Degree Name

PhD (Doctor of Philosophy)

Program

Biomedical Sciences

Date of Award

December 1997

Abstract

The effects of oxidatively modified low density lipoprotein (oxLDL) on atherogenesis may be partly mediated by alterations in nitric oxide (NO) production by macrophages. A major goal of this study was to identify the lipid components in oxLDL modulating NO production. The effect of a water soluble antioxidants (N-acetylcysteine) and lipid soluble antioxidant (alpha-tocopherol) on NO production in macrophages was also determined. A second goal was to determine if the effects of oxLDL occurred at the transcriptional level. Human LDL was oxidized using an azo-initiator 2,2$\sp\prime$-azobis (2-amidinopropane) HCI (ABAP). OxLDL markedly decreased the production of NO in LPS stimulated RAW264.7 macrophages. This inhibition depended on the levels of LOOH formed in oxLDL and was not due to oxLDL cytotoxicity. In contrast, acetylated LDL (AcLDL) and native LDL showed only minor inhibition. Lipid hydroperoxides (LOOH) and lysophosphatidylcholine (lysoPC) are the primary products formed during LDL oxidation. 13-Hydroperoxyl octadecadienoic acid (13-HPODE) markedly inhibited NO production, whereas lysoPC showed only slight inhibition. Furthermore, the effects of 13-HPODE and lysoPC did not require their uptake in an AcLDL carrier. Pre-treatment of macrophages with alpha-tocopherol attenuated the inhibition due to oxLDL. Similarly, pre-treatment with N-acetylcysteine attenuated the inhibition caused by oxLDL or 13-HPODE. OxLDL was found to decrease iNOS protein and mRNA levels in RAW264.7 macrophages induced by LPS. The activation of NF-$\kappa$B was slightly suppressed after 45 minutes of treatment. 13-HPODE showed much stronger reduction of iNOS protein levels than lysoPC. These results suggest that oxLDL may inhibit NO production in macrophages at transcriptional level. 13-HPODE is likely to be the most important lipid component in oxLDL for the inhibitory effect. Antioxidants were found to preserve NO production in macrophages treated with either oxLDL or 13-HPODE. The physiological consequences of decreased NO production in macrophages caused by oxLDL are discussed with respect to atherosclerosis.

Document Type

Dissertation - Open Access

Included in

Cell Biology Commons

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