RÉSUMÉ
Establishing preclinical models of Alzheimer's disease that predict clinical outcomes remains a critically important, yet to date not fully realized, goal. Models derived from human cells offer considerable advantages over non-human models, including the potential to reflect some of the inter-individual differences that are apparent in patients. Here we report an approach using induced pluripotent stem cell-derived cortical neurons from people with early symptomatic Alzheimer's disease where we sought a match between individual disease characteristics in the cells with analogous characteristics in the people from whom they were derived. We show that the response to amyloid-ß burden in life, as measured by cognitive decline and brain activity levels, varies between individuals and this vulnerability rating correlates with the individual cellular vulnerability to extrinsic amyloid-ß in vitro as measured by synapse loss and function. Our findings indicate that patient-induced pluripotent stem cell-derived cortical neurons not only present key aspects of Alzheimer's disease pathology but also reflect key aspects of the clinical phenotypes of the same patients. Cellular models that reflect an individual's in-life clinical vulnerability thus represent a tractable method of Alzheimer's disease modelling using clinical data in combination with cellular phenotypes.
RÉSUMÉ
Human induced Pluripotent Stem Cell (hiPSC) models are a valuable new tool for research into neurodegenerative diseases. Neuroinflammation is now recognized as a key process in neurodegenerative disease and aging, and microglia are central players in this. A plethora of hiPSC-derived microglial models have been published recently to explore neuroinflammation, ranging from monoculture through to xenotransplantation. However, combining physiological relevance, reproducibility, and scalability into one model is still a challenge. We examine key features of the in vitro microglial environment, especially media composition, extracellular matrix, and co-culture, to identify areas for improvement in current hiPSC-microglia models.
Sujet(s)
Techniques de culture cellulaire , Microenvironnement cellulaire , Cellules souches pluripotentes induites/cytologie , Microglie/cytologie , Modèles biologiques , Animaux , Cellules cultivées , Techniques de coculture , Milieux de culture/pharmacologie , Milieux de culture sans sérum/pharmacologie , Hétérogreffes , Humains , Cellules souches pluripotentes induites/effets des médicaments et des substances chimiques , Cellules souches pluripotentes induites/transplantation , Inflammation/immunologie , Souris , Microglie/effets des médicaments et des substances chimiques , Maladies neurodégénératives/immunologie , Maladies neurodégénératives/anatomopathologieRÉSUMÉ
Astrocytes influence neuronal maturation and function by providing trophic support, regulating the extracellular environment, and modulating signaling at synapses. The emergence of induced pluripotent stem cell (iPSC) technology offers a human system with which to validate and re-evaluate insights from animal studies. Here, we set out to examine interactions between human astrocytes and neurons derived from a common cortical progenitor pool, thereby recapitulating aspects of in vivo cortical development. We show that the cortical iPSC-derived astrocytes exhibit many of the molecular and functional hallmarks of astrocytes. Furthermore, optogenetic and electrophysiological co-culture experiments reveal that the iPSC-astrocytes can actively modulate ongoing synaptic transmission and exert pro-maturational effects upon developing networks of iPSC-derived cortical neurons. Finally, transcriptomic analyses implicate synapse-associated extracellular signaling in the astrocytes' pro-maturational effects upon the iPSC-derived neurons. This work helps lay the foundation for future investigations into astrocyte-to-neuron interactions in human health and disease.
Sujet(s)
Astrocytes/cytologie , Différenciation cellulaire , Cortex cérébral/cytologie , Cellules souches pluripotentes induites/cytologie , Neurones/cytologie , Animaux , Marqueurs biologiques/métabolisme , Signalisation calcique , Lignée cellulaire , Techniques de coculture , Humains , Agents neuromédiateurs/métabolisme , Rats , Synapses/métabolisme , Transmission synaptique , Transcriptome/génétiqueRÉSUMÉ
In the motor system, force gradation is achieved by recruitment of motoneurons and rate modulation of their firing frequency. Classical experiments investigating the relationship between injected current to the soma during intracellular recording and the firing frequency (the I-f relation) in cat spinal motoneurons identified two clear ranges: a primary range and a secondary range. Recent work in mice, however, has identified an additional range proposed to be exclusive to rodents, the subprimary range (SPR), due to the presence of mixed mode oscillations of the membrane potential. Surprisingly, fully summated tetanic contractions occurred in mice during SPR frequencies. With the mouse now one of the most popular models to investigate motor control, it is crucial that such discrepancies between observations in mice and basic principles that have been widely accepted in larger animals are resolved. To do this, we have reinvestigated the I-f relation using ramp current injections in spinal motoneurons in both barbiturate-anesthetized and decerebrate (nonanesthetized) cats and mice. We demonstrate the presence of the SPR and mixed mode oscillations in both species and show that the SPR is enhanced by barbiturate anesthetics. Our measurements of the I-f relation in both cats and mice support the classical opinion that firing frequencies in the higher end of the primary range are necessary to obtain a full summation. By systematically varying the leg oil pool temperature (from 37°C to room temperature), we found that only at lower temperatures can maximal summation occur at SPR frequencies due to prolongation of individual muscle twitches.SIGNIFICANCE STATEMENT This work investigates recent revelations that mouse motoneurons behave in a fundamentally different way from motoneurons of larger animals with respect to the importance of rate modulation of motoneuron firing for force gradation. The current study systematically addresses the proposed discrepancies between mice and larger species (cats) and demonstrates that mouse motoneurons, in fact, use rate modulation as a mechanism of force modulation in a similar manner to the classical descriptions in larger animals.
Sujet(s)
Motoneurones/physiologie , Contraction musculaire/physiologie , Muscles squelettiques/physiologie , Moelle spinale/physiologie , Animaux , Chats , Stimulation électrique/méthodes , Femelle , Mâle , Souris , Souris de lignée C57BL , Muscles squelettiques/innervation , Spécificité d'espèce , Moelle spinale/cytologieRÉSUMÉ
Amyotrophic lateral sclerosis (ALS) is a lethal neurodegenerative disease, which selectively affects upper and lower motoneurones. The underlying pathophysiology of the disease is complex but electrophysiological studies of peripheral nerves in ALS patients as well as human autopsy studies indicate that a potassium channel dysfunction/loss is present early in the symptomatic phase. It remains unclear to what extent potassium channel abnormalities reflect a specific pathogenic mechanism in ALS. The aim of this study was therefore to investigate the temporal changes in the expression and/or function of potassium channels in motoneurones in the adult G127X SOD1 mouse model of ALS, a model which has a very long presymptomatic phase. Evidence from animal models indicates that the early progressive motoneurone dysfunction and degeneration can be largely compensated by motor unit remodeling, delaying the clinical symptom onset. Experiments were therefore performed both before and after symptom onset. Immunohistochemistry of motor axons in the ventral roots of G127X SOD1 mice, was used to investigate juxta-paranodal Kv1.2 potassium channels along with nodal Nav1.6 and the paranodal scaffolding protein Caspr. This allowed an investigation of changes in the distribution of Kv1.2 relative to the general structure of the nodal-paranodal-juxta-paranodal complex. This revealed that the motor axons in the ventral roots of presymptomatic G127X SOD1 mice, already show a disruption in juxta-paranodal Kv1.2 potassium channels. The axonal Kv1.2 disruption was preceded by abnormalities in the distribution of the paranodal scaffolding protein Caspr with the nodal arrangement of Nav1.6 appearing relatively preserved even in symptomatic mice. These changes were accompanied by axon swelling and a slowing of conduction in the peripheral motor axons in symptomatic mice. In vivo electrophysiological intracellular recordings of individual spinal motoneurones revealed that central potassium channel function was preserved or even enhanced with higher amplitude and longer duration after-hyperpolarisations in the G127X SOD1 mice. Our data suggest that the potassium channel abnormalities observed in presymptomatic G127X, rather than representing a specific pathophysiological mechanism targeting potassium channels, most likely reflect early axonal degenerative changes, consistent with the "dying-back" phenomenon observed in other ALS models.
Sujet(s)
Sclérose latérale amyotrophique/métabolisme , Axones/anatomopathologie , Motoneurones/métabolisme , Nerfs périphériques/métabolisme , Canaux potassiques/métabolisme , Racines des nerfs spinaux/métabolisme , Sclérose latérale amyotrophique/anatomopathologie , Animaux , Axones/métabolisme , Modèles animaux de maladie humaine , Souris transgéniques , Motoneurones/anatomopathologie , Racines des nerfs spinaux/anatomopathologieRÉSUMÉ
Lysosomes have traditionally been viewed as degradative organelles, although a growing body of evidence suggests that they can function as Ca2+ stores. Here we examined the function of these stores in hippocampal pyramidal neurons. We found that back-propagating action potentials (bpAPs) could elicit Ca2+ release from lysosomes in the dendrites. This Ca2+ release triggered the fusion of lysosomes with the plasma membrane, resulting in the release of Cathepsin B. Cathepsin B increased the activity of matrix metalloproteinase 9 (MMP-9), an enzyme involved in extracellular matrix (ECM) remodelling and synaptic plasticity. Inhibition of either lysosomal Ca2+ signaling or Cathepsin B release prevented the maintenance of dendritic spine growth induced by Hebbian activity. This impairment could be rescued by exogenous application of active MMP-9. Our findings suggest that activity-dependent exocytosis of Cathepsin B from lysosomes regulates the long-term structural plasticity of dendritic spines by triggering MMP-9 activation and ECM remodelling.
Sujet(s)
Calcium/métabolisme , Cathepsine B/métabolisme , Épines dendritiques/métabolisme , Exocytose/physiologie , Lysosomes/métabolisme , Matrix metalloproteinase 9/métabolisme , Plasticité neuronale/physiologie , Cellules pyramidales/métabolisme , Animaux , Dendrites/métabolisme , Épines dendritiques/physiologie , Hippocampe/cytologie , Mâle , Techniques de patch-clamp , Cellules pyramidales/cytologie , Cellules pyramidales/physiologie , Rats , Rat Wistar , Transduction du signalRÉSUMÉ
The metalloproteinase ADAM17 activates ErbB signalling by releasing ligands from the cell surface, a key step underlying epithelial development, growth and tumour progression. However, mechanisms acutely controlling ADAM17 cell-surface availability to modulate the extent of ErbB ligand release are poorly understood. Here, through a functional genome-wide siRNA screen, we identify the sorting protein PACS-2 as a regulator of ADAM17 trafficking and ErbB signalling. PACS-2 loss reduces ADAM17 cell-surface levels and ADAM17-dependent ErbB ligand shedding, without apparent effects on related proteases. PACS-2 co-localizes with ADAM17 on early endosomes and PACS-2 knockdown decreases the recycling and stability of internalized ADAM17. Hence, PACS-2 sustains ADAM17 cell-surface activity by diverting ADAM17 away from degradative pathways. Interestingly, Pacs2-deficient mice display significantly reduced levels of phosphorylated EGFR and intestinal proliferation. We suggest that this mechanism controlling ADAM17 cell-surface availability and EGFR signalling may play a role in intestinal homeostasis, with potential implications for cancer biology.
Sujet(s)
Protéines ADAM/métabolisme , Protéines oncogènes v-erbB/métabolisme , Protéines du transport vésiculaire/métabolisme , Protéines ADAM/génétique , Protéine ADAM17 , Animaux , Lignée cellulaire tumorale , Régulation de l'expression des gènes codant pour des enzymes , Étude d'association pangénomique , Humains , Souris , Protéines oncogènes v-erbB/génétique , Transduction du signal/physiologie , Protéines du transport vésiculaire/génétiqueRÉSUMÉ
BACKGROUND: Plasma YKL-40 is increased early in patients with ST-elevation myocardial infarction (STEMI). It is not known whether plasma YKL-40 is related to infarct size and recovery of ventricular function after primary percutaneous coronary intervention (PCI) of STEMI and whether granulocyte colony-stimulating factor (G-CSF) therapy influence plasma YKL-40 concentration. MATERIALS AND METHODS: A total of 72 patients (age: 56 +/- 9 years (mean +/- SD), 56 men and 16 women) with STEMI treated with PCI were included in a double-blind, randomized, placebo-controlled trial with subcutaneous G-CSF or placebo injections from day 1 to 7 after the STEMI. Plasma YKL-40, high-sensitivity C-reactive protein (hs-CRP) and CK-MB concentrations were measured at baseline and during the first month. Infarct size and left ventricular ejection fraction (LVEF) were measured by magnetic resonance imaging at baseline and after 6 months. RESULTS: Baseline plasma YKL-40 was increased (median 92 microg/L) compared to healthy subjects (median 34 microg/L, p <0.001). In the placebo group hs-CRP and YKL-40 correlated at baseline (p = 0.04) and day 3 (p = 0.01), but not at day 7 and 30. Moreover, YKL-40 correlated negatively to LVEF recovery (p = 0.04) but not infarct size. G-CSF injections increased YKL-40 compared to placebo (p <0.001), but were not associated with infarct size or LVEF recovery. CONCLUSION: Plasma YKL-40 was significantly increased in STEMI patients at admission and G-CSF treatment caused a further increase in YKL-40. Plasma YKL-40 may be an indirect marker of LVEF recovery, independent of hs-CRP, and higher plasma YKL-40 indicates a lower recovery.