Doctor of Philosophy, Zhejiang University (2019)
Bachelor of Science, Zhejiang University (2014)
Tony Wyss-Coray, Postdoctoral Faculty Sponsor
Blood-brain barrier (BBB) dysfunction has been suggested to play an important role in epilepsy. However, the mechanism mediating the transition from cerebrovascular damage to epilepsy remains unknown. Here, we report that endothelial cyclin-dependent kinase 5 (CDK5) is a central regulator of neuronal excitability. Endothelial-specific Cdk5 knockout led to spontaneous seizures in mice. Knockout mice showed increased endothelial chemokine (C-X-C motif) ligand 1 (Cxcl1) expression, decreased astrocytic glutamate reuptake through the glutamate transporter 1 (GLT1), and increased glutamate synaptic function. Ceftriaxone restored astrocytic GLT1 function and inhibited seizures in endothelial Cdk5-deficient mice, and these effects were also reversed after silencing Cxcl1 in endothelial cells and its receptor chemokine (C-X-C motif) receptor 2 (Cxcr2) in astrocytes, respectively, in the CA1 by AAV transfection. These results reveal a previously unknown link between cerebrovascular factors and epileptogenesis and provide a rationale for targeting endothelial signaling as a potential treatment for epilepsy.
View details for DOI 10.1084/jem.20180992
View details for PubMedID 31699822
View details for PubMedCentralID PMC7037235
The proper interactions between blood vessels and neurons are critical for maintaining the strength of neural circuits and cognitive function. However, the precise molecular events underlying these interactions remain largely unknown. Here, we report that the selective knockout of semaphorin 3G (Sema3G) in endothelial cells impaired hippocampal-dependent memory and reduced dendritic spine density in CA1 neurons in mice; these effects were reversed after restoration of Sema3G levels in the hippocampus by AAV transfection. We further show that Sema3G increased excitatory synapse density via neuropilin-2/PlexinA4 signaling and through activation of Rac1. These results provide the first evidence that, in the central nervous system, endothelial Sema3G serves as a vascular-derived synaptic organizer that regulates synaptic plasticity and hippocampal-dependent memory. Our findings highlight the role of vascular endothelial cells in regulating cognitive function through intercellular communication with neurons in the hippocampus.
View details for DOI 10.1016/j.neuron.2018.12.036
View details for PubMedID 30685224
Grb2-associated-binding protein 1 (Gab1) is a docking/scaffolding molecule known to play an important role in cell growth and survival. Here, we report that Gab1 is decreased in cholinergic neurons in Alzheimer's disease (AD) patients and in a mouse model of AD. In mice, selective ablation of Gab1 in cholinergic neurons in the medial septum impaired learning and memory and hippocampal long-term potentiation. Gab1 ablation also inhibited SK channels, leading to an increase in firing in septal cholinergic neurons. Gab1 overexpression, on the other hand, improved cognitive function and restored hippocampal CaMKII autorphosphorylation in AD mice. These results suggest that Gab1 plays an important role in the pathophysiology of AD and may represent a novel therapeutic target for diseases involving cholinergic dysfunction.
View details for DOI 10.1093/cercor/bhx141
View details for PubMedID 28591834
This communication reports on a fluorescent probe (PPI-P) for imaging active peptidyl-prolyl cis/trans isomerases in live cells. PPI-P is capable of responding to both recombinant and cellular PPIases fluorogenically, and has been shown to specifically image active PPIases in live cells.
View details for DOI 10.1039/c7cc09135d
View details for PubMedID 29387835
Dysfunction of neurovascular pericytes underlies breakdown of the blood-brain barrier, but the molecular mechanisms are largely unknown. In this study, we evaluated the role of the transient receptor potential melastatin-related 2 (TRPM2) channel and autophagy during brain pericyte injury both in vitro and in vivo.A rapid induction in autophagy in human brain vascular pericytes, in the zinc oxide nanoparticles (ZnO-NP)-induced cell stress model, was paralleled with an increase in the expression of the TRPM2-S truncated isoform, which was abolished by treatment with a nitric oxide synthase inhibitor and a peroxynitrite scavenger. Furthermore, Y1485 in the C-terminus of the TRPM2 protein was identified as the tyrosine nitration substrate by mass spectrometry. Overexpression of the Y1485S TRPM2 mutant reduced LC3-II accumulation and pericyte injury induced by ZnO-NP. Consistently, LC3-II accumulation was reduced and pericytes were better preserved in intact brain microvessels of the TRPM2 knockout mice after ZnO-NP-induced vascular injury. Innovation and Conclusions: Our present study has revealed a novel mechanism of autophagy disturbance secondary to nitrosative stress-induced tyrosine nitration of TRPM2 during pericyte injury. Antioxid. Redox Signal. 27, 1297-1316.
View details for DOI 10.1089/ars.2016.6873
View details for PubMedID 28292196
The receptor tyrosine kinase ErbB4 is present throughout the primate brain and has a distinct functional profile. In this study, we investigate the potential role of endothelial ErbB4 receptor signaling in the brain.Here, we show that the endothelial cell-specific deletion of ErbB4 induces decreased exploratory behavior in adult mice. However, the water maze task for spatial memory and the memory reconsolidation test reveal no changes; additionally, we observe no impairment in CaMKII phosphorylation in Cdh5Cre;ErbB4f/f mice, which indicates that the endothelial ErbB4 deficit leads to decreased exploratory activity rather than direct memory deficits. Furthermore, decreased brain metabolism, which was measured using micro-positron emission tomography, is observed in the Cdh5Cre;ErbB4f/f mice. Consistently, the immunoblot data demonstrate the downregulation of brain Glut1, phospho-ULK1 (Ser555), and TIGAR in the endothelial ErbB4 conditional knockout mice. Collectively, our findings suggest that endothelial ErbB4 plays a critical role in regulating brain function, at least in part, through maintaining normal brain energy homeostasis.Targeting ErbB4 or the modulation of endothelial ErbB4 signaling may represent a rational pharmacological approach to treat neurological disorders.
View details for DOI 10.1111/cns.12695
View details for PubMedID 28421673
View details for PubMedCentralID PMC6492688
Severe hypoglycemia has a detrimental impact on the cerebrovasculature, but the molecular events that lead to the disruption of the integrity of the tight junctions remain unclear. Here, we report that the microvessel integrity was dramatically compromised (59.41% of wild-type mice) in TP53-induced glycolysis and apoptosis regulator (TIGAR) transgenic mice stressed by hypoglycemia. Melatonin, a potent antioxidant, protects against hypoglycemic stress-induced brain endothelial tight junction injury in the dosage of 400 nmol/L in vitro. FRET (fluorescence resonance energy transfer) imaging data of endothelial cells stressed by low glucose revealed that TIGAR couples with calmodulin to promote TIGAR tyrosine nitration. A tyrosine 92 mutation interferes with the TIGAR-dependent NADPH generation (55.60% decreased) and abolishes its protective effect on tight junctions in human brain microvascular endothelial cells. We further demonstrate that the low-glucose-induced disruption of occludin and Caludin5 as well as activation of autophagy was abrogated by melatonin-mediated blockade of nitrosative stress in vitro. Collectively, we provide information on the detailed molecular mechanisms for the protective actions of melatonin on brain endothelial tight junctions and suggest that this indole has translational potential for severe hypoglycemia-induced neurovascular damage.
View details for DOI 10.1111/jpi.12440
View details for PubMedID 28776759
View details for PubMedCentralID PMC5656838
Clinical treatment for vascular dementia still remains a challenge mainly due to the blood-brain barrier (BBB). Here, a micelle based on polysialic acid (PSA), which is a hydrophilic and endogenous carbohydrate polymer, was designed to deliver calmodulin antagonist for therapy of vascular dementia. PSA was first chemically conjugated with octadecylamine (ODA), and the obtained PSA-ODA copolymer could self-assemble into micelle in aqueous solution with a 120.0 ?g/mL critical micelle concentration. The calmodulin antagonist loaded PSA-ODA micelle, featuring sustained drug release behavior over a period of 72 h with a 3.6% (w/w) drug content and a 107.0 ± 4.0 nm size was then fabricated. The PSA-ODA micelle could cross the BBB mainly via active endocytosis by brain endothelial cells followed by transcytosis. In a water maze test for spatial learning, calmodulin antagonist loaded PSA-ODA micelle significantly reduced the escape latencies of right unilateral common carotid arteries occlusion (rUCCAO) mice with dosage significantly reduced versus free drug. The decrease of hippocampal phospho-CaMKII (Thr286/287) and phospho-synapsin I (Ser603) was partially restored in rUCCAO mice following calmodulin antagonist loaded PSA-ODA micelle treatment. Consistent with the restored CaMKII phosphorylation, the elevation of BrdU/NeuN double-positive cells in the same context was also observed. Overall, the PSA-ODA micelle developed from the endogenous material might promote the development of therapeutic approaches for improving the efficacy of brain-targeted drug delivery and have great potential for vascular dementia treatment.
View details for DOI 10.1021/acsami.6b13052
View details for PubMedID 27750011
Septic encephalopathy (SE) is a critical factor determining sepsis mortality. Vascular inflammation is known to be involved in SE, but the molecular events that lead to the development of encephalopathy remain unclear. Using time-lapse in vivo two-photon laser scanning microscopy, we provide the first direct evidence that cecal ligation and puncture in septic mice induces microglial trafficking to sites adjacent to leukocyte adhesion on inflamed cerebral microvessels. Our data further demonstrate that septic injury increased the chemokine CXCL1 level in brain endothelial cells by activating endothelial P2RX7 and eventually enhanced the binding of Mac-1 (CD11b/CD18)-expressing leukocytes to endothelial ICAM-1. In turn, leukocyte adhesion upregulated endothelial CX3CL1, thereby triggering microglia trafficking to the injured site. The sepsis-induced increase in endothelial CX3CL1 was abolished in CD18 hypomorphic mutant mice. Inhibition of the P2RX7 pathway not only decreased endothelial ICAM-1 expression and leukocyte adhesion but also prevented microglia overactivation, reduced brain injury, and consequently doubled the early survival of septic mice. These results demonstrate the role of the P2RX7 pathway in linking neurovascular inflammation to brain damage in vivo and provide a rationale for targeting endothelial P2RX7 for neurovascular protection during SE.
View details for DOI 10.1038/cr.2015.61
View details for PubMedID 25998681
View details for PubMedCentralID PMC4456628
Accumulating evidence suggests that formation of peroxynitrite (ONOO(-)) in the cerebral vasculature contributes to the progression of ischemic damage, while the underlying molecular mechanisms remain elusive. To fully understand ONOO(-) biology, efficient tools that can realize the real-time tracing of endogenous ONOO(-) fluxes are indispensable. While a few ONOO(-) fluorescent probes have been reported, direct visualization of ONOO(-) fluxes in the cerebral vasculature of live mice remains a challenge. Herein, we present a fluorescent switch-on probe (NP3) for ONOO(-) imaging. NP3 exhibits good specificity, fast response, and high sensitivity toward ONOO(-) both in vitro and in vivo. Moreover, NP3 is two-photon excitable and readily blood-brain barrier penetrable. These desired photophysical and pharmacokinetic properties endow NP3 with the capability to monitor brain vascular ONOO(-) generation after injury with excellent temporal and spatial resolution. As a proof of concept, NP3 has enabled the direct visualization of neurovascular ONOO(-) formation in ischemia progression in live mouse brain by use of two-photon laser scanning microscopy. Due to these favorable properties, NP3 holds great promise for visualizing endogenous peroxynitrite fluxes in a variety of pathophysiological progressions in vitro and in vivo.
View details for DOI 10.1021/jacs.5b06865
View details for PubMedID 26352914
Although there is accumulating evidence that increased formation of reactive nitrogen species in cerebral vasculature contributes to the progression of ischemic damage, but the underlying molecular mechanisms remain elusive. Peroxiredoxin 1 (Prx1) can initiate the antioxidant response by scavenging free radicals. Therefore, we tested the hypothesis that Prx1 regulates the susceptibility to nitrosative stress damage during cerebral ischemia in vitro and in vivo.Proteomic analysis in endothelial cells revealed that Prx1 was upregulated after stress-related oxygen-glucose deprivation (OGD). Although peroxynitrite upregulated Prx1 rapidly, this was followed by its polyubiquitination within 6 h after OGD mediated by the E3 ubiquitin ligase E6-associated protein (E6AP). OGD colocalized E6AP with nitrotyrosine in endothelial cells. To assess translational relevance in vivo, mice were studied after middle cerebral artery occlusion (MCAO). This was accompanied by Prx1 ubiquitination and degradation by the activation of E6AP. Furthermore, brain delivery of a lentiviral vector encoding Prx1 in mice inhibited blood-brain barrier leakage and neuronal damage significantly following MCAO.Nitrosative stress during ischemic insult activates E6AP E3 ubiquitin ligase that ubiquitinates Prx1 and subsequently worsens cerebral damage. Thus, targeting the Prx1 antioxidant defense pathway may represent a novel treatment strategy for neurovascular protection in stroke.
View details for DOI 10.1089/ars.2013.5381
View details for PubMedID 24295341
View details for PubMedCentralID PMC4048580
The present study was designed to investigate the role of autophagy-lysosome signaling in the brain after application of nanoparticles. Here, lipid nanoparticles (LNs) induced elevations of Atg5, P62, LC3 and cathepsin B in mice brain. The transmission electron microscopy revealed a dramatic elevation of lysosome vacuoles colocalized with LNs cluster inside the neurons in mice brain. Immunoblot data revealed abnormal expression of cathepsin B in brain cortex following LNs injection, whereas its expression was further elevated in Atg5(+/-) mice. The importance of Atg5 in the LNs-induced autophagy-lysosome cascade was further supported by our finding that neurovascular response was exaggerated in Atg5(+/-) mice. In addition, the siRNA knockdown of Atg5 significantly blunted the increasing of LC3 and P62 in LNs-treated Neuro-2a cells. Taken together, we propose that LNs induce autophagy-lysosome signaling and neurovascular response at least partially via an Atg5-dependent pathway.These authors investigated autophagy-lysosome signaling in the mouse brain after application of lipid nanoparticles and report that these nanoparticles induce autophagy-lysosome signaling and neurovascular response at least partially via an Atg5-dependent pathway.
View details for DOI 10.1016/j.nano.2014.03.019
View details for PubMedID 24768629