylation as a critical regulator of myelination and delineates the potential pathogenic mechanisms involved in CMT mutations related to neddylation.

External validation of biomarkers for immune-related adverse events after immune checkpoint inhibition.

Immune checkpoint inhibitors have revolutionized treatment of advanced melanoma, but commonly cause serious immune-mediated complications. The clinical ambition of reserving more aggressive therapies for patients least likely to experience immune-related adverse events (irAE) has driven an extensive search for predictive biomarkers. Here, we externally validate the performance of 59 previously reported markers of irAE risk in a new cohort of 110 patients receiving Nivolumab (anti-PD1) and Ipilimumab (anti-CTLA-4) therapy. Alone or combined, the discriminatory value of these routine clinical parameters and flow cytometry biomarkers was poor. Unsupervised clustering of flow cytometry data returned four T cell subsets with higher discriminatory capacity for colitis than previously reported populations, but they cannot be considered as reliable classifiers. Although mechanisms predisposing some patients to particular irAEs have been described, we are presently unable to capture adequate information from pre-therapy flow cytometry and clinical data to reliably predict risk of irAE in most cases.

GABA Receptor Agonists Protect From Excitotoxic Damage Induced by AMPA in Oligodendrocytes.

Oligodendrocytes are the myelin forming cells of the central nervous system, and their vulnerability to excitotoxicity induced by glutamate contributes to the pathogenesis of neurological disorders including brain ischemia and neurodegenerative diseases, such as multiple sclerosis. In addition to glutamate receptors, oligodendrocytes express GABA receptors (GABAR) that are involved in their survival and differentiation. The interactions between glutamate and GABAergic systems are well documented in neurons, under both physiological and pathological conditions, but this potential crosstalk in oligodendrocytes has not been studied in depth. Here, we evaluated the protective effect of GABAR agonists, baclofen (GABAB) and muscimol (GABAA), against AMPA-induced excitotoxicity in cultured rat oligodendrocytes. First, we observed that both baclofen and muscimol reduced cell death and caspase-3 activation after AMPA insult, proving their oligoprotective potential. Interestingly, analysis of the cell-surface expression of calcium-impermeable GluR2 subunits in oligodendrocytes revealed that GABAergic agonists significantly reverted GluR2 internalization induced by AMPA. We determined that baclofen and muscimol also impaired AMPA-induced intracellular calcium increase and subsequent mitochondrial membrane potential alteration, ROS generation, and calpain activation. However, AMPA-triggered activation of Src, Akt, JNK and CREB was not affected by baclofen or muscimol. Overall, our results suggest that GABAR activation initiates alternative molecular mechanisms that attenuate AMPA-mediated apoptotic excitotoxicity in oligodendrocytes by interfering with expression of GluR subunits in membranes and with calcium-dependent intracellular signaling pathways. Together, these findings provide evidence of GABAR agonists as potential oligodendroglial protectants in central nervous system disorders.

GABAB receptor agonist baclofen promotes central nervous system remyelination.

Promoting remyelination is considered as a potential neurorepair strategy to prevent/limit the development of permanent neurological disability in patients with multiple sclerosis (MS). To this end, a number of clinical trials are investigating the potential of existing drugs to enhance oligodendrocyte progenitor cell (OPC) differentiation, a process that fails in chronic MS lesions. We previously reported that oligodendroglia express GABAB receptors (GABAB Rs) both in vitro and in vivo, and that GABAB R-mediated signaling enhances OPC differentiation and myelin protein expression in vitro. Our goal here was to evaluate the pro-remyelinating potential of GABAB R agonist baclofen (Bac), a clinically approved drug to treat spasticity in patients with MS. We first demonstrated that Bac increases myelin protein production in lysolecithin (LPC)-treated cerebellar slices. Importantly, Bac administration to adult mice following induction of demyelination by LPC injection in the spinal cord resulted in enhanced OPC differentiation and remyelination. Thus, our results suggest that Bac repurposing should be considered as a potential therapeutic strategy to stimulate remyelination in patients with MS.

Genetically modified macrophages accelerate myelin repair.

Preventing neurodegeneration-associated disability progression in patients with multiple sclerosis (MS) remains an unmet therapeutic need. As remyelination prevents axonal degeneration, promoting this process in patients might enhance neuroprotection. In demyelinating mouse lesions, local overexpression of semaphorin 3F (Sema3F), an oligodendrocyte progenitor cell (OPC) attractant, increases remyelination. However, molecular targeting to MS lesions is a challenge. A clinically relevant paradigm for delivering Sema3F to demyelinating lesions could be to use blood-derived macrophages as vehicles. Thus, we chose transplantation of genetically modified hematopoietic stem cells (HSCs) as means of obtaining chimeric mice with circulating Sema3F-overexpressing monocytes. We demonstrated that Sema3F-transduced HSCs stimulate OPC migration in a neuropilin 2 (Nrp2, Sema3F receptor)-dependent fashion, which was conserved in middle-aged OPCs. While demyelinating lesions induced in mice with Sema3F-expressing blood cells showed no changes in inflammation and OPC survival, OPC recruitment was enhanced which accelerated the onset of remyelination. Our results provide a proof of concept that blood cells, particularly monocytes/macrophages, can be used to deliver pro-remyelinating agents "at the right time and place," suggesting novel means for remyelination-promoting strategies in MS.

Development of a Flow Cytometry Assay to Predict Immune Checkpoint Blockade-Related Complications.

Treatment of advanced melanoma with combined immune checkpoint inhibitor (ICI) therapy is complicated in up to 50% of cases by immune-related adverse events (irAE) that commonly include hepatitis, colitis and skin reactions. We previously reported that pre-therapy expansion of cytomegalovirus (CMV)-reactive CD4+ effector memory T cells (TEM) predicts ICI-related hepatitis in a subset of patients with Stage IV melanoma given αPD-1 and αCTLA-4. Here, we develop and validate a 10-color flow cytometry panel for reliably quantifying CD4+ TEM cells and other biomarkers of irAE risk in peripheral blood samples. Compared to previous methods, our new panel performs equally well in measuring CD4+ TEM cells (agreement = 98%) and is superior in resolving CD4+ CD197+ CD45RA- central memory T cells (TCM) from CD4+ CD197+ CD45RA+ naive T cells (Tnaive). It also enables us to precisely quantify CD14+ monocytes (CV = 6.6%). Our new "monocyte and T cell" (MoT) assay predicts immune-related hepatitis with a positive predictive value (PPV) of 83% and negative predictive value (NPV) of 80%. Our essential improvements open the possibility of sharing our predictive methods with other clinical centers. Furthermore, condensing measurements of monocyte and memory T cell subsets into a single assay simplifies our workflows and facilitates computational analyses.

Virus-specific memory T cell responses unmasked by immune checkpoint blockade cause hepatitis.

Treatment of advanced melanoma with combined PD-1/CTLA-4 blockade commonly causes serious immune-mediated complications. Here, we identify a subset of patients predisposed to immune checkpoint blockade-related hepatitis who are distinguished by chronic expansion of effector memory CD4+ T cells (TEM cells). Pre-therapy CD4+ TEM cell expansion occurs primarily during autumn or winter in patients with metastatic disease and high cytomegalovirus (CMV)-specific serum antibody titres. These clinical features implicate metastasis-dependent, compartmentalised CMV reactivation as the cause of CD4+ TEM expansion. Pre-therapy CD4+ TEM expansion predicts hepatitis in CMV-seropositive patients, opening possibilities for avoidance or prevention. 3 of 4 patients with pre-treatment CD4+ TEM expansion who received αPD-1 monotherapy instead of αPD-1/αCTLA-4 therapy remained hepatitis-free. 4 of 4 patients with baseline CD4+ TEM expansion given prophylactic valganciclovir and αPD-1/αCTLA-4 therapy remained hepatitis-free. Our findings exemplify how pathogen exposure can shape clinical reactions after cancer therapy and how this insight leads to therapeutic innovations.

Salud mental, procesos de crianza e infancias / Mental health, parenting processes and childhoods

INTRODUCCIÓN estudio que aborda nueves experiencias comunitarias vinculadas a la salud mental y los procesos de crianza de las infancias en las provincias de Neuquén, Río Negro y mLa Pampa. OBJETIVO comprender experiencias comunitarias vinculadas a la salud mental y los crianza de las infancias en las provincias de Neuquén, Río Negro y La Pampa durante el período 2020-2022; las transformaciones por ellas vivenciadas en el contexto de la pandemia y las medidas de distanciamiento y aislamiento social implementadas; y los conocimientos que surgen de ellas en términos del diseño y revisión de las políticas públicas vinculadas a la salud mental y los procesos de crianza de las infancias. METODOLOGÍA estudio descriptivo transversal con un enfoque eminentemente cualitativo que incluyó los aportes de la etnografía y la etnografía virtual siendo que los instrumentos de recolección de datos utilizados fueron múltiples. El análisis de datos se basó en el concepto de triangulación metodológica y estará inspirado en la teoría fundamentada y en el método de comparación constante. RESULTADOS en ellos se presenta la caracterización de cada una de las nueve experiencias comunitarias estudiadas desde las dimensiones de análisis propuestas en el marco teórico objetivos, acciones, sentidos, emocionalidades, enseñanzas para las políticas públicas; transformaciones vivenciadas durante el tiempo de pandemia; etc. DISCUSIÓN si bien las experiencias investigadas son muy diferentes entre sí, todas coinciden en haber partido de la escucha de las necesidades y las voces de las infancias y adolescencias orientando sus acciones a dar visibilidad y a validar otras maneras de ser, conocer y sentir este mundo al tiempo que todas han generado un tipo de politicidad en clave femenina que sigue un camino anfibio planteando luchas tanto desde fuera del Estado como desde él.

Expression and Function of GABA Receptors in Myelinating Cells.

Myelin facilitates the fast transmission of nerve impulses and provides metabolic support to axons. Differentiation of oligodendrocyte progenitor cells (OPCs) and Schwann cell (SC) precursors is critical for myelination during development and myelin repair in demyelinating disorders. Myelination is tightly controlled by neuron-glia communication and requires the participation of a wide repertoire of signals, including neurotransmitters such as glutamate, ATP, adenosine, or γ-aminobutyric acid (GABA). GABA is the main inhibitory neurotransmitter in the central nervous system (CNS) and it is also present in the peripheral nervous system (PNS). The composition and function of GABA receptors (GABARs) are well studied in neurons, while their nature and role in glial cells are still incipient. Recent studies demonstrate that GABA-mediated signaling mechanisms play relevant roles in OPC and SC precursor development and function, and stand out the implication of GABARs in oligodendrocyte (OL) and SC maturation and myelination. In this review, we highlight the evidence supporting the novel role of GABA with an emphasis on the molecular identity of the receptors expressed in these glial cells and the possible signaling pathways involved in their actions. GABAergic signaling in myelinating cells may have potential implications for developing novel reparative therapies in demyelinating diseases.

Clinical characterization of delayed alcohol-induced headache: A study of 1,108 participants.

OBJECTIVE: To evaluate the International Classification of Headache Disorders (ICHD) criteria and to characterize the clinical phenotype of delayed alcohol-induced headache (DAIH). METHODS: We conducted a cross-sectional study of university students who voluntarily consumed alcohol and experienced headache. Participants completed a survey that included demographic and clinical data. We analyzed the phenotype of the headache, validated ICHD phenotype criteria for DAIH, and analyzed whether participants fulfilled criteria for low-CSF-pressure headache or migraine. RESULTS: A total of 1,108 participants were included (58% female, mean age 23 years, 41% with headache history). Mean alcohol intake was 158 g; spirits were consumed by 60% of the participants; beer was consumed by 41%; and wine was consumed by 18%. The ICHD criteria for DAIH were met in 95% of the participants. Headache duration (mean, 6.7 hours) correlated with total grams of alcohol consumed (r = 0.62, p = 0.03). Pain was bilateral in 85% of patients with predominantly frontal topography (43%). Pain quality was mainly pressing (60%) or pulsatile (39%) and was aggravated by physical activity in 83% of participants. ICHD low-CSF pressure-headache criteria were fulfilled in 58% of patients, and migraine criteria were fulfilled by 36%. CONCLUSIONS: DAIH is a moderate-intensity headache, is typically bilateral, and presents with frontal predominance and a pressing quality. The phenotype of DAIH combines features of both migraine and low-CSF-pressure headaches.

Oligodendrocyte Differentiation and Myelination Is Potentiated via GABAB Receptor Activation.

Differentiation of oligodendrocyte precursor cells (OPCs) into mature oligodendrocytes (OLs) is a key event for axonal myelination in the central nervous system (CNS). Several growth factors and neurotransmitters like GABA are postulated as important regulators of that process, and different protein kinases may also participate in OL differentiation and myelination. However, the molecular mechanisms underlying the regulation of myelination by neurotransmitters are only partially known. In the present study, we provide evidence showing that GABA receptors (GABARs) play an important role in OL differentiation. First, we observed that OPCs and OLs synthesize GABA and expressed GABAR and transporters, both in vitro and in vivo and, in contrast to GABAARs, the subunits GABAB1R and GABAB2R are expressed in OLs over time. Then, we found that exogenous GABA increases the number of myelin segments and MBP expression in DRG-OPC cocultures, indicating that GABA regulates myelination when OLs are in contact with axons. Notably, in purified rat OPC cultures, chronic treatment with GABA and baclofen, specific GABABR agonist, accelerates OPC differentiation by enhancing the processes branching and myelin protein expression, effects that are reverted in presence of GABABR specific antagonist CGP55845. Exposure of OPCs to baclofen promotes the Src-phosphorylation, and the baclofen-induced maturation is attenuated in presence of the Src-family kinases inhibitor PP2. None of these effects are mediated by the GABAAR agonist muscimol. Together, these results highlight the relevance of the GABAergic system in OL differentiation, and indicate that this functional role is mediated through GABABR involving the participation of Src-family kinases. This article is part of a Special Issue entitled: Honoring Ricardo Miledi - outstanding neuroscientist of XX-XXI centuries.

Solid Lipid Nanoparticles Surface Modification Modulates Cell Internalization and Improves Chemotoxic Treatment in an Oral Carcinoma Cell Line.

Solid lipid nanoparticles (SLN) present low toxicity, versatility to incorporate both lipophilic and hydrophilic drugs, controlled drug release and they are easy to scale-up. It is well known that the endocytosis pathway by which SLN are taken up and the subsequent subcellular distribution are crucial for the biological effect of the incorporated drug. In addition, interactions between SLN and cells depend on many factors, such as, the composition of nanoparticle surface. In this work different amounts of phosphatidylethanolamine polyethylene glycol (PE⁻PEG) were added to SLN composed of stearic acid, Epikuron 200 and sodium taurodeoxycholate. Characterization of obtained nanoparticle suspensions were performed by the analysis of particle size, polydispersity index, ζ-potential, cell toxicity and cell internalization pathway. We have observed that the presence of PE⁻PEG improves active cell internalization of the nanoparticles in an oral adenocarcinoma cell line, reducing non-specific internalization mechanisms. Finally, we have tested the effect of surface coating on the efficiency of incorporated drugs using all-trans retinoic acid as a model drug. We have observed that delivery of this drug into PE⁻PEG coated SLN increases its chemotoxic effect compared to non-coated SLN. Therefore, it can be concluded that surface modification with PE⁻PEG improves the efficiency and the specificity of the SLN-loaded drug.

Application of Solid Lipid Nanoparticles to Improve the Efficiency of Anticancer Drugs.

Drug delivery systems have opened new avenues to improve the therapeutic effects of already-efficient molecules. Particularly, Solid Lipid Nanoparticles (SLNs) have emerged as promising nanocarriers in cancer therapy. SLNs offer remarkable advantages such as low toxicity, high bioavailability of drugs, versatility of incorporation of hydrophilic and lipophilic drugs, and feasibility of large-scale production. Their molecular structure is crucial to obtain high quality SLN preparations and it is determined by the relationship between the composition and preparation method. Additionally, SLNs allow overcoming several physiological barriers that hinder drug delivery to tumors and are also able to escape multidrug resistance mechanisms, characteristic of cancer cells. Focusing on cell delivery, SLNs can improve drug delivery to target cells by different mechanisms, such as passive mechanisms that take advantage of the tumor microenvironment, active mechanisms by surface modification of SLNs, and codelivery mechanisms. SLNs can incorporate many different drugs and have proven to be effective in different types of tumors (i.e., breast, lung, colon, liver, and brain), corroborating their potential. Finally, it has to be taken into account that there are still some challenges to face in the application of SLNs in anticancer treatments but their possibilities seem to be high.

Buenas prácticas en salud mental infantil / Good practices in child mental health

INTRODUCCIÓN Estudio cualitativo multicéntrico acerca de las modalidades de atención y actividades de salud mental orientadas a niñ@s desarrolladas en los Sistemas Públicos de Salud de las Provincias de Río Negro y Neuquén, período 2016-2017. OBJETIVO Caracterizar en profundidad un conjunto específico de dispositivos de salud mental/psicosocial orientados a niñ@s seleccionados siguiendo criterios de buenas prácticas en salud. MÉTODO Los instrumentos de recolección de datos fueron; la revisión de documentación sanitaria existente; la realización de entrevistas en profundidad a los equipos profesionales responsables de los dispositivos y a los beneficiarios de los mismos; y la realización de observaciones de las distintas experiencias con apoyatura de registro fotográfico y videos; el análisis de datos se basó en el concepto de triangulación metodológica y estuvo inspirado en la teoría fundamentada y en el método de comparación constante. RESULTADOS Se caracterizaron un total de catorce dispositivos; en Río Negro; Admisión e Interconsultas, Hospital Cipolletti; Taller de Niños Divertidos, ADANIL, Hospital General Roca; Consejo de Niños, Salud Mental, Hospital General Roca; La Huerta para Compartir, Hospital Villa Regina; Futbol Callejero, El Bolsón; en Neuquén: Grupo de Padres de Bebés Prematuros Internados, Hospital Castro Rendón; Murguita Trapitos de Colores, CAPS Confluencia; Taller de Crianza, Hospital Bouquet Roldán; La Casita Itinerante, CAPS Parque Industrial; Red Interinstitucional, CAPS Don Bosco; Actividad Sala de Espera y Taller de Salud Sexual y Afectiva, Hospital Mariano Moreno; Grupo Psicoterapéutico de Niños, Hospital Horacio Heller. DISCUSIÓN A partir de la caracterización realizada, se re-conceptualizaron los criterios de buenas prácticas en salud mental infantil adquiriendo especial énfasis la consideración de l@s niñ@s como sujet@s y la necesidad del abordaje del entorno inmediato en el que el niñ@ vive como criterios de buenas prácticas

Variation revealed by SNP genotyping and morphology provides insight into the origin of the tomato.

Tomato, Solanum lycopersicum, is divided into two widely distributed varieties: the cultivated S. lycopersicum var. lycopersicum, and the weedy S. lycopersicum var. cerasiforme. Solanum pimpinellifolium is the most closely related wild species of tomato.The roles of S. pimpinellifolium and S. l. cerasiforme during the domestication of tomato are still under debate. Some authors consider S. l. cerasiforme to be the ancestor, whereas others think that S. l. cerasiforme is an admixture of S. pimpinellifolium and the cultivated S. l. lycopersicum. It is also not clear whether the domestication occurred in the Andean region or in Mesoamerica. We characterized 272 accessions (63 S. pimpinellifolium, 106 S. l. cerasiforme, 95 S. l. lycopersicum and 8 derived from hybridization processes) were morphologically and genetically using the SolCap platform (7,414 SNPs). The two species were distinguished in a PCA analysis and displayed a rich geographic structure. Solanum lycopersicum var. cerasiforme and S. l. lycopersicum were also differentiated in the PCA and Structure analyses, which supports maintaining them as different varieties. Solanum pimpinellifolium and the Andean S. l. cerasiforme were more diverse than the non-Andean S. lycopersicum. Solanum lycopersicum var. cerasiforme was morphologically and molecularly intermediate between S. pimpinellifolium and tomato. Solanum lycopersicum var. cerasiforme, with the exception of several Ecuadorian and Mexican accessions, is composed of the products of admixture processes according to the Structure analysis. The non-admixtured S. l. cerasiforme might be similar to the ancestral cultivars from which the cultivated tomato originated, and presents remarkable morphological diversity, including fruits of up to 6 cm in diameter. The data obtained would fit a model in which a pre-domestication took place in the Andean region, with the domestication being completed in Mesoamerica. Subsequently, the Spaniards took plants from Mesoamerica to Spain and from there they were exported to the rest of the world.

Internación por cuidados progresivos: propuesta metodológica para su implementación en un hospital general de agudos / Progressive patient care

Implementar un sistema de cuidados progresivos y atención interdisciplinaria en los hospitales generales del subsector público representa un desafío para el sistema de atención de la salud. El modelo, ya presente en un importante número de establecimientos del subsector privado y de la seguridad social, ubica a los pacientes en sectores de internación según sus necesidades de cuidado y dependencia, y no según las especialidades médicas tradicionales. En el presente artículo se presenta el marco conceptual y una propuesta metodológica para su efectiva implementación en un hospital público venciendo las naturales resistencias al cambio.

Implementation of progressive care patients and interdisciplinary care models in General Hospitals has been challengin for Public Health Care Sistems. The model actualy present in some Social Security and Private Hospitals place the patients in areas or units on the basis of their needs for care as deter­minated by the degree of illnes rather than on the basis of a medical specialty. This article describes the models theory and how it is posible to implement in a Public General Hospital despite of the natural resistence to change.

Transformation of Mucor miehei results in plasmid deletion and phenotypic instability.

Mucor miehei transformants resistant to geneticin have been obtained by treatment of protoplasts with different plasmids and by Agrobacterium-mediated DNA transfer. All transformants exhibited a very unstable phenotype whose maintenance required continuous selective pressure. Molecular analysis of transformants showed that the plasmid DNA was extensively modified and maintained in very low amount. Our results indicate that M. miehei reluctance to transformation is due to different causes, including the coenocytic nature of its mycelium and the existence of specific mechanisms for the detection and elimination of foreign DNA.