83 results on '"Hye Kyung Lee"'
Search Results
2. Enhancing early numeracy skills with a tablet-based math game intervention: a study in Tanzania
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Hye Kyung Lee and Ahram Choi
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050101 languages & linguistics ,medicine.medical_specialty ,biology ,05 social sciences ,Subtraction ,050301 education ,biology.organism_classification ,behavioral disciplines and activities ,Session (web analytics) ,Education ,Test (assessment) ,Tanzania ,Numeracy ,Intervention (counseling) ,Item response theory ,Early numeracy ,Physical therapy ,medicine ,0501 psychology and cognitive sciences ,0503 education - Abstract
The study presents results of a tablet-based math game intervention to enhance early numeracy skills of children in Tanzania. Standard level 1 children (n = 122), attending a rural primary school, were randomly allocated to either intervention or control group. The intervention group participated in a daily intervention session for 46 days. Children’s performances in number identification, quantity discrimination, addition, subtraction, and missing number tasks were measured before and after the intervention with randomly selected children from both groups (treatment = 30, control = 31). Score gains in the intervention group were substantially greater than those in the control group. In particular, statistically significant effects of the intervention were identified in quantity discrimination, addition, and subtraction tasks. Item-level analyses using Item Response Theory showed that addition and subtraction items involving regrouping and most missing number items were too difficult even after the intervention. The study also identified which games were played the most or least during the sessions from play-log data and analyzed associations between children’s test performances and gameplays.
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- 2020
3. The interdependence of mammary-specific super-enhancers and their native promoters facilitates gene activation during pregnancy
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Lothar Hennighausen, Xianke Zeng, Chaochen Wang, Precious Achikeh, Chengyu Liu, and Hye Kyung Lee
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Transcriptional Activation ,Clinical Biochemistry ,TBRG4 ,QD415-436 ,Biology ,Biochemistry ,Article ,Mice ,Pregnancy ,Animals ,natural sciences ,Promoter Regions, Genetic ,Enhancer ,Molecular Biology ,Gene ,Transcription factor ,Regulation of gene expression ,Binding Sites ,Promoter ,Chromatin ,Gene regulation ,Cell biology ,Enhancer Elements, Genetic ,Histone ,Gene Expression Regulation ,Next-generation sequencing ,biology.protein ,Chromatin Immunoprecipitation Sequencing ,Medicine ,Molecular Medicine ,Female ,Protein Binding - Abstract
Lineage-specific genetic programs rely on cell-restricted super-enhancers, which are platforms for high-density transcription factor occupation. It is not known whether super-enhancers synergize specifically with their native promoters or provide autonomous and independent regulatory platforms. Here, we investigated the ability of the mammary Wap super-enhancer to activate the promoter of the juxtaposed and ubiquitously expressed Tbrg4 gene in the mouse mammary gland. The Wap super-enhancer was fused, alone or in combination with the Wap promoter, to the Tbrg4 gene. While the super-enhancer increased the expression of the Tbrg4 promoter five-fold, the combination of the super-enhancer and promoter resulted in 80-fold gene upregulation, demonstrating lineage-specific promoter–enhancer synergy. Employing ChIP-seq profiling to determine transcription factor binding and identify activating histone marks, we uncovered a chromatin platform that enables the high-level expression of the native promoter–enhancer but not the heterologous promoter. Taken together, our data reveal that lineage-specific enhancer–promoter synergy is critical for mammary gene regulation during pregnancy and lactation., Gene regulation: linking enhancers to specific promoters Super-enhancers are regions of DNA that highly activate the expression of specific genes. Research led by H.K.L. and L.H. at the National Institutes of Health, USA, now demonstrate that a super-enhancer is much more effective when operating in synergy with its associated promoter, another gene-controlling element. They investigated super-enhancer and promoter synergy in mouse mammary gland cells. A mammary super-enhancer achieved a 5-fold activation of a gene with a common promoter, in contrast to an 80-fold activation with its own promoter together. The findings reveal that this synergy is critical for gene regulation in the mammary gland during pregnancy and lactation. They also help resolve the general issue of whether super-enhancers act independently or, as found here, can work together with a specific promoters in cell-specific genetic programs.
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- 2020
4. Robust immune response to the BNT162b mRNA vaccine in an elderly population vaccinated 15 months after recovery from COVID-19
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Hye Kyung Lee, Ludwig Knabl, Sebastian Kapferer, Birgit Pateter, Mary Walter, Priscilla A. Furth, and Lothar Hennighausen
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Messenger RNA ,education.field_of_study ,Coronavirus disease 2019 (COVID-19) ,biology ,business.industry ,Population ,Booster dose ,Peripheral blood mononuclear cell ,Article ,Vaccination ,Immune system ,Immunology ,biology.protein ,Medicine ,Antibody ,business ,education - Abstract
Knowledge about the impact of prior SARS-CoV-2 infection of the elderly on mRNA vaccination response is needed to appropriately address the need for booster vaccination in this vulnerable population. To address this, we investigated antibody and genomic immune responses in 16 elderly (avg. 81 yrs.) individuals that had received a single booster dose of BNT162b vaccine 15 months after recovering from COVID-19. Spike-specific IgG antibody levels increased in each of the study participants from an average of 710 U/ml prior to the vaccination to more than 40,000 U/ml within ten weeks after the vaccination. In contrast, anti-spike-specific IgG antibody levels averaged 2,190 U/ml in 14 healthy SARS-CoV-2-naïve individuals (avg. 58 yrs.) ten weeks after the second dose of BNT162b. RNA-seq conducted on PBMCs demonstrated the activation of interferon-activated genetic programs in both cohorts within one day. Unlike their transient induction in the younger naïve population, persistent activity and the initiation of additional cell cycle regulated programs were obtained in the older COVID-19 recovered population. Here we show that the elderly, a high-risk population, can mount a strong antibody and a persistent molecular immune response upon receiving a single dose of mRNA vaccine 15 months after recovery from COVID-19.
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- 2021
5. JAK inhibitors dampen activation of interferon-activated transcriptomes and the SARS-CoV-2 receptor ACE2 in human renal proximal tubules
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Hye Kyung Lee, Lothar Hennighausen, Julia Wilflingseder, and Jakub Jankowski
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Gene isoform ,Ruxolitinib ,Kidney ,Cell biology ,Multidisciplinary ,medicine.medical_treatment ,Science ,Acute kidney injury ,Biology ,medicine.disease ,Pathophysiology ,Article ,Cytokine ,medicine.anatomical_structure ,Interferon ,Virology ,Cancer research ,medicine ,Janus kinase ,Receptor ,medicine.drug - Abstract
SARS-CoV-2 infections initiate cytokine storms and activate genetic programs leading to progressive hyperinflammation in multiple organs of patients with COVID-19. While it is known that COVID-19 impacts kidney function, leading to increased mortality, cytokine response of renal epithelium has not been studied in detail. Here, we report on the genetic programs activated in human primary proximal tubule (HPPT) cells by interferons and their suppression by ruxolitinib, a Janus kinase (JAK) inhibitor used in COVID-19 treatment. Integration of our data with those from patients with acute kidney injury and COVID-19, as well as other tissues, permitted the identification of kidney-specific interferon responses. Additionally, we investigated the regulation of the recently discovered isoform (dACE2) of the angiotensin-converting enzyme 2 (ACE2), the SARS-CoV-2 receptor. Using ChIP-seq, we identified candidate interferon-activated enhancers controlling the ACE2 locus, including the intronic dACE2 promoter. Taken together, our study provides an in-depth understanding of genetic programs activated in kidney cells., Graphical abstract, Pathophysiology; Virology; Cell biology
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- 2021
6. Redundant and non-redundant cytokine-activated enhancers control Csn1s2b expression in the lactating mouse mammary gland
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Michaela Willi, Lothar Hennighausen, Hye Kyung Lee, Chengyu Liu, and Tyler Kuhns
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0301 basic medicine ,Science ,Transcriptional regulatory elements ,General Physics and Astronomy ,Article ,General Biochemistry, Genetics and Molecular Biology ,Mice ,03 medical and health sciences ,Mammary Glands, Animal ,Receptors, Glucocorticoid ,0302 clinical medicine ,Pregnancy ,Gene expression ,Genetics ,STAT5 Transcription Factor ,Animals ,Lactation ,Promoter Regions, Genetic ,Enhancer ,Transcription factor ,STAT5 ,Regulation of gene expression ,Multidisciplinary ,biology ,Promoter ,General Chemistry ,Chromatin ,Computational biology and bioinformatics ,Cell biology ,NFI Transcription Factors ,Enhancer Elements, Genetic ,030104 developmental biology ,Histone ,Gene Expression Regulation ,NFIB ,biology.protein ,Cytokines ,Female ,030217 neurology & neurosurgery ,Signal Transduction - Abstract
Enhancers are transcription factor platforms that synergize with promoters to control gene expression. Here, we investigate enhancers that activate gene expression several hundred-fold exclusively in the lactating mouse mammary gland. Using ChIP-seq for activating histone marks and transcription factors, we identify two candidate enhancers and one super-enhancer in the Csn1s2b locus. Through experimental mouse genetics, we dissect the lactation-specific distal enhancer bound by the mammary-enriched transcription factors STAT5 and NFIB and the glucocorticoid receptor. While deletions of canonical binding motifs for NFIB and STAT5, individually or combined, have a limited biological impact, a non-canonical STAT5 site is essential for enhancer activity during lactation. In contrast, the intronic enhancer contributes to gene expression only in late pregnancy and early lactation, possibly by interacting with the distal enhancer. A downstream super-enhancer, which physically interacts with the distal enhancer, is required for the functional establishment of the Csn1s2b promoter and gene activation. Lastly, NFIB binding in the promoter region fine-tunes Csn1s2b expression. Our study provides comprehensive insight into the anatomy and biology of regulatory elements that employ the JAK/STAT signaling pathway and preferentially activate gene expression during lactation., Enhancers and promoters work together to actively regulate gene expression affecting several biological processes. Here, the authors provide molecular insights into the regulation of enhancers and super-enhancers in the Csn1s2b locus during lactation.
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- 2021
7. Platycodon grandiflorum Extract Ameliorates Cyclophosphamide-Induced Immunosuppression in Mice
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Eun Byeol Lee, Yang-Gyu Park, Hyun-Cheol Jeong, Hae-Jeung Lee, Sung-Jin Lee, Shin-Young Park, Jihye Choi, Gyeong-A Hwang, Choi Ae Jin, Jung-Suk Choe, Hwan Hee Jang, In Guk Hwang, Ji Soo Kim, Hye Kyung Lee, and Sung Hyen Lee
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Cyclophosphamide ,medicine.medical_treatment ,medicine ,Immunosuppression ,Biology ,Pharmacology ,medicine.drug - Published
- 2019
8. Gastrodin exerts robust neuroprotection in the postischemic brain via its protective effect against Zn2+-toxicity and its anti-oxidative effects in astrocytes
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Seung Woo Kim, Hahnbie Lee, Il-Doo Kim, Hye-Kyung Lee, Ja-Kyeong Lee, and Lidan Luo
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0301 basic medicine ,Pharmacology ,Neuroprotection ,General Biochemistry, Genetics and Molecular Biology ,Gastrodin ,03 medical and health sciences ,chemistry.chemical_compound ,astrocyte ,0302 clinical medicine ,medicine ,lcsh:QH301-705.5 ,chemistry.chemical_classification ,lcsh:R5-920 ,Reactive oxygen species ,biology ,biology.organism_classification ,anti-Zn2+-toxicity ,Gastrodia elata ,030104 developmental biology ,medicine.anatomical_structure ,lcsh:Biology (General) ,chemistry ,Apoptosis ,Toxicity ,neuroprotection ,Animal Science and Zoology ,Anti oxidative ,MCAO ,lcsh:Medicine (General) ,human activities ,030217 neurology & neurosurgery ,Astrocyte - Abstract
Gastrodin (GAS) is a predominant bioactive constituent of the Chinese herbal medicine Tianma (Gastrodia elata Blume). Many authors have reported GAS has the beneficial effect on diverse diseases of the CNS, including epilepsy, Alzheimer’s disease, Parkinson’s disease, and cerebral ischemia. Here, we report GAS exhibited a robust neuroprotective effect in an Sprague-Dawley rat model of stroke (transient middle cerebral artery occlusion, tMCAO), and show that the underlying molecular mechanism involves its protective effect against Zn2+-toxicity and its anti-oxidative effects in astrocytes. Intraperitoneal administration of GAS (40 mg/kg) after MCAO reduced mean infarct volume to 30.1 ± 5.9% of that of MCAO controls and this neuroprotective effect was accompanied by neurological function recoveries which was measured using modified neurological severity score (mNSS). Interestingly, GAS induced up-regulation and nuclear translocation of Nrf2, and subsequently increased the expressions of anti-oxidative genes, such as, HO-1 and GCLM, in astrocytes. Furthermore, GAS co- or pre-treatment markedly suppressed Zn2+-induced cell death caused by excessive ROS production and PARP-1 induction. We found that GAS suppressed p67 expression and PAR formation in astrocytes, which might underlie the anti- Zn2+-toxicity and anti-oxidative effects of GAS in astrocytes. These findings indicate GAS protects astrocytes from Zn2+-induced toxicity and oxidative stress and these effects contribute to its neuroprotective effects in the postischemic brain.
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- 2018
9. Taurine Protects against Postischemic Brain Injury via the Antioxidant Activity of Taurine Chloramine
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Song-I Seol, Chaekyun Kim, Hyun Jae Kim, Ja-Kyeong Lee, Eun Bi Choi, Hye-Kyung Lee, and In Soon Kang
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0301 basic medicine ,Taurine ,Antioxidant ,Hypochlorous acid ,Physiology ,medicine.medical_treatment ,Clinical Biochemistry ,Central nervous system ,Pharmacology ,Biochemistry ,Neuroprotection ,Article ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,neutrophils ,antioxidant enzymes ,medicine ,Molecular Biology ,Heme ,chemistry.chemical_classification ,middle cerebral artery occlusion (MCAO) ,biology ,lcsh:RM1-950 ,Cell Biology ,myeloperoxidase (MPO) ,030104 developmental biology ,Enzyme ,medicine.anatomical_structure ,lcsh:Therapeutics. Pharmacology ,chemistry ,Myeloperoxidase ,taurine chloramine (Tau-Cl) ,biology.protein ,taurine ,030217 neurology & neurosurgery - Abstract
Taurine is ubiquitously distributed in mammalian tissues and is highly concentrated in the heart, brain, and leukocytes. Taurine exerts neuroprotective effects in various central nervous system diseases and can suppress infarct formation in stroke. Taurine reacts with myeloperoxidase (MPO)-derived hypochlorous acid (HOCl) to produce taurine chloramine (Tau-Cl). We investigated the neuroprotective effects of taurine using a rat middle cerebral artery occlusion (MCAO) model and BV2 microglial cells. Although intranasal administration of taurine (0.5 mg/kg) had no protective effects, the same dose of Tau-Cl significantly reduced infarct volume and ameliorated neurological deficits and promoted motor function, indicating a robust neuroprotective effect of Tau-Cl. There was neutrophil infiltration in the post-MCAO brains, and the MPO produced by infiltrating neutrophils might be involved in the taurine to Tau-Cl conversion. Tau-Cl significantly increased the levels of antioxidant enzymes glutamate–cysteine ligase, heme oxygenase-1, NADPH:quinone oxidoreductase 1, and peroxiredoxin-1 in BV2 cells, whereas taurine slightly increased some of them. Antioxidant enzyme levels were increased in the post-MCAO brains, and Tau-Cl further increased the level of MCAO-induced antioxidant enzymes. These results suggest that the neutrophils infiltrate the area of ischemic injury area, where taurine is converted to Tau-Cl, thus protecting from brain injury by scavenging toxic HOCl and increasing antioxidant enzyme expression.
- Published
- 2021
10. Interferon-regulated genetic programs and JAK/STAT pathway activate the intronic promoter of the short ACE2 isoform in renal proximal tubules
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Lothar Hennighausen, Hye Kyung Lee, Jakub Jankowski, and Julia Wilflingseder
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Gene isoform ,medicine.medical_treatment ,dACE2 ,ACE2 ,JAK-STAT signaling pathway ,interferon ,Biology ,Article ,Cell biology ,Cytokine ,Downregulation and upregulation ,Interferon ,proximal tubule ,Gene expression ,medicine ,biology.protein ,regulatory elements ,STAT1 ,Gene ,hormones, hormone substitutes, and hormone antagonists ,medicine.drug - Abstract
SummaryRecently, a short, interferon-inducible isoform of Angiotensin-Converting Enzyme 2 (ACE2), dACE2 was identified. ACE2 is a SARS-Cov-2 receptor and changes in its renal expression have been linked to several human nephropathies. These changes were never analyzed in context ofdACE2, as its expression was not investigated in the kidney. We used Human Primary Proximal Tubule (HPPT) cells to show genome-wide gene expression patterns after cytokine stimulation, with emphasis on theACE2/dACE2locus. Putative regulatory elements controllingdACE2expression were identified using ChIP-seq and RNA-seq. qRT-PCR differentiating betweenACE2anddACE2revealed 300- and 600-fold upregulation ofdACE2by IFNα and IFNβ, respectively, while full lengthACE2expression was almost unchanged. JAK inhibitor ruxolitinib ablatedSTAT1anddACE2expression after interferon treatment. Finally, with RNA-seq, we identified a set of genes, largely immune-related, induced by cytokine treatment. These gene expression profiles provide new insights into cytokine response of proximal tubule cells.
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- 2021
11. Immune Transcriptomes from Hospitalized Patients Infected with the SARS-CoV-2 Variants B.1.1.7 and B.1.1.7 Carrying the E484K Escape Mutation
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Hye Kyung Lee, Priscilla A. Furth, August Zabernigg, Lothar Hennighausen, Norbert Kaiser, Ludwig Knabl, Manuel Wieser, Knabl Sr. Ludwig, Jana Schumacher, and Anna Mur
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Adult ,Male ,Lineage (genetic) ,Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) ,medicine.disease_cause ,Peripheral blood mononuclear cell ,Article ,Transcriptome ,Young Adult ,Immune system ,Diabetes mellitus ,2',5'-Oligoadenylate Synthetase ,medicine ,Humans ,Gene Regulatory Networks ,Longitudinal Studies ,Aged ,Mutation ,biology ,SARS-CoV-2 ,business.industry ,COVID-19 ,Middle Aged ,Institutional review board ,medicine.disease ,Vaccination ,Immunology ,biology.protein ,Female ,Antibody ,business - Abstract
Fast-spreading variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) energize the COVID-19 pandemic. B.1.1.7 (VOC-202012/01) has become the predominant variant in many countries and a new lineage (VOC-202102/02) harboring the E484K escape mutation in the B.1.1.7 background emerged in February 2021. This variant is of concern due to reduced neutralizing activity by vaccine-elicited antibodies. However, it is not known whether this single amino acid change leads to an altered immune response. Here, we investigate differences in the immune transcriptome in hospitalized patients infected with either B.1.1.7 (n=28) or B.1.1.7+E484K (n=12). RNA-seq conducted on PBMCs isolated within five days after the onset of COVID symptoms demonstrated elevated activation of specific immune pathways, including JAK-STAT signaling, in B.1.1.7+E484K patients as compared to B.1.1.7. Longitudinal transcriptome studies demonstrated a delayed dampening of interferon-activated pathways in B.1.1.7+E484K patients. Prior vaccination with BNT162b vaccine (n=8 one dose; n=1 two doses) reduced the transcriptome inflammatory response to B.1.1.7+E484K infection relative to unvaccinated patients. Lastly, the immune transcriptome of patients infected with additional variants (B.1.258, B.1.1.163 and B.1.7.7) displayed a reduced activation compared to patients infected with B.1.1.7. Acquisition of the E484K substitution in the B.1.1.7 background elicits an altered immune response, which could impact disease progression. Funding: This work was supported by the Intramural Research Programs (IRPs) of National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) and utilized the computational resources of the NIH HPC Biowulf cluster (http://hpc.nih.gov). Declaration of Interest: The authors declare no competing financial interests. Ethical Approval: This study was approved by the Institutional Review Board (IRB) of the Office of Research Oversight / Regulatory Affairs, Medical University of Innsbruck, Austria (EC numbers: 1064/2021).
- Published
- 2021
12. Activation of Interferon-Stimulated Transcriptomes and ACE2 Isoforms in Human Airway Epithelium Is Curbed by Janus Kinase Inhibitors
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Lothar Hennighausen, Hye Kyung Lee, and Olive Jung
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biology ,RNA ,RNA polymerase II ,Epithelium ,Histone ,medicine.anatomical_structure ,Interferon ,biology.protein ,medicine ,Cancer research ,Janus kinase ,Enhancer ,Tropism ,medicine.drug - Abstract
SARS-CoV-2 infection of human airway epithelium activates genetic programs that lead to progressive hyperinflammation in COVID-19 patients. Here we report on genetic programs activated by interferons and the suppression by Janus kinase (JAK) inhibitors. The angiotensin-converting enzyme 2 (ACE2) is the receptor for SARS-CoV-2 and deciphering its regulation is paramount for understanding the cell tropism of SARS-CoV-2 infection. We identified candidate regulatory elements in the ACE2 locus in human primary airway cells and lung tissue. Activating histone and promoter marks and Pol II loading characterize the intronic dACE2 and define novel candidate enhancers distal to the genuine ACE2 promoter and within additional introns. dACE2, and to a lesser extent ACE2, RNA levels increased in primary cells treated with interferons and this induction was mitigated by JAK inhibitors that are used therapeutically in COVID-19 patients. Our analyses provide insight into ACE2 regulatory elements and highlight JAK inhibitors as suitable tools to suppress interferon-activated genetic programs in bronchial cells.
- Published
- 2020
13. Regulation of the ACE2 locus in human airways cells
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Hye Kyung Lee, Lothar Hennighausen, and Olive Jung
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Histone ,biology ,biology.protein ,Intron ,Respiratory epithelium ,RNA polymerase II ,Janus kinase ,Enhancer ,Gene ,hormones, hormone substitutes, and hormone antagonists ,Tropism ,Cell biology - Abstract
The angiotensin-converting enzyme 2 (ACE2) receptor is the gateway for SARS-CoV-2 to airway epithelium1,2 and the strong inflammatory response after viral infection is a hallmark in COVID-19 patients. Deciphering the regulation of the ACE2 gene is paramount for understanding the cell tropism of SARS-CoV-2 infection. Here we identify candidate regulatory elements in the ACE2 locus in human primary airway cells and lung tissue. Activating histone and promoter marks and Pol II loading characterize the intronic dACE2 and define novel candidate enhancers distal to the genuine ACE2 promoter and within additional introns. dACE2, and to a lesser extent ACE2, RNA levels increased in primary bronchial cells treated with interferons and this induction was mitigated by Janus kinase (JAK) inhibitors that are used therapeutically in COVID-19 patients. Our analyses provide insight into regulatory elements governing the ACE2 locus and highlight that JAK inhibitors are suitable tools to suppress interferon-activated genetic programs in bronchial cells.
- Published
- 2020
14. Activation of the SARS-CoV-2 Receptor Ace2 through JAK/STAT-Dependent Enhancers during Pregnancy
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Lothar Hennighausen and Hye Kyung Lee
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0301 basic medicine ,medicine.medical_treatment ,ACE2 ,RNA polymerase II ,Biology ,stat ,General Biochemistry, Genetics and Molecular Biology ,JAK-STAT pathway ,03 medical and health sciences ,0302 clinical medicine ,Report ,medicine ,cytokine ,Enhancer ,Transcription factor ,STAT5 ,Messenger RNA ,mammary alveolar epithelium ,SARS-CoV-2 ,JAK-STAT signaling pathway ,Cell biology ,030104 developmental biology ,Cytokine ,biology.protein ,enhancer ,030217 neurology & neurosurgery ,hormones, hormone substitutes, and hormone antagonists - Abstract
ACE2 binds the coronavirus SARS-CoV-2 and facilitates its cellular entry. Interferons activate ACE2 expression in pneumocytes, suggesting a critical role of cytokines in SARS-CoV-2 target cells. Viral RNA was detected in breast milk in at least seven studies, raising the possibility that ACE2 is expressed in mammary tissue during lactation. Here, we show that Ace2 expression in mouse mammary tissue is induced during pregnancy and lactation, which coincides with the activation of intronic enhancers. These enhancers are occupied by the prolactin-activated transcription factor STAT5 and additional regulatory factors, including RNA polymerase II. Deletion of Stat5a results in decommissioning of the enhancers and an 83% reduction of Ace2 mRNA. We also demonstrate that Ace2 expression increases during lactation in lung, but not in kidney and intestine. JAK/STAT components are present in a range of SARS-CoV-2 target cells, opening the possibility that cytokines contribute to the viral load and extrapulmonary pathophysiology., Graphical Abstract, Highlights • Ace2 expression is induced in the mammary glands of pregnant and lactating mice • Gene enhancers are activated by the prolactin-activated transcription factors STAT5A/B • Deletion of the Stat5a gene mitigates enhancer formation and Ace2 expression • Ace2 levels also increase in lung tissue during lactation, Hennighausen and Lee find that increased expression of the SARS-CoV-2 receptor Ace2 in mammary tissue of mice during pregnancy and lactation is controlled through the hormone-activated JAK/STAT5 signaling pathway. Their findings suggest the possibility of vertical transmission of SARS-CoV-2 through breast milk and non-pulmonary tissue damage.
- Published
- 2020
15. The transcription factor STAT5 binds to distinct super-enhancer sites and controls Lrrc32 expression in a prominent autoimmune and allergic disease risk locus
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Hye Kyung Lee and Lothar Hennighausen
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Super-enhancer ,biology ,Regulatory sequence ,biology.protein ,Locus (genetics) ,Enhancer ,Gene ,Transcription factor ,STAT5 ,Cell biology ,Chromatin - Abstract
SummaryGenetic variants associated with diseases are enriched in genomic sequences linked to regulatory regions, such as enhancers, super-enhancers and possibly repressors, that control nearby and distant genes. A known allergic and autoimmune risk locus at chromosome 11q13.51,2 is associated with the LRRC32 gene, which encodes GARP, a protein critical for TGF-β delivery3. This region coincides with a candidate enhancer that was predicted by the presence of activating chromatin marks and contains a polymorphism significantly associated with GARP expression on CD4+CD127-CD25+ Treg cells4. In the mouse, binding of the cytokine-induced transcription factor STAT5 was detected at two sites within the expansive candidate enhancer region and a 2.3 kb deletion resulted in reduced Lrrc32 expression4. However, a clear definition of the enhancer units controlled by STAT5 and a functional understanding of STAT5 in the regulation of Lrrc32 are needed. Here we use high-resolution ChIP-seq and identify three STAT5 binding sites within the Lrrc32 super-enhancer, one shared between Treg cells and mammary epithelium and one specific to each respective cell type. Using mice that express only 10% of normal STAT5 levels we demonstrate the defining contribution of STAT5 in the activation of the Lrrc32 super-enhancer.
- Published
- 2020
16. Regulatory plasticity within a complex cytokine-sensing mammary enhancer during lactation
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Lothar Hennighausen, Chengyu Liu, and Hye Kyung Lee
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DNA binding site ,Transcription (biology) ,Gene expression ,biology.protein ,Promoter ,Biology ,Enhancer ,Gene ,Transcription factor ,STAT5 ,Cell biology - Abstract
Enhancers are transcription factor platforms that synergize with promoters to activate gene expression up to several-thousand-fold. While genome-wide structural studies are used to predict enhancers, the in vivo significance is less clear. Specifically, the biological importance of individual transcription factors within enhancer complexes remains to be understood. Here we investigate the structural and biological importance of individual transcription factor binding sites and redundancy among transcription components within a complex enhancer in vivo. The Csn1s2b gene is expressed exclusively in mammary tissue and activated several thousand-fold during pregnancy and lactation. Using ChIP-seq we identified a complex lactation-specific candidate enhancer that binds multiple transcription factors and coincides with activating histone marks. Using experimental mouse genetics, we determined that deletion of canonical binding motifs for the transcription factors NFIB and STAT5, individually and combined, had a limited biological impact. Loss of these sites led to a shift of transcription factor binding to juxtaposed sites, suggesting exceptional plasticity that does not require direct protein-DNA interactions. Additional deletions revealed the critical importance of a non-canonical STAT5 binding site for enhancer activity. Our data also suggest that enhancer RNAs are not required for the activity of this specific enhancer. While ChIP-seq experiments predicted an additional candidate intronic enhancer, its deletion did not adversely affect gene expression, emphasizing the limited biological information provided by structural data. Our study provides comprehensive insight into the anatomy and biology of a composite mammary enhancer that activates its target gene several hundred-fold during lactation.
- Published
- 2020
17. Activation of the SARS-CoV-2 receptorAce2by cytokines through pan JAK-STAT enhancers
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Lothar Hennighausen and Hye Kyung Lee
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medicine.medical_treatment ,Peptidyl-Dipeptidase A ,Kidney ,Article ,Alveolar cells ,Mice ,Pregnancy ,Transcription (biology) ,STAT5 Transcription Factor ,medicine ,Animals ,Humans ,Lactation ,STAT1 ,Intestinal Mucosa ,Mammary Glands, Human ,Enhancer ,Lung ,Transcription factor ,STAT5 ,Janus Kinases ,biology ,Chemistry ,JAK-STAT signaling pathway ,Cell biology ,Mice, Inbred C57BL ,Enhancer Elements, Genetic ,Cytokine ,medicine.anatomical_structure ,biology.protein ,Female ,Angiotensin-Converting Enzyme 2 ,hormones, hormone substitutes, and hormone antagonists ,Signal Transduction - Abstract
ACE2, in concert with the protease TMPRSS2, binds the novel coronavirus SARS-CoV-2 and facilitates its cellular entry. The ACE2 gene is expressed in SARS-CoV-2 target cells, including Type II Pneumocytes (Ziegler, 2020), and is activated by interferons. Viral RNA was also detected in breast milk (Wu et al., 2020), raising the possibility that ACE2 expression is under the control of cytokines through the JAK-STAT pathway. Here we show that Ace2 expression in mammary tissue is induced during pregnancy and lactation, which coincides with the establishment of a candidate enhancer. The prolactin-activated transcription factor STAT5 binds to tandem sites that coincide with activating histone enhancer marks and additional transcription components. The presence of pan JAK-STAT components in mammary alveolar cells and in Type II Pneumocytes combined with the autoregulation of both STAT1 and STAT5 suggests a prominent role of cytokine signaling pathways in cells targeted by SARS-CoV-2. Funding: This work was supported by the Intramural Research Program (IRP) of the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)and utilized the computational resources of the NIH HPC Biowulf cluster (http://hpc.nih.gov) Declaration of Interest: The authors declare not competing interests.
- Published
- 2020
18. Cytosine base editor 4 but not adenine base editor generates off-target mutations in mouse embryos
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Lothar Hennighausen, Hye Kyung Lee, Harold E. Smith, Michaela Willi, and Chengyu Liu
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Male ,Medicine (miscellaneous) ,Computational biology ,Biology ,Genome ,Article ,General Biochemistry, Genetics and Molecular Biology ,Cytosine ,Mice ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Animals ,Point Mutation ,Sequencing ,lcsh:QH301-705.5 ,030304 developmental biology ,Subgenomic mRNA ,Gene Editing ,0303 health sciences ,Base Sequence ,Adenine ,Point mutation ,Embryo ,Embryo, Mammalian ,Base (topology) ,lcsh:Biology (General) ,chemistry ,Gene Targeting ,Mutation ,Next-generation sequencing ,Female ,RNA Editing ,Mammalian genome ,CRISPR-Cas Systems ,General Agricultural and Biological Sciences ,030217 neurology & neurosurgery ,RNA, Guide, Kinetoplastida - Abstract
Deaminase base editing has emerged as a tool to install or correct point mutations in the genomes of living cells in a wide range of organisms. However, the genome-wide off-target effects introduced by base editors in the mammalian genome have been examined in only one study. Here, we have investigated the fidelity of cytosine base editor 4 (BE4) and adenine base editors (ABE) in mouse embryos using unbiased whole-genome sequencing of a family-based trio cohort. The same sgRNA was used for BE4 and ABE. We demonstrate that BE4-edited mice carry an excess of single-nucleotide variants and deletions compared to ABE-edited mice and controls. Therefore, an optimization of cytosine base editors is required to improve its fidelity. While the remarkable fidelity of ABE has implications for a wide range of applications, the occurrence of rare aberrant C-to-T conversions at specific target sites needs to be addressed., Hye Kyung Lee, Harold E. Smith et al. examined the fidelity of cytosine base editor 4 (BE4) and adenine base editors (ABEs) in mouse embryos using whole-genome sequencing of a family-based trio cohort. They show that BE4-edited mice carry more single-nucleotide variants and deletions than ABE-edited mice.
- Published
- 2020
19. Activation of the SARS-CoV-2 Receptor Ace2 by Cytokines Through Pan JAK-STAT Enhancers
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Hye Kyung Lee and Lothar Hennighausen
- Subjects
biology ,medicine.medical_treatment ,JAK-STAT signaling pathway ,Cell biology ,Alveolar cells ,medicine.anatomical_structure ,Cytokine ,Transcription (biology) ,biology.protein ,medicine ,STAT1 ,Enhancer ,Transcription factor ,STAT5 - Abstract
ACE2, in concert with the protease TMPRSS2, binds the novel coronavirus SARS-CoV-2 and facilitates its cellular entry. The ACE2 gene is expressed in SARS-CoV-2 target cells, including Type II Pneumocytes (Ziegler, 2020), and is activated by interferons. Viral RNA was also detected in breast milk (Wu et al., 2020), raising the possibility that ACE2 expression is under the control of cytokines through the JAK-STAT pathway. Here we show that Ace2 expression in mammary tissue is induced during pregnancy and lactation, which coincides with the establishment of a candidate enhancer. The prolactin-activated transcription factor STAT5 binds to tandem sites that coincide with activating histone enhancer marks and additional transcription components. The presence of pan JAK-STAT components in mammary alveolar cells and in Type II Pneumocytes combined with the autoregulation of both STAT1 and STAT5 suggests a prominent role of cytokine signaling pathways in cells targeted by SARS-CoV-2. Funding: This work was supported by the Intramural Research Program (IRP) of the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)and utilized the computational resources of the NIH HPC Biowulf cluster (http://hpc.nih.gov) Declaration of Interest: The authors declare not competing interests.
- Published
- 2020
20. Azole Resistance Caused by Increased Drug Efflux in Candida glabrata Isolated from the Urinary Tract of a Dog with Diabetes Mellitus
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Won Hee Jung, Inhyung Lee, Sun Young Hwang, Hye Kyung Lee, and Min-Chul Kim
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0301 basic medicine ,Drug ,media_common.quotation_subject ,Urinary system ,Candida glabrata ,Drug resistance ,Microbiology ,03 medical and health sciences ,Diabetes mellitus ,medicine ,Dog ,Gene ,media_common ,Urinary tract ,biology ,Diabetes ,biology.organism_classification ,medicine.disease ,Ketoacidosis ,Research Note ,030104 developmental biology ,Infectious Diseases ,Azole resistance ,Drug efflux ,Efflux - Abstract
A yeast-like organism was isolated from a urine sample of a 6-year-old neutered male miniature poodle dog with urinary tract infection, diabetes ketoacidosis, and acute pancreatitis. We identified the yeast-like organism to be Candida glabrata and found that this fungus was highly resistant to azole antifungal drugs. To understand the mechanism of azole resistance in this isolate, the sequences and expression levels of the genes involved in drug resistance were analyzed. The results of our analysis showed that increased drug efflux, mediated by overexpression of ATP transporter genes CDR1 and PDH1, is the main cause of azole resistance of the C. glabrata isolated here.
- Published
- 2017
21. Neuroprotective and Anti-inflammatory Effects of a Dodecamer Peptide Harboring Ninjurin 1 Cell Adhesion Motif in the Postischemic Brain
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Seung Woo Kim, Hahnbie Lee, Lidan Luo, Il-Doo Kim, Ja-Kyeong Lee, and Hye-Kyung Lee
- Subjects
Male ,rac1 GTP-Binding Protein ,0301 basic medicine ,Small interfering RNA ,Neutrophils ,Cell Adhesion Molecules, Neuronal ,Amino Acid Motifs ,Anti-Inflammatory Agents ,Neuroscience (miscellaneous) ,HL-60 Cells ,Cell Communication ,Motor Activity ,Biology ,Brain Ischemia ,Proinflammatory cytokine ,Rats, Sprague-Dawley ,Brain ischemia ,Phosphatidylinositol 3-Kinases ,03 medical and health sciences ,Cellular and Molecular Neuroscience ,0302 clinical medicine ,Immune system ,Cell Movement ,Cell Adhesion ,Human Umbilical Vein Endothelial Cells ,medicine ,Animals ,Humans ,Amino Acid Sequence ,Nerve Growth Factors ,Cell adhesion ,Inflammation ,Innate immune system ,Tumor Necrosis Factor-alpha ,Cell adhesion molecule ,Brain ,Infarction, Middle Cerebral Artery ,Recovery of Function ,medicine.disease ,Molecular biology ,Cell biology ,Disease Models, Animal ,Neuroprotective Agents ,030104 developmental biology ,Neurology ,Gene Knockdown Techniques ,Peptides ,Infiltration (medical) ,030217 neurology & neurosurgery - Abstract
It has been reported that the innate immune response plays important roles in brain ischemia and that the infiltration of blood-derived immune cells is a key initiator of this response. Nerve injury-induced protein 1 (Ninjurin 1, Ninj1) is a cell adhesion molecule responsible for cell-to-cell interactions between immune cells and endothelial cells. In the present study, we investigated the proinflammatory and neuroprotective effects of Ninj1 and a dodecamer peptide harboring Ninj1 N-terminal adhesion motif (N-NAM, Pro26~Asn37) in a rat middle cerebral artery occlusion (MCAO) model of stroke. Ninj1 was predominantly induced in neutrophils and endothelial cells in the ischemic hemispheres around 12 h to 1 day post-MCAO, which coincided with a massive neutrophil influx. We demonstrated that intranasal administration of Ninj1 small interfering RNA (siRNA) or N-NAM significantly blocked neutrophil infiltration in postischemic brains. In addition, intranasal administration of Ninj1 siRNA or N-NAM reduced the mean infarct volume to 46.5 ± 9.2 or 30.6 ± 11.7% of that of the PBS-treated MCAO controls, respectively, which was accompanied by significant amelioration of neurological and motor deficits. We showed that N-NAM or Ninj1 siRNA effectively blocked the adhesion and transendothelial migration of TNF-α-stimulated human myelocytic leukemia cells to human umbilical vein endothelial cells and similarly suppressed adhesion and migration of monocytes. Activations of phosphoinositide 3-kinase and Ras-related C3 botulinum toxin substrate 1 are involved in these Ninj1-mediated processes and can be inhibited by N-NAM or Ninj1 siRNA. These results indicate that Ninj1 plays an important role in neutrophil infiltration in the postischemic brain and N-NAM confers robust neuroprotective and anti-inflammatory effects by inhibiting Ninj1-mediated infiltration of neutrophils.
- Published
- 2017
22. Copine-7 binds to the cell surface receptor, nucleolin, and regulates ciliogenesis and Dspp expression during odontoblast differentiation
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Joo-Cheol Park, Hye Kyung Lee, Su-Jin Park, and You-Mi Seo
- Subjects
0301 basic medicine ,Sialoglycoproteins ,Cellular differentiation ,media_common.quotation_subject ,Odontoblast differentiation ,lcsh:Medicine ,Biology ,Article ,03 medical and health sciences ,stomatognathic system ,Ciliogenesis ,Humans ,KIF3A ,Cilia ,Internalization ,lcsh:Science ,media_common ,Regulation of gene expression ,Extracellular Matrix Proteins ,Multidisciplinary ,Odontoblasts ,lcsh:R ,Membrane Proteins ,RNA-Binding Proteins ,Cell Differentiation ,Phosphoproteins ,Molecular biology ,Endocytosis ,Cell biology ,Protein Transport ,stomatognathic diseases ,030104 developmental biology ,Odontoblast ,Gene Expression Regulation ,Calcium ,lcsh:Q ,Nucleolin - Abstract
Tooth development is a progressive process regulated by interactions between epithelial and mesenchymal tissues. Our previous studies showed that copine-7 (Cpne7), a dental epithelium-derived protein, is a signalling molecule that is secreted by preameloblasts and regulates the differentiation of preodontoblasts into odontoblasts. However, the mechanisms involved in the translocation of Cpne7 from preameloblasts to preodontoblasts and the functions of Cpne7 during odontogenesis are poorly understood. Here, we showed that the internalization of Cpne7 was mediated primarily by caveolae. This process was initiated by Cpne7 binding to the cell surface protein, nucleolin. Treatment with recombinant Cpne7 protein (rCpne7) in human dental pulp cells (hDPCs) caused an increase in the number of ciliated cells. The expression level of cilium components, Ift88 and Kif3a, and Dspp were increased by rCpne7. Treatment with Ift88 siRNA in hDPCs and MDPC-23 cells significantly down-regulated the expression of Dspp, an odontoblastic differentiation marker gene. Furthermore, the treatment with nucleolin siRNA in MDPC-23 cells decreased the expression of Dmp1, Dspp, and cilium components. Our findings suggested that the binding of Cpne7 with its receptor, nucleolin, has an important function involving Cpne7 internalization into preodontoblasts and regulation of Dspp expression through ciliogenesis during odontoblast differentiation.
- Published
- 2017
23. Functional assessment of CTCF sites at cytokine-sensing mammary enhancers using CRISPR/Cas9 gene editing in mice
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Lothar Hennighausen, Michaela Willi, Harold E. Smith, Chul Min Yang, Hye Kyung Lee, Chengyu Liu, and Chaochen Wang
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0301 basic medicine ,CCCTC-Binding Factor ,Enhancer RNAs ,Locus (genetics) ,Biology ,Genome ,Mice ,03 medical and health sciences ,Mammary Glands, Animal ,Genetics ,Animals ,Promoter Regions, Genetic ,Enhancer ,Gene ,Gene Editing ,Zinc finger ,CRISPR interference ,Binding Sites ,Caseins ,Genomics ,Repressor Proteins ,Enhancer Elements, Genetic ,030104 developmental biology ,Gene Expression Regulation ,Genetic Loci ,CTCF ,Cytokines ,Female ,CRISPR-Cas Systems ,Sulfotransferases ,Protein Binding - Abstract
The zinc finger protein CTCF has been invoked in establishing boundaries between genes, thereby controlling spatial and temporal enhancer activities. However, there is limited genetic evidence to support the concept that these boundaries restrict the search space of enhancers. We have addressed this question in the casein locus containing five mammary and two non-mammary genes under the control of at least seven putative enhancers. We have identified two CTCF binding sites flanking the locus and two associated with a super-enhancer. Individual deletion of these sites from the mouse genome did not alter expression of any of the genes. However, deletion of the border CTCF site separating the Csn1s1 mammary enhancer from neighboring genes resulted in the activation of Sult1d1 at a distance of more than 95 kb but not the more proximal and silent Sult1e1 gene. Loss of this CTCF site led to de novo interactions between the Sult1d1 promoter and several enhancers in the casein locus. Our study demonstrates that only one out of the four CTCF sites in the casein locus had a measurable in vivo activity. Studies on additional loci are needed to determine the biological role of CTCF sites associated with enhancers.
- Published
- 2017
24. Cytosine but not adenine base editor generates mutations in mice
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Harold E. Smith, Lothar Hennighausen, Chengyu Liu, Hye Kyung Lee, and Michaela Willi
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Whole genome sequencing ,Genetics ,chemistry.chemical_compound ,chemistry ,Point mutation ,Biology ,Base (topology) ,Genome ,Cytosine - Abstract
Deaminase base editing has emerged as a tool to install or correct point mutations in the genomes of living cells in a wide range of organisms and its ultimate success therapeutically depends on its accuracy. Here we have investigated the fidelity of cytosine base editor 4 (BE4) and adenine base editor (ABE) in mouse embryos using unbiased whole genome sequencing of a family-based trio cohort. We demonstrate that BE4-edited mice carry an excess of single-nucleotide variants and deletions compared to ABE-edited mice and controls.
- Published
- 2019
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25. Neutrophil extracellular trap induced by HMGB1 exacerbates damages in the ischemic brain
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Hahnbie Lee, Ja-Kyeong Lee, Hye-Kyung Lee, Il-Doo Kim, and Seung Woo Kim
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0301 basic medicine ,Programmed cell death ,Inflammation ,HMGB1 ,CXCR4 ,lcsh:RC346-429 ,Pathology and Forensic Medicine ,03 medical and health sciences ,Cellular and Molecular Neuroscience ,0302 clinical medicine ,Parenchyma ,medicine ,Receptor ,lcsh:Neurology. Diseases of the nervous system ,biology ,Chemistry ,Research ,NETosis ,Neutrophil extracellular traps ,Cell biology ,030104 developmental biology ,Permanent ischemia ,TLR4 ,biology.protein ,Neurology (clinical) ,MCAO ,medicine.symptom ,030217 neurology & neurosurgery - Abstract
It has been reported that neutrophil extracellular traps (NETs) play important roles in non-infectious diseases. In ischemic stroke, neutrophils infiltrate damaged brain tissue soon after injury and aggravate inflammation. Using a rat permanent MCAO model, we showed citrullinated histone H3+ (CitH3, a marker of NETosis) induction in neutrophils in leptomeninges and in peripheral blood soon after MCAO. Entry of CitH3+ cells occurred through leptomeninges after 6 h of MCAO and these cells were observed in cerebral cortex from 12 h and subsequently in striatum. It is interesting to note that CitH3+ induction began in circulating neutrophils before they migrated to brain parenchyma and they were detected as intact or lysed form. High mobility group box 1 (HMGB1), a danger associated molecular pattern (DAMP) molecule, was accumulated massively in serum after permanent MCAO and plays a critical role in CitH3 inductions in neutrophils in brain parenchyma and in peripheral blood. Both the all-thiol and disulfide types of HMGB1 induced CitH3 via their specific receptors, CXCR4 and TLR4, respectively. Importantly, HMGB1 not only induced NETosis but was included as a part of the extruded NETs, and contribute to NETosis-mediated neuronal death. Therefore, it would appear a vicious cycle exists between neuronal cell death and NETosis and HMGB1 mediates detrimental effects exerted by this cycle. When NETosis was suppressed by a PAD inhibitor in MCAO animals, delayed immune cell infiltrations were markedly suppressed and damages in blood vessels were significantly mitigated. The study shows NETosis with the involvement of HMGB1 as a mediator in a vicious cycle aggravates inflammation and subsequent damage in the ischemic brain. Electronic supplementary material The online version of this article (10.1186/s40478-019-0747-x) contains supplementary material, which is available to authorized users.
- Published
- 2019
26. Simultaneous targeting of linked loci in mouse embryos using base editing
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Michaela Willi, David R. Liu, Shannon M. Miller, Hye Kyung Lee, Chengyu Liu, Lothar Hennighausen, and Harold E. Smith
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0301 basic medicine ,Male ,Zygote ,lcsh:Medicine ,Sequence Homology ,Locus (genetics) ,Biology ,Genome ,Receptor Activity-Modifying Protein 3 ,Article ,03 medical and health sciences ,Mice ,0302 clinical medicine ,Genome editing ,Cytidine Deaminase ,CRISPR ,Animals ,DNA Breaks, Double-Stranded ,lcsh:Science ,Enhancer ,Gene ,Genetics ,Gene Editing ,Multidisciplinary ,Base Sequence ,Cas9 ,lcsh:R ,Embryo, Mammalian ,Milk Proteins ,DNA binding site ,Mice, Inbred C57BL ,030104 developmental biology ,Genetic Loci ,Mutation ,lcsh:Q ,Female ,CRISPR-Cas Systems ,030217 neurology & neurosurgery - Abstract
A particular challenge in genome engineering has been the simultaneous introduction of mutations into linked (located on the same chromosome) loci. Although CRISPR/Cas9 has been widely used to mutate individual sites, its application in simultaneously targeting of linked loci is limited as multiple nearby double-stranded DNA breaks created by Cas9 routinely result in the deletion of sequences between the cleavage sites. Base editing is a newer form of genome editing that directly converts C∙G-to-T∙A, or A∙T-to-G∙C, base pairs without introducing double-stranded breaks, thus opening the possibility to generate linked mutations without disrupting the entire locus. Through the co-injection of two base editors and two sgRNAs into mouse zygotes, we introduced C∙G-to-T∙A transitions into two cytokine-sensing transcription factor binding sites separated by 9 kb. We determined that one enhancer activates the two flanking genes in mammary tissue during pregnancy and lactation. The ability to introduce linked mutations simultaneously in one step into the mammalian germline has implications for a wide range of applications, including the functional analysis of linked cis-elements creating disease models and correcting pathogenic mutations.
- Published
- 2019
27. Targeting fidelity of adenine and cytosine base editors in mouse embryos
- Author
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Lothar Hennighausen, Sojung Kim, Hye Kyung Lee, Chengyu Liu, Michaela Willi, Shannon M. Miller, and David R. Liu
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0301 basic medicine ,Microinjections ,Zygote ,Base pair ,Science ,media_common.quotation_subject ,General Physics and Astronomy ,Fidelity ,Computational biology ,Biology ,Sensitivity and Specificity ,Genome ,Article ,General Biochemistry, Genetics and Molecular Biology ,Animals, Genetically Modified ,Cytosine ,Mice ,03 medical and health sciences ,chemistry.chemical_compound ,CRISPR-Associated Protein 9 ,Animals ,Point Mutation ,lcsh:Science ,Alleles ,media_common ,Gene Editing ,Multidisciplinary ,Adenine ,Point mutation ,Reproducibility of Results ,RNA ,Embryo ,General Chemistry ,Embryo, Mammalian ,Base (topology) ,030104 developmental biology ,chemistry ,Genetic Loci ,lcsh:Q ,CRISPR-Cas Systems ,RNA, Guide, Kinetoplastida - Abstract
Base editing directly converts a target base pair into a different base pair in the genome of living cells without introducing double-stranded DNA breaks. While cytosine base editors (CBE) and adenine base editors (ABE) are used to install and correct point mutations in a wide range of organisms, the extent and distribution of off-target edits in mammalian embryos have not been studied in detail. We analyze on-target and proximal off-target editing at 13 loci by a variety of CBEs and ABE in more than 430 alleles generated from mouse zygotic injections using newly generated and published sequencing data. ABE predominantly generates anticipated A•T-to-G•C edits. Among CBEs, SaBE3 and BE4, result in the highest frequencies of anticipated C•G-to-T•A products relative to editing byproducts. Together, these findings highlight the remarkable fidelity of ABE in mouse embryos and identify preferred CBE variants when fidelity in vivo is critical., Understanding the risks of bystander and off-target editing is essential for genome engineering applications. Here, the authors analyze the fidelity of adenine and cytosine base editors in vivo.
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- 2018
28. Progressing super-enhancer landscape during mammary differentiation controls tissue-specific gene regulation
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Ha Youn Shin, Lothar Hennighausen, Hye Kyung Lee, Chengyu Liu, and Michaela Willi
- Subjects
0301 basic medicine ,Chromatin Immunoprecipitation ,Transgene ,Mice, Transgenic ,Regulatory Sequences, Nucleic Acid ,Biology ,Mice ,03 medical and health sciences ,Mammary Glands, Animal ,Super-enhancer ,Pregnancy ,Lactation ,STAT5 Transcription Factor ,Genetics ,medicine ,Animals ,Enhancer ,Transcription factor ,Regulation of gene expression ,Gene Expression Regulation, Developmental ,Cell Differentiation ,Genomics ,Chromatin ,Cell biology ,Mice, Inbred C57BL ,030104 developmental biology ,medicine.anatomical_structure ,Organ Specificity ,Female ,Chromatin immunoprecipitation - Abstract
The mammary luminal lineage relies on the common cytokine-sensing transcription factor STAT5 to establish super-enhancers during pregnancy and initiate a genetic program that activates milk production. As pups grow, the greatly increasing demand for milk requires progressive differentiation of mammary cells with advancing lactation. Here we investigate how persistent hormonal exposure during lactation shapes an evolving enhancer landscape and impacts the biology of mammary cells. Employing ChIP-seq, we uncover a changing transcription factor occupancy at mammary enhancers, suggesting that their activities evolve with advancing differentiation. Using mouse genetics, we demonstrate that the functions of individual enhancers within the Wap super-enhancer evolve as lactation progresses. Most profoundly, a seed enhancer, which is mandatory for the activation of the Wap super-enhancer during pregnancy, is not required during lactation, suggesting compensatory flexibility. Combinatorial deletions of structurally equivalent constituent enhancers demonstrated differentiation-specific compensatory activities during lactation. We also demonstrate that the Wap super-enhancer, which is built on STAT5 and other common transcription factors, retains its exquisite mammary specificity when placed into globally permissive chromatin, suggesting a limited role of chromatin in controlling cell specificity. Our studies unveil a previously unrecognized progressive enhancer landscape where structurally equivalent components serve unique and differentiation-specific functions.
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- 2018
29. Upregulation of Nrf2–p300 mediates anti-inflammatory effects of curcumin in microglia by downregulating p65–p300
- Author
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Hye-Kyung Lee, Lidan Luo, Hahnbie Lee, Ja-Kyeong Lee, Il-Doo Kim, and Seung Woo Kim
- Subjects
0301 basic medicine ,Microglia ,Biology ,General Biochemistry, Genetics and Molecular Biology ,Cell biology ,Ferritin light chain ,Heme oxygenase ,Nitric oxide synthase ,03 medical and health sciences ,chemistry.chemical_compound ,030104 developmental biology ,medicine.anatomical_structure ,Biochemistry ,Downregulation and upregulation ,chemistry ,Curcumin ,medicine ,biology.protein ,Macrophage ,Animal Science and Zoology ,NAD+ kinase - Abstract
Curcumin (CUR) is a hydrophobic polyphenol derived from the rhizome of Curcuma longa. CUR confers protection in various pathological conditions, including many brain-related diseases, such as cerebral ischemia, intracerebral hemorrhage, or Alzheimer’s disease, and these effects have been attributed to its anti-inflammatory and anti-oxidative properties. In the present study, we found CUR induced the nuclear translocation of nuclear factor E2-related factor 2 (Nrf2) in microglia, brain macrophage, and thus upregulated genes downstream of antioxidant response element, such as heme oxygenase 1, NAD(P)H:quinone oxidoreductase 1, glutamate-cysteine ligase modifier subunit, and ferritin light chain, and simultaneously downregulated lipopolysaccharide-induced inducible nitric oxide synthase expression. We showed that the anti-inflammatory effect of CUR in microglia is connected with its anti-oxidative effect in that CUR promotes Nrf2–p300 binding at the expense of p65–p300 binding. Since CUR is a dietary...
- Published
- 2016
30. Neuroprotective effect of ethyl pyruvate against Zn 2+ toxicity via NAD replenishment and direct Zn 2+ chelation
- Author
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Ja-Kyeong Lee, Hye-Kyung Lee, Seung Woo Kim, Sung-Hwa Yoon, and Hyun-Ji Kim
- Subjects
0301 basic medicine ,Programmed cell death ,N-Methylaspartate ,Primary Cell Culture ,Poly (ADP-Ribose) Polymerase-1 ,Neuroprotection ,Rats, Sprague-Dawley ,03 medical and health sciences ,Cellular and Molecular Neuroscience ,chemistry.chemical_compound ,0302 clinical medicine ,medicine ,Animals ,Hypoxia ,Pyruvates ,Chelating Agents ,Neurons ,Pharmacology ,chemistry.chemical_classification ,Reactive oxygen species ,NADPH oxidase ,Cell Death ,biology ,Neurotoxicity ,NAD ,medicine.disease ,Molecular biology ,Rats ,Zinc ,Glucose ,Neuroprotective Agents ,030104 developmental biology ,Biochemistry ,chemistry ,biology.protein ,NAD+ kinase ,Pyruvic acid ,Reactive Oxygen Species ,030217 neurology & neurosurgery ,Intracellular - Abstract
Ethyl pyruvate (EP) is a simple aliphatic ester of pyruvic acid and has been shown to have robust protective effect in various pathological conditions. A variety of mechanisms have been reported to underlie the protective effects of EP, which include anti-inflammatory, anti-oxidative, and anti-apoptotic functions. Recently, we reported that EP suppressed high mobility group box 1 (HMGB1) release from primary microglial cells via direct Ca(2+) chelation. In the present study, we investigated whether and how EP chelates Zn(2+) in neurons when it is present at toxic levels. In cortical neurons treated with 40 μM of Zn(2+) for 24 h, both EP and pyruvate significantly suppressed neuronal cell death, although the potency of pyruvate was greater than that of EP, and that NAD replenishment contributed to the neuroprotective effects of both pyruvate and EP. However, when cortical neurons were exposed to acute treatment of Zn(2+) (400 μM, 15 min), EP, but not pyruvate, significantly suppressed neuronal death, despite the fact that NAD replenishment by EP was weaker than that by pyruvate. Spectrophotometric studies revealed that EP directly chelates Zn(2+), and this was confirmed in physiological contexts, such as, NMDA-treated primary cortical cultures and OGD-subjected hippocampal slice cultures, in which EP suppressed intracellular Zn(2+) elevation and neuronal cell death. In addition, EP markedly reduced the expressions of PARP-1 and of the NADPH oxidase subunit in Zn(2+)-treated primary cortical neurons, well known Zn(2+)-induced downstream processes. Together, these results show EP suppresses Zn(2+) induced neurotoxicity via dual functions, chelating Zn(2+) and promoting NAD replenishment.
- Published
- 2016
31. Odontoblastic inductive potential of epithelial cells derived from human deciduous dental pulp
- Author
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Joo-Cheol Park, You-Mi Seo, Ji-Won Park, Hyun-Sook Bae, Ha Hoon Kim, Gene Lee, and Hye Kyung Lee
- Subjects
Transcriptional Activation ,0301 basic medicine ,Bone sialoprotein ,Pathology ,medicine.medical_specialty ,Histology ,Physiology ,Cellular differentiation ,Gene Expression ,Odontoblast differentiation ,03 medical and health sciences ,0302 clinical medicine ,stomatognathic system ,medicine ,Humans ,Promoter Regions, Genetic ,Cells, Cultured ,Dental Pulp ,Odontoblasts ,biology ,Chemistry ,Mesenchymal stem cell ,Cell Differentiation ,Epithelial Cells ,Cell Biology ,General Medicine ,DMP1 ,Cell biology ,stomatognathic diseases ,030104 developmental biology ,Odontoblast ,030220 oncology & carcinogenesis ,biology.protein ,Pulp (tooth) ,Stem cell ,Biomarkers - Abstract
For the dentin regeneration, dental epithelial cells are indispensible and must possess odontoblastic induction capability. Epithelial cell-like stem cells were recently identified in human deciduous dental pulp (DPESCs). However, their cellular characteristics remain poorly defined. The purpose of this study was to characterize DPESCs compared to HAT-7 ameloblastic cells. Expression levels of ameloblast-specific markers [odontogenic ameloblast-associated protein (Odam), matrix metalloproteinase (Mmp)-20, amelogenin, and ameloblastin] were detected in DPESCs. Co-culturing odontoblastic MDPC-23 cells with DPESCs increased expression of odontoblast differentiation markers (Dmp1 and Dspp) from days 4 to 10, while the expression of bone sialoprotein rapidly decreased. MDPC-23 cells cultured in DPESC-conditioned medium (CM) showed increased Dspp promoter activity compared with control MDPC-23 cultures. Mineralization was first observed in the CM groups from day 4 and proceeded rapidly until day 14, whereas mineralized nodules were found from day 7 in control media-cultured cells. In conclusion, DPESCs in human deciduous pulp possess ameloblast-like characteristics and differentiation properties, and substances derived from DPESCs promote odontoblastic differentiation. Thus, our results indicate that DPESCs can be a realistic epithelial source for use in odontoblastic induction and dentin formation of dental mesenchymal cells.
- Published
- 2016
32. Odontogenic Ameloblast-Associated Protein (Odam) Plays Crucial Roles in Osteoclast Differentiation via Control of Actin Ring Formation
- Author
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Hye Kyung Lee and Joo-Cheol Park
- Subjects
musculoskeletal diseases ,RHOA ,Stress fiber ,biology ,Adhesion ,Osteoclast fusion ,Cell biology ,medicine.anatomical_structure ,Osteoclast ,biology.protein ,medicine ,Cell adhesion ,Ameloblast ,Actin - Abstract
Purpose: In osteoclast differentiation, actin-rich membrane protrusions play a crucial role in cell adhesion. Odontogenic ameloblast-associated protein (Odam) contributes to cell adhesion by inducing actin rearrangement. Odam-mediated RhoA activity may play a significant role in multinucleation of osteoclasts. However, the precise function of Odam in osteoclast cell adhesion and differentiation remains largely unknown. Here, we identify a critical role for Odam in inducing osteoclast adhesion and differentiation. Materials and Methods: The expression of Odam in osteoclasts was evaluated by immunohistochemistry. Primary mouse bone marrow and RAW264.7 cells were used to test the cell adhesion and actin ring formation induced by Odam. Result: Odam was expressed in osteoclasts around alveolar bone. Odam transfection induced actin filament rearrangement and cell adhesion compared with the control or collagen groups. Overexpression of Odam promoted actin stress fiber remodeling and cell adhesion, resulting in increased osteoclast fusion. Conclusion: These results suggest that Odam expression in primary mouse osteoclasts and RAW264.7 cells promotes their adhesion, resulting in the induction of osteoclast differentiation.
- Published
- 2015
33. Alarmin HMGB1 induces systemic and brain inflammatory exacerbation in post-stroke infection rat model
- Author
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Hahnbie Lee, Hye-Kyung Lee, Seung Woo Kim, Il-Doo Kim, Ja-Kyeong Lee, Juli Choi, and Pyung Lim Han
- Subjects
Lipopolysaccharides ,Male ,0301 basic medicine ,Cancer Research ,Immunology ,Nitric Oxide Synthase Type II ,chemical and pharmacologic phenomena ,Inflammation ,Endogeny ,Pharmacology ,HMGB1 ,Article ,Proinflammatory cytokine ,Rats, Sprague-Dawley ,03 medical and health sciences ,Cellular and Molecular Neuroscience ,0302 clinical medicine ,medicine ,Animals ,HMGB1 Protein ,lcsh:QH573-671 ,Receptor ,Behavior, Animal ,biology ,lcsh:Cytology ,business.industry ,Antagonist ,Brain ,Infarction, Middle Cerebral Artery ,Long-term potentiation ,Bacterial Infections ,Cell Biology ,Rats ,Toll-Like Receptor 4 ,Disease Models, Animal ,030104 developmental biology ,Cyclooxygenase 2 ,TLR4 ,biology.protein ,Ketamine ,medicine.symptom ,Peptides ,business ,030217 neurology & neurosurgery - Abstract
Post-stroke infection (PSI) is known to worsen functional outcomes of stroke patients and accounts to one-third of stroke-related deaths in hospital. In our previous reports, we demonstrated that massive release of high-mobility group box protein 1 (HMGB1), an endogenous danger signal molecule, is promoted by N-methyl-d-aspartic acid-induced acute damage in the postischemic brain, exacerbating neuronal damage by triggering delayed inflammatory processes. Moreover, augmentation of proinflammatory function of lipopolysaccharides (LPS) by HMGB1 via direct interaction has been reported. The aim of this study was to investigate the role of HMGB1 in aggravating inflammation in the PSI by exacerbating the function of LPS. PSI animal model was produced by administrating a low-dose LPS at 24 h post-middle cerebral artery occlusion (MCAO). Profound aggravations of inflammation, deterioration of behavioral outcomes, and infarct expansion were observed in LPS-injected MCAO animals, in which serum HMGB1 surge, especially disulfide type, occurred immediately after LPS administration and aggravated brain and systemic inflammations probably by acting in synergy with LPS. Importantly, blockage of HMGB1 function by delayed administrations of therapeutic peptides known to inhibit HMGB1 (HMGB1 A box, HPep1) or by treatment with LPS after preincubation with HMGB1 A box significantly ameliorated damages observed in the rat PSI model, demonstrating that HMGB1 plays a crucial role. Furthermore, administration of Rhodobacter sphaeroides LPS, a selective toll-like receptor 4 antagonist not only failed to exert these effects but blocked the effects of LPS, indicating its TLR4 dependence. Together, these results indicated that alarmin HMGB1 mediates potentiation of LPS function, exacerbating TLR4-dependent systemic and brain inflammation in a rat PSI model and there is a positive-feedback loop between augmentation of LPS function by HMGB1 and subsequent HMGB1 release/serum. Therefore, HMGB1 might be a valuable therapeutic target for preventing post-stroke infection.
- Published
- 2018
34. ODAM inhibits RhoA-dependent invasion in breast cancer
- Author
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Han Wool Choung, Joo-Cheol Park, Young Il Yang, Hye-Jung Yoon, Inae Park, and Hye Kyung Lee
- Subjects
RHOA ,biology ,Clinical Biochemistry ,Cancer ,Cell Biology ,General Medicine ,medicine.disease ,medicine.disease_cause ,Biochemistry ,Metastasis ,Cell biology ,Cancer cell ,medicine ,biology.protein ,Cancer research ,PTEN ,Carcinogenesis ,Cell adhesion ,Protein kinase B - Abstract
Odontogenic ameloblast-associated protein (ODAM) contributes to cell adhesion. In human cancer, ODAM is down-regulated, and the overexpression of ODAM results in a favourable prognosis; however, the molecular mechanisms underlying ODAM-mediated inhibition of cancer invasion and metastasis remain unclear. Here, we identify a critical role for ODAM in inducing cancer cell adhesion. ODAM induced RhoA activity and the expression of downstream factors, including Rho-associated kinase (ROCK). ODAM-mediated RhoA signalling resulted in actin filament rearrangement by activating PTEN and inhibiting the phosphorylation of AKT. When ODAM is overexpressed in MCF7 breast cancer cells and AGS gastric cancer cells that activate RhoA at high levels, it decreases motility, increases adhesion and inhibits the metastasis of MCF7 cells. Conversely, depletion of ODAM in cancer cells inhibits Rho GTPase activation, resulting in increased cancer migration and invasion. These results suggest that ODAM expression in cells maintains their adhesion, resulting in the prevention of their metastasis via the regulation of RhoA signalling in breast cancer cells. SIGNIFICANCE Breast cancer represents the first most frequent cancer, and the ratio of mortality is high in women. Of utmost importance for reducing risk by breast cancer are their anti-invasion mechanisms, particularly in the non-invasive cancer cells because metastasis is the principal cause of death among cancer patients. ODAM induced RhoA activity. ODAM-mediated RhoA signalling resulted in actin filament rearrangement, increased cell adhesion and inhibited the migration/invasion of MCF7 cells. These results suggest that ODAM expression maintains their adhesion, resulting in the prevention of their metastasis via the regulation of RhoA signalling in breast cancer cells.
- Published
- 2015
35. Odontogenic Ameloblast-associated Protein (ODAM) Mediates Junctional Epithelium Attachment to Teeth via Integrin-ODAM-Rho Guanine Nucleotide Exchange Factor 5 (ARHGEF5)-RhoA Signaling
- Author
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Su-Jin Park, Youngnim Choi, Hye Kyung Lee, Shin-Young Park, Suk Ji, Joo-Cheol Park, Hyo-Jung Lee, and Han-Wool Choung
- Subjects
Amyloid ,Integrins ,RHOA ,Integrin ,Epithelial Attachment ,Junctional epithelium ,Biochemistry ,junctional epithelium ,Cell Line ,Mice ,stomatognathic system ,Laminin ,Animals ,Humans ,tooth ,Periodontitis ,Molecular Biology ,biology ,Intracellular Signaling Peptides and Proteins ,Proteins ,Tooth surface ,Cell Biology ,Fibronectins ,Neoplasm Proteins ,Cell biology ,Fibronectin ,stomatognathic diseases ,Immunology ,biology.protein ,Rho signaling ,Signal transduction ,ODAM ,Carrier Proteins ,rhoA GTP-Binding Protein ,Ameloblast ,Rho Guanine Nucleotide Exchange Factors ,Signal Transduction - Abstract
Background: Adhesion of the junctional epithelium (JE) to the tooth surface is crucial for maintaining periodontal health. Results: JE adhesion to the tooth surface is regulated via fibronectin/laminin-integrin-ODAM-ARHGEF5-RhoA signaling. Conclusion: ODAM mediates JE attachment to healthy teeth. Significance: We investigate ODAM function during JE development and regeneration and its functional significance in the initiation and progression of periodontal disease., Adhesion of the junctional epithelium (JE) to the tooth surface is crucial for maintaining periodontal health. Although odontogenic ameloblast-associated protein (ODAM) is expressed in the JE, its molecular functions remain unknown. We investigated ODAM function during JE development and regeneration and its functional significance in the initiation and progression of periodontitis and peri-implantitis. ODAM was expressed in the normal JE of healthy teeth but absent in the pathologic pocket epithelium of diseased periodontium. In periodontitis and peri-implantitis, ODAM was extruded from the JE following onset with JE attachment loss and detected in gingival crevicular fluid. ODAM induced RhoA activity and the expression of downstream factors, including ROCK (Rho-associated kinase), by interacting with Rho guanine nucleotide exchange factor 5 (ARHGEF5). ODAM-mediated RhoA signaling resulted in actin filament rearrangement. Reduced ODAM and RhoA expression in integrin β3- and β6-knockout mice revealed that cytoskeleton reorganization in the JE occurred via integrin-ODAM-ARHGEF5-RhoA signaling. Fibronectin and laminin activated RhoA signaling via the integrin-ODAM pathway. Finally, ODAM was re-expressed with RhoA in regenerating JE after gingivectomy in vivo. These results suggest that ODAM expression in the JE reflects a healthy periodontium and that JE adhesion to the tooth surface is regulated via fibronectin/laminin-integrin-ODAM-ARHGEF5-RhoA signaling. We also propose that ODAM could be used as a biomarker of periodontitis and peri-implantitis.
- Published
- 2015
36. HMGB1-binding heptamer suppresses the synergistic effect of HMGB1 and LPS by interacting directly with HMGB1
- Author
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Hye-Kyung Lee, Il-Doo Kim, Ja-Kyeong Lee, Lidan Luo, and Hahnbie Lee
- Subjects
Lipopolysaccharides ,N-Methylaspartate ,Anti-Inflammatory Agents ,chemical and pharmacologic phenomena ,Peptide ,Endogeny ,Pharmacology ,Nitric Oxide ,HMGB1 ,Neuroprotection ,Proinflammatory cytokine ,law.invention ,Rats, Sprague-Dawley ,Mice ,law ,Animals ,Medicine ,HMGB1 Protein ,Cells, Cultured ,chemistry.chemical_classification ,Microglia ,biology ,business.industry ,General Neuroscience ,Brain ,Recombinant Proteins ,Neuroprotective Agents ,medicine.anatomical_structure ,chemistry ,Biochemistry ,Culture Media, Conditioned ,biology.protein ,Recombinant DNA ,lipids (amino acids, peptides, and proteins) ,Nasal administration ,business ,Oligopeptides ,Protein Binding - Abstract
High mobility group box 1 (HMGB1) is an endogenous danger signal molecule. In the postischemic brain, HMGB1 is massively released during acute damaging process and triggers inflammatory processes. Moreover, it has been reported HMGB1 augments the proinflammatory effect of LPS by direct interaction. In previous studies, the authors showed intranasally delivered a HMGB1 binding heptamer peptide (HBHP; HMSKPVQ) has robust neuroprotective effects in the ischemic brain after middle cerebral artery occlusion and that it exerts an anti-inflammatory effect. In the present study, the authors investigated whether HBHP suppresses the augmentation of the proinflammatory effect of LPS by HMGB1. In primary microglial cultures, low doses of LPS (5 ng/ml) and recombinant HMGB1 (rHMGB1, 20 ng/ml) synergistically activated microglial cells, and HMGB1-LPS binding was detected. In addition, synergistic NO accumulation along with direct HMGB1-LPS binding was also observed when primary microglial cultures were treated with LPS (5 ng/ml) and HMGB1 accumulated in NMDA-conditioned medium (NCM). Co-treatment of microglial cells with HBHP and LPS or rHMGB1 (NCM), or treatment with rHMGB1 or NCM and LPS after pre-incubating rHMGB1 (or NCM) with HBHP markedly suppressed their synergistic activation. Furthermore, interactions between rHMGB1 and LPS or between HMGB1 in NCM and LPS were suppressed dose-dependently by HBHP, indicating that HBHP suppressed the synergism between HMGB1 and LPS and the underlying mechanism involved inhibition of HMGB1-LPS binding. Together these results show HBHP has anti-inflammatory effects, and that it inhibits synergism caused by the binding of HMGB1 and LPS.
- Published
- 2015
37. Dentin sialophosphoprotein expression in enamel is regulated by Copine-7, a preameloblast-derived factor
- Author
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Hye Kyung Lee, You-Mi Seo, Chul Son, Su-Jin Park, Joo-Cheol Park, and Hyun Sook Bae
- Subjects
0301 basic medicine ,Sialoglycoproteins ,Blotting, Western ,Odontoblast differentiation ,Biology ,Real-Time Polymerase Chain Reaction ,03 medical and health sciences ,Mice ,stomatognathic system ,Dentin sialophosphoprotein ,Gene expression ,Ameloblasts ,Animals ,General Dentistry ,Cells, Cultured ,Extracellular Matrix Proteins ,030102 biochemistry & molecular biology ,Gene Expression Profiling ,Gene Expression Regulation, Developmental ,Cell Biology ,General Medicine ,Amelogenesis ,Phosphoproteins ,Immunohistochemistry ,Cell biology ,stomatognathic diseases ,030104 developmental biology ,Odontoblast ,Otorhinolaryngology ,Ameloblast differentiation ,Ameloblast ,Carrier Proteins ,Dentin sialoprotein ,Plasmids - Abstract
Objective Dentin sialophosphoprotein (Dspp) is expressed in odontoblasts and transiently expressed in early ameloblasts. However, the origin of Dspp in ameloblasts remains unclear. Our previous studies demonstrated that copine-7 (CPNE7), a molecule that is secreted by the dental epithelium, is expressed in early ameloblasts and is then translocated to differentiating odontoblasts; its expression levels correlate with odontoblast differentiation under the control of Dspp expression. The objective of this study is to figure out the relationship between CPNE7 and Dspp during amelogenesis. Design The gene expression patterns of CPNE7 and dentin sialoprotein (DSP) were examined by immunohistochemistry, western blot analysis, and real-time polymerase chain reaction. The effects of CPNE7 on Dspp regulation were investigated using luciferase and chromatin immunoprecipitation assays in ameloblastic HAT-7 cells. Results The gene expression pattern of Cpne7 was similar to that of Dspp during ameloblast differentiation. Moreover, Gene expression omnibus profiles indicated that there is a close correlation between Cpne7 and Dspp expression in various normal human tissues. We also confirmed the effects of CPNE7 on the induction of Dspp in ameloblastic HAT-7 cells. Cpne7 overexpression promoted Dspp expression, whereas Dspp expression was down-regulated by Cpne7 inactivation. Conclusions These results suggest that the expression of Dspp in early amelogenesis is linked to CPNE7, a preameloblast-derived factor.
- Published
- 2017
38. RaRF confers RA resistance by sequestering RAR to the nucleolus and regulating MCL1 in leukemia cells
- Author
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Soo-Jong Um, Hye Kyung Lee, Eun-Joo Kim, Ui-Hyun Park, HyeSook Youn, Sohn Hr, and BuHyun Youn
- Subjects
0301 basic medicine ,Acute promyelocytic leukemia ,Cancer Research ,Receptors, Retinoic Acid ,Retinoic acid ,Repressor ,Datasets as Topic ,Tretinoin ,Kaplan-Meier Estimate ,Biology ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Downregulation and upregulation ,Leukemia, Promyelocytic, Acute ,Myeloblast ,Cell Line, Tumor ,Genetics ,medicine ,Humans ,MCL1 ,Granulocyte Precursor Cells ,Molecular Biology ,Regulation of gene expression ,Gene Expression Regulation, Leukemic ,Cell Differentiation ,medicine.disease ,Up-Regulation ,Repressor Proteins ,Leukemia ,030104 developmental biology ,medicine.anatomical_structure ,chemistry ,Drug Resistance, Neoplasm ,030220 oncology & carcinogenesis ,Cancer research ,Myeloid Cell Leukemia Sequence 1 Protein ,Cell Nucleolus - Abstract
Retinoic acid (RA) has broad clinical applications for the treatment of various cancers, particularly acute promyelocytic leukemia. However, RA-based therapy is limited by relapse in patients associated with RA resistance, the mechanism of which is poorly understood. Here, we suggest a new molecular mechanism of RA resistance by a repressor, named RA resistance factor (RaRF). RaRF suppressed transcriptional activity of the RA receptor (RAR) by directly interacting with and sequestering RAR to the nucleolus in response to RA. RaRF was highly expressed in RA-resistant leukemia cells and its expression was strongly correlated with RA sensitivity. MCL1 was upregulated by RA treatment upon RaRF depletion, accompanying leukemic myeloblast differentiation, which is negatively regulated by ectopic RaRF expression. Collectively, we propose that RaRF may be a factor in the resistance mechanism and thus a potential target for leukemia therapy using RA.
- Published
- 2017
39. Anti-inflammatory and anti-excitoxic effects of diethyl oxopropanamide, an ethyl pyruvate bioisoster, exert robust neuroprotective effects in the postischemic brain
- Author
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Il-Doo Kim, Sung-Hwa Yoon, Ja-Kyeong Lee, Hye-Kyung Lee, Ju-Young Park, Seung Woo Kim, and Hahnbie Lee
- Subjects
0301 basic medicine ,Male ,Chemokine ,Programmed cell death ,medicine.drug_class ,Neutrophils ,medicine.medical_treatment ,Anti-Inflammatory Agents ,Pharmacology ,Nitric Oxide ,Neuroprotection ,Anti-inflammatory ,Article ,Brain Ischemia ,Rats, Sprague-Dawley ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,NF-KappaB Inhibitor alpha ,Cell Movement ,medicine ,Cell Adhesion ,Human Umbilical Vein Endothelial Cells ,Animals ,Humans ,Neurons ,Multidisciplinary ,biology ,Microglia ,NF-kappa B ,Transcription Factor RelA ,Brain ,Infarction, Middle Cerebral Artery ,Amides ,Rats ,030104 developmental biology ,Cytokine ,medicine.anatomical_structure ,Neuroprotective Agents ,chemistry ,biology.protein ,Pyruvic acid ,NAD+ kinase ,030217 neurology & neurosurgery - Abstract
Ethyl pyruvate (EP) is a simple aliphatic ester of pyruvic acid and has been shown to have robust neuroprotective effects via its anti-inflammatory, anti-oxidative, and anti-apoptotic functions. In an effort to develop novel EP derivatives with greater protective potencies than EP, we generated four EP isosteres, among them the neuroprotective potency of N,N-diethyl-2-oxopropanamide (DEOPA), in which the ethoxy group of EP was replaced with diethylamine, was far greater than that of EP. When DEOPA was administered intravenously (5 mg/kg) to rat middle cerebral artery occlusion (MCAO) model at 6 hrs post-surgery, it suppressed infarct formation, ameliorated neurological and sensory/motor deficits, and inhibited microglial activation and neutrophil infiltrations in the postischemic brain more effectively than EP. In particular, DEOPA markedly suppressed LPS-induced nitrite production and cytokine/chemokine inductions in microglia, neutrophils, and endothelial cells and these effects are attributable to inhibition of the activity of NF-κB by suppressing IκB-α degradation and p65 to DNA binding. In addition, DEOPA suppressed NMDA-induced neuronal cell death in primary cortical neuron cultures by NAD replenishment and suppression of NF-κB activity. Together, these results indicate DEOPA has multi-modal protective effects against ischemic brain damage targeting numerous cell types in the brain and also against other inflammation-related diseases.
- Published
- 2017
40. Ethyl pyruvate inhibits HMGB1 phosphorylation and secretion in activated microglia and in the postischemic brain
- Author
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Joo-Hyun Shin, Ja-Kyeong Lee, Hye-Kyung Lee, Yinchuan Jin, and Hahnbie Lee
- Subjects
Male ,Programmed cell death ,Anti-Inflammatory Agents ,chemical and pharmacologic phenomena ,Pharmacology ,HMGB1 ,Neuroprotection ,Brain Ischemia ,Rats, Sprague-Dawley ,medicine ,Animals ,Secretion ,HMGB1 Protein ,Phosphorylation ,Pyruvates ,Protein kinase A ,Protein kinase C ,Microglia ,biology ,General Neuroscience ,Brain ,Protein Transport ,Neuroprotective Agents ,medicine.anatomical_structure ,Biochemistry ,biology.protein - Abstract
Ethyl pyruvate (EP) has been shown to have anti-inflammatory effects and confer protective effects in various pathological conditions. For example, EP inhibits secretion of high mobility group box 1 (HMGB1), which is known to be released from activated or dying cells and aggravate inflammatory pathways. In the present study, we investigated whether EP reduces HMGB1 phosphorylation and release in ischemic brain and in cultured microglia. In the postischemic brains (60 min middle cerebral artery occlusion (MCAO)), HMGB1 was released extracellularly, generating dual peaks in cerebrospinal fluid (CSF) around 1 and 7 days after ischemic insult, which were probably generated from damaged neurons and activated inflammatory cells, respectively. We showed that treatment with EP 30 min post-MCAO (5 mg/kg, i.v.), which has been shown to confer a robust neuroprotective effect in the postischemic brain, reduced both peaks. In addition, delayed EP treatment from 4 days post-MCAO reduced HMGB1 accumulation in CSF at 7 day post-MCAO in the absence of accompanying amelioration of ischemic brain damage, indicating that the suppression of HMGB1 release is a direct effect. We also found that EP markedly suppressed the LPS-induced nuclear translocations of protein kinase C alpha and calcium/calmodulin-dependent protein kinase IV, HMGB1 phosphorylation, and subsequent secretion of HMGB1 induced by LPS in BV2 cells and EP-mediated above-mentioned effects were also independent of cell death or survival. These results indicate that EP inhibits HMGB1 phosphorylation and release in activated microglia, which might be responsible for EP-mediated suppression of HMGB1 release in the postischemic brain.
- Published
- 2014
41. Facultative CTCF sites moderate mammary super-enhancer activity and regulate juxtaposed gene in non-mammary cells
- Author
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Michaela Willi, Kyung Hyun Yoo, Tyler Kuhns, F. Reinisch, Chaochen Wang, Lothar Hennighausen, and Hye Kyung Lee
- Subjects
0301 basic medicine ,CCCTC-Binding Factor ,Science ,DNA Mutational Analysis ,General Physics and Astronomy ,Biology ,Insulator (genetics) ,Receptor Activity-Modifying Protein 3 ,General Biochemistry, Genetics and Molecular Biology ,Article ,03 medical and health sciences ,Mice ,Super-enhancer ,Mammary Glands, Animal ,Gene expression ,Animals ,Gene Regulatory Networks ,Regulatory Elements, Transcriptional ,Enhancer ,Promoter Regions, Genetic ,Gene ,Genetics ,Regulation of gene expression ,Multidisciplinary ,Binding Sites ,technology, industry, and agriculture ,General Chemistry ,Milk Proteins ,Chromatin ,030104 developmental biology ,Enhancer Elements, Genetic ,Gene Expression Regulation ,CTCF ,Female - Abstract
Precise spatiotemporal gene regulation is paramount for the establishment and maintenance of cell-specific programmes. Although there is evidence that chromatin neighbourhoods, formed by the zinc-finger protein CTCF, can sequester enhancers and their target genes, there is limited in vivo evidence for CTCF demarcating super-enhancers and preventing cross talk between distinct regulatory elements. Here, we address these questions in the Wap locus with its mammary-specific super-enhancer separated by CTCF sites from widely expressed genes. Mutational analysis demonstrates that the Wap super-enhancer controls Ramp3, despite three separating CTCF sites. Their deletion in mice results in elevated expression of Ramp3 in mammary tissue through augmented promoter–enhancer interactions. Deletion of the distal CTCF-binding site results in loss of Ramp3 expression in non-mammary tissues. This suggests that CTCF sites are porous borders, allowing a super-enhancer to activate a secondary target. Likewise, CTCF sites shield a widely expressed gene from suppressive influences of a silent locus., Chromatin neighbourhoods, formed by CTCF, have been proposed to isolate enhancers and their target genes from other regulatory elements. Here, the authors provide evidence that while CTCF binding does regulates mammary-specific super-enhancers, CTCF sites are relatively porous borders.
- Published
- 2016
42. Proangiogenic functions of an RGD-SLAY-containing osteopontin icosamer peptide in HUVECs and in the postischemic brain
- Author
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Seung Woo Kim, Yinchuan Jin, Ja-Kyeong Lee, Hye-Kyung Lee, Hahnbie Lee, and Il-Doo Kim
- Subjects
0301 basic medicine ,MAPK/ERK pathway ,Male ,Angiogenesis ,MAP Kinase Signaling System ,Clinical Biochemistry ,Integrin ,Amino Acid Motifs ,Biochemistry ,Umbilical vein ,Rats, Sprague-Dawley ,03 medical and health sciences ,0302 clinical medicine ,Cell Movement ,Human Umbilical Vein Endothelial Cells ,Animals ,Humans ,Osteopontin ,Molecular Biology ,Protein kinase B ,RGD motif ,Cell Proliferation ,Tube formation ,biology ,Chemistry ,Brain ,Infarction, Middle Cerebral Artery ,Integrin alphaVbeta3 ,Peptide Fragments ,Cell biology ,030104 developmental biology ,Neuroprotective Agents ,biology.protein ,Molecular Medicine ,Angiogenesis Inducing Agents ,Original Article ,Oligopeptides ,030217 neurology & neurosurgery - Abstract
Osteopontin (OPN) is a phosphorylated glycoprotein secreted into body fluids by various cell types. OPN contains arginine-glycine-aspartate (RGD) and serine-leucine-alanine-tyrosine (SLAY) motifs that bind to several integrins and mediate a wide range of cellular processes. In the present study, the proangiogenic effects of a 20-amino-acid OPN peptide (OPNpt20) containing RGD and SLAY motifs were examined in human umbilical vein endothelial cells (HUVECs) and in a rat focal cerebral ischemia model. OPNpt20 exerted robust proangiogenic effects in HUVECs by promoting proliferation, migration and tube formation. These effects were significantly reduced in OPNpt20-RAA (RGD->RAA)-treated cells, but only slightly reduced in OPNpt20-SLAA (SLAY->SLAA)-treated cells. Interestingly, a mutant peptide without both motifs failed to induce these proangiogenic processes, indicating that the RGD motif is crucial and that SLAY also has a role. In OPNpt20-treated HUVEC cultures, AKT and ERK signaling pathways were activated, but activation of these pathways and tube formation were suppressed by anti-αvβ3 antibody, indicating that OPNpt20 stimulates angiogenesis via the αvβ3-integrin/AKT and ERK pathways. The proangiogenic function of OPNpt20 was further confirmed in a rat middle cerebral artery occlusion model. Total vessel length and vessel densities were markedly greater in OPNpt20-treated ischemic brains, accompanied by induction of proangiogenic markers. Together, these results demonstrate that the 20-amino-acid OPN peptide containing RGD and SLAY motifs exerts proangiogenic effects, wherein both motifs have important roles, and these effects appear to contribute to the neuroprotective effects of this peptide in the postischemic brain.
- Published
- 2016
43. Up-down Regulation of HO-1 and iNOS Gene Expressions by Ethyl Pyruvate via Recruiting p300 to Nrf2 and Depriving It from p65
- Author
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Seung Woo Kim, Joo-Hyun Shin, Hye-Kyung Lee, and Ja-Kyeong Lee
- Subjects
NF-E2-Related Factor 2 ,Immunoblotting ,Anti-Inflammatory Agents ,Nitric Oxide Synthase Type II ,Transfection ,Biochemistry ,Antioxidants ,Cell Line ,Mice ,chemistry.chemical_compound ,Physiology (medical) ,medicine ,Animals ,Immunoprecipitation ,p300-CBP Transcription Factors ,RNA, Small Interfering ,CREB-binding protein ,Pyruvates ,Reporter gene ,Gene knockdown ,biology ,Reverse Transcriptase Polymerase Chain Reaction ,Transcription Factor RelA ,Membrane Proteins ,NF-κB ,Immunohistochemistry ,Antioxidant Response Elements ,Cell biology ,Protein Transport ,Cytosol ,Neuroprotective Agents ,Glutathione S-transferase ,medicine.anatomical_structure ,Gene Expression Regulation ,chemistry ,Cell culture ,biology.protein ,Nucleus ,Heme Oxygenase-1 - Abstract
Ethyl pyruvate (EP), a simple ester of pyruvic acid, has been shown to exert robust neuroprotection in various neuropathological conditions, such as, cerebral ischemia and KA-induced seizure animal models. The neuroprotective effect of EP is attributable to the anti-inflammatory, anti-oxidative, and anti-apoptotic effects. In the present study, we investigated convergence of anti-inflammatory and anti-oxidative functions of EP and present a novel molecular mechanism underlying anti-inflammatory effects of EP, which is conveyed by p300, a transcriptional co-activator for both Nuclear factor E2-related factor 2 (Nrf2) and p65. In BV2 cells, a microglia cell line, EP induced translocation of Nrf2 from the cytosol to the nucleus and enhanced the expression of hemeoxygenase 1 (HO-1) in a dose-dependent manner and 1h incubation with 10mM EP increased HO-1 to 4.9-fold. Nrf2 was found to translocate from the cytosol to the nucleus beginning 30 min after EP-treatment and binds to the antioxidant response element (ARE) located on HO-1 promoter. Interestingly, LPS-induced inducible NO synthase (iNOS) induction was substantially suppressed in EP-pre-treated BV2 cells and it was reverted by Nrf2 knockdown. We found that EP-induced Nrf2 accumulation in the nucleus recruits p300, a transcriptional co-activator of both Nrf2 and p65, inhibiting p65-p300 interaction. Competition between Nrf2 and p65 for p300 binding was confirmed by glutathione S-transferase (GST) pull down assay and reporter gene analysis. These results demonstrate that EP induced nuclear translocation of Nrf2 which binds to ARE along with p300 and hampers iNOS expression via depleting p300 from p65. This is a novel anti-inflammatory mechanism conveyed by EP, which enhances protective effect by converging anti-inflammatory and anti-oxidative effects and might be applicable to various Nrf2-activating agents, such as phytochemicals.
- Published
- 2013
44. The utility of CD44 and D2-40 as a prognostic predictor in invasive carcinomas of the breast
- Author
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Byungmo Lee, Woo Yong Lee, Jung Whan Chun, Seong Woo Hong, Hye Kyung Lee, and Yeo Goo Chang
- Subjects
Oncology ,medicine.medical_specialty ,Disease free survival ,Invasive carcinoma ,biology ,Lymphovascular invasion ,business.industry ,CD44 ,medicine.disease ,Lymphatic system ,Breast cancer ,Internal medicine ,Carcinoma ,medicine ,biology.protein ,Immunohistochemistry ,business - Abstract
Purpose: Local recurrence and distant metastasis are major prognostic factors associated with breast cancer. Lymphovascular invasion is an important pathway of metastatic spread. CD44 and D2-40 are assumed to be related with invasion of malignant cells to the adjacent lymphatic or vascular structures by different mechanisms. This study was conducted to examine whether CD44 and D2-40 expression together have prognostic value in invasive carcinoma of the breast. Methods: A total of 46 surgically resected tissue samples of invasive carcinoma of the breast were analyzed for immunohistochemical expression of CD44 and D2-40, along with other clinicopathologic factors. Association between patient related factors and disease free survival was evaluated by statistical analysis. Results: Disease free survival was shorter in CD44-positive patients (median, 66.5 months) than in CD44-negative patients (median, 112.6 months; P Conclusion: We concluded that expression of CD44 and D2-40 is associated with unfavorable prognosis. Concurrent positivity of both markers may be useful in predicting malignant relapse following surgical resection with shorter disease free survival in patients with invasive carcinoma of the breast.
- Published
- 2013
45. CRISPR/Cas9 targeting events cause complex deletions and insertions at 17 sites in the mouse genome
- Author
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Lothar Hennighausen, Kyung Hyun Yoo, Ha Youn Shin, Chul Min Yang, Tyler Kuhns, Hye Kyung Lee, Xianke Zeng, Teresa Mohr, Chaochen Wang, and Chengyu Liu
- Subjects
0301 basic medicine ,Male ,Genotype ,Science ,General Physics and Astronomy ,Mice, Transgenic ,Biology ,Genome ,General Biochemistry, Genetics and Molecular Biology ,Article ,03 medical and health sciences ,Mice ,Genome editing ,CRISPR ,Animals ,Guide RNA ,Genotyping ,Gene ,Sequence (medicine) ,Sequence Deletion ,Genetics ,Gene Editing ,Multidisciplinary ,Cas9 ,General Chemistry ,Mice, Inbred C57BL ,030104 developmental biology ,Female ,CRISPR-Cas Systems ,RNA, Guide, Kinetoplastida - Abstract
Although CRISPR/Cas9 genome editing has provided numerous opportunities to interrogate the functional significance of any given genomic site, there is a paucity of data on the extent of molecular scars inflicted on the mouse genome. Here we interrogate the molecular consequences of CRISPR/Cas9-mediated deletions at 17 sites in four loci of the mouse genome. We sequence targeted sites in 632 founder mice and analyse 54 established lines. While the median deletion size using single sgRNAs is 9 bp, we also obtain large deletions of up to 600 bp. Furthermore, we show unreported asymmetric deletions and large insertions of middle repetitive sequences. Simultaneous targeting of distant loci results in the removal of the intervening sequences. Reliable deletion of juxtaposed sites is only achieved through two-step targeting. Our findings also demonstrate that an extended analysis of F1 genotypes is required to obtain conclusive information on the exact molecular consequences of targeting events., CRISPR/Cas9 gene editing has been used to generate mutations in several mouse genes. Here, the authors show that targeting events using single guide RNAs cause large deletions at 17 sites in the mouse genome, suggesting that careful genotyping is needed and sequential targeting may avoid such deletions.
- Published
- 2016
46. A Case of H. pylori-associated Granulomatous Gastritis with Hypertrophic Gastropathy
- Author
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Hye Kyung Lee, Seung-Woo Lee, Yeon Soo Kim, Soon Woo Nam, Dong Soo Lee, Jong Ok Kim, and Sang Beom Kang
- Subjects
medicine.medical_specialty ,Hypertrophic gastropathy ,Pathology ,Helicobacter pylori ,Hepatology ,biology ,business.industry ,Gastroenterology ,Rapid urease test ,Case Report ,Histology ,biology.organism_classification ,Granulomatous gastritis ,Chronic granulomatous gastritis ,Hypertrophic gastritis ,Giant cell ,Internal medicine ,medicine ,business ,Granulomatous Gastritis ,Antrum - Abstract
A 46-year-old man had chronic granulomatous gastritis characterized by giant gastric folds with noncaseating epithelioid granulomas including giant cells in the corpus. No definite etiologic factors were detected. Histology and the rapid urease test indicated that H. pylori was present in both the antrum and corpus. The granulomatous gastritis with giant gastric folds improved after H. pylori eradication. This case suggests an association between isolated granulomatous gastritis and H. pylori infection.
- Published
- 2009
47. Quality Characteristics of Abalone Porridge with Viscera
- Author
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Koth-Bong-Woo-Ri Kim, Mi-Jung Kim, Jae-Hun Kim, Myung-Woo Byun, Kyung-A Lee, Hye-Kyung Lee, Ju-Woon Lee, Dong-Hyun Ahn, Eun-Su Shin, and Eun-Soon Lyu
- Subjects
body regions ,Nutrition and Dietetics ,Abalone ,food and beverages ,Food science ,Biology ,Quality characteristics ,Food Science - Abstract
To develop an optimal composite recipe for a functional abalone porridge including abalone viscera, this study investigated the effects of added viscera on the physical, textural and sensory characteristics of the porridge. Several kinds of abalone porridge were prepared with 1%, 2%, 3%, 4%, or 5% of the viscera (w/w) and with round rice, half rice, or ground rice. pH of the porridge with half and ground rice decreased with increasing amounts of the viscera. TBARS value increased with increasing size of rice and increased with increasing amount of the viscera in the porridge with round and ground rice. Yellowness of the porridge increased significantly by the addition of the viscra. Among the four textural properties, consistency and firmness increased with increasing size of rice; in contrast, viscosity and cohesivness decreased with increasing size of rice. However, textural properties of the porridge were not significantly different by the addition of the viscera. In the sensory evaluation, sensory scores of the porridge with round and half rice were higher than with round rice for texture, taste and total. In conclusion, concerning overall sensory evaluation, round rice porridge with 2% viscera, half rice porridge with 3% viscera and ground rice porridge with 4% viscera showed the best results, implying that developing functional abalone porridge using the viscera may be worthy.
- Published
- 2008
48. MED25 is distinct from TRAP220/MED1 in cooperating with CBP for retinoid receptor activation
- Author
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Eun-Joo Kim, Hye Kyung Lee, Soo-Jong Um, and Ui-Hyun Park
- Subjects
Transcriptional Activation ,Chromatin Immunoprecipitation ,Receptors, Retinoic Acid ,Retinoic acid ,Retinoid receptor ,Tretinoin ,Biology ,Retinoid X receptor ,Article ,General Biochemistry, Genetics and Molecular Biology ,Mediator Complex Subunit 1 ,Mice ,chemistry.chemical_compound ,Animals ,Humans ,RNA, Small Interfering ,Molecular Biology ,Transcription factor ,Mediator Complex ,Thyroid hormone receptor ,Base Sequence ,General Immunology and Microbiology ,organic chemicals ,General Neuroscience ,3T3 Cells ,Molecular biology ,body regions ,Retinoic acid receptor ,Nuclear receptor ,chemistry ,embryonic structures ,Trans-Activators ,RNA Interference ,Carrier Proteins ,Corticosterone ,Chromatin immunoprecipitation ,Transcription Factors - Abstract
We isolated MED25, which associates with retinoic acid (RA)-bound retinoic acid receptor (RAR) through the C-terminal nuclear hormone receptor (NR) box/LxxLL motif, and increases RAR/RXR-mediated transcription. When tethered to a promoter, MED25 showed intrinsic transcriptional activity in its PTOV domain, which is likely accomplished by direct association with CBP. Reporter assays using dominant negatives of MED25 demonstrated the importance of the N-terminal Mediator-binding and C-terminal domains in CBP and RAR/RXR binding, which affect MED25 activity. Downregulation of MED25 specifically reduced RAR but not thyroid hormone receptor (TR) activity. Stimulation of RAR by MED25 was correlated with enhanced RA cytotoxicity in vivo. Chromatin immunoprecipitation (ChIP) assays revealed the RA-dependent recruitment of MED25 to the RARbeta2 promoter. Recruitment of CBP and TRAP220 was diminished by the overexpression of a MED25 NR box deletion mutant, and by treatment with MED25 siRNA. Time-course ChIP assays indicated that CBP, together with RAR and MED25, is recruited early, whereas TRAP220 is recruited later to the promoter. Our data suggest that MED25, in cooperation with CBP and Mediators through its distinct domains, imposes a selective advantage on RAR/RXR activation.
- Published
- 2007
49. Preameloblast-Derived Factors Mediate Osteoblast Differentiation of Human Bone Marrow Mesenchymal Stem Cells by Runx2-Osterix-BSP Signaling
- Author
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Joo-Cheol Park, Won-Jun Shon, Han-Wool Choung, Hye Kyung Lee, and Dong-Seol Lee
- Subjects
0301 basic medicine ,Bone sialoprotein ,Pathology ,medicine.medical_specialty ,Cellular differentiation ,Biomedical Engineering ,Bioengineering ,Bone Marrow Cells ,Core Binding Factor Alpha 1 Subunit ,Biochemistry ,Biomaterials ,03 medical and health sciences ,Mice ,0302 clinical medicine ,Dental pulp stem cells ,medicine ,Ameloblasts ,Animals ,Humans ,Integrin-Binding Sialoprotein ,Osteoblasts ,biology ,Chemistry ,Regeneration (biology) ,Mesenchymal stem cell ,Osteoblast ,Cell Differentiation ,Mesenchymal Stem Cells ,Cell biology ,Rats ,RUNX2 ,030104 developmental biology ,medicine.anatomical_structure ,Sp7 Transcription Factor ,030220 oncology & carcinogenesis ,Culture Media, Conditioned ,biology.protein ,Ameloblast ,Signal Transduction ,Transcription Factors - Abstract
Epithelial-mesenchymal interaction occurs during development of various tissues, including teeth and bone. Recently, a preameloblast-conditioned medium (PA-CM) from mouse apical bud cells (ABCs), a type of dental epithelial cell, was found to induce odontogenic differentiation of dental pulp stem cells and promote dentin formation. The aims of the present study were to investigate the effects of PA-CM on human bone marrow mesenchymal stem cells (hBMSCs) in vitro, and to investigate the bone regenerative capacity in vivo through epithelial-mesenchymal interactions of developmental osteogenesis. Coculturing with ABCs and PA-CM treatment upregulated osteoblast differentiation markers of hBMSCs compared to cells cultured alone. PA-CM accelerated mineralized nodule formation and also increased bone sialoprotein promoter activity in hBMSCs. PA-CM facilitated the migration of hBMSCs, but did not significantly influence proliferation. PA-CM promoted bone formation of hBMSCs in vivo. Radiographic and histologic findings showed that PA-CM induced the bony regeneration at calvarial defects in rat. Taken together, these data show that PA-CM enhances the migration and osteogenic differentiation of hBMSCs in vitro and induces bone formation in vivo.
- Published
- 2015
50. Novel MMP20 and KLK4 Mutations in Amelogenesis Imperfecta
- Author
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Elif Bahar Tuna, Hye Kyung Lee, D.-S. Lee, Koray Gençay, Figen Seymen, Yae Jean Kim, Merve Bayram, Joo-Cheol Park, Zang Hee Lee, J.-W. Kim, Mine Koruyucu, and Kyung-Ah Lee
- Subjects
Genotype ,Amelogenesis Imperfecta ,Mutant ,Blotting, Western ,Biology ,Polymerase Chain Reaction ,Polymorphism, Single Nucleotide ,Consanguinity ,stomatognathic system ,Radiography, Panoramic ,medicine ,Humans ,Amelogenesis imperfecta ,Child ,General Dentistry ,Gene ,Exome sequencing ,Genetics ,Enamel paint ,MMP20 ,Homozygote ,Sequence Analysis, DNA ,medicine.disease ,Tooth enamel ,Pedigree ,stomatognathic diseases ,medicine.anatomical_structure ,Matrix Metalloproteinase 20 ,visual_art ,Mutation ,visual_art.visual_art_medium ,Female ,Kallikreins ,Amelogenin - Abstract
In order to achieve highly mineralized tooth enamel, enamel proteinases serve the important function of removing the remaining organic matrix in the mineralization and maturation of the enamel matrix. Mutations in the kallikrein 4 ( KLK4), enamelysin ( MMP20), and WDR72 genes have been identified as causing hypomaturation enamel defects in an autosomal-recessive hereditary pattern. In this report, 2 consanguineous families with a hypomaturation-type enamel defect were recruited, and mutational analysis was performed to determine the molecular genetic etiology of the disease. Whole exome sequencing and autozygosity mapping identified novel homozygous mutations in the KLK4 (c.620_621delCT, p.Ser207Trpfs*38) and MMP20 (c.1054G>A, p.Glu352Lys) genes. Further analysis on the effect of the mutations on the translation, secretion, and function of KLK4 and MMP20 revealed that mutant KLK4 was degraded intracellularly and became inactive while mutant MMP20 was expressed at a normal level but secreted only minimally with proteolytic function.
- Published
- 2015
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