Abstract
Ouabain (OUA) is a cardiac glycoside that binds to Na+,K+-ATPase (NKA), a conserved membrane protein that controls cell transmembrane ionic concentrations and requires ATP hydrolysis. At nM concentrations, OUA activates signaling pathways that are not related to its typical inhibitory effect on the NKA pump. Activation of these signaling pathways protects against some types of injury of the kidneys and central nervous system. There are 4 isoforms of the alpha subunit of NKA, which are differentially distributed across tissues and may have different physiological roles. Glial cells are important regulators of injury and inflammation in the brain and express the α1 and α2 NKA isoforms. This study investigated the role of α2 NKA in OUA modulation of the neuroinflammatory response induced by lipopolysaccharide (LPS) in mouse primary glial cell cultures. LPS treatment increased lactate dehydrogenase release, while OUA did not decrease cell viability and blocked LPS-induced NF-κB activation. Silencing α2 NKA prevented ERK and NF-κB activation by LPS. α2 NKA also regulates TNF-α and IL-1β levels. The data reported here indicate a significant role of α2 NKA in regulating central LPS effects, with implications in the associated neuroinflammatory processes.
Highlights
Na+,K+-ATPase (NKA) is an essential membrane protein due to its maintenance of cellular resting potential and osmotic balance[1]
For a range of OUA concentrations (1 nM–300 μM), Lactate dehydrogenase (LDH) release was not different after 24 hours of treatment compared with control, while the MTT response increased at some OUA concentrations (1 nM, 3 nM and 300 nM)
Glial cells play an important role in inflammatory responses, with microglia regarded as brain macrophages in part due to the similarity of their responses and their participation in central nervous system (CNS) adaptive and innate immune responses[36]
Summary
Na+,K+-ATPase (NKA) is an essential membrane protein due to its maintenance of cellular resting potential and osmotic balance[1]. Pre-treatment of glial cells with concentrations of 1, 10 and 100 μM OUA for 15 minutes, 1 hour and 6 hours, followed by challenge with LPS at 1 hour and 6 hours (Fig. 2) confirmed increased RelA nuclear translocation by LPS vs controls at both times.
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