Investigation of the inhibitory action mechanisms of 7HQ and several synthetic chemical compounds on nitric oxide production in murine macrophage RAW 264.7 cells

• Main Authors: Yu-Chun Huang, 黃煜珺
• Other Authors: Che-Ming Teng
• Format: Others
• Language: zh-TW
• Published: 1999
• Online Access: http://ndltd.ncl.edu.tw/handle/70723819600655819873

碩士 === 國立臺灣大學 === 藥理學研究所 === 87 === Nitric oxide (NO) is an important biological mediator associated with multiple pathophysiological phenomena, such as platelet aggregation, vasodilatation, septic shock, and autoimmune diseases. In addition, NO is also of great benefit in homeostasis. However, excessive production of NO may cause detrimental effects. The active component of endotoxin, lipopolysaccharide (LPS), promotes NO production in inflammatory states. In this study, we demonstrated the effects of several synthetic chemical compounds, such as 7HQ derivatives, DCDC and DHNC on LPS-induced NO formation in murine RAW 264.7 cells, a macrophage-like cell line. LPS significantly induced iNOS expression and nitrite production in RAW 264.7 cells. Pre-incubated with the above compounds, the iNOS expression and nitrite accumulation by LPS were inhibited in a dose-dependent manner. However, the direct iNOS activity was only slightly affected by these compounds. Thus, the relationship between iNOS activity and NO formation was not very relevant. NF-kB, a transcription factor, served as a key point in the modulation of iNOS gene expression. In unstimulated cells, it located in the cytoplasma and associated with an inhibitory protein, IkB. After activation, NF-kB translocated into the nucleus and initiated the gene transcription. According to our electrophoretic mobility shift assay (EMSA) results, we have demonstrated that these synthetic chemical compounds could efficiently inhibit the activation of NF-kB by preventing its translocation. Furthermore, reactive oxygen species may involve in the activation of NF-kB. Analysis by 1,1-diphenyl-2-picryl-hydrazyl (DPPH) scavenging test, we found that DCDC and DHNC were powerful as free radical scavengers. We have also determined the effects of these compounds on COX-2 activity and found that DCDC exerted a profoundly inhibitory effect on this enzyme activity. These results suggest that these synthetic chemical compounds exhibit inhibitory effects on LPS-induced NO production by decreasing iNOS expression through inhibition of IkB-a degradation and NF-kB activation.