Abstract
Ischemic injury to white matter tracts is increasingly recognized to play a key role in age-related cognitive decline, vascular dementia, and Alzheimer’s disease. Knowledge of the effects of ischemic axonal injury on cortical neurons is limited yet critical to identifying molecular pathways that link neurodegeneration and ischemia. Using a mouse model of subcortical white matter ischemic injury coupled with retrograde neuronal tracing, we employed magnetic affinity cell sorting with fluorescence-activated cell sorting to capture layer-specific cortical neurons and performed RNA-sequencing. With this approach, we identified a role for microtubule reorganization within stroke-injured neurons acting through the regulation of tau. We find that subcortical stroke-injured Layer 5 cortical neurons up-regulate the microtubule affinity-regulating kinase, Mark4, in response to axonal injury. Stroke-induced up-regulation of Mark4 is associated with selective remodeling of the apical dendrite after stroke and the phosphorylation of tau in vivo. In a cell-based tau biosensor assay, Mark4 promotes the aggregation of human tau in vitro. Increased expression of Mark4 after ischemic axonal injury in deep layer cortical neurons provides new evidence for synergism between axonal and neurodegenerative pathologies by priming of tau phosphorylation and aggregation.
Highlights
Understanding the consequences of axonal injury on cortical neurons, subcortically projecting Layer 5 cortical neurons, has wide-reaching implications in a variety of neurologic diseases
Seven days after ischemic induction in wild-type mice, stroke-injured fluororuby+/CTIP2+ neurons (Fig. 1b, lower panels) within the overlying sensory and motor cortex were increased compared to sham (Additional file 1: Figure S1a-b) with subcortical stroke labeling an average of 0.24% ± 0.02% of the total CTIP2+ Layer 5 cortical neuron population in ipsilateral sensorimotor cortex overlying the ischemic lesions (Additional file 1: Figure S1c)
To verify that our layer-specific MACSFACS-seq approach enriched for Layer 5 cortical neurons, we examined average fpkm for reported layer-specific cortical neuron marker genes [4] (Fig. 1c)
Summary
Understanding the consequences of axonal injury on cortical neurons, subcortically projecting Layer 5 cortical neurons, has wide-reaching implications in a variety of neurologic diseases. Models of axonal transection and crush injury in the peripheral, cranial, and optic nerves have provided tremendous insight into the molecular response of neurons to distal axonal injury, including the identification of dual leucine zipper kinase (DLK), Jun kinase and other molecular pathways [15]. These pathways are being studied outside the context of peripheral nerve regrowth and explored in models of neurodegeneration. Jun kinase is implicated in neurodegeneration following injury and can directly phosphorylate the microtubule associated protein tau and promote the formation of neurofibrillary tangles that drive
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