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Perinatal asphyxia (PA) is a medical condition associated with a high short-term morbimortality and different long-term neurological diseases. In previous works, we have shown that neuronal and synaptic changes in rat striatum lead to ubi-protein accumulation in post-synaptic density (PSD) after six months of sub-severe PA. However, very little is known about the synaptic and related structural modifications induced by PA in young rats. In the present work, we studied neuronal cytoskeleton modifications in striatum induced by subsevere PA in 30-day-old rats. We observed a significant decrease in the number of neurons, in particular calbindin immunoreactive neurons after PA. In addition, it was also observed that actin cytoskeleton was highly modified in the PSD as well as an increment of F-actin staining by Phalloidin-alexa488 in the striatum of PA rats. Using correlative fluorescence-electron microscopy photooxidation, we confirmed and extended confocal observations. F-actin staining augmentation was mostly related with an increment in the number of mushroom-shaped spines. Consistent with microscopic data, Western blot analysis revealed a β-actin increment in PSD in PA rats. On the other hand, MAP-2 immunostaining was decreased after PA, being NF-200 expression unmodified. Although neuronal death was observed, signs of generalized neurodegeneration were absent. Taken together these results showed early post-synaptic F-actin cytoskeleton changes induced by PA with slightly modifications in the other components of the neuronal cytoskeleton, suggesting that F-actin accumulation in the dendritic spines could be involved in the neuronal loss induced by PA. Synapse, 2012. © 2011 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

Engrafting organoids into vascularized tissues in model animals, such as the immunodeficient mouse or chick embryo chorioallantoic membrane (CAM), has proven efficient for neovascularization modeling. The CAM is a richly vascularized extraembryonic membrane, which shows limited immunoreactivity, thus becoming an excellent hosting model for human origin cell transplants. This paper describes the strategy to engraft human brain organoids differentiated at multiple maturation stages into the CAM. The cellular composition of brain organoids changes with time, reflecting the milestones of human brain development. We grafted brain organoids at relevant maturation stages: neuroepithelial expansion (18 DIV), early neurogenesis (60 DIV), and early gliogenesis (180 DIV) into the CAM of embryonic day (E)7 chicken embryos. Engrafted brain organoids were harvested 5 days later and their histological features were analyzed. No histological signs of neovascularization in the grafted organoids or abnormal blood vessels adjacent to the graftings were detected. Moreover, remarkable changes were observed in the cellular composition of the grafted organoids, namely, an increase in the number of glial fibrillary acidic protein-positive-reactive astrocytes. However, the cytoarchitectural changes were dependent on the organoid maturation stage. Altogether, these results suggest that brain organoids can grow in the CAM, and they show differences in the cytoarchitecture depending on their maturation stage at grafting.

Alzheimer's disease (AD) is the primary cause of dementia, to date. The urgent need to understand the biological and biochemical processes related to this condition, as well as the demand for reliable in vitro models for drug screening, has led to the development of novel techniques, among which stem cell methods are of utmost relevance for AD research, particularly the development of human brain organoids. Brain organoids are three-dimensional cellular aggregates derived from induced pluripotent stem cells (iPSCs) that recreate different neural cell interactions and tissue characteristics in culture. Here, we describe the protocol for the generation of brain organoids derived from AD patients and for the analysis of AD-derived pathology. AD organoids can recapitulate beta-amyloid and tau pathological features, making them a promising model for studying the molecular mechanisms underlying disease and for in vitro drug testing., (© 2023. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Eosinophils are found in the mucosa of the healthy gastrointestinal tract, but they also often accompany gastrointestinal diseases. We hypothesized that a positive correlation exists between blood eosinophil count and colonic eosinophil mucosal density in children. Electronic health records regarding 181 colonoscopies, performed with biopsy in the years 2019-2022, were screened for information on blood and colonic eosinophil count, age, sex, diagnoses, weight, height, white blood cell (WBC) count, serum C-reactive protein (CRP), and total IgE concentration. The median age (IQR) of the 107 included children (109 colonoscopies) was 12.4 years (8.1-15.5); 32 presented with blood eosinophilia (29.3%). The median eosinophil density/high-power field in the colonic mucosa was 22.5 (9-31). We found a weak correlation between colonic mucosal eosinophil density and blood eosinophil count (r = 0.295, 95% CI 0.108-0.462, p = 0.0018). This association was more pronounced in patients with elevated CRP (r = 0.529, 95% CI 0.167-0.766, p = 0.0054) and older than 12.4 years (r = 0.448, 95% CI 0.197-0.644, p = 0.00068). Peripheral blood eosinophilia might hint at increased mucosal colonic eosinophil density, especially in older children and in the presence of systemic inflammation. However, it seems unlikely that blood and colonic eosinophilia are strongly linked in younger children. Studies in adults are warranted.

Dyrk1a triplication in Down's syndrome and its overexpression in Alzheimer's disease suggest a role for increased DYRK1A activity in the abnormal metabolism of APP. Transport defects are early phenotypes in the progression of Alzheimer's disease, which lead to APP processing impairments. However, whether DYRK1A regulates the intracellular transport and delivery of APP in human neurons remains unknown. From a proteomic dataset of human cerebral organoids treated with harmine, a DYRK1A inhibitor, we found expression changes in protein clusters associated with the control of microtubule-based transport and in close interaction with the APP vesicle. Live imaging of APP axonal transport in human-derived neurons treated with harmine or overexpressing a dominant negative DYRK1A revealed a reduction in APP vesicle density and enhanced the stochastic behavior of retrograde vesicle transport. Moreover, harmine increased the fraction of slow segmental velocities and changed speed transitions supporting a DYRK1A-mediated effect in the exchange of active motor configuration. Contrarily, the overexpression of DYRK1A in human polarized neurons increased the axonal density of APP vesicles and enhanced the processivity of retrograde APP. In addition, increased DYRK1A activity induced faster retrograde segmental velocities together with significant changes in slow to fast anterograde and retrograde speed transitions, suggesting the facilitation of the active motor configuration. Our results highlight DYRK1A as a modulator of the axonal transport machinery driving APP intracellular distribution in neurons, and stress DYRK1A inhibition as a putative therapeutic intervention to restore APP axonal transport in Down's syndrome and Alzheimer's disease. SIGNIFICANCE STATEMENT Axonal transport defects are early events in the progression of neurodegenerative diseases, such as Alzheimer's disease. However, the molecular mechanisms underlying transport defects remain elusive. Dyrk1a kinase is triplicated in Down's syndrome and overexpressed in Alzheimer's disease, suggesting that DYRK1A dysfunction affects molecular pathways leading to early-onset neurodegeneration. Here, we show by live imaging of human-derived neurons that DYRK1A activity differentially regulates the intracellular trafficking of APP. Further, single-particle analysis revealed DYRK1A as a modulator of axonal transport and the configuration of active motors within the APP vesicle. Our work highlights DYRK1A as a regulator of APP axonal transport and metabolism, supporting DYRK1A inhibition as a therapeutic strategy to restore intracellular dynamics in Alzheimer's disease., (Copyright © 2022 the authors.)

Thioredoxin family proteins are key modulators of cellular redox regulation and have been linked to several physiological functions, including the cellular response to hypoxia-ischemia. During perinatal hypoxia-ischemia (PHI), the central nervous system is subjected to a fast decrease in O 2 and nutrients with a subsequent reoxygenation that ultimately leads to the production of reactive species impairing physiological redox signaling. Particularly, the retina is one of the most affected tissues, due to its high oxygen consumption and exposure to light. One of the main consequences of PHI is retinopathy of prematurity, comprising changes in retinal neural and vascular development, with further compensatory mechanisms that can ultimately lead to retinal detachment and blindness. In this study, we have analyzed long-term changes that occur in the retina using two well established in vivo rat PHI models (perinatal asphyxia and carotid ligation model), as well as the ARPE-19 cell line that was exposed to hypoxia and reoxygenation. We observed significant changes in the protein levels of the cytosolic oxidoreductase thioredoxin 1 (Trx1) in both animal models and a cell model. Knock-down of Trx1 in ARPE-19 cells affected cell morphology, proliferation and the levels of specific differentiation markers. Administration of recombinant Trx1 decreased astrogliosis and improved delayed neurodevelopment in animals exposed to PHI. Taken together, our results suggest therapeutical implications for Trx1 in retinal damage induced by hypoxia-ischemia during birth., (Copyright © 2019 IBRO. Published by Elsevier Ltd. All rights reserved.)

Cerebral hypoxia-ischemia damages synaptic proteins, resulting in cytoskeletal alterations, protein aggregation and neuronal death. In the previous works, we have shown neuronal and synaptic changes in rat neostriatum subjected to hypoxia that leads to ubi-protein accumulation. Recently, we also showed that, changes in F-actin organization could be related to early alterations induced by hypoxia in the Central Nervous System. However, little is known about effective treatment to diminish the damage. The main aim of this work is to study the effects of birth hypothermia on the actin cytoskeleton of neostriatal post-synaptic densities (PSD) in 60 days olds rats by immunohistochemistry, photooxidation and western blot. We used 2 different protocols of hypothermia: (a) intrahypoxic hypothermia at 15°C and (b) post-hypoxia hypothermia at 32°C. Consistent with previous data at 30 days, staining with phalloidin-Alexa(488) followed by confocal microscopy analysis showed an increase of F-actin fluorescent staining in the neostriatum of hypoxic animals. Correlative photooxidation electron microscopy confirmed these observations showing an increment in the number of mushroom-shaped F-actin staining spines in neostriatal excitatory synapses in rats subjected to hypoxia. In addition, western blot revealed β-actin increase in PSDs in hypoxic animals. The optic relative density measurement showed a significant difference between controls and hypoxic animals. When hypoxia was induced under hypothermic conditions, the changes observed in actin cytoskeleton were blocked. Post-hypoxic hypothermia showed similar answer but actin cytoskeleton modifications were not totally reverted as we observed at 15°C. These data suggest that the decrease of the body temperature decreases the actin modifications in dendritic spines preventing the neuronal death., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Background: The oxidoreductases of the thioredoxin (Trx) family of proteins play a major role in the cellular response to oxidative stress. Redox imbalance is a major feature of brain damage. For instance, neuronal damage and glial reaction induced by a hypoxic-ischemic episode is highly related to glutamate excitotoxicity, oxidative stress and mitochondrial dysfunction. Most animal models of hypoxia-ischemia in the central nervous system (CNS) use rats to study the mechanisms involved in neuronal cell death, however, no comprehensive study on the localization of the redox proteins in the rat CNS was available., Methods: The aim of this work was to study the distribution of the following proteins of the thioredoxin and glutathione/glutaredoxin (Grx) systems in the rat CNS by immunohistochemistry: Trx1, Trx2, TrxR1, TrxR2, Txnip, Grx1, Grx2, Grx3, Grx5, and γ-GCS, peroxiredoxin 1 (Prx1), Prx2, Prx3, Prx4, Prx5, and Prx6. We have focused on areas most sensitive to a hypoxia-ischemic insult: Cerebellum, striatum, hippocampus, spinal cord, substantia nigra, cortex and retina., Results and Conclusions: Previous studies implied that these redox proteins may be distributed in most cell types and regions of the CNS. Here, we have observed several remarkable differences in both abundance and regional distribution that point to a complex interplay and crosstalk between the proteins of this family., General Significance: We think that these data might be helpful to reveal new insights into the role of thiol redox pathways in the pathogenesis of hypoxia-ischemia insults and other disorders of the CNS. This article is part of a Special Issue entitled Human and Murine Redox Protein Atlases., (Copyright © 2010. Published by Elsevier B.V.)