In Vivo Effects of Intracellular Reactive Oxygen Species and Anti-protease Action on NF-kB Activation in Rats
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Authors
Desai, Prashant J.
Issue Date
1995
Type
Thesis
Language
en_US
Keywords
Alternative Title
Abstract
Reactive oxygen species (ROS) have recently been suggested (in vitro) to
play a role as second messengers in the signal transduction pathway leading to
nuclear factor-kappa B (NF-kB) activation, a critical step for effective
inflammatory responses, both beneficial and harmful. To better understand the
involvement of ROS in the NF-kB activation cascade and explore possible ways
to modulate its undesired activation in vivo, a series of experiments were
performed involving chemically-induced intracellular ROS and their effect on
NF-kB activation. Redox-cycling agent diquat (DQ) (0.1 mmoI/kg i.p.) was
administered in male Fischer rats by itself, as well as in rats pretreated with
glutathione-depleting agent phorone (PH) (200 mg/kg i.p.). Rats were also
treated with endotoxin (ET) (0.01 mg/kg i.v.) and DQ together. Hepatic
glutathione concentrations and plasma alanine aminotransferase (ALT) activities
were measured to determine the extent of the induced intracellular oxidative
stress (caused by ROS). Since proteases are thought to be involved in the NF-kB
activation cascade, we explored the role of anti-proteases such as N-tosyl-L-Iysylchloromethylketone (TLCK) in the attenuation of NF-kB activation in rat
hepatocytes. NF-kB activation was determined by running nuclear extracts
through electrophoretic mobility shift assays (EMSA). Increases in the
percentage of hepatic glutathione disulfide (GSSG) concentrations and plasma
alanine aminotransferase (ALT) activities indicated the presence of intracellular
oxidative stress in both DQ only and PH + DQ treated animals. PH pretreatment
resulted in greater intracellular oxidative stress as indicated by higher ALT
activities. NF-kB was activated in both groups. ET pretreatment alone had little
effect on intracellular oxidative stress but highly activated NF-kB, whereas the
addition of DQ greatly enhanced intracellular oxidative stress without any
additive effect on NF-kB activation demonstrating the lack of modulatory,
additive effect of ROS on NF-kB activation. The anti-proteases, TLCK decreased
endotoxin-induced NF-kB activation. This in vivo study appears to support the
hypothesis of ROS as (direct) second messengers in NF-kB activation and further
suggests a lack of modulatory (additive) effect of ROS on NF-kB activation. In
addition, the study also demonstrates possible regulation of NF-kB activation by
anti-protease (TLCK) action.
With honors.
With honors.
Description
iv, 39 p.
Citation
Publisher
Kalamazoo College
License
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