title: A versatile high throughput screening system for the simultaneous identification of anti-inflammatory and neuroprotective compounds.
abstract: In many chronic neurodegenerative diseases including Frontotemporal Dementia and Alzheimer's disease (AD), microglial activation is suggested to be involved in pathogenesis or disease progression. Activated microglia secrete a variety of cytokines, including interleukin-1beta, interleukin-6, and tumor necrosis factor as well as reactive oxygen and nitrogen species (ROS/RNS). ROS and RNS contribute to alterations in neuronal glucose uptake, inhibition of mitochondrial enzymes, a decrease in mitochondrial membrane potential, impaired axonal transport, and synaptic signaling. In addition, ROS act as signaling molecules in pro-inflammatory redox-active signal transduction pathways. To establish a high throughput screening system for anti-inflammatory and neuroprotective compounds, we have constructed an "Enhanced Green Fluorescent protein" (EGFP) expressing neuronal cell line and set up a murine microglia/neuron co-culture system with these EGFP expressing neuronal cells. We show that microglia activation leads to neuronal cell death, which can be conveniently measured by loss of neuronal EGFP fluorescence. Moreover, we used this system to test selected polyphenolic compounds for their ability to downregulate inflammatory markers and to protect neurons against microglial insult. We suggest that this system might allow accelerated drug discovery for the treatment of inflammation-mediated neurodegenerative diseases.
authors.affiliation: Department of Biochemistry and Molecular Biology & Comparative Genomics Centre, School of Pharmacy and Molecular Sciences, James Cook University, Townsville, Australia.
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Hansen
authors.forename: Martina
authors.initials: M
authors.lastname: Krautwald
authors.forename: Annette E
authors.initials: AE
authors.lastname: Maczurek
authors.forename: Grant
authors.initials: G
authors.lastname: Stuchbury
authors.forename: Phillip
authors.initials: P
authors.lastname: Fromm
authors.forename: Megan
authors.initials: M
authors.lastname: Steele
authors.forename: Oliver
authors.initials: O
authors.lastname: Schulz
authors.forename: Obdulio Benavente
authors.initials: OB
authors.lastname: Garcia
authors.forename: Julian
authors.initials: J
authors.lastname: Castillo
authors.forename: Heinrich
authors.initials: H
authors.lastname: Körner
authors.forename: Gerald
authors.initials: G
authors.lastname: Münch
mesh_terms: D000818:Animals; D000893:Anti-Inflammatory Agents / Q000378:metabolism* / Q000494:pharmacology*; D016923:Cell Death / Q000187:drug effects; D002461:Cell Line, Transformed; D002470:Cell Survival / Q000502:physiology; D018920:Coculture Techniques / Q000379:methods; D016207:Cytokines / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004353:Drug Evaluation, Preclinical; D005434:Flow Cytometry / Q000379:methods; D005786:Gene Expression Regulation / Q000187:drug effects; D049452:Green Fluorescent Proteins / Q000235:genetics; D051379:Mice; D017628:Microglia / Q000187:drug effects; D009474:Neurons / Q000187:drug effects; D018696:Neuroprotective Agents / Q000378:metabolism* / Q000494:pharmacology*; D009498:Neurotoxins / Q000633:toxicity; D017382:Reactive Oxygen Species / Q000378:metabolism; D014162:Transfection / Q000379:methods
chemical_list: D000893:Anti-Inflammatory Agents; D016207:Cytokines; D018696:Neuroprotective Agents; D009498:Neurotoxins; D017382:Reactive Oxygen Species; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins