Clinical magnetic resonance imaging evaluation of glymphatic function.

The glymphatic system is a system of specialized perivascular spaces in the brain that facilitates removal of toxic waste solutes from the brain. Evaluation of glymphatic system function by means of magnetic resonance imaging (MRI) has thus far been largely focused on rodents because of the limitations of intrathecal delivery of gadolinium-based contrast agents to humans. This review discusses MRI methods that can be employed clinically for glymphatic-related measurements intended for early diagnosis, prevention, and the treatment of various neurological conditions. Although glymphatic system-based MRI research is in its early stages, recent studies have identified promising noninvasive MRI markers associated with glymphatic system alterations in neurological diseases. However, further optimization in data acquisition, validation, and modeling are needed to investigate the glymphatic system within the clinical setting.

Velocity of cerebrospinal fluid in the aqueduct measured by phase-contrast MRI in rat.

Cerebrospinal fluid (CSF) circulation plays a key role in cerebral waste clearance via the glymphatic system. Although CSF flow velocity is an essential component of CSF dynamics, it has not been sufficiently characterized, and particularly, in studies of the glymphatic system in rat. To investigate the relationship between the flow velocity of CSF in the brain aqueduct and the glymphatic waste clearance rate, using phase-contrast MRI we performed the first measurements of CSF velocity in rats. Phase-contrast MRI was performed using a 7 T system to map mean velocity of CSF flow in the aqueduct in rat brain. The effects of age (3 months old versus 18 months old), gender, strain (Wistar, RNU, Dark Agouti), anesthetic agents (isoflurane versus dexmedetomidine), and neurodegenerative disorder (Alzheimer' disease in Fischer TgF344-AD rats, males and females) on CSF velocity were investigated in eight independent groups of rats (12 rats per group). Our results demonstrated that quantitative velocities of CSF flow in the aqueduct averaged 5.16 ± 0.86 mm/s in healthy young adult male Wistar rats. CSF flow velocity in the aqueduct was not altered by rat gender, strain, and the employed anesthetic agents in all rats, also age in the female rats. However, aged (18 months) Wistar male rats exhibited significantly reduced the CSF flow velocity in the aqueduct (4.31 ± 1.08 mm/s). In addition, Alzheimer's disease further reduced the CSF flow velocity in the aqueduct of male and female rats.

Ablation of Argonaute 2 in Schwann cells accelerates the progression of diabetic peripheral neuropathy.

Schwann cells (SCs) form myelin and provide metabolic support for axons, and are essential for normal nerve function. Identification of key molecules specific to SCs and nerve fibers may provide new therapeutic targets for diabetic peripheral neuropathy (DPN). Argonaute2 (Ago2) is a key molecular player that mediates the activity of miRNA-guided mRNA cleavage and miRNA stability. Our study found that Ago2 knockout (Ago2-KO) in proteolipid protein (PLP) lineage SCs in mice resulted in a significant reduction of nerve conduction velocities and impairments of thermal and mechanical sensitivities. Histopathological data revealed that Ago2-KO significantly induced demyelination and neurodegeneration. When DPN was induced in both wild-type and Ago2-KO mice, Ago2-KO mice exhibited further decreased myelin thickness and exacerbated neurological outcomes compared with wild-type mice. Deep sequencing analysis of Ago2 immunoprecipitated complexes showed that deregulated miR-206 in Ago2-KO mice is highly related to mitochondrial function. In vitro data showed that knockdown of miR-200 induced mitochondrial dysfunction and apoptosis in SCs. Together, our data suggest that Ago2 in SCs is essential to maintain peripheral nerve function while ablation of Ago2 in SCs exacerbates SC dysfunction and neuronal degeneration in DPN. These findings provide new insight into the molecular mechanisms of DPN.

Extracellular vesicles-Mediators of opioid use disorder?

Opioid use disorder (OUD) is a growing health emergency in the United States leading to an epidemic of overdose deaths. OUD is recognized as an addictive brain disorder resulting in psychological, cognitive and behavioural dysfunction. These observed clinical dysfunctions are a result of cellular changes that occur in the brain. Derangements in inflammation, neurogenesis and synaptic plasticity are observed in the brains of OUD patients. The mechanisms of these derangements are unclear; however, extracellular vesicles (EVs), membrane bound particles containing protein, nucleotides and lipids are currently being investigated as agents that invoke these cellular changes. The primary function of EVs is to facilitate intercellular communication by transfer of cargo (protein, nucleotides and lipids) between cells; however, changes in this cargo have been observed in models of OUD suggesting that EVs may be agents promoting the observed cellular derangements. This review summarizes evidence that altered cargo of EVs, specifically protein and miRNA, in models of OUD promote impairments in neurons, astrocytes and microglial cells. These findings support the premise that opioids alter EVs to detrimentally affect neuro-cellular function resulting in the observed addictive, psychological and neurocognitive deficits in OUD patients.

Circulating Extracellular Vesicles in Stroke Patients Treated With Mesenchymal Stem Cells: A Biomarker Analysis of a Randomized Trial.

BACKGROUND: Mesenchymal stem cells (MSCs) secrete trophic factors and extracellular vesicles (EVs). However, the level and role of EVs after MSC therapy in patients with stroke are unknown. We investigated whether circulating EVs and trophic factors are increased after MSCs and are related to the therapeutic benefits in the STARTING-2 trial (Stem Cell Application Researches and Trials in Neurology-2) participants. METHODS: In this prospective randomized controlled trial, patients with chronic major stroke were assigned, in a 2:1 ratio, to receive autologous MSC intravenous injection (MSC group, n=39) or standard treatment (control group, n=15) and followed for 3 months. Detailed clinical assessment and neuroplasticity on diffusion tensor image and resting-state functional magnetic resonance imaging were evaluated. Serial samples were collected, before/after MSCs therapy. The primary outcome measure was circulating factors that are associated with the clinical improvement in the Fugl-Meyer Assessment (secondary end point of the trial) and neuroplasticity on diffusion tensor image and resting-state functional magnetic resonance imaging. Additional outcome measures were microRNAs and trophic factors enriched in the plasma EVs, obtained using quantitative polymerase chain reaction and ELISA, respectively. RESULTS: Circulating EV levels were increased ≈5-fold (mean±SD, from 2.7×109±2.2×109 to 1.3×1010±1.7×1010 EVs/mL) within 24 hours after injection of MSCs (P=0.001). After adjustment of age, sex, baseline stroke severity, and the time interval from stroke onset to treatment, only the EV number was independently associated with improvement in motor function (odds ratio, 5.718 for EV numberLog [95% CI, 1.144-28.589]; P=0.034). Diffusion tensor image and resting-state functional magnetic resonance imaging showed that integrity of the ipsilesional corticospinal tract and intrahemispheric motor network were significantly correlated with circulating EV levels, respectively (P<0.05). MicroRNAs related to neurogenesis/neuroplasticity (eg, microRNA-18a-5p) were significantly increased in circulating EVs after MSC therapy (P=0.0479). In contrast, trophic factor levels were not changed after MSC therapy. CONCLUSIONS: This trial is the first to show that treatment of ischemic stroke patients with MSCs significantly increases circulating EVs, which were significantly correlated with improvement in motor function and magnetic resonance imaging indices of plasticity. REGISTRATION: URL: https://www. CLINICAL TRIALS: gov; Unique identifier: NCT01716481.

Pharmacological Characterization of a Novel Neuroactive Steroid Prodrug, NTS-104, and Its Neuroprotective Activity in Experimental Stroke Models.

BACKGROUND: Ischemic stroke affects about 700 000 patients per year in the United States, and to date, there are no effective pharmacological agents that promote recovery. Here, we studied the pharmacokinetics, pharmacodynamics, and efficacy of NTS-105, a novel neuroactive steroid, and NTS-104, a prodrug of NTS-105, in 2 models of ischemic stroke. METHODS: The pharmacodynamics and pharmacokinetics of NTS-104/105 were investigated in naive and stroke rats, and models of embolic and transient middle cerebral artery occlusion were used to investigate the dose-related effects of NTS-104. All rats were randomly assigned into the experimental groups, and all outcome measurements were performed blindly. RESULTS: Blood plasma and brain pharmacokinetic analysis revealed that NTS-104 rapidly converted to NTS-105, which reached peak concentration at ≈1 hour after dosing and distributed similarly to normal and ischemic brains. NTS-104 administration 4 hours after embolic middle cerebral artery occlusion led to a dose-dependent improvement of neurological outcomes and a dose-dependent reduction of infarct volumes relative to vehicle-treated animals. A single dose level study confirmed that NTS-104 administered 4 hours after transient middle cerebral artery occlusion was also neuroprotective. Quantitative ELISA revealed that NTS-104 treatment resulted in time- and dose-dependent changes in AKT activation and cytokine levels within the ischemic brain, which included reductions of IL-6, VEGF, ICAM-1, IL-1ß, MCP-1, RAGE, and GM-CSF. Time- and dose-dependent reductions in IL-6 and GM-CSF were also observed in the plasma along with an elevation of galectin-1. CONCLUSIONS: NTS-104 is a novel prodrug that converts to a novel neuroactive steroid, NTS-105, which improves functional outcomes in experimental ischemic stroke models.

Long noncoding RNA mediates stroke-induced neurogenesis.

Neurogenesis contributes to poststroke recovery. Long noncoding RNAs (lncRNAs) participate in the regulation of stem cell self-renewal and differentiation. However, the role of lncRNAs in stroke-induced neurogenesis remains unknown. In this study, we found that H19 was the most highly upregulated lncRNA in neural stem cells (NSCs) of the subventricular zone (SVZ) of rats subjected to focal cerebral ischemia. Deletion of H19 suppressed cell proliferation, promoted cell death, and blocked NSC differentiation. RNA sequencing analysis revealed that genes deregulated by H19 knockdown were those that are involved in transcription, apoptosis, proliferation, cell cycle, and response to hypoxia. H19 knockdown significantly increased the transcription of cell cycle-related genes including p27, whereas overexpression of H19 substantially reduced expression of these genes through the interaction with chromatin remodeling proteins EZH2 and SUZ12. Moreover, H19 regulated neurogenesis-related miRNAs. Inactivation of H19 in NSCs of ischemic rats attenuated spontaneous functional recovery after stroke. Collectively, our data provide novel insights into the epigenetic regulation of lncRNAs in stroke-induced neurogenesis.

Mesenchymal stromal cell-derived exosomes ameliorate peripheral neuropathy in a mouse model of diabetes.

AIMS/HYPOTHESIS: Diabetic peripheral neuropathy (DPN) is one of the major complications of diabetes, which contributes greatly to morbidity and mortality. There is currently no effective treatment for this disease. Exosomes are cell-derived nanovesicles and play an important role in intercellular communications. The present study investigated whether mesenchymal stromal cell (MSC)-derived exosomes improve neurological outcomes of DPN. METHODS: Exosomes were isolated from the medium of cultured mouse MSCs by ultracentrifugation. Diabetic mice (BKS.Cg-m+/+Leprdb/J, db/db) at the age of 20 weeks were used as DPN models. Heterozygous mice (db/m) of the same age were used as the control. MSC-exosomes were administered weekly via the tail vein for 8 weeks. Neurological function was evaluated by testing motor and sensory nerve conduction velocities, and thermal and mechanical sensitivity. Morphometric analysis was performed by myelin sheath staining and immunohistochemistry. Macrophage markers and circulating cytokines were measured by western blot and ELISA. MicroRNA (miRNA) array and bioinformatics analyses were performed to examine the exosomal miRNA profile and miRNA putative target genes involved in DPN. RESULTS: Treatment of DPN with MSC-exosomes markedly decreased the threshold for thermal and mechanical stimuli and increased nerve conduction velocity in diabetic mice. Histopathological analysis showed that MSC-exosomes markedly augmented the density of FITC-dextran perfused blood vessels and increased the number of intraepidermal nerve fibres (IENFs), myelin thickness and axonal diameters of sciatic nerves. Western blot analysis revealed that MSC-exosome treatment decreased and increased M1 and M2 macrophage phenotype markers, respectively. Moreover, MSC-exosomes substantially suppressed proinflammatory cytokines. Bioinformatics analysis revealed that MSC-exosomes contained abundant miRNAs that target the Toll-like receptor (TLR)4/NF-κB signalling pathway. CONCLUSIONS/INTERPRETATION: MSC-derived exosomes alleviate neurovascular dysfunction and improve functional recovery in mice with DPN by suppression of proinflammatory genes.

Ischemic Cerebral Endothelial Cell-Derived Exosomes Promote Axonal Growth.

BACKGROUND AND PURPOSE: Cerebral endothelial cells (CECs) and axons of neurons interact to maintain vascular and neuronal homeostasis and axonal remodeling in normal and ischemic brain, respectively. However, the role of exosomes in the interaction of CECs and axons in brain under normal conditions and after stroke is unknown. METHODS: Exosomes were isolated from CECs of nonischemic rats and is chemic rats (nCEC-exos and isCEC-exos), respectively. A multicompartmental cell culture system was used to separate axons from neuronal cell bodies. RESULTS: Axonal application of nCEC-exos promotes axonal growth of cortical neurons, whereas isCEC-exos further enhance axonal growth than nCEC-exos. Ultrastructural analysis revealed that CEC-exos applied into distal axons were internalized by axons and reached to their parent somata. Bioinformatic analysis revealed that both nCEC-exos and isCEC-exos contain abundant mature miRNAs; however, isCEC-exos exhibit more robust elevation of select miRNAs than nCEC-exos. Mechanistically, axonal application of nCEC-exos and isCEC-exos significantly elevated miRNAs and reduced proteins in distal axons and their parent somata that are involved in inhibiting axonal outgrowth. Blockage of axonal transport suppressed isCEC-exo-altered miRNAs and proteins in somata but not in distal axons. CONCLUSIONS: nCEC-exos and isCEC-exos facilitate axonal growth by altering miRNAs and their target protein profiles in recipient neurons.

Ablation of the microRNA-17-92 cluster in neural stem cells diminishes adult hippocampal neurogenesis and cognitive function.

Impairment of adult neurogenesis in the hippocampus causes cognitive deficits; however, the underlying molecular mechanisms have not been fully elucidated. microRNAs (miRNAs) regulate neural stem cell (NSC) function. With the use of a transgenic mouse line with conditional ablation of the miR-17-92 cluster in nestin lineage NSCs, we tested the hypothesis that the miR-17-92 cluster regulates adult neurogenesis and cognitive function in vivo. Compared with wild-type mice, ablation of the miR-17-92 cluster significantly reduced the number of proliferating NSCs and neuroblasts and neuronal differentiation in the dentate gyrus (DG) of the hippocampus and significantly impaired hippocampal-dependent learning and memory, as assayed by social recognition memory, novel object recognition, and Morris water-maze tests. Statistical analysis showed a highly significant correlation between newly generated neuroblasts in the DG and cognition deficits in miR-17-92 knockout (KO) mice. Western blot analysis showed that conditional KO of the miR-17-92 cluster significantly increased and reduced a cytoskeleton-associated protein, Enigma homolog 1 (ENH1), and its downstream transcription factor, inhibitor of differentiation 1 (ID1), respectively, as well as increased phosphatase and tensin homolog gene. These proteins are related to neuronal differentiation. Our study demonstrates that the miR-17-92 cluster in NSCs is critical for cognitive and behavioral function and regulates neurogenesis and that the miR-17-92 cluster may target ENH1/ID1 signaling.-Pan, W. L., Chopp, M., Fan, B., Zhang, R., Wang, X., Hu, J., Zhang, X. M., Zhang, Z. G., Liu, X. S. Ablation of the microRNA-17-92 cluster in neural stem cells diminishes adult hippocampal neurogenesis and cognitive function.

Epigenetic Mechanisms Underlying Adult Post Stroke Neurogenesis.

Stroke remains the leading cause of adult disability. Post-stroke neurogenesis contributes to functional recovery. As an intrinsic neurorestorative process, it is important to elucidate the molecular mechanism underlying stroke-induced neurogenesis and to develop therapies designed specifically to augment neurogenesis. Epigenetic mechanisms include DNA methylation, histone modification and its mediation by microRNAs and long-non-coding RNAs. In this review, we highlight how epigenetic factors including DNA methylation, histone modification, microRNAs and long-non-coding RNAs mediate stroke-induced neurogenesis including neural stem cell self-renewal and cell fate determination. We also summarize therapies targeting these mechanisms in the treatment of stroke.

ABCA1/ApoE/HDL Signaling Pathway Facilitates Myelination and Oligodendrogenesis after Stroke.

ATP-binding cassette transporter A1 (ABCA1) plays an important role in the regulation of apolipoprotein E (ApoE) and the biogenesis of high-density lipoprotein (HDL) cholesterol in the mammalian brain. Cholesterol is a major source for myelination. Here, we investigate whether ABCA1/ApoE/HDL contribute to myelin repair and oligodendrogenesis in the ischemic brain after stroke. Specific brain ABCA1-deficient (ABCA1-B/-B) and ABCA1-floxed (ABCA1fl/fl) control mice were subjected to permanent distal middle-cerebral-artery occlusion (dMCAo) and were intracerebrally administered (1) artificial mouse cerebrospinal fluid (CSF) as vehicle control, (2) human plasma HDL3, and (3) recombined human ApoE2 starting 24 h after dMCAo for 14 days. All stroke mice were sacrificed 21 days after dMCAo. The ABCA1-B/-B-dMCAo mice exhibit significantly reduced myelination and oligodendrogenesis in the ischemic brain as well as decreased functional outcome 21 days after stroke compared with ABCA1fl/fl mice; administration of human ApoE2 or HDL3 in the ischemic brain significantly attenuates the deficits in myelination and oligodendrogenesis in ABCA1-B/-B-dMCAo mice ( p < 0.05, n = 9/group). In vitro, ABCA1-B/-B reduces ApoE expression and decreases primary oligodendrocyte progenitor cell (OPC) migration and oligodendrocyte maturation; HDL3 and ApoE2 treatment significantly reverses ABCA1-B/-B-induced reduction in OPC migration and oligodendrocyte maturation. Our data indicate that the ABCA1/ApoE/HDL signaling pathway contributes to myelination and oligodendrogenesis in the ischemic brain after stroke.

N-Acetyl-Seryl-Aspartyl-Lysyl-Proline Augments Thrombolysis of tPA (Tissue-Type Plasminogen Activator) in Aged Rats After Stroke.

Background and Purpose- Stroke is a leading cause of disability worldwide, mainly affecting the elderly. However, preclinical studies in aged ischemic animals are limited. N-acetyl-seryl-aspartyl-lysyl-proline (AcSDKP) is a naturally occurring tetrapeptide with vascular-protective properties. The present study investigated the effect of AcSDKP on tPA (tissue-type plasminogen activator)-induced thrombolysis in aged rats after ischemic stroke. Methods- Aged male rats (18 months) were subjected to embolic middle cerebral artery occlusion. Rats subjected to 4 hours of middle cerebral artery occlusion were randomized into the following groups: (1) AcSDKP; (2) tPA; (3) AcSDKP in combination with tPA; and (4) saline. Neurological deficits, cerebral microvascular patency and integrity, and infarction were examined at 1 day and 7 days after middle cerebral artery occlusion. In vitro experiments were performed to examine the effect of AcSDKP on aged cerebral endothelial cell permeability. Results- Compared with saline, AcSDKP, or tPA as monotherapy did not have any therapeutic effects, whereas AcSDKP in combination with tPA significantly reduced cerebral tissue infarction and improved neurological outcome without increasing cerebral hemorrhage. Concurrently, the combination treatment significantly augmented microvascular perfusion and reduced thrombosis and blood-brain barrier leakage. In vitro, compared with cerebral endothelial cells from ischemic adult rats, the endothelial cells from ischemic aged rats exhibited significantly increased leakage. AcSDKP suppressed tPA-induced aged endothelial cell leakage and reduced expression of ICAM-1 (intercellular adhesion molecule 1) and NF (nuclear factor)-κB. Conclusions- The present study provides evidence for the therapeutic efficacy of AcSDKP in combination tPA for the treatment of embolic stroke in aged rats at 4 hours after stroke onset. AcSDKP likely acts on cerebral endothelial cells to enhance the benefits of tPA by increasing tissue perfusion and augmenting the integrity of the blood-brain barrier. Visual Overview- An online visual overview is available for this article.

Vepoloxamer Enhances Fibrinolysis of tPA (Tissue-Type Plasminogen Activator) on Acute Ischemic Stroke.

Background and Purpose- Thrombolytic treatment of acute ischemic stroke with tPA (tissue-type plasminogen activator) is hampered by its narrow therapeutic window and potential hemorrhagic complication. Vepoloxamer is a nonionic surfactant that exerts potent hemorheologic and antithrombotic properties in various thrombotic diseases. The current study investigated the effect of vepoloxamer on tPA treatment in a rat model of embolic stroke. Methods- Male Wistar rats subjected to embolic middle cerebral artery occlusion were treated with the combination of vepoloxamer and tPA, vepoloxamer alone, tPA alone, or saline initiated 4 hours after middle cerebral artery occlusion. Results- Monotherapy with tPA did not reduce infarct volume, and adversely potentiated microvascular thrombosis and vascular leakage compared with the saline treatment. Vepoloxamer monotherapy reduced infarct volume by 25% and improved brain perfusion. However, the combination treatment with vepoloxamer and tPA significantly reduced infarct volume by 32% and improved neurological function, without increasing the incidence of gross hemorrhage. Compared with vepoloxamer alone, the combination treatment with vepoloxamer and tPA robustly reduced secondary thrombosis and tPA-augmented microvascular leakage and further improved brain perfusion, which was associated with substantial reductions of serum active PAI-1 (plasminogen activator inhibitor-1) level and tPA-upregulated PAI-1 in the ischemic brain. Mechanistically, exosomes derived from platelets of ischemic rats treated with tPA-augmented cerebral endothelial barrier permeability and elevated protein levels of PAI-1 and TF (tissue factor) in the endothelial cells, whereas exosomes derived from platelets of rats subjected to the combination treatment with vepoloxamer and tPA diminished endothelial permeability augmented by tPA and fibrin and reduced PAI-1 and TF levels in the endothelial cells. Conclusions- The combination treatment with vepoloxamer and tPA exerts potent thrombolytic effects in rats subjected to acute ischemic stroke. Vepoloxamer reduces tPA-aggravated prothrombotic effect of platelet-derived exosomes on cerebral endothelial cells, which may contribute to the therapeutic effect of the combination treatment.

MiR-126 Mediates Brain Endothelial Cell Exosome Treatment-Induced Neurorestorative Effects After Stroke in Type 2 Diabetes Mellitus Mice.

Background and Purpose- Stroke patients with type 2 diabetes mellitus (T2DM) exhibit increased vascular and white matter damage and have worse prognosis compared with nondiabetic stroke patients. We investigated the neurorestorative effects of exosomes derived from mouse brain endothelial cells (EC-Exo) as treatment for stroke in T2DM mice and investigated the role of miR-126 in mediating EC-Exo-derived therapeutic benefits in T2DM-stroke mice. Methods- Adult, male BKS.Cg-m+/+Leprdb/J (T2DM) mice were subjected to photothrombotic stroke model. T2DM mice were intravenously injected at 3 days after stroke with (1) PBS; (2) liposome mimic (vehicle control, 3×1010); (3) EC-Exo (3×1010); (4) knockdown of miR-126 in EC-Exo (miR-126-/- EC-Exo, 3×1010). Behavioral and cognitive tests were performed, and mice were sacrificed at 28 days after stroke. Results- Compared with non-DM stroke mice, T2DM-stroke mice exhibit significantly decreased serum and brain tissue miR-126 expression. Endothelial cells and EC-Exo contain high levels of miR-126 compared with other cell types or exosomes derived from other types of cells, respectively (smooth muscle cells, astrocytes, and marrow stromal cells). Compared with PBS or liposome mimic treatment, EC-Exo treatment of T2DM-stroke mice significantly improves neurological and cognitive function, increases axon density, myelin density, vascular density, arterial diameter, as well as induces M2 macrophage polarization in the ischemic boundary zone. MiR-126-/- EC-Exo treatment significantly decreases miR-126 expression in serum and brain, as well as attentuates EC-Exo treatment-induced functional improvement and does not significantly increase axon and myelin density, vascular density, arterial diameter or induce M2 macrophage polarization in T2DM-stroke mice. In vitro, EC-Exo treatment significantly increases primary cortical neuron axonal outgrowth and increases endothelial capillary tube formation whereas miR-126-/- EC-Exo attentuates EC-Exo induced capillary tube formation and axonal outgrowth. Conclusions- EC-Exo treatment of stroke promotes neurorestorative effects in T2DM mice. MiR-126 may mediate EC-Exo-induced neurorestorative effects in T2DM mice. Visual Overview- An online visual overview is available for this article.

Modeling glymphatic system of the brain using MRI.

The glymphatic system is functional waste clearance path from the brain parenchyma through dynamic exchange of cerebrospinal fluid (CSF) with interstitial fluid (ISF). Impairment of glymphatic waste clearance is involved in the development of neurodegenerative conditions. Despite many recent studies investigating the glymphatic system, few studies have tried to use a mathematical model to describe this system, quantitatively. In this study, we aim to model the glymphatic system from the kinetics of Gd-DTPA tracer measured using MRI in order to: 1) map the glymphatic system path, 2) derive kinetic parameters of the glymphatic system, and 3) provide quantitative maps of the structure and function of this system. In the proposed model, the brain is clustered to similar regions with respect to the profile of contrast agent (CA) density measured by MRI. Then, each region is described as a two-compartment kinetic model 'derived from' or 'clears to' its neighbors with local input function. We thus fit our model to the local cerebral regions rather than to the averaged time signal curve (TSC) of the whole brain. The estimated parameters showed distinctive differences between diabetes mellitus (DM) and control rats. The results suggest that in a typical DM brain the CSF bulk speed in the para-vasculature network is low. In addition, the resulting maps indicate that there may be increased binding and decreased absorbing of large molecules in a diabetic compared with a non-diabetic brain. The important contribution of this work was to fit the model to the local regions rather than to the averaged time signal curve (TSC) of the whole brain. This enabled us to derive quantitative maps of the glymphatic system from MRI.

MiR-146a promotes oligodendrocyte progenitor cell differentiation and enhances remyelination in a model of experimental autoimmune encephalomyelitis.

The death of mature oligodendrocytes (OLs) leads to demyelination in the central nervous system (CNS) and subsequently to functional deficits. Remyelination requires the differentiation of oligodendrocyte progenitor cells (OPCs) into myelinating OLs, which in the CNS with neurodegenerative diseases such as multiple sclerosis (MS), is often inhibited. Among the inhibitors of OPC differentiation are toll-like receptor 2 (TLR2) and interleukin-1 receptor-associated kinase 1 (IRAK1) signaling, and both are negatively regulated by microRNA-146a (miR-146a). Therefore, we hypothesized that increase of miR-146a level in the CNS would foster OPC differentiation and remyelination by inhibiting the TLR2/IRAK1 signaling pathway. Here, we tested this hypothesis using exogenous miR-146a mimics and a mouse model of MS, experimental autoimmune encephalomyelitis (EAE) induced by immunization with myelin proteolipid protein peptide (PLP139-151). EAE mice were treated by miR-146a mimics or miR-146a mimic negative controls, respectively, which initiated at day 14 post immunization, once a week for 6 consecutive weeks. Neurological function was monitored daily. Immunofluorescent staining, qRT-PCR and Western blot were used to measure the differentiation of OPCs and myelination, and to investigate the underlying mechanisms of action of miR-146a. Using a fluorescence tracing approach, we found that miR-146a mimics crossed the blood brain barrier and targeted OPCs and microglia/macrophages after systemic administration. MiR-146a mimic treatment substantially improved neurological functional outcome, increased the number of newly generated OLs which may facilitate remyelination in the CNS. The cell number, cytokine level and protein levels of M2 phonotype of microglia/macrophages significantly increased, while cytokine and protein levels of the M1 phenotype significantly decreased after miR-146a mimic treatment. Increased OPC differentiation and remyelination were associated with reduction of TLR2/IRAK1 signaling pathway activity by miR-146a mimic treatment. This study provides insight into the cellular and molecular bases for the therapeutic effects of miR-146a on OPC differentiation and remyelination, and suggests the potential of enhancing miR-146a as a treatment of demyelinating disorders.

Exosomes derived from high-glucose-stimulated Schwann cells promote development of diabetic peripheral neuropathy.

Schwann cells actively interact with axons of dorsal root ganglia (DRG) neurons. Exosomes mediate intercellular communication by transferring their biomaterials, including microRNAs (miRs) into recipient cells. We hypothesized that exosomes derived from Schwann cells stimulated by high glucose (HG) exosomes accelerate development of diabetic peripheral neuropathy and that exosomal cargo miRs contribute to this process. We found that HG exosomes contained high levels of miR-28, -31a, and -130a compared to exosomes derived from non-HG-stimulated Schwann cells. In vitro, treatment of distal axons with HG exosomes resulted in reduction of axonal growth, which was associated with elevation of miR-28, -31a, and -130a and reduction of their target proteins of DNA methyltransferase-3α, NUMB (an endocytic adaptor protein), synaptosome associated protein 25, and growth-associated protein-43 in axons. In vivo, administration of HG exosomes to sciatic nerves of diabetic db/db mice at 7 wk of age promoted occurrence of peripheral neuropathy characterized by impairment of nerve conduction velocity and induction of mechanic and thermal hypoesthesia, which was associated with substantial decreases in intraepidermal nerve fibers. Our findings demonstrate a functional role of exosomes derived from HG-stimulated Schwann cells in mediating development of diabetic peripheral neuropathy.-Jia, L., Chopp, M., Wang, L., Lu, X., Szalad, A., Zhang, Z. G. Exosomes derived from high-glucose-stimulated Schwann cells promote development of diabetic peripheral neuropathy.

Brain-Heart Interaction: Cardiac Complications After Stroke.

Neurocardiology is an emerging specialty that addresses the interaction between the brain and the heart, that is, the effects of cardiac injury on the brain and the effects of brain injury on the heart. This review article focuses on cardiac dysfunction in the setting of stroke such as ischemic stroke, brain hemorrhage, and subarachnoid hemorrhage. The majority of post-stroke deaths are attributed to neurological damage, and cardiovascular complications are the second leading cause of post-stroke mortality. Accumulating clinical and experimental evidence suggests a causal relationship between brain damage and heart dysfunction. Thus, it is important to determine whether cardiac dysfunction is triggered by stroke, is an unrelated complication, or is the underlying cause of stroke. Stroke-induced cardiac damage may lead to fatality or potentially lifelong cardiac problems (such as heart failure), or to mild and recoverable damage such as neurogenic stress cardiomyopathy and Takotsubo cardiomyopathy. The role of location and lateralization of brain lesions after stroke in brain-heart interaction; clinical biomarkers and manifestations of cardiac complications; and underlying mechanisms of brain-heart interaction after stroke, such as the hypothalamic-pituitary-adrenal axis; catecholamine surge; sympathetic and parasympathetic regulation; microvesicles; microRNAs; gut microbiome, immunoresponse, and systemic inflammation, are discussed.

MRI investigation of glymphatic responses to Gd-DTPA infusion rates.

The glymphatic system is a newly identified waste clearance pathway in brain discovered and investigated predominately using in vivo two-photon confocal microscopy. Magnetic resonance imaging (MRI), in contrast to two-photon confocal microscopy, provides dynamic and real-time pictures of the glymphatic system in whole brain. We employ MRI to investigate the response of the glymphatic system to the rate of infusion of Gd-DTPA (magnevist). Wistar rats were subjected to a surgery of inserting a tube into the cisterna magna for infusion during MRI. Three infusion rates were chosen for 20 min infusions of diluted magnevist into the cerebrospinal fluid (CSF) of rat brain. Glymphatic response was imaged using dynamic MRI 3D measurement for 5 hr. Robust correlations were found in all ventricles between the peak intensities of image enhancement and infusion rates, with additional correlations between the peak times of MRI image enhancement and infusion rates in the fourth ventricle. An infusion rate of 2.92 µL/min induced an evident accumulation of tracer in the fourth ventricle near the cisterna magna. In hippocampal tissue, image enhancements exhibited low correlation with the infusion rates. However, an infusion rate of 1.67 µL/min provided a high image enhancement, but less tracer accumulation near the cisterna magna. Contrast-enhanced MRI provides a suitable tool for investigating image contrast infusion rate response of the glymphatic system in rat brain. Considering both T1 and T2* effects in response to the infused magnevist into CSF, the infusion rate of 1.67 µL/min appears suitable for MRI study of the glymphatic system in rat.