Abstracts
2000 Digestive Disease Week

# 2268 The Role of Reactive Oxygen Species in Antigen Presentation by Rat Kupffer Cell.
Kosei Maemura, Zhao Li Sun, Tatehiko Wada, Gregory G Bulkley, Andrew S. Klein, Baltimore, MD

Purpose: Kupffer cells (KCs), by virtue of their capability to phagocytose, degrade and present foreign peptides to immunocompetent lymphocytes, play a major role in the local and systemic immune response. We have reported that the generation of reactive oxygen species (ROS) is important for phagocytic killing by KCs. Recently the participation of oxidative mechanism in MHC class II-restricted antigen presentation was suggested in vitro. This study was designed to define the role of ROS and their enzymatic generating systems in the regulation of KCs antigen presentation. Methods: Lewis rat KCs were isolated by a two-step Percoll gradient technique and pretreated with several antioxidants (N-acetylcysteine (NAC), pyrrolidine dithiocarbamate (PDTC) and dimethylthiourea (DMTU)) or enzyme inhibitors (diphenyleneiodonium(inhibitor of NADPH), allopurinol (inhibitor of xanthine oxidase)) before the assay. We quantified, 1) hydrogen peroxide generation from antigen stimulated KCs, 2) the effect of ROS on antigen presenting function of KCs using the in vitroproliferative response of an Ovalbumin (OVA) specific T-cell clone (TOVA; CD4+/CD8-) to OVA presentation by antigen (OVA)-primed KCs, and 3) the effect of ROS on the expression of MHC class II on KCs surface using the flow cytometory. Results: 1)TOVA proliferation was stimulated in response to OVA presentation by KCs. 2) TOVA proliferative response was inhibited by cell permeable antioxidant. 3)Remarkable inhibition of ROS generation KCs was observed by using the inhibitor of NADPH oxidase and/or xanthine oxidase (XO). 4) These two inhibitors suppressed both TOVA proliferative response and MHC class II expression of KCs. Conclusion: 1)KCs effectively and specifically present OVA to the OVAspecific T-cell clone. 2)NADPH- and XO- pathways are both major sources of hydrogen peroxidase generation in KCs. 3)KCs antigen presentation is diminished by antioxidant pretreatment, perhaps via the suppression of MHC class II expression. 4) The generation of ROS by NADPH- and XOdependent pathways plays an important immunomodulately role in process of KCs antigen presentation.