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Frontiers in Neuroscience 2023Axons are processes of neurons, up to a metre long, that form the essential biological cables wiring nervous systems. They must survive, often far away from their cell... (Review)
Review
Axons are processes of neurons, up to a metre long, that form the essential biological cables wiring nervous systems. They must survive, often far away from their cell bodies and up to a century in humans. This requires self-sufficient cell biology including structural proteins, organelles, and membrane trafficking, metabolic, signalling, translational, chaperone, and degradation machinery-all maintaining the homeostasis of energy, lipids, proteins, and signalling networks including reactive oxygen species and calcium. Axon maintenance also involves specialised cytoskeleton including the cortical actin-spectrin corset, and bundles of microtubules that provide the highways for motor-driven transport of components and organelles for virtually all the above-mentioned processes. Here, we aim to provide a conceptual overview of key aspects of axon biology and physiology, and the homeostatic networks they form. This homeostasis can be derailed, causing axonopathies through processes of ageing, trauma, poisoning, inflammation or genetic mutations. To illustrate which malfunctions of organelles or cell biological processes can lead to axonopathies, we focus on axonopathy-linked subcellular defects caused by genetic mutations. Based on these descriptions and backed up by our comprehensive data mining of genes linked to neural disorders, we describe the 'dependency cycle of local axon homeostasis' as an integrative model to explain why very different causes can trigger very similar axonopathies, providing new ideas that can drive the quest for strategies able to battle these devastating diseases.
PubMed: 37564364
DOI: 10.3389/fnins.2023.1236815 -
BioRxiv : the Preprint Server For... Jun 2023Interleukin (IL)-27, a member of the IL-12 family of cytokines, induces human immunodeficiency virus (HIV)-resistant monocyte-derived macrophages and T cells. This...
Interleukin (IL)-27, a member of the IL-12 family of cytokines, induces human immunodeficiency virus (HIV)-resistant monocyte-derived macrophages and T cells. This resistance is mediated via the downregulation of spectrin beta, non-erythrocytic 1 (SPTBN1), induction of autophagy, or suppression of the acetylation of Y-box binding protein-1 (YB-1); however, the role of IL-27 administration during the induction of immature monocyte-derived dendritic cells (iDC) is poorly investigated. In the current study, we investigated the function of IL-27-induced iDC (27DC) on HIV infection. 27DC inhibited HIV infection by 95 ± 3 % without significant changes in the expression of CD4, CCR5, and SPTBN1 expression, autophagy induction and acetylation of YB-1 compared to iDC. An HIV proviral DNA copy number assay displayed that 27DC suppressed reverse transcriptase (RT) reaction without influencing the virus entry. A DNA microarray analysis was performed to identify the differentially expressed genes between 27DC and iDC. Compared to iDC, 51 genes were differentially expressed in 27DC, with more than 3-fold changes in four independent donors. Cross-reference analysis with the reported 2,214 HIV regulatory host genes identified nine genes as potential interests: Ankyrin repeat domain 22, Guanylate binding protein (GBP)-1, -2, -4, -5, Stabilin 1, Serpin family G member 1 (SERPING1), Interferon alpha inducible protein 6, and Interferon-induced protein with tetratricopeptide repeats 3. A knock-down study using si-RNA failed to determine a key factor associated with the anti-HIV activity due to the induction of robust amounts of off-target effects. Overexpression of each protein in cells had no impact on HIV infection. Thus, we could not define the mechanism of the anti-HIV effect in 27DC. However, our findings indicated that IL-27 differentiates monocytes into HIV-resistant DC, and the inhibitory mechanism differs from IL-27-induced HIV-resistant macrophages and T cells.
PubMed: 37546823
DOI: 10.1101/2023.06.12.544550 -
IScience Jul 2023Thrombocytopenia is one of the symptoms of many virus infections which is the "hallmark" in the case of dengue virus. In this study, we show the differential...
Thrombocytopenia is one of the symptoms of many virus infections which is the "hallmark" in the case of dengue virus. In this study, we show the differential localization of existing two forms of dengue virus protease, i.e., NS2BNS3 to the nucleus and NS3 to the nucleus and mitochondria. We also report a nuclear transcription factor, erythroid differentiation regulatory factor 1 (EDRF1), as the substrate for this protease. EDRF1 regulates the expression and activity of GATA1, which in turn controls spectrin synthesis. Both GATA1 and spectrins are required for platelet formation. On the other hand, we found that the mitochondrial activities will be damaged by NS3 localization which cleaves GrpEL1, a co-chaperone of mitochondrial Hsp70. Levels of both EDRF1 and GrpEL1 were found to deteriorate in dengue virus-infected clinical samples. Hence, we conclude that NS2BNS3-mediated EDRF1 cleavage and the NS3-led mitochondrial dysfunction account for thrombocytopenia.
PubMed: 37534186
DOI: 10.1016/j.isci.2023.107024 -
Cells Jul 2023The hypothesis about the role of the cortical cytoskeleton as the primary mechanosensor was tested. oocytes were exposed to simulated microgravity (by 3D clinorotation...
The hypothesis about the role of the cortical cytoskeleton as the primary mechanosensor was tested. oocytes were exposed to simulated microgravity (by 3D clinorotation in random directions with 4 rotations per minute-sµg group) and hypergravity at the 2 g level (by centrifugal force from one axis rotation-hg group) for 30, 90, and 210 min without and with cytochalasin B, colchicine, acrylamide, and calyculin A. Cell stiffness was measured by atomic force microscopy, protein content in the membrane and cytoplasmic fractions by Western blotting, and cellular respiration by polarography. The obtained results indicate that the stiffness of the cortical cytoskeleton of oocytes decreases in simulated micro- (after 90 min) and hypergravity (after 30 min), possibly due to intermediate filaments. The cell stiffness recovered after 210 min in the hg group, but intact microtubules were required for this. Already after 30 min of exposure to sµg, the cross-sectional area of oocytes decreased, which indicates deformation, and the singed protein, which organizes microfilaments into longitudinal bundles, diffused from the cortical cytoskeleton into the cytoplasm. Under hg, after 30 min, the cross-sectional area of the oocytes increased, and the proteins that organize filament networks, alpha-actinin and spectrin, diffused from the cortical cytoskeleton.
Topics: Animals; Drosophila melanogaster; Hypergravity; Cytoskeleton; Oocytes; Mercury
PubMed: 37508484
DOI: 10.3390/cells12141819 -
Membranes Jul 2023Dielectric relaxations at 1.4 MHz (β) and 9 MHz (γ1) on the erythrocyte spectrin network were studied by dielectric spectroscopy using dense suspensions of...
Dielectric relaxations at 1.4 MHz (β) and 9 MHz (γ1) on the erythrocyte spectrin network were studied by dielectric spectroscopy using dense suspensions of erythrocytes and erythrocyte ghost membranes, subjected to extraction with up to 0.2% volume Triton-X-100. The step-wise extraction of up to 60% of membrane lipids preserved γ1 and gradually removed β-relaxation. On increasing the concentration up to 100 mM of NaCl at either side of erythrocyte plasma membranes, the β-relaxation was linearly enhanced, while the strength of γ1-relaxation remained unchanged. In media with NaCl between 100 and 150 mM β-relaxation became slightly inhibited, while γ1-relaxation almost disappeared, possibly due to the decreased electrostatic repulsion allowing erythrocytes to come into closer contact. When these media contained, at concentrations 10-30 mg/mL dextran (MW 7 kDa), polyethylene glycol or polyvinylpyrrolidone (40 kDa), or albumin or homologous plasma with equivalent concentration of albumin, the γ1-relaxation was about tenfold enhanced, while β-relaxation was strengthened or preserved. The results suggest the Maxwell-Vagner accumulation of ions on the lipid bilayer as an energy source for β-relaxation. While β-relaxation appears sensitive to erythrocyte membrane deformability, γ1-relaxation could be a sensitive marker for the inter-membrane interactions between erythrocytes.
PubMed: 37505024
DOI: 10.3390/membranes13070658 -
BioRxiv : the Preprint Server For... Aug 2023Postnatal regulation of dendritic spine formation and refinement in cortical pyramidal neurons is critical for excitatory/inhibitory balance in neocortical networks....
Postnatal regulation of dendritic spine formation and refinement in cortical pyramidal neurons is critical for excitatory/inhibitory balance in neocortical networks. Recent studies have identified a selective spine pruning mechanism in the mouse prefrontal cortex (PFC) mediated by class 3 Semaphorins and the L1-CAM cell adhesion molecules Neuron-glia related CAM (NrCAM), Close Homolog of L1 (CHL1), and L1. L1-CAMs bind Ankyrin B (AnkB), an actin-spectrin adaptor encoded by ( ), a high confidence gene for autism spectrum disorder (ASD). In a new inducible mouse model (Nex1Cre-ERT2: : RCE), deletion in early postnatal pyramidal neurons increased spine density on apical dendrites in PFC layer 2/3 of homozygous and heterozygous -deficient mice. In contrast, deletion in adulthood had no effect on spine density. Sema3F-induced spine pruning was impaired in cortical neuron cultures from AnkB-null mice and was rescued by re-expression of the 220 kDa AnkB isoform but not 440 kDa AnkB. AnkB bound to NrCAM at a cytoplasmic domain motif (FIGQY ), and mutation to FIGQH inhibited binding, impairing Sema3F-induced spine pruning in neuronal cultures. Identification of a novel function for AnkB in dendritic spine regulation provides insight into cortical circuit development, as well as potential molecular deficiencies in ASD.
PubMed: 37503187
DOI: 10.1101/2023.07.11.548527 -
Frontiers in Physiology 2023Transfusion of red blood cell concentrates is the most common medical procedure to treat anaemia. However, their storage is associated with development of storage...
Variability of extracellular vesicle release during storage of red blood cell concentrates is associated with differential membrane alterations, including loss of cholesterol-enriched domains.
Transfusion of red blood cell concentrates is the most common medical procedure to treat anaemia. However, their storage is associated with development of storage lesions, including the release of extracellular vesicles. These vesicles affect viability and functionality of transfused red blood cells and appear responsible for adverse post-transfusional complications. However, the biogenesis and release mechanisms are not fully understood. We here addressed this issue by comparing the kinetics and extents of extracellular vesicle release as well as red blood cell metabolic, oxidative and membrane alterations upon storage in 38 concentrates. We showed that extracellular vesicle abundance increased exponentially during storage. The 38 concentrates contained on average 7 × 10 extracellular vesicles at 6 weeks (w) but displayed a ∼40-fold variability. These concentrates were subsequently classified into 3 cohorts based on their vesiculation rate. The variability in extracellular vesicle release was not associated with a differential red blood cell ATP content or with increased oxidative stress (in the form of reactive oxygen species, methaemoglobin and band3 integrity) but rather with red blood cell membrane modifications, i.e., cytoskeleton membrane occupancy, lateral heterogeneity in lipid domains and transversal asymmetry. Indeed, no changes were noticed in the low vesiculation group until 6w while the medium and the high vesiculation groups exhibited a decrease in spectrin membrane occupancy between 3 and 6w and an increase of sphingomyelin-enriched domain abundance from 5w and of phosphatidylserine surface exposure from 8w. Moreover, each vesiculation group showed a decrease of cholesterol-enriched domains associated with a cholesterol content increase in extracellular vesicles but at different storage time points. This observation suggested that cholesterol-enriched domains could represent a starting point for vesiculation. Altogether, our data reveal for the first time that the differential extent of extracellular vesicle release in red blood cell concentrates did not simply result from preparation method, storage conditions or technical issues but was linked to membrane alterations.
PubMed: 37408586
DOI: 10.3389/fphys.2023.1205493 -
Acta Neuropathologica Communications Jul 2023Oligodendrocyte (OL) injury and loss are central features of evolving lesions in multiple sclerosis. Potential causative mechanisms of OL loss include metabolic stress...
Oligodendrocyte (OL) injury and loss are central features of evolving lesions in multiple sclerosis. Potential causative mechanisms of OL loss include metabolic stress within the lesion microenvironment. Here we use the injury response of primary human OLs (hOLs) to metabolic stress (reduced glucose/nutrients) in vitro to help define the basis for the in situ features of OLs in cases of MS. Under metabolic stress in vitro, we detected reduction in ATP levels per cell that precede changes in survival. Autophagy was initially activated, although ATP levels were not altered by inhibitors (chloroquine) or activators (Torin-1). Prolonged stress resulted in autophagy failure, documented by non-fusion of autophagosomes and lysosomes. Consistent with our in vitro results, we detected higher expression of LC3, a marker of autophagosomes in OLs, in MS lesions compared to controls. Both in vitro and in situ, we observe a reduction in nuclear size of remaining OLs. Prolonged stress resulted in increased ROS and cleavage of spectrin, a target of Ca-dependent proteases. Cell death was however not prevented by inhibitors of ferroptosis or MPT-driven necrosis, the regulated cell death (RCD) pathways most likely to be activated by metabolic stress. hOLs have decreased expression of VDAC1, VDAC2, and of genes regulating iron accumulation and cyclophilin. RNA sequencing analyses did not identify activation of these RCD pathways in vitro or in MS cases. We conclude that this distinct response of hOLs, including resistance to RCD, reflects the combined impact of autophagy failure, increased ROS, and calcium influx, resulting in metabolic collapse and degeneration of cellular structural integrity. Defining the basis of OL injury and death provides guidance for development of neuro-protective strategies.
Topics: Humans; Multiple Sclerosis; Reactive Oxygen Species; Oligodendroglia; Cell Death; Multiple Sclerosis, Chronic Progressive; Adenosine Triphosphate
PubMed: 37408029
DOI: 10.1186/s40478-023-01601-1 -
ELife Jul 2023Hypoplastic left heart syndrome (HLHS) is a severe congenital heart disease (CHD) with a likely oligogenic etiology, but our understanding of the genetic complexities...
Hypoplastic left heart syndrome (HLHS) is a severe congenital heart disease (CHD) with a likely oligogenic etiology, but our understanding of the genetic complexities and pathogenic mechanisms leading to HLHS is limited. We performed whole genome sequencing (WGS) on 183 HLHS patient-parent trios to identify candidate genes, which were functionally tested in the heart model. Bioinformatic analysis of WGS data from an index family of a HLHS proband born to consanguineous parents prioritized 9 candidate genes with rare, predicted damaging homozygous variants. Of them, cardiac-specific knockdown (KD) of mitochondrial MICOS complex subunit resulted in drastically compromised heart contractility, diminished levels of sarcomeric actin and myosin, reduced cardiac ATP levels, and mitochondrial fission-fusion defects. These defects were similar to those inflicted by cardiac KD of ATP synthase subunits of the electron transport chain (ETC), consistent with the MICOS complex's role in maintaining cristae morphology and ETC assembly. Five additional HLHS probands harbored rare, predicted damaging variants in or . Hypothesizing an oligogenic basis for HLHS, we tested 60 additional prioritized candidate genes from these patients for genetic interactions with in sensitized fly hearts. Moderate KD of in combination with (activator of RNA polymerase II), (, E3 ubiquitin ligase), or ( scaffolding protein) caused synergistic heart defects, suggesting the likely involvement of diverse pathways in HLHS. Further elucidation of novel candidate genes and genetic interactions of potentially disease-contributing pathways is expected to lead to a better understanding of HLHS and other CHDs.
Topics: Humans; Hypoplastic Left Heart Syndrome; Actomyosin; Heart Defects, Congenital; Computational Biology; Adenosine Triphosphate; Mitochondrial Proteins
PubMed: 37404133
DOI: 10.7554/eLife.83385 -
Brain Research Oct 2023Damage to the axonal white matter tracts within the brain is a key cause of neurological impairment and long-term disability following traumatic brain injury (TBI)....
Damage to the axonal white matter tracts within the brain is a key cause of neurological impairment and long-term disability following traumatic brain injury (TBI). Understanding how axonal injury develops following TBI requires gyrencephalic models that undergo shear strain and tissue deformation similar to the clinical situation and investigation of the effects of post-injury insults like hypoxia. The aim of this study was to determine the effect of post-traumatic hypoxia on axonal injury and inflammation in a sheep model of TBI. Fourteen male Merino sheep were allocated to receive a single TBI via a modified humane captive bolt animal stunner, or sham surgery, followed by either a 15 min period of hypoxia or maintenance of normoxia. Head kinematics were measured in injured animals. Brains were assessed for axonal damage, microglia and astrocyte accumulation and inflammatory cytokine expression at 4 hrs following injury. Early axonal injury was characterised by calpain activation, with significantly increased SNTF immunoreactivity, a proteolytic fragment of alpha-II spectrin, but not with impaired axonal transport, as measured by amyloid precursor protein (APP) immunoreactivity. Early axonal injury was associated with an increase in GFAP levels within the CSF, but not with increases in IBA1 or GFAP+ve cells, nor in levels of TNFα, IL1β or IL6 within the cerebrospinal fluid or white matter. No additive effect of post-injury hypoxia was noted on axonal injury or inflammation. This study provides further support that axonal injury post-TBI is driven by different pathophysiological mechanisms, and detection requires specific markers targeting multiple injury mechanisms. Treatment may also need to be tailored for injury severity and timing post-injury to target the correct injury pathway.
Topics: Male; Animals; Sheep; Brain Injuries; Brain Injuries, Traumatic; Brain; Hypoxia; Inflammation
PubMed: 37400012
DOI: 10.1016/j.brainres.2023.148475