Dohee Kim

Identification of disulfide cross-linked tau dimer responsible for tau propagation

Recent evidence suggests that tau aggregates are not only neurotoxic, but also propagate in neurons acting as a seed for native tau aggregation. Prion-like tau transmission is now considered as an important pathogenic mechanism driving the progression of tau pathology in the brain. However, prion-like tau species have not been clearly characterized. To identify infectious tau conformers, here we prepared diverse tau aggregates and evaluated the effect on inducing intracellular tau-aggregation. Among tested, tau dimer containing P301L-mutation is identified as the most infectious form to induce tau pathology. Biochemical analysis reveals that P301L-tau dimer is covalently cross-linked with a disulfide bond. The relatively small and covalently cross-linked tau dimer induced tau pathology efficiently in primary neurons and also in tau-transgenic mice. So far, the importance of tau disulfide cross-linking has been overlooked in the study of tau pathology. Here our results suggested that tau disulfide cross-linking might play critical role in tau propagation by producing structurally stable and small tau conformers.

Gamma Linolenic Acid Exerts Anti-Inflammatory and Anti-Fibrotic Effects in Diabetic Nephropathy

Purpose This study was undertaken to investigate the effects of gamma linolenic acid (GLA) on inflammation and extracellular matrix (ECM) synthesis in mesangial and tubular epithelial cells under diabetic conditions. Materials and Methods Sprague-Dawley rats were intraperitoneally injected with either a diluent [n=16, control (C)] or streptozotocin [n=16, diabetes (DM)], and eight rats each from the control and diabetic groups were treated with evening primrose oil by gavage for three months. Rat mesangial cells and NRK-52E cells were exposed to medium containing 5.6 mM glucose and 30 mM glucose (HG), with or without GLA (10 or 100 µM). Intercellular adhesion molecule-1 (ICAM-1), monocyte chemoattractant protein-1 (MCP-1), and fibronectin (FN) mRNA and protein expression levels were evaluated. Results Twenty-four-hour urinary albumin excretion was significantly increased in DM compared to C rats, and GLA treatment significantly reduced albuminuria in DM rats. ICAM-1, MCP-1, FN mRNA and protein expression levels were significantly higher in DM than in C kidneys, and these increases were significantly abrogated by GLA treatment. In vitro, GLA significantly inhibited increases in MCP-1 mRNA expression and protein levels under high glucose conditions in HG-stimulated mesangial and tubular epithelial cells (p<0.05, respectively). ICAM-1 and FN expression showed a similar pattern to the expression of MCP-1. Conclusion GLA attenuates not only inflammation by inhibiting enhanced MCP-1 and ICAM-1 expression, but also ECM accumulation in diabetic nephropathy.

Inhibition of tau aggregation by a rosamine derivative that blocks tau intermolecular disulfide cross-linking

Abstract Abnormal tau aggregates are presumed to be neurotoxic and are an important therapeutic target for multiple neurodegenerative disorders including Alzheimer’s disease. Growing evidence has shown that tau intermolecular disulfide cross-linking is critical in generating tau oligomers that serve as a building block for higher-order aggregates. Here we report that a small molecule inhibitor prevents tau aggregation by blocking the generation of disulfide cross-linked tau oligomers. Among the compounds tested, a rosamine derivative bearing mild thiol reactivity selectively labeled tau and effectively inhibited oligomerization and fibrillization processes in vitro. Our data suggest that controlling tau oxidation status could be a new therapeutic strategy for prevention of abnormal tau aggregation.

Cell-based Models To Investigate Tau Aggregation

Accumulation of abnormal tau aggregates in neuron is an important pathological signature in multiple neurodegenerative disorders including Alzheimers disease. Tau is a neuron specific microtubule-associated protein that regulates microtubule stability, which is critical for axonal outgrowth and synaptic plasticity. In a pathological condition, tau dissociates from microtubules and forms insoluble aggregates called neurofibrillary tangles (NFTs). The accumulation of NFTs in neuron directly correlates with microtubule dysfunction and neuronal degeneration. Due to the pathophysiological importance of tau, great efforts have been made to understand tau aggregation processes and find therapeutics to halt or reverse the processes. However, progress has been slow due to the lack of a suitable method for monitoring tau aggregation. In this mini-review, we will review the conventional methods for studying tau aggregation, and introduce recent cell-based sensor approaches that allow monitoring tau aggregation in living cells.

Age-dependent inverse correlations in CSF and plasma amyloid-β(1-42) concentrations prior to amyloid plaque deposition in the brain of 3xTg-AD mice.

Amyloid-β (Aβ) plays a critical role as a biomarker in Alzheimer’s disease (AD) diagnosis. In addition to its diagnostic potential in the brain, recent studies have suggested that changes of Aβ level in the plasma can possibly indicate AD onset. In this study, we found that plasma Aβ(1–42) concentration increases with age, while the concentration of Aβ(1–42) in the cerebrospinal fluid (CSF) decreases in APPswe, PS1M146V and TauP301L transgenic (3xTg-AD) mice, if measurements were made before formation of ThS-positive plaques in the brain. Our data suggests that there is an inverse correlations between the plasma and CSF Aβ(1–42) levels until plaques form in transgenic mice’s brains and that the plasma Aβ concentration possesses the diagnostic potential as a biomarker for diagnosis of early AD stages.

MEMS neural probe array for multiple-site optical stimulation with low-loss optical waveguide by using thick glass cladding layer

We present a MEMS neural probe array for multiple-site optical stimulation with low-loss SU-8 optical waveguides. An embedded 20-μm-thick cladding layer was formed by glass reflow process; due to this embedded structure, no additional thickness was required. In addition, optical loss was reduced by using the thick cladding layer and integrating a thick SU-8 layer as a core layer. The low-loss optical waveguide enables multiple-site stimulation with two-step optical splitters. Using the presented probe array, we also demonstrate a successful in-vivo optical stimulation through recording of neural signals from the hippocampus of a transgenic ChR2-YFP mouse. Recorded neural signals were synchronized with light pulses, which confirms that neurons were successfully stimulated by the blue light and the integrated electrode array successfully recorded the neural signals from activated neurons.

Discovery of 2-aryloxy-4-amino-quinazoline derivatives as novel protease-activated receptor 2 (PAR2) antagonists

Protease-activated receptor 2 (PAR2) is a member of G protein-coupled receptor and its activation initiates diverse inflammatory responses. Recent studies suggest that antagonists of PAR2 may provide a novel therapeutic strategy for inflammatory diseases. In this study, we have developed a series of 2-aryloxy-4-amino-quinazoline derivatives as PAR2 antagonists and examined their effects against LPS-induced inflammatory responses in RAW 264.7 macrophages. Among these derivatives, compound 2f displayed the greatest antagonistic activity with the IC50 value of 2.8μM. Binding modes of the newly identified PAR2 antagonists were analyzed by molecular docking using IFD/MM-GBSA methods in the putative binding site of PAR2 homology model. Moreover, 2f demonstrated significant inhibitory effects on the LPS-activated pro-inflammatory mediators including nitric oxide (NO), prostaglandin E2 (PGE2), interleukin-1β (IL-1β), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) through the regulation of various intracellular signaling pathways involving nuclear factor-κB (NF-κB), activator protein 1 (AP-1) and the mitogen-activated protein kinases (MAPK). Furthermore, administration of 2f significantly reduced the mortality of LPS-induced sepsis in mice. These results provide useful insights into the development of novel PAR2 antagonists with anti-inflammatory activity in vitro and in vivo.

Glioblastoma-secreted soluble CD44 activates tau pathology in the brain

During aggressive tumor growth and migration, glioblastoma cells secrete diverse molecules and adhesion proteins to the extracellular matrix. Yet, the biochemical effects of the glioblastoma secretome in the brain remain largely unknown. Here we show that soluble CD44 secreted from glioblastoma cells induces neuronal degeneration through the activation of tau pathology in the brain. Glioblastoma-xenograft tissues showed a number of degenerating neurons bearing highly phosphorylated tau. Through a series of secretome-analyses, we identified that soluble CD44 was the responsible protein inducing tau phosphorylation and aggregation (EC50 = 19.1 ng/mL). The treatment of sCD44 to primary hippocampal neurons-induced tau hyperphosphorylation, leading to neuronal degeneration. Also, the injection of sCD44 into the brains of tau transgenic mice induced tau hyper-phosphorylation in hippocampal neurons. Altogether, our data suggest a neurodegenerative role of sCD44 in promoting tau pathology and serving as a molecular link between glioblastoma and neurodegeneration.Brain tumors: Protein prompt for brain degenerationA protein secreted by aggressive brain tumors triggers the degeneration of neurons in surrounding brain tissues. The most aggressive brain tumors are formed by glioblastoma cells, which secrete molecules that infiltrate surrounding brain tissues, leading to loss of memory, communication and motor functions. Researchers led by Cheolju Lee and Yun Kyung Kim at the Korea Institute of Science and Technology in Seoul have shown that the CD44 protein, secreted by glioblastoma cells, is responsible for triggering this neurodegeneration. They discovered that sCD44 activates another process known as tau pathology, which is characteristic of multiple neuro-degnerative disorders such as Alzheimer’s disease. The tau protein usually stabilizes internal cellular structures, but when it is modified by abnormal activity such as the elevated levels of sCD44 found in this study, it forms insoluble masses, disrupting neuronal structure and function.

Nootropic nanocomplex with enhanced blood-brain barrier permeability for treatment of traumatic brain injury-associated neurodegeneration

ABSTRACT Traumatic brain injury (TBI) is an intracranial injury which can induce immediate neuroinflammation and long‐term neurological deficits. Methylene blue (MB) as a nootropic has a great potential to treat neurodegeneration after TBI because of its anti‐inflmmatory and neuroprotective functions. However, its limited accumulation to the brain across the blood‐brain barrier (BBB) remains a major hurdle to be overcome. In this paper, we present a polymer surfactant‐encapsulated nanocomplex of MB as a delivery system with high BBB permeability for efficacious treatment of TBI‐induced neurodegeneration. MB was formulated via electrostatically/hydrophobically directed assembly with fatty acid and Pluronic surfactant (F‐127 or F‐68) to construct nanocomplexes of two different colloidal sizes (<10 nm and ˜108 nm in hydrodynamic diameter for NanoMB‐127 and NanoMB‐68, respectively). Compared to uncomplexed free MB, formulation into the ultrasmall nanocomplex (NanoMB‐127) significantly enhanced the uptake of MB by blood‐brain vascular endothelial bEnd3 cells in vitro, and indeed improved its BBB penetration upon systemic administration to normal mice in vivo. However, large‐size NanoMB‐68 showed negligible BBB crossing despite the efficient bEnd3 cell internalization in vitro, probably due to the unfavorable pharmacokinetic profile associated with its large particle size. By virtue of the efficient BBB penetration and cellular uptake, ultrasmall NanoMB‐127 was shown to distinctively reduce the expression level of an inflammatory cytokine with no notable toxicity in vitro and also considerably prevent the neurodegeneration after TBI in mice at much lower doses than free MB. Overall, the Pluronic‐supported nanocomplexation method allows efficient brain delivery of MB, offering a novel way of enhancing the efficacy of neurotherapeutics to treat brain diseases. Graphical abstract Figure. No caption available.

Image-Based Analysis of Intracellular Tau Aggregation by Using Tau-BiFC Cell Model.

Abnormal tau aggregation is a pathological hallmark of neurodegenerative disease classified as tauopathy. Preventing tau aggregation becomes an important therapeutic strategy to cure tau-mediated neurodegeneration. Here, we describe a method to investigate intracellular tau aggregation by using a recently developed tau aggregation cell-based model named tau-BiFC. High-throughput and high-contents screening method for quantifying intracellular tau aggregation would expedite the discovery of drugs that inhibit tau aggregation.