1.Adenovirus-induced Reactive Astrogliosis Exacerbates the Pathology of Parkinson’s Disease
Heeyoung AN ; Hyowon LEE ; Seulkee YANG ; Woojin WON ; C. Justin LEE ; Min-Ho NAM
Experimental Neurobiology 2021;30(3):222-231
Parkinson’s disease (PD) is the most prevalent neurodegenerative motor disorder. While PD has been attributed to dopaminergic neuronal death in substantia nigra pars compacta (SNpc), accumulating lines of evidence have suggested that reactive astrogliosis is critically involved in PD pathology. These pathological changes are associated with α-synuclein aggregation, which is more prone to be induced by an A53T mutation. Therefore, the overexpression of A53T-mutated α-synuclein (A53T-α-syn) has been utilized as a popular animal model of PD. However, this animal model only shows marginal-to-moderate extents of reactive astrogliosis and astrocytic α-synuclein accumulation, while these phenomena are prominent in human PD brains. Here we show that Adeno-GFAP-GFP virus injection into SNpc causes severe reactive astrogliosis and exacerbates the A53Tα-syn-mediated PD pathology. In particular, we demonstrate that AAV-CMV-A53T-α-syn injection, when combined with Adeno-GFAP-GFP, causes more significant loss of dopaminergic neuronal tyrosine hydroxylase level and gain of astrocytic GFAP and GABA levels. Moreover, the combination of AAV-CMV-A53T-α-syn and Adeno-GFAP-GFP causes an extensive astrocytic α-syn expression, just as in human PD brains. These results are in marked contrast to previous reports that AAV-CMV-A53T-α-syn alone causes α-syn expression mostly in neurons but rarely in astrocytes. Furthermore, the combination causes a severe PD-like motor dysfunction as assessed by rotarod and cylinder tests within three weeks from the virus injection, whereas Adeno-GFAP-GFP alone or AAV-CMV-A53T-α-syn alone does not. Our findings implicate that inducing reactive astrogliosis exacerbates PD-like pathologies and propose the virus combination as an advanced strategy for developing a new animal model of PD.
2.Adenovirus-induced Reactive Astrogliosis Exacerbates the Pathology of Parkinson’s Disease
Heeyoung AN ; Hyowon LEE ; Seulkee YANG ; Woojin WON ; C. Justin LEE ; Min-Ho NAM
Experimental Neurobiology 2021;30(3):222-231
Parkinson’s disease (PD) is the most prevalent neurodegenerative motor disorder. While PD has been attributed to dopaminergic neuronal death in substantia nigra pars compacta (SNpc), accumulating lines of evidence have suggested that reactive astrogliosis is critically involved in PD pathology. These pathological changes are associated with α-synuclein aggregation, which is more prone to be induced by an A53T mutation. Therefore, the overexpression of A53T-mutated α-synuclein (A53T-α-syn) has been utilized as a popular animal model of PD. However, this animal model only shows marginal-to-moderate extents of reactive astrogliosis and astrocytic α-synuclein accumulation, while these phenomena are prominent in human PD brains. Here we show that Adeno-GFAP-GFP virus injection into SNpc causes severe reactive astrogliosis and exacerbates the A53Tα-syn-mediated PD pathology. In particular, we demonstrate that AAV-CMV-A53T-α-syn injection, when combined with Adeno-GFAP-GFP, causes more significant loss of dopaminergic neuronal tyrosine hydroxylase level and gain of astrocytic GFAP and GABA levels. Moreover, the combination of AAV-CMV-A53T-α-syn and Adeno-GFAP-GFP causes an extensive astrocytic α-syn expression, just as in human PD brains. These results are in marked contrast to previous reports that AAV-CMV-A53T-α-syn alone causes α-syn expression mostly in neurons but rarely in astrocytes. Furthermore, the combination causes a severe PD-like motor dysfunction as assessed by rotarod and cylinder tests within three weeks from the virus injection, whereas Adeno-GFAP-GFP alone or AAV-CMV-A53T-α-syn alone does not. Our findings implicate that inducing reactive astrogliosis exacerbates PD-like pathologies and propose the virus combination as an advanced strategy for developing a new animal model of PD.
3.The Pathological Role of Astrocytic MAOB in Parkinsonism Revealed by Genetic Ablation and Over-expression of MAOB
Heeyoung AN ; Jun Young HEO ; C. Justin LEE ; Min-Ho NAM
Experimental Neurobiology 2021;30(2):113-119
The cause of Parkinson’s disease has been traditionally believed to be the dopaminergic neuronal death in the substantia nigra pars compacta (SNpc).This traditional view has been recently challenged by the proposal that reactive astrocytes serve as key players in the pathology of Parkinson’s disease through excessive GABA release. This aberrant astrocytic GABA is synthesized by the enzymatic action of monoamine oxidase B (MAOB), whose pharmacological inhibition and gene-silencing are reported to significantly alleviate parkinsonian motor symptoms in animal models of Parkinson’s disease. However, whether genetic ablation and over-expression of MAOB can bidirectionally regulate parkinsonian motor symptoms has not been tested. Here we demonstrate that genetic ablation of MAOB blocks the MPTP-induced augmentation of astrocytic GABA-mediated tonic inhibition of neighboring dopaminergic neurons as well as parkinsonian motor symptoms, indicating the necessity of MAOB for parkinsonian motor symptoms. Furthermore, we demonstrate that GFAP-MAOB transgenic mice, in which MAOB is over-expressed under the GFAP promoter for astrocyte-specific over-expression, display exacerbated MPTP-induced tonic inhibition and parkinsonian motor symptoms compared to wild-type mice, indicating the importance of astrocytic MAOB for parkinsonian motor symptoms. Our study provides genetic pieces of evidence for the causal link between the pathological role of astrocytic MAOB-dependent tonic GABA synthesis and parkinsonian motor symptoms.
4.Differential Proximity of Perisynaptic Astrocytic Best1at the Excitatory and Inhibitory Tripartite Synapses in APP/PS1 and MAOB-KO Mice Revealed by Lattice Structured Illumination Microscopy
Heeyoung AN ; Wuhyun KOH ; SeungHee KANG ; Min-Ho NAM ; C. Justin LEE
Experimental Neurobiology 2021;30(3):213-221
Bestrophin-1 (Best1) is a GABA- and glutamate-permeable, Ca 2+ -activated Cl - channel, which is mainly expressed in astrocytes and localized at the microdomain or perisynaptic junction of the tripartite synapse. Distribution of Best1 is dramatically changed in pathological conditions such as Alzheimer’s disease. However, it is still unknown whether Best1 is located at the glutamatergic or GABAergic tripartite synapses. Here, we utilized the Lattice structured illumination microscopy (Lattice SIM) to visualize Best1 expression at the perisynaptic junctions of the tripartite synapses in CA1 of mouse hippocampus. We performed co-labeling with antibodies against 1) Best1 and vesicular glutamate transporter-2 (vGLUT2) or 2) Best1 and vesicular GABA transporter (vGAT) to measure the proximity of Best1-containing perisynapse to glutamatergic or GABAergic presynapse, respectively. In addition, we examined two transgenic mouse lines of 1) APP/PS1 mouse showing high astrocytic MAOB activity and cytosolic GABA and 2) MAOB-KO mouse showing low astrocytic GABA. Lattice SIM images were further processed by Imaris, which allowed 3Drendering and spot identification. We found that astrocytic Best1 was distributed closer to the glutamatergic synapses than GABAergic synapses in the wild-type mice. In APP/PS1 mice, Best1 distribution was significantly changed by moving away from the glutamatergic synapses while moving closer to the GABAergic synapses. On the contrary, in MAOB-KO mice, the Best1 distribution was dramatically changed by moving closer to the glutamatergic synapses and moving far away from the GABAergic synapses. Our findings propose that the proximity of Best1-containing perisynapses to presynapses dynamically changes according to the level of astrocytic cytosolic GABA.
5.The Pathological Role of Astrocytic MAOB in Parkinsonism Revealed by Genetic Ablation and Over-expression of MAOB
Heeyoung AN ; Jun Young HEO ; C. Justin LEE ; Min-Ho NAM
Experimental Neurobiology 2021;30(2):113-119
The cause of Parkinson’s disease has been traditionally believed to be the dopaminergic neuronal death in the substantia nigra pars compacta (SNpc).This traditional view has been recently challenged by the proposal that reactive astrocytes serve as key players in the pathology of Parkinson’s disease through excessive GABA release. This aberrant astrocytic GABA is synthesized by the enzymatic action of monoamine oxidase B (MAOB), whose pharmacological inhibition and gene-silencing are reported to significantly alleviate parkinsonian motor symptoms in animal models of Parkinson’s disease. However, whether genetic ablation and over-expression of MAOB can bidirectionally regulate parkinsonian motor symptoms has not been tested. Here we demonstrate that genetic ablation of MAOB blocks the MPTP-induced augmentation of astrocytic GABA-mediated tonic inhibition of neighboring dopaminergic neurons as well as parkinsonian motor symptoms, indicating the necessity of MAOB for parkinsonian motor symptoms. Furthermore, we demonstrate that GFAP-MAOB transgenic mice, in which MAOB is over-expressed under the GFAP promoter for astrocyte-specific over-expression, display exacerbated MPTP-induced tonic inhibition and parkinsonian motor symptoms compared to wild-type mice, indicating the importance of astrocytic MAOB for parkinsonian motor symptoms. Our study provides genetic pieces of evidence for the causal link between the pathological role of astrocytic MAOB-dependent tonic GABA synthesis and parkinsonian motor symptoms.
6.Differential Proximity of Perisynaptic Astrocytic Best1at the Excitatory and Inhibitory Tripartite Synapses in APP/PS1 and MAOB-KO Mice Revealed by Lattice Structured Illumination Microscopy
Heeyoung AN ; Wuhyun KOH ; SeungHee KANG ; Min-Ho NAM ; C. Justin LEE
Experimental Neurobiology 2021;30(3):213-221
Bestrophin-1 (Best1) is a GABA- and glutamate-permeable, Ca 2+ -activated Cl - channel, which is mainly expressed in astrocytes and localized at the microdomain or perisynaptic junction of the tripartite synapse. Distribution of Best1 is dramatically changed in pathological conditions such as Alzheimer’s disease. However, it is still unknown whether Best1 is located at the glutamatergic or GABAergic tripartite synapses. Here, we utilized the Lattice structured illumination microscopy (Lattice SIM) to visualize Best1 expression at the perisynaptic junctions of the tripartite synapses in CA1 of mouse hippocampus. We performed co-labeling with antibodies against 1) Best1 and vesicular glutamate transporter-2 (vGLUT2) or 2) Best1 and vesicular GABA transporter (vGAT) to measure the proximity of Best1-containing perisynapse to glutamatergic or GABAergic presynapse, respectively. In addition, we examined two transgenic mouse lines of 1) APP/PS1 mouse showing high astrocytic MAOB activity and cytosolic GABA and 2) MAOB-KO mouse showing low astrocytic GABA. Lattice SIM images were further processed by Imaris, which allowed 3Drendering and spot identification. We found that astrocytic Best1 was distributed closer to the glutamatergic synapses than GABAergic synapses in the wild-type mice. In APP/PS1 mice, Best1 distribution was significantly changed by moving away from the glutamatergic synapses while moving closer to the GABAergic synapses. On the contrary, in MAOB-KO mice, the Best1 distribution was dramatically changed by moving closer to the glutamatergic synapses and moving far away from the GABAergic synapses. Our findings propose that the proximity of Best1-containing perisynapses to presynapses dynamically changes according to the level of astrocytic cytosolic GABA.
7.The predictive value of resting heart rate in identifying undiagnosed diabetes in Korean adults: Korea National Health and Nutrition Examination Survey
Dong-Hyuk PARK ; Wonhee CHO ; Yong-Ho LEE ; Sun Ha JEE ; Justin Y. JEON
Epidemiology and Health 2022;44(1):e2022009-
OBJECTIVES:
The purpose of this study was (1) to examine whether the addition of resting heart rate (RHR) to the existing undiagnosed diabetes mellitus (UnDM) prediction model would improve predictability, and (2) to develop and validate UnDM prediction models by using only easily assessable variables such as gender, RHR, age, and waist circumference (WC).
METHODS:
Korea National Health and Nutrition Examination Survey (KNHANES) 2010, 2012, 2014, 2016 data were used to develop the model (model building set, n=19,675), while the data from 2011, 2013, 2015, 2017 were used to validate the model (validation set, n=19,917). UnDM was defined as a fasting glucose level ≥126 mg/dL or glycated hemoglobin ≥6.5%; however, doctors have not diagnosed it. Statistical package for the social sciences logistic regression analysis was used to determine the predictors of UnDM.
RESULTS:
RHR, age, and WC were associated with UnDM. When RHR was added to the existing model, sensitivity was reduced (86 vs. 73%), specificity was increased (49 vs. 65%), and a higher Youden index (35 vs. 38) was expressed. When only gender, RHR, age, and WC were used in the model, a sensitivity, specificity, and Youden index of 70%, 67%, and 37, respectively, were observed.
CONCLUSIONS
Adding RHR to the existing UnDM prediction model improved specificity and the Youden index. Furthermore, when the prediction model only used gender, RHR, age, and WC, the outcomes were not inferior to those of the existing prediction model.
8.Evaluation of the periodontal regenerative properties of patterned human periodontal ligament stem cell sheets.
Joong Hyun KIM ; Seok Yeong KO ; Justin Ho LEE ; Deok Ho KIM ; Jeong Ho YUN
Journal of Periodontal & Implant Science 2017;47(6):402-415
PURPOSE: The aim of this study was to determine the effects of patterned human periodontal ligament stem cell (hPDLSC) sheets fabricated using a thermoresponsive substratum. METHODS: In this study, we fabricated patterned hPDLSC sheets using nanotopographical cues to modulate the alignment of the cell sheet. RESULTS: The hPDLSCs showed rapid monolayer formation on various surface pattern widths. Compared to cell sheets grown on flat surfaces, there were no significant differences in cell attachment and growth on the nanopatterned substratum. However, the patterned hPDLSC sheets showed higher periodontal ligamentogenesis-related gene expression in early stages than the unpatterned cell sheets. CONCLUSIONS: This experiment confirmed that patterned cell sheets provide flexibility in designing hPDLSC sheets, and that these stem cell sheets may be candidates for application in periodontal regenerative therapy.
Cues
;
Extracellular Matrix
;
Gene Expression
;
Humans*
;
Periodontal Ligament*
;
Pliability
;
Regeneration
;
Stem Cells*
;
Tissue Engineering
9.Ca2+ Entry is Required for Mechanical Stimulation-induced ATP Release from Astrocyte.
Jaekwang LEE ; Ye Eun CHUN ; Kyung Seok HAN ; Jungmoo LEE ; Dong Ho WOO ; C Justin LEE
Experimental Neurobiology 2015;24(1):17-23
Astrocytes and neurons are inseparable partners in the brain. Neurotransmitters released from neurons activate corresponding G protein-coupled receptors (GPCR) expressed in astrocytes, resulting in release of gliotransmitters such as glutamate, D-serine, and ATP. These gliotransmitters in turn influence neuronal excitability and synaptic activities. Among these gliotransmitters, ATP regulates the level of network excitability and is critically involved in sleep homeostasis and astrocytic Ca2+ oscillations. ATP is known to be released from astrocytes by Ca2+-dependent manner. However, the precise source of Ca2+, whether it is Ca2+ entry from outside of cell or from the intracellular store, is still not clear yet. Here, we performed sniffer patch to detect ATP release from astrocyte by using various stimulation. We found that ATP was not released from astrocyte when Ca2+ was released from intracellular stores by activation of Galpha(q)-coupled GPCR including PAR1, P2YR, and B2R. More importantly, mechanical stimulation (MS)-induced ATP release from astrocyte was eliminated when external Ca2+ was omitted. Our results suggest that Ca2+ entry, but not release from intracellular Ca2+ store, is critical for MS-induced ATP release from astrocyte.
Adenosine Triphosphate*
;
Astrocytes*
;
Brain
;
Glutamic Acid
;
Homeostasis
;
Neurons
;
Neurotransmitter Agents
10.Isolation and Expression Profile of the Ca(2+)-Activated Chloride Channel-like Membrane Protein 6 Gene in Xenopus laevis.
Ra Mi LEE ; Rae Hyung RYU ; Seong Won JEONG ; Soo Jin OH ; Hue HUANG ; Jin Soo HAN ; Chi Ho LEE ; C Justin LEE ; Lily Yeh JAN ; Sang Min JEONG
Laboratory Animal Research 2011;27(2):109-116
To clone the first anion channel from Xenopus laevis (X. laevis), we isolated a calcium-activated chloride channel (CLCA)-like membrane protein 6 gene (CMP6) in X. laevis. As a first step in gene isolation, an expressed sequence tags database was screened to find the partial cDNA fragment. A putative partial cDNA sequence was obtained by comparison with rat CLCAs identified in our laboratory. First stranded cDNA was synthesized by reverse transcription polymerase-chain reaction (RT-PCR) using a specific primer designed for the target cDNA. Repeating the 5' and 3' rapid amplification of cDNA ends, full-length cDNA was constructed from the cDNA pool. The full-length CMP6 cDNA completed via 5'- and 3'-RACE was 2,940 bp long and had an open reading frame (ORF) of 940 amino acids. The predicted 940 polypeptides have four major transmembrane domains and showed about 50% identity with that of rat brain CLCAs in our previously published data. Semi-quantification analysis revealed that CMP6 was most abundantly expressed in small intestine, colon and liver. However, all tissues except small intestine, colon and liver had undetectable levels. This result became more credible after we did real-time PCR quantification for the target gene. In view of all CLCA studies focused on human or murine channels, this finding suggests a hypothetical protein as an ion channel, an X. laevis CLCA.
Amino Acids
;
Animals
;
Brain
;
Chloride Channels
;
Clone Cells
;
Colon
;
DNA, Complementary
;
Expressed Sequence Tags
;
Humans
;
Intestine, Small
;
Ion Channels
;
Liver
;
Membrane Proteins
;
Membranes
;
Open Reading Frames
;
Peptides
;
Rats
;
Real-Time Polymerase Chain Reaction
;
Resin Cements
;
Reverse Transcription
;
Staphylococcal Protein A
;
Tissue Distribution
;
Xenopus
;
Xenopus laevis