Sialometabolism in Brain Health and Alzheimer’s Disease

Sialic acids refer to a unique family of acidic sugars with a 9-carbon backbone that are mostly found as terminal residues in glycan structures of glycoconjugates including both glycoproteins and glycolipids. The highest levels of sialic acids are expressed in the brain where they regulate neuronal sprouting and plasticity, axon myelination and myelin stability, as well as remodeling of mature neuronal connections. Moreover, sialic acids are the sole ligands for microglial Siglecs (sialic acid-binding immunoglobulin-type lectins), and sialic acid-Siglec interactions have been indicated to play a critical role in the regulation of microglial homeostasis in a healthy brain. The recent discovery of CD33, a microglial Siglec, as a novel genetic risk factor for late-onset Alzheimer’s disease (AD), highlights the potential role of sialic acids in the development of microglial dysfunction and neuroinflammation in AD. Apart from microglia, sialic acids have been found to be involved in several other major changes associated with AD. Elevated levels of serum sialic acids have been reported in AD patients. Alterations in ganglioside (major sialic acid carrier) metabolism have been demonstrated as an aggravating factor in the formation of amyloid pathology in AD. Polysialic acids are linear homopolymers of sialic acids and have been implicated to be an important regulator of neurogenesis that contributes to neuronal repair and recovery from neurodegeneration such as in AD. In summary, this article reviews current understanding of neural functions of sialic acids and alterations of sialometabolism in aging and AD brains. Furthermore, we discuss the possibility of looking at sialic acids as a promising novel therapeutic target for AD intervention.

Abstract P794: Neuregulin-1 Induction of a Distinct Neuronal Growth Program Profile Post-Ischemia

Stroke is ranked as the fifth leading cause of death and the leading cause of adult disability in the United States. Tissue Plasminogen Activator (tPA) is the only FDA approved stroke treatment. Unfortunately, tPA is only administered to 2-5% of patients due to risk of hemorrhage, and it has no therapeutic effect on neuroprotection or neurological recovery. Thus, there is no treatment capable of addressing progressive adult disability. However, it has been shown that exogenous Neuregulin-1 (NRG-1) significantly decreases acute neuronal death and neuroinflammation after ischemic injury. Furthermore, NRG-1 has demonstrated a neuroregenerative effect with increased neuronal sprouting, synapse formation markers, and functional recovery after ischemia. However, the spatiotemporal regenerative effects of NRG-1 following ischemia have not been clearly elucidated. This study aimed to define the endogenous and NRG-1 treatment induced neuronal growth program (NGP) by spatiotemporally characterizing neuroplasticity and neuronal gene expression after ischemic stroke. Male and Female Thy1-eGFP+ transgenic mice were given middle cerebral artery occlusion (MCAO). Mice were treated daily for 10 days, I.P., with vehicle (0.1% BSA, endogenous) or NRG-1 (5ug/kg, treatment-induced) starting 24 hours after MCAO and sacrificed at 10dpi. Confocal microscopy data indicate that NRG-1 daily I.P. treatment protects Thy1-eGFP+ cortical neurons and exhibits more processes and less debris within the peri-infarct tissue. NRG-1 treated mice also exhibited an increase in BCAS1+/Olig2+ oligodendrocytes within the ipsilateral cortex and Olig2+ cells in the ipsilateral subcortex and contralateral cortex compared to vehicle treated mice 24 hours after MCAO. Additional studies including digital spatial profiling (DSP) suggest a spatiotemporal regulation of transcriptional profiles initiated by endogenous and NRG-1 treatment-induced NGP. These studies will help determine whether NRG-1 amplifies endogenous neuroplasticity or induces a distinct and/or region-specific neuroplastic pathway.

The Human ApoE4 Variant Reduces Functional Recovery and Neuronal Sprouting After Incomplete Spinal Cord Injury in Male Mice

Spinal cord injury (SCI) is a devastating form of neurotrauma. Patients who carry one or two apolipoprotein E (ApoE)4 alleles show worse functional outcomes and longer hospital stays after SCI, but the cellular and molecular underpinnings for this genetic link remain poorly understood. Thus, there is a great need to generate animal models to accurately replicate the genetic determinants of outcomes after SCI to spur development of treatments that improve physical function. Here, we examined outcomes after a moderate contusion SCI of transgenic mice expressing human ApoE3 or ApoE4. ApoE4 mice have worse locomotor function and coordination after SCI. Histological examination revealed greater glial staining in ApoE4 mice after SCI associated with reduced levels of neuronal sprouting markers. Bulk RNA sequencing revealed that subcellular processes (SCPs), such as extracellular matrix organization and inflammatory responses, were highly ranked among upregulated genes at 7 days after SCI in ApoE4 variants. Conversely, SCPs related to neuronal action potential and neuron projection development were increased in ApoE3 mice at 21 days. In summary, our results reveal a clinically relevant SCI mouse model that recapitulates the influence of ApoE genotypes on post SCI function in individuals who carry these alleles and suggest that the mechanisms underlying worse recovery for ApoE4 animals involve glial activation and loss of sprouting and synaptic activity.

The human ApoE4 variant reduces functional recovery and neuronal sprouting after incomplete spinal cord injury in male mice

Spinal cord injury (SCI) is a devastating form of neurotrauma. Patients who carry one or two ApoE4 alleles show worse functional outcomes and longer hospital stays after SCI but the cellular and molecular underpinnings for this genetic link remain poorly understood. Thus, there is a great need to generate animal models to accurately replicate the genetic determinants of outcomes after SCI to spur development of treatments that improve physical function. Here, we examined outcomes after a moderate contusion SCI of transgenic mice expressing human ApoE3 or ApoE4. ApoE4 mice have worse locomotor function and coordination after SCI. Histological examination revealed greater glial staining in ApoE4 mice after SCI associated with reduced levels of neuronal sprouting markers. Bulk RNA sequencing revealed that subcellular processes (SCPs), such as extracellular matrix organization and inflammatory responses, were highly-ranked among upregulated genes at 7 days after SCI in ApoE4 variants. Conversely, SCPs related to neuronal action potential and neuron projection development were increased in ApoE3 mice at 21 days. In summary, our results reveal a clinically relevant SCI mouse model that recapitulates the influence of ApoE genotypes on post-SCI function in individuals who carry these alleles and suggest that the mechanisms underlying worse recovery for ApoE4 animals involve glial activation and loss of sprouting and synaptic activity.

let-7-Complex MicroRNAs Regulate Broad-Z3, Which Together with Chinmo Maintains Adult Lineage Neurons in an Immature State

During Drosophila melanogaster metamorphosis, arrested immature neurons born during larval development differentiate into their functional adult form. This differentiation coincides with the downregulation of two zinc-finger transcription factors, Chronologically Inappropriate Morphogenesis (Chinmo) and the Z3 isoform of Broad (Br-Z3). Here, we show that br-Z3 is regulated by two microRNAs, let-7 and miR-125, that are activated at the larval-to-pupal transition and are known to also regulate chinmo. The br-Z3 3′UTR contains functional binding sites for both let-7 and miR-125 that confers sensitivity to both of these microRNAs, as determined by deletion analysis in reporter assays. Forced expression of let-7 and miR-125 miRNAs leads to early silencing of Br-Z3 and Chinmo and is associated with inappropriate neuronal sprouting and outgrowth. Similar phenotypes were observed by the combined but not separate depletion of br-Z3 and chinmo. Because persistent Br-Z3 was not detected in let-7-C mutants, this work suggests a model in which let-7 and miR-125 activation at the onset of metamorphosis may act as a failsafe mechanism that ensures the coordinated silencing of both br-Z3 and chinmo needed for the timely outgrowth of neurons arrested during larval development. The let-7 and miR-125 binding site sequences are conserved across Drosophila species and possibly other insects as well, suggesting that this functional relationship is evolutionarily conserved.

Abstract WP157: Neuregulin-1 Stimulates Breast Carcinoma Amplified Sequence 1+ Pre-Myelinating Oligodendrocytes Following Ischemic Stroke

Stroke is ranked as the fifth leading cause of death and the leading cause of adult disability in the United States. Tissue Plasminogen Activator (tPA) is the only FDA approved stroke treatment. Unfortunately, tPA is only administered to 2-5% of patients due to risk of hemorrhage, and it has no therapeutic effect on neuroprotection or neurological recovery. Thus, there is no treatment capable of addressing progressive adult disability. However, it has been shown that exogenous Neuregulin-1 (NRG-1) significantly decreases acute neuronal death and neuroinflammation after ischemic injury. Furthermore, NRG-1 has demonstrated a neuroregenerative effect with increased neuronal sprouting, synapse formation markers, and functional recovery after ischemia. NRG-1 also plays critical roles in oligodendrocyte differentiation, survival, and maturation, suggesting that treatment would further enhance remyelination after ischemic injury. In this study, we examined the effect of NRG-1 treatment on a transient pre-myelinating BCAS1+ oligodendrocyte population. BCAS1 is expressed along oligodendrocyte processes when actively myelinating and exhibits a bushy morphology. Mice were given permanent middle cerebral artery occlusion (MCAO) and administered intra-arterial NRG-1(10ug/kg) or equivalent vehicle (0.1% BSA) at the time of reperfusion. Mice were sacrificed 24 hours post-stroke and ischemic lesion was exhibited with Triphenyl tetrazolium chloride (TTC). Immunohistochemistry with FluoroJade B and TUNEL was used to quantify the infarct size. Oligodendrocytes were evaluated with anti-Olig2 and anti-BCAS1. We showed that pre-myelinating BCAS1+ oligodendrocyte populations are present within the sham striatum and cortex but disappear in the injured striatum and cortex after stroke. However, this population increases within the ipsilateral peri-infarct cortex and throughout the contralateral hemisphere. NRG-1 treated mice exhibited an increase in BCAS1+ oligodendrocytes within the injured cortex compared to vehicle treated mice after 24 hours. Thus, NRG-1 may stimulate localized oligodendrocyte differentiation and early remyelination after stroke, suggesting a role as a regenerative agent.

δ-Catenin (CTNND2) missense mutation in familial cortical myoclonic tremor and epilepsy

Objective:To identify the causative gene in a large Dutch family with familial cortical myoclonic tremor and epilepsy (FCMTE).Methods:We performed exome sequencing for 3 patients of our FCMTE family. Next, we performed knock-down (shRNA) and rescue experiments by overexpressing wild-type and mutant human δ-catenin (CTNND2) proteins in cortical mouse neurons and compared the results with morphologic abnormalities in the postmortem FCMTE brain.Results:We identified a missense mutation, p.Glu1044Lys, in theCTNND2gene that cosegregated with the FCMTE phenotype. The knock-down ofCtnnd2in cultured cortical mouse neurons revealed increased neurite outgrowth that was rescued by overexpression of wild-type, but not mutant, CTNND2 and was reminiscent of the morphologic abnormalities observed in cerebellar Purkinje cells from patients with FCMTE.Conclusions:We proposeCTNND2as the causal gene in FCMTE3. Functional testing of the mutant protein revealed abnormal neuronal sprouting, consistent with the abnormal cerebellar Purkinje cell morphology in patients with FCMTE.

Effects of long-term fluoxetine treatment on adrenergic plasticity in rat vas deferens

Chronic antidepressant administration increases neurotrophin levels in the central and peripheral nervous system, leading to an increase of neuronal sprouting, reestablishment of neural networks and neurotransmitter levels. Injured peripheral nerves regenerate at very slow rates. However, the recovery of the hypogastric nerve in rodents after injury is significantly improved with neurotrophin administration. Accordingly, our goal was to determine whether treatment with the antidepressant fluoxetine affects catecholamine levels and neuronal function, after surgical denervation of the rat vas deferens. Noradrenaline levels in the denervated vas deferens were higher in fluoxetine-treated animals than in the vehicle-treated group, as measured by high performance liquid chromatography. In functional studies of smooth muscle contraction, the responses induced by phenylephrine or ATP, as well as pre-synaptic α2-adrenoceptor reactivity, were not modified by chronic treatment with the antidepressant. However, the contraction mediated by neuronal release of noradrenaline induced by tyramine was increased on days 7 and 21 after denervation in rats treated with fluoxetine. These data indicate that fluoxetine can improve functional recovery after rat vas deferens denervation.