A versatile high throughput screening system for the simultaneous identification of anti-inflammatory and neuroprotective compounds.
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.
Hansen
Elizabeth
E
Department of Biochemistry and Molecular Biology & Comparative Genomics Centre, School of Pharmacy and Molecular Sciences, James Cook University, Townsville, Australia.
Krautwald
Martina
M
Maczurek
Annette E
AE
Stuchbury
Grant
G
Fromm
Phillip
P
Steele
Megan
M
Schulz
Oliver
O
Garcia
Obdulio Benavente
OB
Castillo
Julian
J
Körner
Heinrich
H
Münch
Gerald
G
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
D000893:Anti-Inflammatory Agents; D016207:Cytokines; D018696:Neuroprotective Agents; D009498:Neurotoxins; D017382:Reactive Oxygen Species; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins