Melittin inhibits inflammatory target gene expression and mediator generation via interaction with IkappaB kinase |
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Authors: | Park Hye Ji Son Dong Ju Lee Chung Woo Choi Myoung Suk Lee Ung Soo Song Ho Sueb Lee Jeong Min Hong Jin Tae |
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Affiliation: | College of Pharmacy, Chungbuk National University, 12 Gaesin-dong, Heungduk-gu, Cheongju, Chungbuk 361-763, Republic of Korea. |
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Abstract: | We previously found that bee venom (BV) and melittin (a major component of BV) has anti-inflammatory effect by reacting with the sulfhydryl group of p50 of NF-kappaB. Since the sulfhydryl group is present in IkappaB kinase (IKKalpha and IKKbeta), anti-inflammatory effect of melittin via interaction with IKKs was investigated. We first examined binding of melittin to IKKs using surface plasmon resonance analyzer. Melittin binds to IKKalpha (K(d) = 1.34 x 10(-9) M) and IKKbeta (K(d) = 1.01 x 10(-9) M). Consistent with the high binding affinity, melittin (5 and 10 microg/ml) and BV (0.5, 1 and 5 microg/ml) suppressed sodium nitroprusside, TNF-alpha and LPS induced-IKKbeta and IKKbeta activities, IkappaB release, and NF-kappaB activity as well as the expressions of iNOS and COX-2, and the generation of nitric oxide (NO) and prostaglandin E(2) (PGE(2)) in Raw 264.7 mouse macrophages and synoviocytes obtained from rheumatoid arthritis patients. The binding affinities of melittin to mutant IKKs, was reduced, and the inhibitory effect of melittin on IKK and NF-kappaB activities, and NO and PGE(2) generation were abrogated by the reducing agents or in Raw 264.7 transfected with mutant plasmid IKKalpha (C178A) or IKKbeta (C179A). These results suggest that melittin binding to the sulfhydryl group of IKKs resulted in reduced IKK activities, IkappaB release, NF-kappaB activity and generation of inflammatory mediators, indicating that IKKs may be also anti-inflammatory targets of BV. |
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Keywords: | BV, bee venom COX, cyclooxygenase cPLA2, cytosolic phospholipase A2 DTT, dithiothreitol EMSA, electrophoretic mobility shift assay GSH, glutathione IκBs, inhibitors of κB IKK, IκB kinase iNOS, inducible nitric oxide synthase LPS, lipopolysaccharide NF-κB, nuclear factor-kappa B NO, nitric oxide PGs, prostaglandins RA, rheumatoid arthritis SNP, sodium nitroprusside TNF-α, tumor necrosis factor-α |
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