Microglial NADPH Oxidase Mediates Leucine Enkephalin Dopaminergic Neuroprotection
Abstract: Here, we report that leucine enkephalin (LE) is neuroprotective to dopaminergic (DA) neurons at femtomolar concentrations through anti-inflammatory properties. Mesencephalic neuron-glia cultures pretreated with femtomolar concentrations of LE (10?15-10?13 M) protected DA neurons from lipopolysaccharide (LPS)-induced DA neurotoxicity, as determined by DA uptake assay and tyrosine hydroxylase (TH) immunocytochemistry (ICC). However, des-tyrosine leucine enkephalin (DTLE), an LE analogue that is missing the tyrosine residue required for binding to the kappa opioid receptor, was also neuroprotective (10?15-10?13 M), as determined by DA uptake assay and TH ICC. Both LE and DTLE (10?15-10?13 M) reduced LPS-induced superoxide production from microglia-enriched cultures. Further, both LE and DTLE (10?14, 10?13 M) reduced the LPS-induced tumor necrosis factor-alpha (TNF?) mRNA and TNF? protein from PHOX+/+ microglia, as determined by quantitative real-time RT-PCR and ELISA analysis in mesencephalic neuron-glia cultures, respectively. However, both peptides failed to inhibit TNF? expression in PHOX?/? cultures, which are unable to produce extracellular superoxide in response to LPS. Additionally, LE and DTLE (10?14, 10?13 M) failed to show any neuroprotection against LPS in PHOX?/? cultures. Together, these data indicate that LE and DTLE are neuroprotective at femtomolar concentrations through the inhibition of oxidative insult associated with microglial NADPH oxidase and the attenuation of the ROS-mediated amplification of TNF? gene expression in microglia.
microglial nadph oxidase neuroprotection leucine enkephalin (le) neurotoxicity des-tyrosine leucine enkephalin (dtle) tumor necrosis factor-alpha (tnf?) lipopolysaccharide (lps)