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Safe S, Oany AR, Upadhyay S, Tsui WN, Hailemariam A, Latka S, Landua J, Scherer S, Welm AL, Villanueva H, Lewis M. Orphan Nuclear Receptor 4A1 (NR4A1) and NR4A2are Endogenous Regulators of CD71 and TheirLigands Induce Ferroptosis in Breast Cancer. RESEARCH SQUARE 2025:rs.3.rs-6214709. [PMID: 40313760 PMCID: PMC12045456 DOI: 10.21203/rs.3.rs-6214709/v1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Indexed: 05/03/2025]
Abstract
Ferroptosis is an iron-dependent cell death pathway that involves multiple genes including the transferrin receptor (TFRC/CD71), glutathione peroxidase 4 (GPX4) and SLC7A11. This study is based on the hypothesis that orphan nuclear receptor 4A1 (NR4A1) and NR4A2 maintain low levels of ferroptosis in triple negative breast cancer (TNBC) cells and that bis-indole derived (CDIM) compounds act as NR4A1/2 ligands that induce ferroptosis by enhancing CD71 expression. 1,1-Bis(3'-indolyl)-1-(3,5-disubstitutedphenyl)methane (DIM-3,5) analogs were investigated for their cytotoxicity and effects on NR4A1 and NR4A2 regulated genes and induction of ferroptosis by cytotoxicity, western blot and RT-PCR. Several assays also determined enhanced lipoperoxidation, reactive oxygen species and malondialdehyde formation in TNBC cells. Knockdown of NR4A1, NR4A2, Sp1 and Sp4 was carried out by RNA interference. Molecular mechanisms of NR4A1/2-mediated regulation of CD71 expression were determined using CD71-luciferase promoter constructs, overexpression of Sp1 and chromatin immunoprecipitation (ChIP) assays. Initial studies show that DIM-3,5 act as an inverse NR4A1/NR4A2 agonist that downregulated the pro-oncogenic responses/gene products regulated by both receptors in TNBC cells. DIM-3,5 analogs also induced ROS, malondialdehyde and lipoperoxide formation in TNBC cells and this was accompanied by indicators of ferroptosis that include decreased expression of GPX4 and SLC7A11 and induction of CD71. Induction of CD71, an important biomarker of ferroptosis was observed after treatment of TNBC cells with DIM-3,5 analogs, knockdown of NR4A1, NR4A2, Sp1 or Sp4 demonstrating that induction of CD71 was coregulated by both receptors. Moreover, both promoter and ChIP analysis indicated that NR4A1 and NR4A2 acted as ligand-dependent cofactors of Sp1/4-mediated expression of CD71 in TNBC cells. CD71, a key biomarker of ferroptosis is an NR4A1/2/Sp regulated gene that can be directly targeted by DIM-3,5 inverse NR4A1/2 agonists to induce ferroptosis in TNBC cells.
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Affiliation(s)
| | | | | | | | | | | | | | - Sandra Scherer
- University of Utah, Huntsman Cancer Institute and Department of Oncological Sciences
| | - Alana L Welm
- Department of Oncological Sciences, Huntsman Cancer Institute, University of Utah, Salt Lake City, UT 84112, USA
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2
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Upadhyay S, Lee M, Zhang L, Oany AR, Mikheeva SA, Mikheev AM, Rostomily RC, Safe S. Dual nuclear receptor 4A1 (NR4A1/NR4A2) ligands inhibit glioblastoma growth and target TWIST1. Mol Pharmacol 2025; 107:100009. [PMID: 40023516 PMCID: PMC11881746 DOI: 10.1016/j.molpha.2024.100009] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/12/2024] [Accepted: 11/18/2024] [Indexed: 03/04/2025] Open
Abstract
1,1-Bis(3'-indolyl)-1-(3,5-disubstitutedphenyl)methane (DIM-3,5) compounds are dual receptor ligands that bind both orphan nuclear receptor 4A1 (NR4A1) and NR4A2. Knockdown of NR4A1 or NR4A2 by RNA interference in glioblastoma (GBM) cells decreased growth and induced apoptosis and comparable effects were observed for DIM-3,5 analogs, which exhibit inverse agonist activity and inhibit NR4A1- and NR4A2-mediated pro-oncogenic activity. Knockdown of NR4A1 or NR4A2 or treatment with DIM-3,5 analogs also decreased expression of TWIST1 mRNA and protein in GBM cells by 40%-90%.The proximal region of the TWIST1 gene promoter contains functional GC-rich binding sites that bind Sp1 and Sp4, and knockdown of these transcription factors also decreased TWIST1 expression in GBM cells. Further analysis by chromatin immunoprecipitation, protein-protein coimmunoprecipitation, and binding assays demonstrated that NR4A1/NR4A2 coregulate TWIST1 gene expression as ligand-dependent cofactors of Sp1 and Sp4, which interact with cis proximal GC-rich sites in the TWIST1 gene promoter. In vivo studies show that DIM-3,5 dual NR4A1/2 inverse agonists also reduced intratumoral TWIST1 expression while significantly prolonging survival of mice in a syngeneic mouse model of GBM, demonstrating that these ligands are promising new agents for targeting TWIST1 and treating GBM. SIGNIFICANCE STATEMENT: The TWIST1 gene is a pro-oncogenic factor that regulates epithelial-to-mesenchymal transition in glioblastoma cells. This paper shows that the orphan nuclear receptor 4A1 (NR4A1) and NR4A2 regulate TWIST1 expression, which can be targeted by bis-indole-derived dual NR4A1/2 inverse agonists.
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MESH Headings
- Twist-Related Protein 1/metabolism
- Twist-Related Protein 1/genetics
- Twist-Related Protein 1/antagonists & inhibitors
- Glioblastoma/drug therapy
- Glioblastoma/metabolism
- Glioblastoma/pathology
- Glioblastoma/genetics
- Humans
- Animals
- Nuclear Proteins/metabolism
- Nuclear Proteins/genetics
- Nuclear Proteins/antagonists & inhibitors
- Nuclear Receptor Subfamily 4, Group A, Member 1/metabolism
- Nuclear Receptor Subfamily 4, Group A, Member 1/genetics
- Nuclear Receptor Subfamily 4, Group A, Member 1/agonists
- Nuclear Receptor Subfamily 4, Group A, Member 2/metabolism
- Nuclear Receptor Subfamily 4, Group A, Member 2/genetics
- Nuclear Receptor Subfamily 4, Group A, Member 2/agonists
- Cell Line, Tumor
- Ligands
- Mice
- Indoles/pharmacology
- Cell Proliferation/drug effects
- Mice, Nude
- Apoptosis/drug effects
- Xenograft Model Antitumor Assays
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Affiliation(s)
- Srijana Upadhyay
- Department of Veterinary Physiology and Pharmacology, College of Veterinary Medicine, Texas A&M University, College Station, Texas
| | - Miok Lee
- Department of Biochemistry and Biophysics, College of Agricultural and Life Sciences, Texas A&M University, College Station, Texas
| | - Lei Zhang
- Department of Veterinary Physiology and Pharmacology, College of Veterinary Medicine, Texas A&M University, College Station, Texas
| | - Arafat Rahman Oany
- Department of Veterinary Physiology and Pharmacology, College of Veterinary Medicine, Texas A&M University, College Station, Texas
| | - Svetlana A Mikheeva
- Department of Neurosurgery, Houston Methodist Hospital and Research Institute, Houston, Texas
| | - Andrei M Mikheev
- Department of Neurosurgery, Houston Methodist Hospital and Research Institute, Houston, Texas
| | - Robert C Rostomily
- Department of Neurosurgery, Houston Methodist Hospital and Research Institute, Houston, Texas
| | - Stephen Safe
- Department of Veterinary Physiology and Pharmacology, College of Veterinary Medicine, Texas A&M University, College Station, Texas.
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3
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Göksu AY. A review article on the development of dopaminergic neurons and establishment of dopaminergic neuron-based in vitro models by using immortal cell lines or stem cells to study and treat Parkinson's disease. Int J Dev Neurosci 2024; 84:817-842. [PMID: 39379284 DOI: 10.1002/jdn.10383] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/13/2023] [Revised: 09/09/2024] [Accepted: 09/12/2024] [Indexed: 10/10/2024] Open
Abstract
The primary pathological hallmark of Parkinson's disease (PD) is the degeneration of dopaminergic (DA) neurons in the substantia nigra pars compacta, a critical midbrain region. In vitro models based on DA neurons provide a powerful platform for investigating the cellular and molecular mechanisms of PD and testing novel therapeutic strategies. A deep understanding of DA neuron development, including the signalling pathways and transcription factors involved, is essential for advancing PD research. This article first explores the differentiation and maturation processes of DA neurons in the midbrain, detailing the relevant signalling pathways. It then compares various in vitro models, including primary cells, immortalized cell lines, and stem cell-based models, focusing on the advantages and limitations of each. Special attention is given to the role of immortalized and stem cell models in PD research. This review aims to guide researchers in selecting the most appropriate model for their specific research goals. Ethical considerations and clinical implications of using stem cells in PD research are also discussed.
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Affiliation(s)
- Azize Yasemin Göksu
- Department of Histology and Embryology, Department of Gene and Cell Therapy, Akdeniz University, School of Medicine, Antalya, Turkey
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Johnson S, Yu Z, Li X, Zarei M, Vaziri-Gohar A, Lee M, Upadhyay S, Du H, Zarei M, Safe S. A novel NR4A2-HuR axis promotes pancreatic cancer growth and tumorigenesis that is inhibited by NR4A2 antagonists. Am J Cancer Res 2024; 14:4337-4352. [PMID: 39417168 PMCID: PMC11477821 DOI: 10.62347/kcpn6689] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/02/2024] [Accepted: 08/18/2024] [Indexed: 10/19/2024] Open
Abstract
Pancreatic ductal adenocarcinoma (PDAC) patients' express higher levels of the orphan Nuclear Receptor 4A2 (NR4A2, NURR1) compared to normal pancreas and NR4A2 is a prognostic factor for patient survival. Knockdown of NR4A2 by RNA interference (RNAi) inhibited cell proliferation, invasion, and migration. RNA sequencing performed in NR4A2(+/+) and NR4A2(-/-) MiaPaCa2 cells demonstrated that NR4A2 played a significant role in cellular metabolism. Human antigen R (HuR) and isocitrate dehydrogenase 1 (IDH1) were identified as NR4A2 target genes. HuR is a pro-oncogenic RNA binding protein and silencing of HuR by RNAi significantly downregulated expression of NR4A2. Expression of HuR and IDH1 were significantly downregulated after treatment with NR4A2 inverse agonist, 1,1-bis(3'-indolyl)-1-(p-chlorophenyl)methane resulting in significant inhibition of tumor growth in an athymic nude mouse xenograft model. This study demonstrates that NR4A2 and HuR regulate genes and signaling pathways that enhance tumorigenesis and targeting NR4A2 and HuR expression with an NR4A2 inverse agonist represents a novel regimen for treating PDAC.
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Affiliation(s)
- Sneha Johnson
- Department of Veterinary Physiology and Pharmacology, Texas A&M UniversityCollege Station, TX, USA
| | - Zuhua Yu
- Department of Veterinary Physiology and Pharmacology, Texas A&M UniversityCollege Station, TX, USA
- Henan University of Science and TechnologyLuoyang, Henan, P. R. China
| | - Xi Li
- Department of Veterinary Physiology and Pharmacology, Texas A&M UniversityCollege Station, TX, USA
| | - Mehrdad Zarei
- Department of Surgery, University HospitalsCleveland, OH, USA
- School of Medicine, Case Western UniversityCleveland, OH, USA
| | - Ali Vaziri-Gohar
- Department of Surgery, University HospitalsCleveland, OH, USA
- School of Medicine, Case Western UniversityCleveland, OH, USA
| | - Miok Lee
- Department of Veterinary Physiology and Pharmacology, Texas A&M UniversityCollege Station, TX, USA
- Department of Biochemistry and Biophysics, Texas A&M UniversityCollege Station, TX, USA
| | - Srijana Upadhyay
- Department of Veterinary Physiology and Pharmacology, Texas A&M UniversityCollege Station, TX, USA
| | - Heng Du
- Department of Medicine, Brigham and Women’s Hospital and Harvard Medical SchoolBoston, MA, USA
| | - Mahsa Zarei
- Department of Veterinary Physiology and Pharmacology, Texas A&M UniversityCollege Station, TX, USA
- Department of Medicine, Brigham and Women’s Hospital and Harvard Medical SchoolBoston, MA, USA
| | - Stephen Safe
- Department of Veterinary Physiology and Pharmacology, Texas A&M UniversityCollege Station, TX, USA
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Gatica-Garcia B, Bannon MJ, Martínez-Dávila IA, Soto-Rojas LO, Reyes-Corona D, Escobedo L, Maldonado-Berny M, Gutierrez-Castillo ME, Espadas-Alvarez AJ, Fernandez-Parrilla MA, Mascotte-Cruz JU, Rodríguez-Oviedo CP, Valenzuela-Arzeta IE, Luna-Herrera C, Lopez-Salas FE, Santoyo-Salazar J, Martinez-Fong D. Unilateral rNurr1-V5 transgene expression in nigral dopaminergic neurons mitigates bilateral neuropathology and behavioral deficits in parkinsonian rats with α-synucleinopathy. Neural Regen Res 2024; 19:2057-2067. [PMID: 38227536 DOI: 10.4103/1673-5374.391190] [Citation(s) in RCA: 2] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/14/2023] [Accepted: 11/28/2023] [Indexed: 01/17/2024] Open
Abstract
JOURNAL/nrgr/04.03/01300535-202409000-00039/figure1/v/2024-01-16T170235Z/r/image-tiff Parkinsonism by unilateral, intranigral β-sitosterol β-D-glucoside administration in rats is distinguished in that the α-synuclein insult begins unilaterally but spreads bilaterally and increases in severity over time, thus replicating several clinical features of Parkinson's disease, a typical α-synucleinopathy. As Nurr1 represses α-synuclein, we evaluated whether unilateral transfected of rNurr1-V5 transgene via neurotensin-polyplex to the substantia nigra on day 30 after unilateral β-sitosterol β-D-glucoside lesion could affect bilateral neuropathology and sensorimotor deficits on day 30 post-transfection. This study found that rNurr1-V5 expression but not that of the green fluorescent protein (the negative control) reduced β-sitosterol β-D-glucoside-induced neuropathology. Accordingly, a bilateral increase in tyrosine hydroxylase-positive cells and arborization occurred in the substantia nigra and increased tyrosine hydroxylase-positive ramifications in the striatum. In addition, tyrosine hydroxylase-positive cells displayed less senescence marker β-galactosidase and more neuron-cytoskeleton marker βIII-tubulin and brain-derived neurotrophic factor. A significant decrease in activated microglia (positive to ionized calcium-binding adaptor molecule 1) and neurotoxic astrocytes (positive to glial fibrillary acidic protein and complement component 3) and increased neurotrophic astrocytes (positive to glial fibrillary acidic protein and S100 calcium-binding protein A10) also occurred in the substantia nigra. These effects followed the bilateral reduction in α-synuclein aggregates in the nigrostriatal system, improving sensorimotor behavior. Our results show that unilateral rNurr1-V5 transgene expression in nigral dopaminergic neurons mitigates bilateral neurodegeneration (senescence and loss of neuron-cytoskeleton and tyrosine hydroxylase-positive cells), neuroinflammation (activated microglia, neurotoxic astrocytes), α-synuclein aggregation, and sensorimotor deficits. Increased neurotrophic astrocytes and brain-derived neurotrophic factor can mediate the rNurr1-V5 effect, supporting its potential clinical use in the treatment of Parkinson's disease.
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Affiliation(s)
- Bismark Gatica-Garcia
- Departamento de Fisiología, Biofísica y Neurociencias, Centro de Investigación y de Estudios Avanzados, Ciudad de México, México
| | - Michael J Bannon
- Department of Pharmacology, Wayne State University School of Medicine, Detroit, MI, USA
| | - Irma Alicia Martínez-Dávila
- Departamento de Fisiología, Biofísica y Neurociencias, Centro de Investigación y de Estudios Avanzados, Ciudad de México, México
| | - Luis O Soto-Rojas
- Laboratorio de Patogénesis Molecular, Carrera Médico Cirujano, Facultad de Estudios Superiores Iztacala, Universidad Nacional Autónoma de México, Tlalnepantla de Baz, México
- Red de Medicina para la Educación y Desarrollo y la Investigación Científica de Iztacala (Red MEDICI), Carrera Médico Cirujano, Facultad de Estudios Superiores Iztacala, Universidad Nacional Autónoma de México, Tlalnepantla de Baz, México
| | | | - Lourdes Escobedo
- Departamento de Fisiología, Biofísica y Neurociencias, Centro de Investigación y de Estudios Avanzados, Ciudad de México, México
| | - Minerva Maldonado-Berny
- Departamento de Fisiología, Biofísica y Neurociencias, Centro de Investigación y de Estudios Avanzados, Ciudad de México, México
| | - M E Gutierrez-Castillo
- Departamento de Biociencias e Ingeniería, Centro Interdisciplinario de Investigaciones y Estudios sobre Medio Ambiente y Desarrollo, Instituto Politécnico Nacional, Ciudad de México, México
| | - Armando J Espadas-Alvarez
- Departamento de Biociencias e Ingeniería, Centro Interdisciplinario de Investigaciones y Estudios sobre Medio Ambiente y Desarrollo, Instituto Politécnico Nacional, Ciudad de México, México
| | | | - Juan U Mascotte-Cruz
- Departamento de Fisiología, Biofísica y Neurociencias, Centro de Investigación y de Estudios Avanzados, Ciudad de México, México
| | | | - Irais E Valenzuela-Arzeta
- Departamento de Fisiología, Biofísica y Neurociencias, Centro de Investigación y de Estudios Avanzados, Ciudad de México, México
| | - Claudia Luna-Herrera
- Departamento de Fisiología, Escuela Nacional de Ciencias Biológicas, Instituto Politécnico Nacional, Ciudad de México, México
| | - Francisco E Lopez-Salas
- Departamento de Biología Molecular y Biotecnología, Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, Ciudad de México, México
| | - Jaime Santoyo-Salazar
- Departamento de Física, Centro de Investigación y de Estudios Avanzados, Ciudad de México, México
| | - Daniel Martinez-Fong
- Departamento de Fisiología, Biofísica y Neurociencias, Centro de Investigación y de Estudios Avanzados, Ciudad de México, México
- Nanoparticle Therapy Institute, Aguascalientes, México
- Programa de Nanociencias y Nanotecnología, Centro de Investigación y de Estudios Avanzados, Ciudad de México, México
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6
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Qasim R, Thiab TA, Alhindi T, Al-Hunaiti A, Imraish A. The Nurr1 ligand indole acetic acid hydrazide loaded onto ZnFe2O4 nanoparticles suppresses proinflammatory gene expressions in SimA9 microglial cells. Sci Rep 2024; 14:13987. [PMID: 38886466 PMCID: PMC11183088 DOI: 10.1038/s41598-024-64820-z] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/13/2024] [Accepted: 06/13/2024] [Indexed: 06/20/2024] Open
Abstract
The nuclear receptor-related factor 1 (Nurr1), an orphan nuclear receptor in microglia, has been recognized as a major player in attenuating the transcription of the pro-inflammatory genes to maintain CNS homeostasis. In this study, we investigate Nurr1 trans-repression activity by targeting this receptor with one of the indole derivatives 3-Indole acetic acid hydrazide (IAAH) loaded onto zinc iron oxide (ZnFe2O4) NPs coated with PEG. XRD, SEM, FTIR, UV-Vis spectroscopy, and DLS were used to characterize the synthesized IAAH-NPs. The anti-inflammatory properties of IAAH-NPs on LPS-stimulated SimA9 microglia were assayed by measuring pro-inflammatory cytokine gene expressions and protein levels using RT-PCR and ELISA, respectively. As a result, IAAH-NPs showed an ability to suppress pro-inflammatory genes, including IL-6, IL-1β, and TNF-α in LPS-stimulated SimA9 via targeting Nurr1. The current study suggests that ZnFe2O4 NPs as a delivery system can increase the efficiency of cellular uptake and enhance the IAAH ability to inhibit the pro-inflammatory cytokines. Collectively, we demonstrate that IAAH-NPs is a potential modulator of Nurr1 that combines nanotechnology as a delivery system to suppress neuroinflammation in CNS which opens a window for possible ambitious neuroprotective therapeutic approaches to neuro disorders.
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Affiliation(s)
- Raneen Qasim
- Department of Biological Sciences, School of Science, The University of Jordan, Queen Rania Al-Abdullah Street, Amman, 11942 Jordan
| | - Tuqa Abu Thiab
- Department of Biological Sciences, School of Science, The University of Jordan, Queen Rania Al-Abdullah Street, Amman, 11942 Jordan
| | - Tareq Alhindi
- Department of Biological Sciences, School of Science, The University of Jordan, Queen Rania Al-Abdullah Street, Amman, 11942 Jordan
| | - Afnan Al-Hunaiti
- Department of Chemistry, School of Science, The University of Jordan, Queen Rania Al-Abdullah Street, Amman, 11942 Jordan
| | - Amer Imraish
- Department of Biological Sciences, School of Science, The University of Jordan, Queen Rania Al-Abdullah Street, Amman, 11942 Jordan
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7
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Safe S. Natural products and synthetic analogs as selective orphan nuclear receptor 4A (NR4A) modulators. Histol Histopathol 2024; 39:543-556. [PMID: 38116863 PMCID: PMC11267491 DOI: 10.14670/hh-18-689] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2023]
Abstract
Although endogenous ligands for the orphan nuclear receptor 4A1 (NR4A1, Nur77), NR4A2 (Nurr1), and NR4A3 (Nor-1) have not been identified, several natural products and synthetic analogs bind NR4A members. These studies are becoming increasingly important since members of the NR4A subfamily of 3 receptors are potential drug targets for treating cancer and non-cancer endpoints and particularly those conditions associated with inflammatory diseases. Ligands that bind NR4A1, NR4A2, and NR4A3 including Cytosporone B, celastrol, bis-indole derived (CDIM) compounds, tryptophan/indolic, metabolites, prostaglandins, resveratrol, piperlongumine, fatty acids, flavonoids, alkaloids, peptides, and drug families including statins and antimalarial drugs. The structural diversity of NR4A ligands and their overlapping and unique effects on NR4A1, NR4A2, and NR4A3 suggest that NR4A ligands are selective NR4A modulators (SNR4AMs) that exhibit tissue-, structure-, and response-specific activities. The SNR4AM activities of NR4A ligands are exemplified among the Cytosporone B analogs where n-pentyl-2-[3,5-dihydroxy-2-(nonanoyl)]phenyl acetate (PDNPA) binds NR4A1, NR4A2 and NR4A3 but activates only NR4A1 and exhibits significant functional differences with other Cytosporone B analogs. The number of potential clinical applications of agents targeting NR4A is increasing and this should spur future development of SNR4AMs as therapeutics that act through NR4A1, NR4A2 and NR4A3.
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Affiliation(s)
- Stephen Safe
- Department of Veterinary Physiology and Pharmacology, Texas A&M University, College Station, TX, USA.
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8
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Rocha SM, Gustafson DL, Safe S, Tjalkens RB. Comparative safety, pharmacokinetics, and off-target assessment of 1,1-bis(3'-indolyl)-1-( p-chlorophenyl) methane in mouse and dog: implications for therapeutic development. Toxicol Res (Camb) 2024; 13:tfae059. [PMID: 38655145 PMCID: PMC11033559 DOI: 10.1093/toxres/tfae059] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/11/2023] [Revised: 03/28/2024] [Accepted: 04/03/2024] [Indexed: 04/26/2024] Open
Abstract
The modified phytochemical derivative, 1,1-bis(3'-indolyl)-1-(p-chlorophenyl) methane (C-DIM12), has been identified as a potential therapeutic platform based on its capacity to improve disease outcomes in models of neurodegeneration and cancer. However, comprehensive safety studies investigating pathology and off-target binding have not been conducted. To address this, we administered C-DIM12 orogastrically to outbred male CD-1 mice for 7 days (50 mg/kg/day, 200 mg/kg/day, and 300 mg/kg/day) and investigated changes in hematology, clinical chemistry, and whole-body tissue pathology. We also delivered a single dose of C-DIM12 (1 mg/kg, 5 mg/kg, 25 mg/kg, 100 mg/kg, 300 mg/kg, 1,000 mg/kg) orogastrically to male and female beagle dogs and investigated hematology and clinical chemistry, as well as plasma pharmacokinetics over 48-h. Consecutive in-vitro off-target binding through inhibition was performed with 10 μM C-DIM12 against 68 targets in tandem with predictive off-target structural binding capacity. These data show that the highest dose C-DIM12 administered in each species caused modest liver pathology in mouse and dog, whereas lower doses were unremarkable. Off-target screening and predictive modeling of C-DIM12 show inhibition of serine/threonine kinases, calcium signaling, G-protein coupled receptors, extracellular matrix degradation, and vascular and transcriptional regulation pathways. Collectively, these data demonstrate that low doses of C-DIM12 do not induce pathology and are capable of modulating targets relevant to neurodegeneration and cancer.
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Affiliation(s)
- Savannah M Rocha
- Department of Environmental and Radiological Health Sciences, Colorado State University, 1680 Campus Delivery Fort Collins, CO 80523, USA
| | - Daniel L Gustafson
- Department of Clinical Sciences, Colorado State University, 1678 Campus Delivery Fort Collins, CO 80523, USA
| | - Stephen Safe
- Department of Veterinary Physiology and Pharmacology, Texas A&M School of Veterinary, Medicine & Biomedical Sciences, 4466 TAMU College Station, TX 77843-4466, USA
| | - Ronald B Tjalkens
- Department of Environmental and Radiological Health Sciences, Colorado State University, 1680 Campus Delivery Fort Collins, CO 80523, USA
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9
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Upadhyay S, Hailemariam AE, Mariyam F, Hafiz Z, Martin G, Kothari J, Farkas E, Sivaram G, Bell L, Tjalkens R, Safe S. Bis-Indole Derivatives as Dual Nuclear Receptor 4A1 (NR4A1) and NR4A2 Ligands. Biomolecules 2024; 14:284. [PMID: 38540704 PMCID: PMC10967861 DOI: 10.3390/biom14030284] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/06/2024] [Revised: 02/22/2024] [Accepted: 02/23/2024] [Indexed: 06/27/2024] Open
Abstract
Bis-indole derived compounds such as 1,1-bis(3'-indolyl)-1-(3,5-disubstitutedphenyl) methane (DIM-3,5) and the corresponding 4-hydroxyl analogs (DIM8-3,5) are NR4A1 ligands that act as inverse NR4A1 agonists and are potent inhibitors of tumor growth. The high potency of several DIM-3,5 analogs (IC50 < 1 mg/kg/day), coupled with the >60% similarity of the ligand-binding domains (LBDs) of NR4A1 and NR4A2 and the pro-oncogenic activities of both receptors lead us to hypothesize that these compounds may act as dual NR4A1 and NR4A2 ligands. Using a fluorescence binding assay, it was shown that 22 synthetic DIM8-3,5 and DIM-3,5 analogs bound the LBD of NR4A1 and NR4A2 with most KD values in the low µM range. Moreover, the DIM-3,5 and DIM8-3,5 analogs also decreased NR4A1- and NR4A2-dependent transactivation in U87G glioblastoma cells transfected with GAL4-NR4A1 or GAL4-NR4A2 chimeras and a UAS-luciferase reporter gene construct. The DIM-3,5 and DIM8-3,5 analogs were cytotoxic to U87 glioblastoma and RKO colon cancer cells and the DIM-3,5 compounds were more cytotoxic than the DIM8-3,5 compounds. These studies show that both DIM-3,5 and DIM8-3,5 compounds previously identified as NR4A1 ligands bind both NR4A1 and NR4A2 and are dual NR4A1/2 ligands.
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Affiliation(s)
- Srijana Upadhyay
- Department of Veterinary Physiology, Texas A&M University, College Station, TX 77843, USA; (S.U.); (A.E.H.); (F.M.); (G.M.); (E.F.)
| | - Amanuel Esayas Hailemariam
- Department of Veterinary Physiology, Texas A&M University, College Station, TX 77843, USA; (S.U.); (A.E.H.); (F.M.); (G.M.); (E.F.)
| | - Fuada Mariyam
- Department of Veterinary Physiology, Texas A&M University, College Station, TX 77843, USA; (S.U.); (A.E.H.); (F.M.); (G.M.); (E.F.)
| | - Zahin Hafiz
- Department of Chemical Engineering, Texas A&M University, College Station, TX 77843, USA; (Z.H.); (J.K.)
| | - Gregory Martin
- Department of Veterinary Physiology, Texas A&M University, College Station, TX 77843, USA; (S.U.); (A.E.H.); (F.M.); (G.M.); (E.F.)
| | - Jainish Kothari
- Department of Chemical Engineering, Texas A&M University, College Station, TX 77843, USA; (Z.H.); (J.K.)
| | - Evan Farkas
- Department of Veterinary Physiology, Texas A&M University, College Station, TX 77843, USA; (S.U.); (A.E.H.); (F.M.); (G.M.); (E.F.)
| | - Gargi Sivaram
- Department of Biochemistry and Biophysics, Texas A&M University, College Station, TX 77843, USA;
| | - Logan Bell
- Department of Chemistry, University of La Verne, La Verne, CA 91750, USA;
| | - Ronald Tjalkens
- Department of Environmental and Radiological Health Sciences, College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, CO 80526, USA;
| | - Stephen Safe
- Department of Veterinary Physiology, Texas A&M University, College Station, TX 77843, USA; (S.U.); (A.E.H.); (F.M.); (G.M.); (E.F.)
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10
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Solís-Barbosa MA, Santana E, Muñoz-Torres JR, Segovia-Gamboa NC, Patiño-Martínez E, Meraz-Ríos MA, Samaniego R, Sánchez-Mateos P, Sánchez-Torres C. The nuclear receptor Nurr1 is preferentially expressed in human pro-inflammatory macrophages and limits their inflammatory profile. Int Immunol 2024; 36:111-128. [PMID: 38066638 DOI: 10.1093/intimm/dxad048] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/21/2023] [Accepted: 12/02/2023] [Indexed: 02/22/2024] Open
Abstract
Nurr1 is a member of the orphan nuclear receptor family NR4A (nuclear receptor subfamily 4 group A) that modulates inflammation in several cell lineages, both positively and negatively. Macrophages are key regulators of inflammatory responses, yet information about the role of Nurr1 in human macrophages is scarce. Here we examined Nurr1 expression and activity in steady state and activated human macrophages. Pro- and anti-inflammatory macrophages were generated in vitro by culture of blood monocytes with granulocyte/macrophage colony-stimulating factor (GM-CSF) and macrophage colony-stimulating factor (M-CSF), respectively. Nurr1 expression was predominant in macrophages with the pro-inflammatory phenotype. Nurr1 activation with the agonists 1,1-bis(3'-indolyl)-1-(p-chlorophenyl) methane (C-DIM12) or isoxazolo-pyridinone 7e (IP7e) did not globally modify the polarization status of pro-inflammatory macrophages, but they decreased their production of TNF, IL-1β, IL-6, IL-8, IL-12 p40, CCL2, IFN-β, and reactive oxygen species, with variable potencies. Conversely, Nurr1 deficient macrophages increased the expression of transcripts encoding inflammatory mediators, particularly that of IL6, IFNB1, and CCL2. Mechanistically, endogenous Nurr1 interacted with NF-κB p65 in basal conditions and upon lipopolysaccharide (LPS)-mediated activation. C-DIM12 stabilized those complexes in cells exposed to LPS and concurrently decreased NF-κB transcriptional activity and p65 nuclear translocation. Expression of high levels of Nurr1 was associated with a subset of dermal macrophages that display enhanced levels of TNF and lower expression of the anti-inflammatory marker CD163L1 in skin lesions from patients with bullous pemphigoid (BP), a chronic inflammatory autoimmune blistering disorder. These results suggest that Nurr1 expression is linked with the pro-inflammatory phenotype of human macrophages, both in vivo and in vitro, where it may constitute a brake to attenuate the synthesis of inflammatory mediators.
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Affiliation(s)
- Miguel A Solís-Barbosa
- Department of Molecular Biomedicine, Center for Research and Advanced Studies of the National Polytechnic Institute (CINVESTAV-IPN), 07360 Mexico City, Mexico
| | - Eduardo Santana
- Department of Molecular Biomedicine, Center for Research and Advanced Studies of the National Polytechnic Institute (CINVESTAV-IPN), 07360 Mexico City, Mexico
| | - José R Muñoz-Torres
- Department of Molecular Biomedicine, Center for Research and Advanced Studies of the National Polytechnic Institute (CINVESTAV-IPN), 07360 Mexico City, Mexico
| | - Norma C Segovia-Gamboa
- Department of Molecular Biomedicine, Center for Research and Advanced Studies of the National Polytechnic Institute (CINVESTAV-IPN), 07360 Mexico City, Mexico
| | - Eduardo Patiño-Martínez
- Department of Molecular Biomedicine, Center for Research and Advanced Studies of the National Polytechnic Institute (CINVESTAV-IPN), 07360 Mexico City, Mexico
| | - Marco A Meraz-Ríos
- Department of Molecular Biomedicine, Center for Research and Advanced Studies of the National Polytechnic Institute (CINVESTAV-IPN), 07360 Mexico City, Mexico
| | - Rafael Samaniego
- Confocal Microscopy Unit, Instituto de Investigación Sanitaria Gregorio Marañón (IiSGM), 28007 Madrid, Spain
| | - Paloma Sánchez-Mateos
- Immuno-Oncology Laboratory, Instituto de Investigación Sanitaria Gregorio Marañón (IiSGM), 28007 Madrid, Spain
| | - Carmen Sánchez-Torres
- Department of Molecular Biomedicine, Center for Research and Advanced Studies of the National Polytechnic Institute (CINVESTAV-IPN), 07360 Mexico City, Mexico
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11
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Nassar A, Kodi T, Satarker S, Gurram PC, Fayaz SM, Nampoothiri M. Astrocytic transcription factors REST, YY1, and putative microRNAs in Parkinson's disease and advanced therapeutic strategies. Gene 2024; 892:147898. [PMID: 37832803 DOI: 10.1016/j.gene.2023.147898] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/13/2023] [Revised: 07/10/2023] [Accepted: 10/10/2023] [Indexed: 10/15/2023]
Abstract
Transcription factors (TF) and microRNAs are regulatory factors in astrocytes and are linked to several Parkinson's disease (PD) progression causes, such as disruption of glutamine transporters in astrocytes and concomitant disrupted glutamine uptake and inflammation. REST, a crucial TF, has been documented as an epigenetic repressor that limits the expression of neuronal genes in non-neural cells. REST activity is significantly linked to its corepressors in astrocytes, specifically histone deacetylases (HDACs), CoREST, and MECP2. Another REST-regulating TF, YY1, has been studied in astrocytes, and its interaction with REST has been investigated. In this review, the molecular processes that support the astrocytic control of REST and YY1 in terms of the regulation of glutamate transporter EAAT2 were addressed in a more detailed and comprehensive manner. Both TFs' function in astrocytes and how astrocyte abnormalities cause PD is still a mystery. Moreover, microRNAs (short non-coding RNAs) are key regulators that have been correlated to the expression and regulation of numerous genes linked to PD. The identification of numerous miRs that are engaged in astrocyte dysfunction that triggers PD has been shown. The term "Gut-brain axis" refers to the two systems' mutual communication. Gut microbial dysbiosis, which mediates an imbalance of the gut-brain axis, might contribute to neurodegenerative illnesses through altered astrocytic regulation. New treatment approaches to modify the gut-brain axis and prevent astrocytic repercussions have also been investigated in this review.
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Affiliation(s)
- Ajmal Nassar
- Department of Pharmacology, Manipal College of Pharmaceutical Sciences, Manipal Academy of Higher Education, Manipal 576104, Karnataka, India.
| | - Triveni Kodi
- Department of Pharmacology, Manipal College of Pharmaceutical Sciences, Manipal Academy of Higher Education, Manipal 576104, Karnataka, India.
| | - Sairaj Satarker
- Department of Pharmacology, Manipal College of Pharmaceutical Sciences, Manipal Academy of Higher Education, Manipal 576104, Karnataka, India.
| | - Prasada Chowdari Gurram
- Department of Pharmacology, Manipal College of Pharmaceutical Sciences, Manipal Academy of Higher Education, Manipal 576104, Karnataka, India.
| | - S M Fayaz
- Department of Biotechnology, Manipal Institute of Technology, Manipal Academy of Higher Education, Manipal 576104, Karnataka, India.
| | - Madhavan Nampoothiri
- Department of Pharmacology, Manipal College of Pharmaceutical Sciences, Manipal Academy of Higher Education, Manipal 576104, Karnataka, India.
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12
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Rocha SM, Kirkley KS, Chatterjee D, Aboellail TA, Smeyne RJ, Tjalkens RB. Microglia-specific knock-out of NF-κB/IKK2 increases the accumulation of misfolded α-synuclein through the inhibition of p62/sequestosome-1-dependent autophagy in the rotenone model of Parkinson's disease. Glia 2023; 71:2154-2179. [PMID: 37199240 PMCID: PMC10330367 DOI: 10.1002/glia.24385] [Citation(s) in RCA: 16] [Impact Index Per Article: 8.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/09/2022] [Revised: 05/02/2023] [Accepted: 05/05/2023] [Indexed: 05/19/2023]
Abstract
Parkinson's disease (PD) is the most common neurodegenerative movement disorder worldwide, with a greater prevalence in men than women. The etiology of PD is largely unknown, although environmental exposures and neuroinflammation are linked to protein misfolding and disease progression. Activated microglia are known to promote neuroinflammation in PD, but how environmental agents interact with specific innate immune signaling pathways in microglia to stimulate conversion to a neurotoxic phenotype is not well understood. To determine how nuclear factor kappa B (NF-κB) signaling dynamics in microglia modulate neuroinflammation and dopaminergic neurodegeneration, we generated mice deficient in NF-κB activation in microglia (CX3CR1-Cre::IKK2fl/fl ) and exposed them to 2.5 mg/kg/day of rotenone for 14 days, followed by a 14-day post-lesioning incubation period. We postulated that inhibition of NF-κB signaling in microglia would reduce overall inflammatory injury in lesioned mice. Subsequent analysis indicated decreased expression of the NF-κB-regulated autophagy gene, sequestosome 1 (p62), in microglia, which is required for targeting ubiquitinated α-synuclein (α-syn) for lysosomal degradation. Knock-out animals had increased accumulation of misfolded α-syn within microglia, despite an overall reduction in neurodegeneration. Interestingly, this occurred more prominently in males. These data suggest that microglia play key biological roles in the degradation and clearance of misfolded α-syn and this process works in concert with the innate immune response associated with neuroinflammation. Importantly, the accumulation of misfolded α-syn protein aggregates alone did not increase neurodegeneration following exposure to rotenone but required the NF-κB-dependent inflammatory response in microglia.
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Affiliation(s)
- Savannah M. Rocha
- Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, CO 80523
- Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, CO 80523
| | - Kelly S. Kirkley
- Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, CO 80523
| | - Debotri Chatterjee
- Jefferson Comprehensive Parkinson’s Center, Vickie & Jack Farber Institute for Neuroscience, Thomas Jefferson University, Philadelphia, PA 19107
| | - Tawfik A. Aboellail
- Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, CO 80523
| | - Richard J. Smeyne
- Jefferson Comprehensive Parkinson’s Center, Vickie & Jack Farber Institute for Neuroscience, Thomas Jefferson University, Philadelphia, PA 19107
| | - Ronald B. Tjalkens
- Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, CO 80523
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13
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Systems level analysis of sex-dependent gene expression changes in Parkinson's disease. NPJ Parkinsons Dis 2023; 9:8. [PMID: 36681675 PMCID: PMC9867746 DOI: 10.1038/s41531-023-00446-8] [Citation(s) in RCA: 15] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/10/2022] [Accepted: 01/03/2023] [Indexed: 01/22/2023] Open
Abstract
Parkinson's disease (PD) is a heterogeneous disorder, and among the factors which influence the symptom profile, biological sex has been reported to play a significant role. While males have a higher age-adjusted disease incidence and are more frequently affected by muscle rigidity, females present more often with disabling tremors. The molecular mechanisms involved in these differences are still largely unknown, and an improved understanding of the relevant factors may open new avenues for pharmacological disease modification. To help address this challenge, we conducted a meta-analysis of disease-associated molecular sex differences in brain transcriptomics data from case/control studies. Both sex-specific (alteration in only one sex) and sex-dimorphic changes (changes in both sexes, but with opposite direction) were identified. Using further systems level pathway and network analyses, coordinated sex-related alterations were studied. These analyses revealed significant disease-associated sex differences in mitochondrial pathways and highlight specific regulatory factors whose activity changes can explain downstream network alterations, propagated through gene regulatory cascades. Single-cell expression data analyses confirmed the main pathway-level changes observed in bulk transcriptomics data. Overall, our analyses revealed significant sex disparities in PD-associated transcriptomic changes, resulting in coordinated modulations of molecular processes. Among the regulatory factors involved, NR4A2 has already been reported to harbor rare mutations in familial PD and its pharmacological activation confers neuroprotective effects in toxin-induced models of Parkinsonism. Our observations suggest that NR4A2 may warrant further research as a potential adjuvant therapeutic target to address a subset of pathological molecular features of PD that display sex-associated profiles.
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A Nurr1 ligand C-DIM12 attenuates brain inflammation and improves functional recovery after intracerebral hemorrhage in mice. Sci Rep 2022; 12:11009. [PMID: 35773404 PMCID: PMC9246855 DOI: 10.1038/s41598-022-15178-7] [Citation(s) in RCA: 7] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/29/2022] [Accepted: 06/20/2022] [Indexed: 11/08/2022] Open
Abstract
We have previously reported that amodiaquine, a compound that binds to the ligand-binding domain of a nuclear receptor Nurr1, attenuates inflammatory responses and neurological deficits after intracerebral hemorrhage (ICH) in mice. 1,1-Bis(3'-indolyl)-1-(p-chlorophenyl)methane (C-DIM12) is another Nurr1 ligand that recognizes a domain of Nurr1 different from the ligand-binding domain. In the present study, mice were treated daily with C-DIM12 (50 or 100 mg/kg, p.o.) or amodiaquine (40 mg/kg, i.p.), or twice daily with 1400 W (20 mg/kg, i.p.), an inducible nitric oxide synthase (iNOS) inhibitor, from 3 h after ICH induction by microinjection of collagenase into the striatum. C-DIM12 improved the recovery of neurological function and prevented neuron loss in the hematoma, while suppressed activation of microglia/macrophages and expression of inflammatory mediators interleukin-6 and CC chemokine ligand 2. In addition, C-DIM12 as well as amodiaquine preserved axonal structures in the internal capsule and axonal transport function. We also found that C-DIM12 and amodiaquine suppressed the increases of iNOS mRNA expression after ICH. Moreover, 1400 W improved neurological function and prevented neuron loss, activation of microglia/macrophages and axonal transport dysfunction. These results suggest that suppression of iNOS induction contributes to several features of the therapeutic effects of Nurr1 ligands.
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15
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Li L, Ho PWL, Liu H, Pang SYY, Chang EES, Choi ZYK, Malki Y, Kung MHW, Ramsden DB, Ho SL. Transcriptional Regulation of the Synaptic Vesicle Protein Synaptogyrin-3 (SYNGR3) Gene: The Effects of NURR1 on Its Expression. Int J Mol Sci 2022; 23:ijms23073646. [PMID: 35409005 PMCID: PMC8998927 DOI: 10.3390/ijms23073646] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/07/2022] [Revised: 03/24/2022] [Accepted: 03/24/2022] [Indexed: 11/24/2022] Open
Abstract
Synaptogyrin-3 (SYNGR3) is a synaptic vesicular membrane protein. Amongst four homologues (SYNGR1 to 4), SYNGR1 and 3 are especially abundant in the brain. SYNGR3 interacts with the dopamine transporter (DAT) to facilitate dopamine (DA) uptake and synaptic DA turnover in dopaminergic transmission. Perturbed SYNGR3 expression is observed in Parkinson’s disease (PD). The regulatory elements which affect SYNGR3 expression are unknown. Nuclear-receptor-related-1 protein (NURR1) can regulate dopaminergic neuronal differentiation and maintenance via binding to NGFI-B response elements (NBRE). We explored whether NURR1 can regulate SYNGR3 expression using an in silico analysis of the 5′-flanking region of the human SYNGR3 gene, reporter gene activity and an electrophoretic mobility shift assay (EMSA) of potential cis-acting sites. In silico analysis of two genomic DNA segments (1870 bp 5′-flanking region and 1870 + 159 bp of first exon) revealed one X Core Promoter Element 1 (XCPE1), two SP1, and three potential non-canonical NBRE response elements (ncNBRE) but no CAAT or TATA box. The longer segment exhibited gene promoter activity in luciferase reporter assays. Site-directed mutagenesis of XCPE1 decreased promoter activity in human neuroblastoma SH-SY5Y (↓43.2%) and human embryonic kidney HEK293 cells (↓39.7%). EMSA demonstrated NURR1 binding to these three ncNBRE. Site-directed mutagenesis of these ncNBRE reduced promoter activity by 11–17% in SH-SY5Y (neuronal) but not in HEK293 (non-neuronal) cells. C-DIM12 (Nurr1 activator) increased SYNGR3 protein expression in SH-SY5Y cells and its promoter activity using a real-time luciferase assay. As perturbed vesicular function is a feature of major neurodegenerative diseases, inducing SYNGR3 expression by NURR1 activators may be a potential therapeutic target to attenuate synaptic dysfunction in PD.
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Affiliation(s)
- Lingfei Li
- Division of Neurology, Department of Medicine, University of Hong Kong, Hong Kong SAR, China; (L.L.); (P.W.-L.H.); (H.L.); (S.Y.-Y.P.); (E.E.-S.C.); (Z.Y.-K.C.); (Y.M.); (M.H.-W.K.)
| | - Philip Wing-Lok Ho
- Division of Neurology, Department of Medicine, University of Hong Kong, Hong Kong SAR, China; (L.L.); (P.W.-L.H.); (H.L.); (S.Y.-Y.P.); (E.E.-S.C.); (Z.Y.-K.C.); (Y.M.); (M.H.-W.K.)
| | - Huifang Liu
- Division of Neurology, Department of Medicine, University of Hong Kong, Hong Kong SAR, China; (L.L.); (P.W.-L.H.); (H.L.); (S.Y.-Y.P.); (E.E.-S.C.); (Z.Y.-K.C.); (Y.M.); (M.H.-W.K.)
| | - Shirley Yin-Yu Pang
- Division of Neurology, Department of Medicine, University of Hong Kong, Hong Kong SAR, China; (L.L.); (P.W.-L.H.); (H.L.); (S.Y.-Y.P.); (E.E.-S.C.); (Z.Y.-K.C.); (Y.M.); (M.H.-W.K.)
| | - Eunice Eun-Seo Chang
- Division of Neurology, Department of Medicine, University of Hong Kong, Hong Kong SAR, China; (L.L.); (P.W.-L.H.); (H.L.); (S.Y.-Y.P.); (E.E.-S.C.); (Z.Y.-K.C.); (Y.M.); (M.H.-W.K.)
| | - Zoe Yuen-Kiu Choi
- Division of Neurology, Department of Medicine, University of Hong Kong, Hong Kong SAR, China; (L.L.); (P.W.-L.H.); (H.L.); (S.Y.-Y.P.); (E.E.-S.C.); (Z.Y.-K.C.); (Y.M.); (M.H.-W.K.)
| | - Yasine Malki
- Division of Neurology, Department of Medicine, University of Hong Kong, Hong Kong SAR, China; (L.L.); (P.W.-L.H.); (H.L.); (S.Y.-Y.P.); (E.E.-S.C.); (Z.Y.-K.C.); (Y.M.); (M.H.-W.K.)
| | - Michelle Hiu-Wai Kung
- Division of Neurology, Department of Medicine, University of Hong Kong, Hong Kong SAR, China; (L.L.); (P.W.-L.H.); (H.L.); (S.Y.-Y.P.); (E.E.-S.C.); (Z.Y.-K.C.); (Y.M.); (M.H.-W.K.)
| | - David Boyer Ramsden
- Institute of Metabolism and Systems Research, University of Birmingham, Birmingham B15 2TT, UK
- Correspondence: (D.B.R.); (S.-L.H.)
| | - Shu-Leong Ho
- Division of Neurology, Department of Medicine, University of Hong Kong, Hong Kong SAR, China; (L.L.); (P.W.-L.H.); (H.L.); (S.Y.-Y.P.); (E.E.-S.C.); (Z.Y.-K.C.); (Y.M.); (M.H.-W.K.)
- Correspondence: (D.B.R.); (S.-L.H.)
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16
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Exosomes isolated during dopaminergic neuron differentiation suppressed neuronal inflammation in a rodent model of Parkinson's disease. Neurosci Lett 2021; 771:136414. [PMID: 34954117 DOI: 10.1016/j.neulet.2021.136414] [Citation(s) in RCA: 18] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/11/2021] [Revised: 12/15/2021] [Accepted: 12/20/2021] [Indexed: 02/08/2023]
Abstract
Our previous investigation showed Wnt signal pathway was significantly activated during DA neuron differentiation of epiblast-derived stem cells. In this study, we next attempt to examine the therapeutic potential of the purified exosomes derived bone marrow mesenchymal stem cells (BMSCs) by administrating exosomes into the rat striatum of parkinson's disease (PD) animal model. Results revealed that the protein levels of interleukin (IL)-6, IL-1β, tumor necrosis factor-alpha (TNF-α), and reactive oxygen species (ROS) in the substantia nigra of PD rats were down regulated after injection of BMSC induced-Exosomes into the striatum of PD model compared to BMSC quiescent-Exosomes. In addition, the expression of ionized calcium binding adaptor molecule 1 (Iba1) mRNA was significantly decreased, while the expression of tyrosine hydroxylase (TH) mRNA was increased after injection of BMSC induced-Exosomes. Injection of BMSC induced-Exosomes into the striatum rescued the rotation behavior and climbing speed in the PD rats. More importantly, Wnt5a was found to be enriched in BMSC induced Exosomes, which could be effectively transferred to the substantia nigra of PD rats. In conclusion, these findings demonstrated that exosomes isolated during dopaminergic neuron differentiation could rescue the pathogenic features of Parkinson's disease by reshaping the inflammatory microenvironment in the substantia nigra and repairing the injury to DA nerves.
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17
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Bantle CM, Rocha SM, French CT, Phillips AT, Tran K, Olson KE, Bass TA, Aboellail T, Smeyne RJ, Tjalkens RB. Astrocyte inflammatory signaling mediates α-synuclein aggregation and dopaminergic neuronal loss following viral encephalitis. Exp Neurol 2021; 346:113845. [PMID: 34454938 PMCID: PMC9535678 DOI: 10.1016/j.expneurol.2021.113845] [Citation(s) in RCA: 13] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/14/2021] [Revised: 08/13/2021] [Accepted: 08/23/2021] [Indexed: 12/13/2022]
Abstract
Viral infection of the central nervous system (CNS) can cause lasting neurological decline in surviving patients and can present with symptoms resembling Parkinson's disease (PD). The mechanisms underlying postencephalitic parkinsonism remain unclear but are thought to involve increased innate inflammatory signaling in glial cells, resulting in persistent neuroinflammation. We therefore studied the role of glial cells in regulating neuropathology in postencephalitic parkinsonism by studying the involvement of astrocytes in loss of dopaminergic neurons and aggregation of α-synuclein protein following infection with western equine encephalitis virus (WEEV). Infections were conducted in both wildtype mice and in transgenic mice lacking NFκB inflammatory signaling in astrocytes. For 2 months following WEEV infection, we analyzed glial activation, neuronal loss and protein aggregation across multiple brain regions, including the substantia nigra pars compacta (SNpc). These data revealed that WEEV induces loss of SNpc dopaminergic neurons, persistent activation of microglia and astrocytes that precipitates widespread aggregation of α-synuclein in the brain of C57BL/6 mice. Microgliosis and macrophage infiltration occurred prior to activation of astrocytes and was followed by opsonization of ⍺-synuclein protein aggregates in the cortex, hippocampus and midbrain by the complement protein, C3. Astrocyte-specific NFκB knockout mice had reduced gliosis, α-synuclein aggregate formation and neuronal loss. These data suggest that astrocytes play a critical role in initiating PD-like pathology following encephalitic infection with WEEV through innate immune inflammatory pathways that damage dopaminergic neurons, possibly by hindering clearance of ⍺-synuclein aggregates. Inhibiting glial inflammatory responses could therefore represent a potential therapy strategy for viral parkinsonism.
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Affiliation(s)
- Collin M Bantle
- Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, CO 80523, United States of America
| | - Savannah M Rocha
- Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO 80523, United States of America
| | - C Tenley French
- Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, CO 80523, United States of America
| | - Aaron T Phillips
- Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, CO 80523, United States of America; Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO 80523, United States of America
| | - Kevin Tran
- Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, CO 80523, United States of America
| | - Kenneth E Olson
- Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO 80523, United States of America
| | - Todd A Bass
- Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO 80523, United States of America
| | - Tawfik Aboellail
- Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO 80523, United States of America
| | - Richard J Smeyne
- Jefferson Comprehensive Parkinson's Center, Vickie & Jack Farber Institute for Neuroscience, Thomas Jefferson University, Philadelphia, PA 19107, United States of America
| | - Ronald B Tjalkens
- Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, CO 80523, United States of America.
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18
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Katsuki H. Nuclear receptors of NR1 and NR4 subfamilies in the regulation of microglial functions and pathology. Pharmacol Res Perspect 2021; 9:e00766. [PMID: 34676987 PMCID: PMC8532137 DOI: 10.1002/prp2.766] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/22/2021] [Revised: 03/01/2021] [Accepted: 03/15/2021] [Indexed: 12/15/2022] Open
Abstract
This review provides an overview of researches on the NR1 and NR4 nuclear receptors involved in the regulation of microglial functions. Nuclear receptors are attractive candidates for drug targets in the therapies of the central nervous system disorders, because the activation of these receptors is expected to regulate the functions and the phenotypes of microglia, by controlling the expression of specific gene subsets and also by regulating the cellular signaling mechanisms in a nongenomic manner. Several members of NR1 nuclear receptor subfamily have been examined for their ability to regulate microglial functions. For example, stimulation of vitamin D receptor inhibits the production of pro-inflammatory factors and increases the production of anti-inflammatory cytokines. Similar regulatory actions of nuclear receptor ligands on inflammation-related genes have also been reported for other NR1 members such as retinoic acid receptors, peroxisome proliferator-activated receptors (PPARs), and liver X receptors (LXRs). In addition, stimulation of PPARγ and LXRs may also result in increased phagocytic activities of microglia. Consistent with these actions, the agonists at nuclear receptors of NR1 subfamily are shown to produce therapeutic effects on animal models of various neurological disorders such as experimental allergic encephalomyelitis, Alzheimer's disease, Parkinson's disease, and ischemic/hemorrhagic stroke. On the other hand, increasing lines of evidence suggest that the stimulation of NR4 subfamily members of nuclear receptors such as Nur77 and Nurr1 also regulates microglial functions and alleviates neuropathological events in several disease models. Further advancement of these research fields may prove novel therapeutic opportunities.
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Affiliation(s)
- Hiroshi Katsuki
- Department of Chemico‐Pharmacological SciencesGraduate School of Pharmaceutical SciencesKumamoto UniversityKumamotoJapan
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A Low-Cost, Autonomous Gait Detection and Estimation System for Analyzing Gait Impairments in Mice. JOURNAL OF HEALTHCARE ENGINEERING 2021; 2021:9937904. [PMID: 34804462 PMCID: PMC8604610 DOI: 10.1155/2021/9937904] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 03/12/2021] [Revised: 09/03/2021] [Accepted: 10/27/2021] [Indexed: 11/18/2022]
Abstract
With the advancement in imaging technology, many commercial systems have been developed for performing motion analysis in mice. However, available commercial systems are expensive and use proprietary software. In this paper, we describe a low-cost, camera-based design of an autonomous gait acquisition and analysis system for inspecting gait deficits in C57BL/6 mice. Our system includes video acquisition, autonomous gait-event detection, gait-parameter extraction, and result visualization. We provide a simple, user-friendly, step-by-step detailed methodology to apply well-known image processing techniques for detecting mice footfalls and calculating various gait parameters for analyzing gait abnormalities in healthy and neurotraumatic mice. The system was used in a live animal study for assessing recovery in a mouse model of Parkinson's disease. Using the videos acquired in the study, we validate the performance of our system with receiver operating characteristic (ROC) and Hit : Miss : False (H : M : F) detection analyses. Our system correctly detected the mice footfalls with an average H : M : F score of 92.1 : 2.3 : 5.6. The values for the area under an ROC curve for all the ROC plots are above 0.95, which indicates an almost perfect detection model. The ROC and H : M : F analyses show that our system produces accurate gait detection. The results observed from the gait assessment study are in agreement with the known literature. This demonstrates the practical viability of our system as a gait analysis tool.
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20
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Gagnon J, Caron V, Gyenizse L, Tremblay A. Atypic SUMOylation of Nor1/NR4A3 regulates neural cell viability and redox sensitivity. FASEB J 2021; 35:e21827. [PMID: 34383980 DOI: 10.1096/fj.202100395r] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/05/2021] [Revised: 06/30/2021] [Accepted: 07/15/2021] [Indexed: 11/11/2022]
Abstract
Neuron-derived orphan receptor 1, NR4A3 (Nor1)/NR4A3 is an orphan nuclear receptor involved in the transcriptional control of developmental and neurological functions. Oxidative stress-induced conditions are primarily associated with neurological defects in humans, yet the impact on Nor1-mediated transcription of neuronal genes remains with unknown mechanism. Here, we demonstrate that Nor1 is a non-conventional target of SUMO2/3 conjugation at Lys-137 contained in an atypic ψKxSP motif referred to as the pSuM. Nor1 pSuM SUMOylation differs from the canonical process with the obligate phosphorylation of Ser-139 by Ras signaling to create the required negatively charged interface for SUMOylation. Additional phosphorylation at sites flanking the pSuM is also mediated by the coordinated action of protein kinase casein kinase 2 to function as a small ubiquitin-like modifier enhancer, regulating Nor1-mediated transcription and proteasomal degradation. Nor1 responsive genes involved in cell proliferation and metabolism, such as activating transcription factor 3, cyclin D1, CASP8 and FADD-like apoptosis regulator, and enolase 3 were upregulated in response to pSuM disruption in mouse HT-22 hippocampal neuronal cells and human neuroblastoma SH-SY5Y cells. We also identified critical antioxidant genes, such as catalase, superoxide dismutase 1, and microsomal glutathione S-transferase 2, as responsive targets of Nor1 under pSuM regulation. Nor1 SUMOylation impaired gene transcription through less effective Nor1 chromatin binding and reduced enrichment of histone H3K27ac marks to gene promoters. These effects resulted in decreased neuronal cell growth, increased apoptosis, and reduced survival to oxidative stress damage, underlying the role of pSuM-modified Nor1 in redox homeostasis. Our findings uncover a hierarchical post-translational mechanism that dictates Nor1 non-canonical SUMOylation, disrupting Nor1 transcriptional competence, and neuroprotective redox sensitivity.
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Affiliation(s)
- Jonathan Gagnon
- Research Center, CHU Sainte-Justine, Montréal, Québec, Canada
- Department of Biochemistry and Molecular Medicine, Faculty of Medicine, University of Montreal, Montréal, Québec, Canada
| | - Véronique Caron
- Research Center, CHU Sainte-Justine, Montréal, Québec, Canada
| | - Laurent Gyenizse
- Research Center, CHU Sainte-Justine, Montréal, Québec, Canada
- Department of Biochemistry and Molecular Medicine, Faculty of Medicine, University of Montreal, Montréal, Québec, Canada
| | - André Tremblay
- Research Center, CHU Sainte-Justine, Montréal, Québec, Canada
- Department of Biochemistry and Molecular Medicine, Faculty of Medicine, University of Montreal, Montréal, Québec, Canada
- Centre de Recherche en Reproduction et Fertilité, University of Montreal, Saint-Hyacinthe, Québec, Canada
- Department of Obstetrics & Gynecology, Faculty of Medicine, University of Montreal, Montréal, Québec, Canada
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21
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Hammond SL, Bantle CM, Popichak KA, Wright KA, Thompson D, Forero C, Kirkley KS, Damale PU, Chong EKP, Tjalkens RB. NF-κB Signaling in Astrocytes Modulates Brain Inflammation and Neuronal Injury Following Sequential Exposure to Manganese and MPTP During Development and Aging. Toxicol Sci 2021; 177:506-520. [PMID: 32692843 DOI: 10.1093/toxsci/kfaa115] [Citation(s) in RCA: 28] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/01/2023] Open
Abstract
Chronic exposure to manganese (Mn) is associated with neuroinflammation and extrapyramidal motor deficits resembling features of Parkinson's disease. Activation of astrocytes and microglia is implicated in neuronal injury from Mn but it is not known whether early life exposure to Mn may predispose glia to more severe inflammatory responses during aging. We therefore examined astrocyte nuclear factor kappa B (NF-κB) signaling in mediating innate immune inflammatory responses during multiple neurotoxic exposures spanning juvenile development into adulthood. MnCl2 was given in drinking water for 30-day postweaning to both wildtype mice and astrocyte-specific knockout (KO) mice lacking I kappa B kinase 2, the central upstream activator of NF-κB. Following juvenile exposure to Mn, mice were subsequently administered 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) at 4 months of age. Animals were evaluated for behavioral alterations and brain tissue was analyzed for catecholamine neurotransmitters. Stereological analysis of neuronal and glial cell counts from multiple brain regions indicated that juvenile exposure to Mn amplified glial activation and neuronal loss from MPTP exposure in the caudate-putamen and globus pallidus, as well as increased the severity of neurobehavioral deficits in open field activity assays. These alterations were prevented in astrocyte-specific I kappa B kinase 2 KO mice. Juvenile exposure to Mn increased the number of neurotoxic A1 astrocytes expressing C3 as well as the number of activated microglia in adult mice following MPTP challenge, both of which were inhibited in KO mice. These results demonstrate that exposure to Mn during juvenile development heightens the innate immune inflammatory response in glia during a subsequent neurotoxic challenge through NF-κB signaling in astrocytes.
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Affiliation(s)
- Sean L Hammond
- Toxicology Program, Department of Environmental and Radiological Health Sciences, College of Veterinary Medicine and Biomedical Sciences
| | - Collin M Bantle
- Toxicology Program, Department of Environmental and Radiological Health Sciences, College of Veterinary Medicine and Biomedical Sciences
| | - Katriana A Popichak
- Toxicology Program, Department of Environmental and Radiological Health Sciences, College of Veterinary Medicine and Biomedical Sciences
| | - Katie A Wright
- Toxicology Program, Department of Environmental and Radiological Health Sciences, College of Veterinary Medicine and Biomedical Sciences
| | - Delaney Thompson
- Toxicology Program, Department of Environmental and Radiological Health Sciences, College of Veterinary Medicine and Biomedical Sciences
| | - Catalina Forero
- Toxicology Program, Department of Environmental and Radiological Health Sciences, College of Veterinary Medicine and Biomedical Sciences
| | - Kelly S Kirkley
- Toxicology Program, Department of Environmental and Radiological Health Sciences, College of Veterinary Medicine and Biomedical Sciences
| | - Pranav U Damale
- Department of Electrical and Computer Engineering, College of Engineering, Colorado State University, Fort Collins, Colorado 80523-1680
| | - Edwin K P Chong
- Department of Electrical and Computer Engineering, College of Engineering, Colorado State University, Fort Collins, Colorado 80523-1680
| | - Ronald B Tjalkens
- Toxicology Program, Department of Environmental and Radiological Health Sciences, College of Veterinary Medicine and Biomedical Sciences
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Kholodar SA, Lang G, Cortopassi WA, Iizuka Y, Brah HS, Jacobson MP, England PM. Analogs of the Dopamine Metabolite 5,6-Dihydroxyindole Bind Directly to and Activate the Nuclear Receptor Nurr1. ACS Chem Biol 2021; 16:1159-1163. [PMID: 34165961 DOI: 10.1021/acschembio.1c00326] [Citation(s) in RCA: 12] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022]
Abstract
The nuclear receptor-related 1 protein, Nurr1, is a transcription factor critical for the development and maintenance of dopamine-producing neurons in the substantia nigra pars compacta, a cell population that progressively loses the ability to make dopamine and degenerates in Parkinson's disease. Recently, we demonstrated that Nurr1 binds directly to and is regulated by the endogenous dopamine metabolite 5,6-dihydroxyindole (DHI). Unfortunately, DHI is an unstable compound, and thus a poor tool for studying Nurr1 function. Here, we report that 5-chloroindole, an unreactive analog of DHI, binds directly to the Nurr1 ligand binding domain with micromolar affinity and stimulates the activity of Nurr1, including the transcription of genes governing the synthesis and packaging of dopamine.
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Willems S, Zaienne D, Merk D. Targeting Nuclear Receptors in Neurodegeneration and Neuroinflammation. J Med Chem 2021; 64:9592-9638. [PMID: 34251209 DOI: 10.1021/acs.jmedchem.1c00186] [Citation(s) in RCA: 39] [Impact Index Per Article: 9.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Nuclear receptors, also known as ligand-activated transcription factors, regulate gene expression upon ligand signals and present as attractive therapeutic targets especially in chronic diseases. Despite the therapeutic relevance of some nuclear receptors in various pathologies, their potential in neurodegeneration and neuroinflammation is insufficiently established. This perspective gathers preclinical and clinical data for a potential role of individual nuclear receptors as future targets in Alzheimer's disease, Parkinson's disease, and multiple sclerosis, and concomitantly evaluates the level of medicinal chemistry targeting these proteins. Considerable evidence suggests the high promise of ligand-activated transcription factors to counteract neurodegenerative diseases with a particularly high potential of several orphan nuclear receptors. However, potent tools are lacking for orphan receptors, and limited central nervous system exposure or insufficient selectivity also compromises the suitability of well-studied nuclear receptor ligands for functional studies. Medicinal chemistry efforts are needed to develop dedicated high-quality tool compounds for the therapeutic validation of nuclear receptors in neurodegenerative pathologies.
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Affiliation(s)
- Sabine Willems
- Institute of Pharmaceutical Chemistry, Goethe University Frankfurt, Max-von-Laue-Strasse 9, 60438 Frankfurt, Germany
| | - Daniel Zaienne
- Institute of Pharmaceutical Chemistry, Goethe University Frankfurt, Max-von-Laue-Strasse 9, 60438 Frankfurt, Germany
| | - Daniel Merk
- Institute of Pharmaceutical Chemistry, Goethe University Frankfurt, Max-von-Laue-Strasse 9, 60438 Frankfurt, Germany
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de Souza FRO, Ribeiro FM, Lima PMD. Implications of VIP and PACAP in Parkinson's Disease: What do we Know So Far? Curr Med Chem 2021; 28:1703-1715. [PMID: 32196442 DOI: 10.2174/0929867327666200320162436] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/30/2019] [Revised: 01/30/2020] [Accepted: 02/03/2020] [Indexed: 11/22/2022]
Abstract
BACKGROUND Parkinson's disease is one of the most common neurodegenerative disorders and although its aetiology is not yet fully understood, neuroinflammation has been identified as a key factor in the progression of the disease. Vasoactive intestinal peptide and pituitary adenylate-cyclase activating polypeptide are two neuropeptides that exhibit anti-inflammatory and neuroprotective properties, modulating the production of cytokines and chemokines and the behaviour of immune cells. However, the role of chemokines and cytokines modulated by the endogenous receptors of the peptides varies according to the stage of the disease. METHODS We present an overview of the relationship between some cytokines and chemokines with vasoactive intestinal peptide, pituitary adenylate cyclase activating polypeptide and their endogenous receptors in the context of Parkinson's disease neuroinflammation and oxidative stress, as well as the modulation of microglial cells by the peptides in this context. RESULTS The two peptides exhibit neuroprotective and anti-inflammatory properties in models of Parkinson's disease, as they ameliorate cognitive functions, decrease the level of neuroinflammation and promote dopaminergic neuronal survival. The peptides have been tested in a variety of in vivo and in vitro models of Parkinson's disease, demonstrating the potential for therapeutic application. CONCLUSION More studies are needed to establish the clinical use of vasoactive intestinal peptide and pituitary adenylate cyclase activating polypeptide as safe candidates for treating Parkinson's disease, as the use of the peptides in different stages of the disease could produce different results concerning effectiveness.
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Affiliation(s)
- Filipe Resende Oliveira de Souza
- Laboratory of Immunology and Microbiology, Department of Natural Sciences, Federal University of Sao Joao Del Rei, Praca Dom Helvecio, n. 74, Fabricas, 36301160, Sao Joao Del Rei, MG, Brazil
| | - Fabiola Mara Ribeiro
- Laboratory of Neurobiochemistry, Department of Biochemistry and Immunology, Federal University of Minas Gerais, MG, Brazil
| | - Patrícia Maria d'Almeida Lima
- Laboratory of Immunology and Microbiology, Department of Natural Sciences, Federal University of Sao Joao Del Rei, Praca Dom Helvecio, n. 74, Fabricas, 36301160, Sao Joao Del Rei, MG, Brazil
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25
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Safe S, Karki K. The Paradoxical Roles of Orphan Nuclear Receptor 4A (NR4A) in Cancer. Mol Cancer Res 2021; 19:180-191. [PMID: 33106376 PMCID: PMC7864866 DOI: 10.1158/1541-7786.mcr-20-0707] [Citation(s) in RCA: 78] [Impact Index Per Article: 19.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/13/2020] [Revised: 09/22/2020] [Accepted: 10/19/2020] [Indexed: 11/16/2022]
Abstract
The three-orphan nuclear receptor 4A genes are induced by diverse stressors and stimuli, and there is increasing evidence that NR4A1 (Nur77), NR4A2 (Nurr1), and NR4A3 (Nor1) play an important role in maintaining cellular homeostasis and in pathophysiology. In blood-derived tumors (leukemias and lymphomas), NR4A expression is low and NR4A1-/-/NR4A3-/- double knockout mice rapidly develop acute myelocytic leukemia, suggesting that these receptors exhibit tumor suppressor activity. Treatment of leukemia and most lymphoma cells with drugs that induce expression of NR4A1and NR4A3 enhances apoptosis, and this represents a potential clinical application for treating this disease. In contrast, most solid tumor-derived cell lines express high levels of NR4A1 and NR4A2, and both receptors exhibit pro-oncogenic activities in solid tumors, whereas NR4A3 exhibits tumor-specific activities. Initial studies with retinoids and apoptosis-inducing agents demonstrated that their cytotoxic activity is NR4A1 dependent and involved drug-induced nuclear export of NR4A1 and formation of a mitochondrial proapoptotic NR4A1-bcl-2 complex. Drug-induced nuclear export of NR4A1 has been reported for many agents/biologics and involves interactions with multiple mitochondrial and extramitochondrial factors to induce apoptosis. Synthetic ligands for NR4A1, NR4A2, and NR4A3 have been identified, and among these compounds, bis-indole derived (CDIM) NR4A1 ligands primarily act on nuclear NR4A1 to inhibit NR4A1-regulated pro-oncogenic pathways/genes and similar results have been observed for CDIMs that bind NR4A2. Based on results of laboratory animal studies development of NR4A inducers (blood-derived cancers) and NR4A1/NR4A2 antagonists (solid tumors) may be promising for cancer therapy and also for enhancing immune surveillance.
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Affiliation(s)
- Stephen Safe
- Department of Veterinary Physiology and Pharmacology, Texas A&M University, College Station, Texas.
| | - Keshav Karki
- Department of Veterinary Physiology and Pharmacology, Texas A&M University, College Station, Texas
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26
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Munoz-Tello P, Lin H, Khan P, de Vera IMS, Kamenecka TM, Kojetin DJ. Assessment of NR4A Ligands That Directly Bind and Modulate the Orphan Nuclear Receptor Nurr1. J Med Chem 2020; 63:15639-15654. [PMID: 33289551 PMCID: PMC8006468 DOI: 10.1021/acs.jmedchem.0c00894] [Citation(s) in RCA: 42] [Impact Index Per Article: 8.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/18/2022]
Abstract
Nurr1/NR4A2 is an orphan nuclear receptor transcription factor implicated as a drug target for neurological disorders including Alzheimer's and Parkinson's diseases. Previous studies identified small-molecule NR4A nuclear receptor modulators, but it remains unclear if these ligands affect transcription via direct binding to Nurr1. We assessed 12 ligands reported to affect NR4A activity for Nurr1-dependent and Nurr1-independent transcriptional effects and the ability to bind the Nurr1 ligand-binding domain (LBD). Protein NMR structural footprinting data show that amodiaquine, chloroquine, and cytosporone B bind the Nurr1 LBD; ligands that do not bind include C-DIM12, celastrol, camptothecin, IP7e, isoalantolactone, ethyl 2-[2,3,4-trimethoxy-6-(1-octanoyl)phenyl]acetate (TMPA), and three high-throughput screening hit derivatives. Importantly, ligands that modulate Nurr1 transcription also show Nurr1-independent effects on transcription in a cell type-specific manner, indicating that care should be taken when interpreting the functional response of these ligands in transcriptional assays. These findings should help focus medicinal chemistry efforts that desire to optimize Nurr1-binding ligands.
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Affiliation(s)
- Paola Munoz-Tello
- Department of Integrative Structural and Computational Biology, The Scripps Research Institute, Jupiter, FL, 33458, USA
| | - Hua Lin
- Department of Molecular Medicine, The Scripps Research Institute, Jupiter, Florida 33458, USA
| | - Pasha Khan
- Department of Molecular Medicine, The Scripps Research Institute, Jupiter, Florida 33458, USA
| | - Ian Mitchelle S. de Vera
- Department of Integrative Structural and Computational Biology, The Scripps Research Institute, Jupiter, FL, 33458, USA
| | - Theodore M. Kamenecka
- Department of Molecular Medicine, The Scripps Research Institute, Jupiter, Florida 33458, USA
| | - Douglas J. Kojetin
- Department of Integrative Structural and Computational Biology, The Scripps Research Institute, Jupiter, FL, 33458, USA
- Department of Molecular Medicine, The Scripps Research Institute, Jupiter, Florida 33458, USA
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27
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Jeon SG, Yoo A, Chun DW, Hong SB, Chung H, Kim JI, Moon M. The Critical Role of Nurr1 as a Mediator and Therapeutic Target in Alzheimer's Disease-related Pathogenesis. Aging Dis 2020; 11:705-724. [PMID: 32489714 PMCID: PMC7220289 DOI: 10.14336/ad.2019.0718] [Citation(s) in RCA: 33] [Impact Index Per Article: 6.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/14/2019] [Accepted: 07/18/2019] [Indexed: 01/16/2023] Open
Abstract
Several studies have revealed that the transcription factor nuclear receptor related 1 (Nurr1) plays several roles not only in the regulation of gene expression related to dopamine synthesis, but also in alternative splicing, and miRNA targeting. Moreover, it regulates cognitive functions and protects against inflammation-induced neuronal death. In particular, the role of Nurr1 in the pathogenesis of Parkinson's disease (PD) has been well investigated; for example, it has been shown that it restores behavioral and histological impairments in PD models. Although many studies have evaluated the connection between Nurr1 and PD pathogenesis, the role of Nurr1 in Alzheimer's disease (AD) remain to be studied. There have been several studies describing Nurr1 protein expression in the AD brain. However, only a few studies have examined the role of Nurr1 in the context of AD. Therefore, in this review, we highlight the overall effects of Nurr1 under the neuropathologic conditions related to AD. Furthermore, we suggest the possibility of using Nurr1 as a therapeutic target for AD or other neurodegenerative disorders.
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Affiliation(s)
- Seong Gak Jeon
- Department of Biochemistry, College of Medicine, Konyang University, Daejeon, 35365, Republic of Korea
| | - Anji Yoo
- Department of Biochemistry, College of Medicine, Konyang University, Daejeon, 35365, Republic of Korea
| | - Dong Wook Chun
- Department of Biochemistry, College of Medicine, Konyang University, Daejeon, 35365, Republic of Korea
| | - Sang Bum Hong
- Department of Biochemistry, College of Medicine, Konyang University, Daejeon, 35365, Republic of Korea
| | - Hyunju Chung
- Department of Core Research Laboratory, Clinical Research Institute, Kyung Hee University Hospital at Gangdong, Seoul 05278, Republic of Korea
| | - Jin-il Kim
- Department of Nursing, College of Nursing, Jeju National University, Jeju-si 63243, Republic of Korea
| | - Minho Moon
- Department of Biochemistry, College of Medicine, Konyang University, Daejeon, 35365, Republic of Korea
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28
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Karki K, Li X, Jin UH, Mohankumar K, Zarei M, Michelhaugh SK, Mittal S, Tjalkens R, Safe S. Nuclear receptor 4A2 (NR4A2) is a druggable target for glioblastomas. J Neurooncol 2020; 146:25-39. [PMID: 31754919 PMCID: PMC7054911 DOI: 10.1007/s11060-019-03349-y] [Citation(s) in RCA: 23] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/27/2019] [Accepted: 11/15/2019] [Indexed: 01/17/2023]
Abstract
INTRODUCTION The orphan nuclear receptor 4A2 (NR4A2) has been extensively characterized in subcellular regions of the brain and is necessary for the function of dopaminergic neurons. The NR4A2 ligand, 1,1-bis (31-indoly1)-1-(p-chlorophenyl)methane (DIM-C-pPhCl) inhibits markers of neuroinflammation and degeneration in mouse models and in this study we investigated expression and function of NR4A2 in glioblastoma (GBM). METHODS Established and patient-derived cell lines were used as models and the expression and functions of NR4A2 were determined by western blots and NR4A2 gene silencing by antisense oligonucleotides respectively. Effects of NR4A2 knockdown and DIM-C-pPhCl on cell growth, induction of apoptosis (Annexin V Staining) and migration/invasion (Boyden chamber and spheroid invasion assay) and transactivation of NR4A2-regulated reporter genes were determined. Tumor growth was investigated in athymic nude mice bearing U87-MG cells as xenografts. RESULTS NR4A2 knockdown and DIM-C-pPhCl inhibited GBM cell and tumor growth, induced apoptosis and inhibited migration and invasion of GBM cells. DIM-C-pPhCl and related analogs also inhibited NR4A2-regulated transactivation (luciferase activity) confirming that DIM-C-pPhCl acts as an NR4A2 antagonist and blocks NR4A2-dependent pro-oncogenic responses in GBM. CONCLUSION We demonstrate for the first time that NR4A2 is pro-oncogenic in GBM and thus a potential druggable target for patients with tumors expressing this receptor. Moreover, our bis-indole-derived NR4A2 antagonists represent a novel class of anti-cancer agents with potential future clinical applications for treating GBM.
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MESH Headings
- Animals
- Antineoplastic Agents/pharmacology
- Apoptosis
- Biomarkers, Tumor/genetics
- Biomarkers, Tumor/metabolism
- Cell Movement
- Cell Proliferation
- Female
- Gene Expression Regulation, Neoplastic/drug effects
- Glioblastoma/drug therapy
- Glioblastoma/metabolism
- Glioblastoma/pathology
- Humans
- Indoles/pharmacology
- Mice
- Mice, Nude
- Nuclear Receptor Subfamily 4, Group A, Member 2/antagonists & inhibitors
- Nuclear Receptor Subfamily 4, Group A, Member 2/genetics
- Nuclear Receptor Subfamily 4, Group A, Member 2/metabolism
- Prognosis
- RNA, Small Interfering/genetics
- Survival Rate
- Tumor Cells, Cultured
- Xenograft Model Antitumor Assays
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Affiliation(s)
- Keshav Karki
- Department of Veterinary Physiology and Pharmacology, Texas A&M University, College Station, TX, 77843, USA
| | - Xi Li
- Department of Veterinary Physiology and Pharmacology, Texas A&M University, College Station, TX, 77843, USA
| | - Un-Ho Jin
- Department of Veterinary Physiology and Pharmacology, Texas A&M University, College Station, TX, 77843, USA
| | - Kumaravel Mohankumar
- Department of Veterinary Physiology and Pharmacology, Texas A&M University, College Station, TX, 77843, USA
| | - Mahsa Zarei
- Department of Veterinary Physiology and Pharmacology, Texas A&M University, College Station, TX, 77843, USA
| | | | - Sandeep Mittal
- Department of Surgery, Virginia Tech University, Roanoke, VA, 24016, USA
| | - Ronald Tjalkens
- Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, CO, 80523, USA
| | - Stephen Safe
- Department of Veterinary Physiology and Pharmacology, Texas A&M University, College Station, TX, 77843, USA.
- Department of Veterinary Physiology & Pharmacology, College of Veterinary Medicine & Biomedical Sciences, Texas A&M University, 4466 TAMU, College Station, TX, 77843-4466, USA.
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29
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Chatterjee S, Walsh EN, Yan AL, Giese KP, Safe S, Abel T. Pharmacological activation of Nr4a rescues age-associated memory decline. Neurobiol Aging 2020; 85:140-144. [PMID: 31732218 PMCID: PMC6917472 DOI: 10.1016/j.neurobiolaging.2019.10.001] [Citation(s) in RCA: 20] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/24/2019] [Revised: 09/12/2019] [Accepted: 10/01/2019] [Indexed: 12/26/2022]
Abstract
Age-associated cognitive impairments affect an individual's quality of life and are a growing problem in society. Therefore, therapeutic strategies to treat age-related cognitive decline are needed to enhance the quality of life among the elderly. Activation of the Nr4a family of transcription factors has been closely linked to memory formation and dysregulation of these transcription factors is thought to be associated with age-related cognitive decline. Previously, we have shown that Nr4a transcription can be activated by synthetic bisindole-derived compounds (C-DIM). C-DIM compounds enhance synaptic plasticity and long-term contextual fear memory in young healthy mice. In this study, we show that activation of Nr4a2 by 1,1-bis(3'-Indolyl)-1-(p-chlorophenyl) methane (C-DIM12), enhances long-term spatial memory in young mice and rescues memory deficits in aged mice. These findings suggest that C-DIM activators of Nr4a transcription may be suitable to prevent memory deficits associated with aging.
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Affiliation(s)
- Snehajyoti Chatterjee
- Department of Neuroscience and Pharmacology, Iowa Neuroscience Institute Carver College of Medicine, University of Iowa, Iowa City, IA, USA
| | - Emily N Walsh
- Department of Neuroscience and Pharmacology, Iowa Neuroscience Institute Carver College of Medicine, University of Iowa, Iowa City, IA, USA
| | - Amy L Yan
- Department of Neuroscience and Pharmacology, Iowa Neuroscience Institute Carver College of Medicine, University of Iowa, Iowa City, IA, USA
| | - K Peter Giese
- Department of Basic and Clinical Neuroscience, King's College London, London, UK
| | - Stephen Safe
- Department of Veterinary Physiology and Pharmacology, Texas A&M University, College Station, TX, USA
| | - Ted Abel
- Department of Neuroscience and Pharmacology, Iowa Neuroscience Institute Carver College of Medicine, University of Iowa, Iowa City, IA, USA.
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30
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Shen Y, Dai L, Tian H, Xu R, Li F, Li Z, Zhou J, Wang L, Dong J, Sun L. Treatment Of Magnesium-L-Threonate Elevates The Magnesium Level In The Cerebrospinal Fluid And Attenuates Motor Deficits And Dopamine Neuron Loss In A Mouse Model Of Parkinson's disease. Neuropsychiatr Dis Treat 2019; 15:3143-3153. [PMID: 31806980 PMCID: PMC6857673 DOI: 10.2147/ndt.s230688] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/12/2019] [Accepted: 10/28/2019] [Indexed: 12/13/2022] Open
Abstract
PURPOSE Epidemiology research has demonstrated that magnesium (Mg) deficiency is associated with a high incidence of Parkinson's disease (PD). It is known that the systemic administration of MgSO4 is not able to elevate the Mg concentration in cerebrospinal fluid (CSF). This study aims to verify the protective effect of magnesium-L-threonate (MgT) in 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) mouse model. METHODS C57BL/6J mice were orally administered MgT or MgSO4 for 4 weeks, and received MPTP in the third week. After analysis of open-field and rotarod tests on the last day, tyrosine hydroxylase (TH) immunopositive cells and protein levels were quantified in the substantia nigra pars compacta (SNpc) and striatum. The expression of inducible nitric oxide synthase (iNOS) level was evaluated. Mg concentration in serum and CSF was measured after oral administration of MgSO4 or MgT in normal mice. Mg concentration in the CSF was increased in the mice treated with MgT but not MgSO4. RESULTS The total distance and mean speed in open-field tests, and the time spent on rotarod in the MgT group were increased, compared with MPTP group. The MgT treatment but not MgSO4 dose-dependently attenuated the loss of TH-positive neurons, and the reduction of the TH expression in the SNpc. The MgT treatment also inhibited the expression of iNOS as measured by immunohistochemistry and Western blots. Double-immunofluorescence staining of TH and iNOS showed iNOS-positive cells were collocalized for TH-positive cells. CONCLUSION The treatment with MgT is associated with an increase of Mg in the CSF. MgT, rather than MgSO4, can significantly attenuate MPTP-induced motor deficits and dopamine (DA) neuron loss.
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Affiliation(s)
- Yanling Shen
- Department of Guangxi Key Laboratory of Brain and Cognitive Neuroscience, Guilin Medical University, Guilin541004, Guangxi, People’s Republic of China
- Department of Pathology, Affiliated Chenggong Hospital, Xiamen University, Xiamen, Fujian361000, People’s Republic of China
| | - Ling Dai
- Department of Guangxi Key Laboratory of Brain and Cognitive Neuroscience, Guilin Medical University, Guilin541004, Guangxi, People’s Republic of China
| | - Haibo Tian
- Department of Guangxi Key Laboratory of Brain and Cognitive Neuroscience, Guilin Medical University, Guilin541004, Guangxi, People’s Republic of China
- Department of Pathology, Fuling Central Hospital of Chongqing City, Chongqing408099, People’s Republic of China
| | - Runnan Xu
- Department of Guangxi Key Laboratory of Brain and Cognitive Neuroscience, Guilin Medical University, Guilin541004, Guangxi, People’s Republic of China
| | - Fuying Li
- Department of Neurology and Neurological Science, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8519, Japan
| | - Zhuohang Li
- Department of Guangxi Key Laboratory of Brain and Cognitive Neuroscience, Guilin Medical University, Guilin541004, Guangxi, People’s Republic of China
| | - Jeremy Zhou
- School of Pharmacy and Medical Sciences, and UniSA Cancer Research Institute, University of South Australia, Adelaide, SA5001, Australia
| | - Liping Wang
- School of Pharmacy and Medical Sciences, and UniSA Cancer Research Institute, University of South Australia, Adelaide, SA5001, Australia
| | - Jianghui Dong
- Department of Guangxi Key Laboratory of Brain and Cognitive Neuroscience, Guilin Medical University, Guilin541004, Guangxi, People’s Republic of China
- School of Pharmacy and Medical Sciences, and UniSA Cancer Research Institute, University of South Australia, Adelaide, SA5001, Australia
| | - Liyuan Sun
- Department of Guangxi Key Laboratory of Brain and Cognitive Neuroscience, Guilin Medical University, Guilin541004, Guangxi, People’s Republic of China
- Department of Neurology and Neurological Science, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8519, Japan
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31
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Li X, Tjalkens RB, Shrestha R, Safe S. Structure-dependent activation of gene expression by bis-indole and quinoline-derived activators of nuclear receptor 4A2. Chem Biol Drug Des 2019; 94:1711-1720. [PMID: 31102570 PMCID: PMC6791730 DOI: 10.1111/cbdd.13564] [Citation(s) in RCA: 13] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/28/2019] [Revised: 04/26/2019] [Accepted: 05/13/2019] [Indexed: 12/30/2022]
Abstract
Bis-indole derivatives including 1,1-bis(3'-indolyl)-1-(4-chlorophenyl)methane (DIM-C-pPhCl) and substituted quinolines such as chloroquine (CQ) and amodiaquine (AQ) are nuclear receptor 4A2 (NR4A2, Nurr1) ligands, and they exhibit anti-inflammatory activities in mouse and rat models of Parkinson's disease, respectively. However, computational modeling demonstrates that the quinoline derivatives interact with the ligand-binding domain, whereas the bis-indoles preferentially interact with a C-terminal cofactor binding site of NR4A2. In this study, the effects of DIM-C-pPhCl and related analogs were compared with CQ/AQ as inducers of NR4A2-responsive genes including vasoactive intestinal peptide, osteopontin, proopiomelanocortin, and neuropilin 1 in Panc1 and Panc28 pancreatic cancer cells. The results demonstrate that, among the bis-indole analogs, their relative potencies as inducers were structure-gene- and cell context dependent. In contrast, CQ and AQ were significantly less potent than the bis-indole derivatives and, for some of the NR4A2-regulated genes, CQ and AQ were inactive as inducers. These results demonstrate that although bis-indole and quinoline derivatives have been characterized as activators of NR4A2-dependent gene expression, these two classes of compounds exhibit different activities, indicating that they are selective NR4A2 modulators.
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Affiliation(s)
- Xi Li
- Department of Veterinary Physiology and Pharmacology, Texas A&M University, College Station, TX
| | - Ronald B. Tjalkens
- Department of Environmental and Radiological Health Sciences, College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, CO
| | - Rupesh Shrestha
- Department of Biochemistry and Biophysics, Texas A&M University, College Station, TX
| | - Stephen Safe
- Department of Veterinary Physiology and Pharmacology, Texas A&M University, College Station, TX
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Bantle CM, Phillips AT, Smeyne RJ, Rocha SM, Olson KE, Tjalkens RB. Infection with mosquito-borne alphavirus induces selective loss of dopaminergic neurons, neuroinflammation and widespread protein aggregation. NPJ PARKINSONS DISEASE 2019; 5:20. [PMID: 31531390 PMCID: PMC6744428 DOI: 10.1038/s41531-019-0090-8] [Citation(s) in RCA: 69] [Impact Index Per Article: 11.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 09/17/2018] [Accepted: 08/12/2019] [Indexed: 12/28/2022]
Abstract
Neuroinvasive infections with mosquito-borne alphaviruses such as Western equine encephalitis virus (WEEV) can cause post-encephalitic parkinsonism. To understand the mechanisms underlying these neurological effects, we examined the capacity of WEEV to induce progressive neurodegeneration in outbred CD-1 mice following non-lethal encephalitic infection. Animals were experientally infected with recombinant WEEV expressing firefly luciferase or dsRed (RFP) reporters and the extent of viral replication was controlled using passive immunotherapy. WEEV spread along the neuronal axis from the olfactory bulb to the entorhinal cortex, hippocampus and basal midbrain by 4 days post infection (DPI). Infection caused activation of microglia and astrocytes, selective loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc) and neurobehavioral abnormalities. After 8 weeks, surviving mice displayed continued loss of dopamine neurons in the SNpc, lingering glial cell activation and gene expression profiles consistent with a neurodegenerative phenotype. Strikingly, prominent proteinase K-resistant protein aggregates were present in the the entorhinal cortex, hippocampus and basal midbrain that stained positively for phospho-serine129 α-synuclein (SNCA). These results indicate that WEEV may cause lasting neurological deficits through a severe neuroinflammatory response promoting both neuronal injury and protein aggregation in surviving individuals.
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Affiliation(s)
- Collin M Bantle
- 1Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, CO 80523 USA
| | - Aaron T Phillips
- 1Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, CO 80523 USA.,2Arthropod-Borne and Infectious Disease Laboratory, Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO 80523 USA
| | - Richard J Smeyne
- 3Department of Neuroscience, Vickie & Jack Farber Institute for Neuroscience, Thomas Jefferson University, Philadelphia, PA 19107 USA
| | - Savannah M Rocha
- 2Arthropod-Borne and Infectious Disease Laboratory, Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO 80523 USA
| | - Ken E Olson
- 2Arthropod-Borne and Infectious Disease Laboratory, Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO 80523 USA
| | - Ronald B Tjalkens
- 1Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, CO 80523 USA
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Kirkley KS, Popichak KA, Hammond SL, Davies C, Hunt L, Tjalkens RB. Genetic suppression of IKK2/NF-κB in astrocytes inhibits neuroinflammation and reduces neuronal loss in the MPTP-Probenecid model of Parkinson's disease. Neurobiol Dis 2019; 127:193-209. [PMID: 30818064 PMCID: PMC6588478 DOI: 10.1016/j.nbd.2019.02.020] [Citation(s) in RCA: 24] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/02/2018] [Revised: 12/20/2018] [Accepted: 02/22/2019] [Indexed: 12/13/2022] Open
Abstract
Neuroinflammatory activation of glia is considered a pathological hallmark of Parkinson's disease (PD) and is seen in both human PD patients and in animal models of PD; however, the relative contributions of these cell types, especially astrocytes, to the progression of disease is not fully understood. The transcription factor, nuclear factor kappa B (NFκB), is an important regulator of inflammatory gene expression in glia and is activated by multiple cellular stress signals through the kinase complex, IKK2. We sought to determine the role of NFκB in modulating inflammatory activation of astrocytes in a model of PD by generating a conditional knockout mouse (hGfapcre/Ikbk2F/F) in which IKK2 is specifically deleted in astrocytes. Measurements of IKK2 revealed a 70% deletion rate of IKK2 within astrocytes, as compared to littermate controls (Ikbk2F/F). Use of this mouse in a subacute, progressive model of PD through exposure to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine and probenecid (MPTPp) revealed significant protection in exposed mice to direct and progressive loss of dopaminergic neurons in the substantia nigra (SN). hGfapcre/Ikbk2F/F mice were also protected against MPTPp-induced loss in motor activity, loss of striatal proteins, and genomic alterations in nigral NFκB gene expression, but were not protected from loss of striatal catecholamines. Neuroprotection in hGfapcre/Ikbk2F/F mice was associated with inhibition of MPTPp-induced astrocytic expression of inflammatory genes and protection against nitrosative stress and apoptosis in neurons. These data indicate that deletion of IKK2 within astrocytes is neuroprotective in the MPTPp model of PD and suggests that reactive astrocytes directly contribute the potentiation of dopaminergic pathology.
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Affiliation(s)
- Kelly S Kirkley
- Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, CO, USA 80523
| | - Katriana A Popichak
- Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, CO, USA 80523
| | - Sean L Hammond
- Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, CO, USA 80523
| | - Cecilia Davies
- Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, CO, USA 80523
| | - Lindsay Hunt
- Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, CO, USA 80523
| | - Ronald B Tjalkens
- Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, CO, USA 80523; Program in Molecular, Cellular and Integrative Neuroscience, Colorado State University, Fort Collins, CO, USA 80523.
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34
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Jiao Z, Zhang W, Chen C, Zhu X, Chen X, Zhou M, Peng G, Liu H, Qiu J, Lin Y, Huang S, Mo M, Yang X, Qu S, Xu P. Gene Dysfunction Mediates Immune Response to Dopaminergic Degeneration in Parkinson's Disease. ACS Chem Neurosci 2019; 10:803-811. [PMID: 30289236 DOI: 10.1021/acschemneuro.8b00373] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022] Open
Abstract
Many publications reported that genetic dysfunction mediates abnormal immune responses in the brain, which is important for the development of neurodegenerative diseases, especially for Parkinson's disease (PD). This immune disorder results in subsequent inflammatory reaction, which stimulates microglia or other immune cells to secrete cytokines and chemokines and disturbs the proportion of peripheral blood lymphocyte subsets contributing to dopaminergic (DA) neuron apoptosis. Furthermore, the abnormal immune related signal pathways caused by genetic variants promote chronic inflammation destroying the blood-brain barrier, which allows infiltration of different molecules and blood cells into the central nervous system (CNS) exerting toxicity on DA neurons. As a result, the inflammatory reaction in the CNS accelerates the progression of Parkinson's disease and promotes α-synuclein aggregation and diffusion among DA neurons in the procession of Parkinson's disease. Thus, for disease evaluation, the genetic mediated abnormal immune response in PD may be assessed based on the multiple immune molecules and inflammatory factors, as well as the ratio of lymphocyte subsets from PD patient's peripheral blood as potential biomarkers.
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Affiliation(s)
- Zhigang Jiao
- Central Laboratory, Shunde Hospital, Southern Medical University, Foshan 528300, China
- Key Laboratory of Mental Health of the Ministry of Education, Southern Medical University, Foshan 528300, China
| | - Wenlong Zhang
- Department of Neurology, the First Affiliated Hospital of Guangzhou Medical University, Guangzhou 510120, China
| | - Chaojun Chen
- Department of Neurology, Guangzhou Chinese Medical Integrated Hospital (Huadu), Guangdong 510800, China
| | - Xiaoqin Zhu
- Guangzhou Medical University, Guangzhou 511436, China
| | - Xiang Chen
- Department of Neurology, the First Affiliated Hospital of Guangzhou Medical University, Guangzhou 510120, China
| | - Miaomiao Zhou
- Department of Neurology, the First Affiliated Hospital of Guangzhou Medical University, Guangzhou 510120, China
| | - Guoyou Peng
- Department of Neurology, the First Affiliated Hospital of Guangzhou Medical University, Guangzhou 510120, China
| | - Hanqun Liu
- Department of Neurology, the First Affiliated Hospital of Guangzhou Medical University, Guangzhou 510120, China
| | - Jiewen Qiu
- Department of Neurology, the First Affiliated Hospital of Guangzhou Medical University, Guangzhou 510120, China
| | - Yuwan Lin
- Department of Neurology, the First Affiliated Hospital of Guangzhou Medical University, Guangzhou 510120, China
| | - Shuxuan Huang
- Department of Neurology, the First Affiliated Hospital of Guangzhou Medical University, Guangzhou 510120, China
| | - Mingshu Mo
- Department of Neurology, the First Affiliated Hospital of Guangzhou Medical University, Guangzhou 510120, China
| | - Xinling Yang
- Department of Neurology, the Third Affiliated Hospital of Xinjiang Medical University, Urumqi 830011, China
| | - Shaogang Qu
- Central Laboratory, Shunde Hospital, Southern Medical University, Foshan 528300, China
- Key Laboratory of Mental Health of the Ministry of Education, Southern Medical University, Foshan 528300, China
| | - Pingyi Xu
- Department of Neurology, the First Affiliated Hospital of Guangzhou Medical University, Guangzhou 510120, China
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35
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Windshügel B. Structural insights into ligand-binding pocket formation in Nurr1 by molecular dynamics simulations. J Biomol Struct Dyn 2019; 37:4651-4657. [PMID: 30582418 DOI: 10.1080/07391102.2018.1559099] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022]
Abstract
The nuclear receptor Nurr1 (NR4A2) has been identified as a potential target for the treatment of Parkinson's disease. In contrast to most other nuclear receptors, the X-ray crystal structure of the Nurr1 ligand-binding domain (LBD) lacks any ligand-binding pocket (LBP). However, NMR spectroscopy measurements have revealed that the known Nurr1 agonist docosahexaenoic acid (DHA) binds to a region within the LBD that corresponds to the classical NR ligand-binding pocket (LBP). In order to investigate the structural dynamics of the Nurr1 LBD and to study potential LBP formation, the conformational space of the receptor was sampled using a molecular dynamics (MD) simulation. Docking of DHA into 50,000 LBD structures extracted from the simulation revealed the existence of a transient LBP that is capable to fully harbor the compound. The location of the identified pocket overlaps with the ligand-binding site suggested by NMR experiments. Structural analysis of the protein-ligand complex showed that only modest structural rearrangements within the Nurr1 LBD are required for LBP formation. These findings may support structure-based drug discovery campaigns for the development of receptor-specific agonists.
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Affiliation(s)
- Björn Windshügel
- Fraunhofer Institute for Molecular Biology and Applied Ecology IME, ScreeningPort , Hamburg , Germany
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36
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Jakaria M, Haque ME, Cho DY, Azam S, Kim IS, Choi DK. Molecular Insights into NR4A2(Nurr1): an Emerging Target for Neuroprotective Therapy Against Neuroinflammation and Neuronal Cell Death. Mol Neurobiol 2019; 56:5799-5814. [PMID: 30684217 DOI: 10.1007/s12035-019-1487-4] [Citation(s) in RCA: 66] [Impact Index Per Article: 11.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/31/2018] [Accepted: 01/10/2019] [Indexed: 01/23/2023]
Abstract
NR4A2 is a nuclear receptor and a transcription factor, with distinctive physiological features. In the cell nuclei of the central nervous system, it is widely expressed and identified as a crucial regulator of dopaminergic (DA) neuronal differentiation, survival, and maintenance. Importantly, it has regulated different genes crucial for dopaminergic signals, and its expression has been diminished in both aged and PD post-mortem brains and reduced in PD patients. In microglia and astrocytes, the expression of NR4A2 has been found where it can be capable of inhibiting the expression of proinflammatory mediators; hence, it protected inflammation-mediated DA neuronal death. In addition, NR4A2 plays neuroprotective role via regulating different signals. However, NR4A2 has been mainly focused on Parkinson's research, but, in recent times, it has been studied in Alzheimer's disease (AD), multiple sclerosis (MS), and stroke. Altered expression of NR4A2 is connected to AD progression, and activation of its may improve cognitive function. It is downregulated in peripheral blood mononuclear cells of MS patients; nonetheless, its role in MS has not been fully clear. miR-145-5p known as a putative regulator of NR4A2 and in a middle cerebral artery occlusion/reperfusion model, anti-miR-145-5p administration promoted neurological outcomes in rat. To date, various activators and modulators of NR4A2 have been discovered and investigated as probable therapeutic drugs in neuroinflammatory and neuronal cell death models. The NR4A2 gene and cell-based therapy are described as promising drug candidates for neurodegenerative diseases. Moreover, microRNA might have a crucial role in neurodegeneration via affecting NR4A2 expression. Herein, we present the role of NR4A2 in neuroinflammation and neuronal cell death focusing on neurodegenerative conditions and display NR4A2 as a promising therapeutic target for the therapy of neuroprotection.
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Affiliation(s)
- Md Jakaria
- Department of Applied Life Sciences and Integrated Bioscience, Graduate School, Konkuk University, Chungju, South Korea
| | - Md Ezazul Haque
- Department of Applied Life Sciences and Integrated Bioscience, Graduate School, Konkuk University, Chungju, South Korea
| | - Duk-Yeon Cho
- Department of Applied Life Sciences and Integrated Bioscience, Graduate School, Konkuk University, Chungju, South Korea
| | - Shofiul Azam
- Department of Applied Life Sciences and Integrated Bioscience, Graduate School, Konkuk University, Chungju, South Korea
| | - In-Su Kim
- Department of Applied Life Sciences and Integrated Bioscience, Graduate School, Konkuk University, Chungju, South Korea.,Department of Integrated Bioscience and Biotechnology, College of Biomedical and Health Sciences and Research Institute of Inflammatory Diseases (RID), Konkuk University, Chungju, South Korea
| | - Dong-Kug Choi
- Department of Applied Life Sciences and Integrated Bioscience, Graduate School, Konkuk University, Chungju, South Korea. .,Department of Integrated Bioscience and Biotechnology, College of Biomedical and Health Sciences and Research Institute of Inflammatory Diseases (RID), Konkuk University, Chungju, South Korea.
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