Biology subjects

Chiang, W.

Publications and source records attributed to Chiang, W..

4 recordsLinked to original sources

bioRxiv2026

Dual kinase inhibition provides early clues to restoration of neuroimmune homeostasis in an AAV-haSyn model of Parkinsons disease

Background Previous studies of drugs targeting mixed linage kinase type 3 (MLK3) and/or leucine rich repeat kinase type 2 (LRRK2) in sporadic Parkinsons disease (PD) have failed to meet clinical endpoints despite multiple lines of evidence implicating these kinases in pathogenesis. This has reduced enthusiasm for MLK3 and LRRK2 as druggable targets. Largely overlooked is the evidence that these targets remain valid but must be considered in combination to achieve a therapeutic outcome. This provides the rationale for this study. Methods We conducted a pilot study modeling early, pre-symptomatic PD with subsequent evaluation of therapeutic potential of a multi-kinase inhibitor (URMC-099) in alleviating disease pathology. Sprague Dawley rats received unilateral intranigral injections of low dose AAV-haSyn. After a two-week recovery period, all animals were administered either once-daily intraperitoneal (i.p.) injection of URMC-099 or its vehicle control until their terminal endpoint. Induction of early PD, and its alleviation with URMC-099, was evaluated by lateralized motor impairment using the cylinder test and immunohistochemical analysis of neuropathology in the substantia nigra (SN) and striatum. Finally, an in vitro microglial PD model exposed to aSyn pre-formed fibrils and treated with URMC-099 was used to further evaluate the mechanism underlying therapeutic activity. Results A unilateral model of early and progressive AAV-haSyn model for PD was used to demonstrate the therapeutic potential of a targeted multi-kinase inhibitor to attenuate PD-associated neuropathology and prevent the manifestation of lateral motor deficits. Immunohistochemical analysis revealed protection of neuronal cellular integrity along the nigral-striatal pathway with once-daily i.p. URMC-099. The mitigation of degenerative neuropathology was linked to targeted inhibition of MLK3 and LRRK2 and complemented by reduced levels of gliosis in astrocytes and microglia in the SN. Single-microglia segmentation analysis suggested URMC-099 treatment preserved dynamic effector state switching in microglia to enhance autophagic flux to reduce the overall burden of pathologic aSyn. Transcriptomic analysis of a complementary in vitro microglia model corroborated the retention of a homeostatic-like effector state with URMC-099 treatment that was unique to MLK3/LRRK2 dual inhibition. Conclusions Unilateral injection of low dose AAV-haSyn successfully resulted in a lateralized model of early, progressive PD. Dual inhibition of aberrant MLK3 and LRRK2 activity with once-daily i.p. URMC-099 therapy significantly mitigated the development of PD pathology and progression in an early manifest phenotype. Our results present a potential mechanism of therapeutic action dependent on the dual inhibition of MLK3 and LRRK2. This presents a novel therapeutic modality that may be beneficial to a broad population of etiologic PD patients.

neuroscience↗

bioRxiv2023

Neutrophilia with damage to the blood-brain barrier and neurovascular unit following acute lung injury

BackgroundThe role of neutrophils in mediating neurovascular vulnerability has been increasingly implicated in various acute inflammatory models of neuroimmune crosstalk between the periphery and the brain. Whether neurovascular vulnerability is similarly modulated in the context of frequent, but not acute, inflammatory activation in the periphery is the aim of our study. Such a model of frequent inflammatory irritation is pertinent to understanding the neurologic risk of constant exposure to aerosolized environmental hazards leading to progressive pulmonary disease. MethodsTo model repeated pulmonary inflammation, we applied a three-dose regimen of intranasal (i.n.) lipopolysaccharide (LPS) in C57BL/6J mice and studied the impact on the inflammatory environment of the brain, with a specific focus on neutrophil dynamics at the neurovascular unit (NVU). Tissue and circulatory inflammatory profiles were screened via bronchoalveolar lavage (BAL) protein content and cellularity, transcript analysis of brain tissue, and flow cytometry of peripheral blood. Intravital two-photon microscopy (2PM) of the brain vasculature identified neutrophil dynamics at the NVU. Immunofluorescence validated neutrophil dynamics and identified neuroinflammatory hallmarks and peripheral immune factor interactions at the NVU. In vivo findings were corroborated and replicated in murine and human microphysiological systems (MPS) modeling the blood-brain barrier as a proxy demonstration of the translational relevance of our findings. Results2PM of tdTomato-Ly6G+ neutrophils demonstrated increased levels of circulating neutrophils and corresponding engagement with the brain vasculature after the three-dose repeated i.n. exposure regimen. Neutrophilia at the NVU was corroborated with increased transcript levels of Ly6G and other pro-inflammatory markers. This coordination between endothelial physiology and neutrophil phenotypes was recapitulated in murine and human MPS models. System-wide neutrophilia in the lung and circulation was found to be cotemporaneous to neutrophilia at the NVU based on the cellularity of BAL and peripheral blood samples collected at the same endpoints. Immunohistochemical analysis of brain tissue implicates temporal coordination between vascular surface adhesion molecules with changes in neutrophil dynamics from adhesion, crawling, stalling, and transmigration. Extravasation of neutrophils was complemented by sustained paravascular deposition of fibrinogen and microgliosis up to 72 hours after the final i.n. dosing. Microglia-associated effector functions for synaptic pruning and regulation of neutrophil activity demonstrated distinct temporal profiles. ConclusionsOur results identify systemic levels of neutrophilia accompanied by ingress and extravascular accumulation in brain parenchyma that correlated with sustained microglial activation. This neutrophil-centric lung-brain interaction is complemented by the observation of paravascular fibrinogen deposition that alters synaptic metabolism. Thus, we propose a mechanistic role for neutrophilia and associated inflammatory dysregulation as essential mediators along the lung-brain neuroimmune axis in a generalizable model of repeated respiratory exposure to inflammatory agents. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=119 SRC="FIGDIR/small/562508v2_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@113d747org.highwire.dtl.DTLVardef@19a8a5corg.highwire.dtl.DTLVardef@f1a1eaorg.highwire.dtl.DTLVardef@1bd23c7_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2023

Hybrid Amyloid Quantum Dot Nanoassemblies to Probe Neuroinflammatory Damage

Various oligomeric species of amyloid-beta have been proposed to play different immunogenic roles in the cellular pathology of Alzheimers Disease. However, investigating the role of a homogenous single oligomeric species has been difficult due to highly dynamic oligomerization and fibril formation kinetics that convert between many species. Here we report the design and construction of a quantum dot mimetic for larger spherical oligomeric amyloid species as an "endogenously" fluorescent proxy for this cytotoxic species to investigate its role in inducing inflammatory and stress response states in neuronal and glial cell types.

neuroscience↗

bioRxiv2022

A Quantum Dot Biomimetic for SARS-CoV-2 to Interrogate Dysregulation of the Neurovascular Unit Relevant to Brain Inflammation

Despite limited evidence for competent infection and viral replication of SARS-CoV-2 in the central nervous system (CNS), neurologic dysfunction is a common post-acute medical condition reported in "recovered" COVID-19 patients. To identify a potential noninfectious route for SARS-CoV-2-mediated neurological damage, we constructed colloidal nanocrystal quantum dots linked to micelles decorated with spike protein (COVID-QDs) as a biomimetic to interrogate how blood-brain barrier (BBB) dysregulation may subsequently induce neuroinflammation in the absence of infection. In transwell co-culture of endothelial bEnd.3 monolayers and primary neuroglia, we exposed only the bEnd.3 monolayers to COVID-QDs and examined by fluorescence microscopy whether such treatment led to (i) increased inflammation and leakage across the bEnd.3 monolayers, (ii) permeability of the COVID-QDs across the monolayers, and (iii) induction of neuroinflammation in neuroglial cultures. The results of our study provide evidence of neuroinflammatory hallmarks in cultured neurons and astrocytes without direct exposure to SARS-CoV-2-like nanoparticles. Additionally, we found that pre-treatment of our co-cultures with a small-molecule, broad-spectrum inhibitor of mixed lineage and leucine rich repeat kinases led to reversal of the observed dysregulation in endothelial monolayers and resulted in neuroglial protection. The results reported here may serve to guide future studies into the potential mechanisms by which SARS-CoV-2 mediates neurologic dysfunction.

neuroscience↗