Biology subjects

Kuan, C.-Y.

Publications and source records attributed to Kuan, C.-Y..

7 recordsLinked to original sources

bioRxiv2026

Meningeal neutrophil infiltration drives inflammation-exacerbated pediatric stroke through IL-36γ signaling

Infection and systemic inflammation have been identified as major risk factors for pediatric stroke; however, the immune mechanisms underlying these clinical observations remain poorly understood. Here, we utilized an LPS-sensitized photothrombotic stroke model (LPS/PT) to investigate the contribution of immune cell infiltration to pediatric stroke pathogenesis and identified a previously unrecognized route of neutrophil entry into the injured brain. Compared with photothrombotic stroke alone (PT), LPS/PT mice exhibited markedly increased neutrophil infiltration accompanied by a hyperactivated phenotype. Depletion of neutrophils, but not monocytes, significantly reduced infarct size in LPS/PT animals, indicating a central role for neutrophils in inflammation-exacerbated pediatric stroke. Notably, we observed that neutrophils first accumulated within the leptomeningeal space before entering the brain parenchyma during the early phase of stroke, suggesting that the meninges may serve as an initial staging site for neutrophil recruitment. Using KikGR photoconvertible reporter mice combined with two-photon imaging, we further demonstrated that neutrophils infiltrate the ischemic brain through a compromised meningeal barrier following stroke. Transcriptomic analysis of infiltrating neutrophils revealed distinct gene expression signatures between meningeal neutrophils and circulating blood-derived neutrophils. Among these, IL-36{gamma} was highly enriched in meninges-associated neutrophils during pediatric stroke. Consistently, single-cell RNA sequencing of meningeal immune cells confirmed elevated IL-36{gamma} expression within the neutrophil cluster in LPS/PT animals. Importantly, intracisternal administration of IL-36 receptor antagonist (IL-36Ra) or anti-ICAM-1 antibody significantly reduced infarct volume in this pediatric stroke model. Together, our findings identify the meningeal barrier as a critical gateway for neutrophil infiltration and reveal IL-36{gamma} as a key inflammatory mediator regulating neuroinflammation in pediatric stroke, highlighting a potential therapeutic target for limiting immune-mediated brain injury.

neuroscience↗

bioRxiv2024

Microglia modulate cerebral blood flow and neurovascular coupling through ectonucleotidase CD39

Microglia and the border-associated macrophages (BAMs) contribute to the modulation of cerebral blood flow (CBF), but the mechanisms have remained ill-defined. Here, we show that microglia regulate the CBF baseline and upsurges after whisker stimulation or intracisternal magna injection of adenosine triphosphate (ATP). Genetic or pharmacological depletion of microglia reduces the activity-dependent hyperemia but not the cerebrovascular responses to adenosine stimulation. Notably, microglia repopulation corrects these CBF reactivity deficits. The microglial-dependent regulation of CBF requires the ATP-sensing P2ry12 receptor and the ectonucleotidase CD39 that initiates the breakdown of extracellular ATP. Pharmacological inhibition or microglia-specific deletion of CD39 simulates the CBF anomalies detected in microglia-deficient mice and reduces the rise of extracellular adenosine after whisker stimulation. Together, these results suggest that the microglial CD39-initiated conversion of extracellular ATP to adenosine is an important step in neurovascular coupling and the regulation of cerebrovascular reactivity.

neuroscience↗

bioRxiv2023

P-tau Inflicts Intracellular Stress Responses That Are Mitigated by Apomorphine

BackgroundAbnormal phosphorylation of the microtubule-binding protein tau in the brain is a key pathological marker for Alzheimers disease and additional neurodegenerative tauopathies. However, how hyperphosphorylated tau causes cellular dysfunction or death that underlie neurodegeneration remains an unsolved question critical for the understanding of disease mechanism and the design of efficacious drugs. MethodsUsing a recombinant hyperphosphorylated tau protein (p-tau) synthesized by the PIMAX approach, we examined how cells responded to the cytotoxic tau and explored means to enhance cellular resistance to tau attack. ResultsUpon p-tau uptake, the intracellular calcium levels rose promptly. Gene expression analyses revealed that p-tau potently triggered endoplasmic reticulum (ER) stress, Unfolded Protein Response (UPR), ER stress-associated apoptosis, and pro-inflammation in cells. Proteomics studies showed that p-tau diminished heme oxygenase-1 (HO-1), an ER stress associated anti-inflammation and anti-oxidative stress regulator, while stimulated the accumulation of MIOS and other proteins. P-tau-induced ER stress-associated apoptosis and pro-inflammation are ameliorated by apomorphine, a brain-permeable prescription drug widely used to treat Parkinsons disease symptoms, and by overexpression of HO-1. ConclusionOur results reveal probable cellular functions targeted by hyperphosphorylated tau. Some of these dysfunctions and stress responses have been linked to neurodegeneration in Alzheimers disease. The observations that the ill effects of p-tau can be mitigated by a small compound and by overexpressing HO-1 that is otherwise diminished in the treated cells inform new directions of Alzheimers disease drug discovery.

biochemistry↗

bioRxiv2021

Hypothermia Suppresses Uncoupling of Oxidative-Phosphorylation after Neonatal Cerebral Hypoxia-Ischemia

Hypoxia-ischemia (HI), which disrupts the oxygen supply-demand balance in the brain by impairing blood oxygen supply and the cerebral metabolic rate of oxygen (CMRO2), is a leading cause of neonatal brain injury. However, it is unclear how post-HI hypothermia helps to restore the balance, as cooling reduces CMRO2. Also, how transient HI leads to secondary energy failure (SEF) in neonatal brains remains elusive. Using photoacoustic microscopy, we examined the effects of HI on CMRO2 in awake 10-day-old mice, supplemented by bioenergetic analysis of purified cortical mitochondria. Our results show that while HI suppresses ipsilateral CMRO2, it sparks a prolonged CMRO2-surge post-HI, associated with increased mitochondrial oxygen consumption, superoxide emission, and reduced mitochondrial membrane potential necessary for ATP synthesis--indicating oxidative phosphorylation (OXPHOS) uncoupling. Post-HI hypothermia prevents the CMRO2-surge by constraining oxygen extraction fraction, reduces mitochondrial oxidative stress, and maintains ATP and N-acetylaspartate levels, resulting in attenuated infarction at 24 hours post-HI. Our findings suggest that OXPHOS-uncoupling induced by the post-HI CMRO2-surge underlies SEF and blocking the surge is a key mechanism of hypothermia protection. Also, our study highlights the potential of optical CMRO2-measurements for detecting neonatal HI brain injury and guiding the titration of therapeutic hypothermia at the bedside.

neuroscience↗

bioRxiv2021

Adropin confers neuroprotection and promotes functional recovery from ischemic stroke

Adropin is a highly-conserved peptide that has been shown to preserve endothelial barrier function. Blood-brain barrier (BBB) disruption is a key pathological event in cerebral ischemia. However, the effects of adropin on ischemic stroke outcomes remain unexplored. Hypothesizing that adropin exerts neuroprotective effects by maintaining BBB integrity, we investigated the role of adropin in stroke pathology utilizing loss- and gain-of-function genetic approaches combined with pharmacological treatment with synthetic adropin peptide. Stroke decreased endogenous adropin levels in the brain and plasma. Adropin treatment or transgenic adropin overexpression robustly reduced brain injury and improved long-term sensorimotor and cognitive function in young and aged mice subjected to ischemic stroke. In contrast, genetic deletion of adropin exacerbated ischemic brain injury. Mechanistically, adropin neuroprotection depends on endothelial nitric oxide synthase and is associated with reduced BBB permeability and neuroinflammation. We identify adropin as a novel neuroprotective peptide with the potential to improve stroke outcomes.

neuroscience↗

bioRxiv2021

Capillary-associated microglia regulate vascular structure and function through PANX1-P2RY12 coupling

Microglia are brain-resident immune cells with a repertoire of functions in the developing, mature and pathological brain. Their wide-ranging roles in physiology include the clearance of cellular debris, elimination of excess synapses, regulation of neuronal activity and contributions to blood vessel development. Despite these known roles for microglia, the extent of their interactions with the vasculature and potential regulation of vascular physiology has been insufficiently explored. Here, using in vivo acute and longitudinal two-photon imaging in transgenic mice combined with electron microscopy, fixed tissue immunohistochemistry, pharmacological treatments and laser speckle imaging, we document the steady-state interactions between ramified CX3CR1+ myeloid cell somata and capillaries in the brain. We first confirm that these myeloid cells are bona fide microglia by molecular, morphological and ultrastructural approaches. Then we give a detailed spatio-temporal characterization of these capillary-associated microglia (CAMs) comparing and contrasting them with parenchymal microglia (PCMs) in their static, dynamic and chronic morphological activities including during microglial depletion and repopulation. Molecularly, we identify microglial-specific purinergic P2RY12 receptors as a receptor regulating CAM interactions under the control of released purines from pannexin 1 (PANX1) channels. Furthermore, to elucidate roles for microglia in vascular structure and function, we eliminated microglia and showed that this triggered capillary dilation, blood flow increase, and impaired vasodilative responses. We find that P2RY12-/- and PANX1-/- mice recapitulate these vascular impairments suggesting purines released through PANX1 channels play important roles in activating microglial P2RY12 receptors to regulate neurovascular structure and function.

neuroscience↗

bioRxiv2018

Adenosine Deaminase Acting on RNA 1 Associates with Orf Virus OV20.0 and Enhances Viral Replication

Orf virus (ORFV) infects sheep and goats and is also an important zoonotic pathogen. The viral protein OV20.0 has been shown to suppress innate immunity by targeting the double-stranded RNA (dsRNA)-activated protein kinase (PKR) by multiple mechanisms. These mechanisms include a direct interaction with PKR and binding with two PKR activators, dsRNA and the cellular PKR activator (PACT), which ultimately leads to the inhibition of PKR activation. In the present study, we identified a novel association between OV20.0 and adenosine deaminase acting on RNA 1 (ADAR1). OV20.0 bound directly to the dsRNA binding domains (RBDs) of ADAR1 in the absence of dsRNA. Additionally, OV20.0 preferentially interacted with RBD1 of ADAR1, which was essential for its dsRNA binding ability and for the homodimerization that is critical for intact adenosine (A)-to-inosine (I) editing activity. Finally, the association with OV20.0 suppressed the A-to-I editing ability of ADAR1, while ADAR1 played a proviral role during ORFV infection by inhibiting PKR phosphorylation. These observations revealed a new strategy used by OV20.0 to evade antiviral responses via PKR.\n\nImportanceViruses evolve specific strategies to counteract host innate immunity. ORFV, an important zoonotic pathogen, encodes OV20.0 to suppress PKR activation via multiple mechanisms, including interactions with PKR and two PKR activators. In this study, we demonstrated that OV20.0 interacts with ADAR1, a cellular enzyme responsible for converting adenosine (A) to inosine (I) in RNA. The RNA binding domains, but not the catalytic domain, of ADAR1 are required for this interaction. The OV20.0-ADAR1 association affects the functions of both proteins; OV20.0 suppressed the A-to-I editing of ADAR1, while ADAR1 elevated OV20.0 expression. The proviral role of ADAR1 is likely due to the inhibition of PKR phosphorylation. As RNA editing by ADAR1 contributes to the stability of the genetic code and the structure of RNA, these observations suggest that in addition to serving as a PKR inhibitor, OV20.0 might modulate ADAR1-dependent gene expression to combat antiviral responses or achieve efficient viral infection.

microbiology↗