Costunolide alleviates atherosclerosis in high-fat diet-fed ApoE^-/- mice through covalently binding to IKK and inhibiting NF-B-mediated inflammation

Costunolide (CTD) is a sesquiterpene lactone isolated from costus root and exhibits various biological activities including anti-inflammation. Since atherosclerosis is a chronic inflammatory disease, we herein investigated the anti-atherosclerotic effects of CTD and the underlying mechanism. Atherosclerosis was induced in ApoE(-/-) mice by feeding them with a high-fat diet (HFD) for 8 weeks, followed by administration of CTD (10, 20 mg.kg(-1).d(-1), i.g.) for 8 weeks. We showed that CTD administration dose-dependently alleviated atherosclerosis in HFD-fed ApoE-/- mice. Furthermore, we found that CTD dose-dependently reduced inflammatory responses in aortas of the mice, as CTD prevented infiltration of inflammatory cells in aortas and attenuated oxLDL uptake in macrophages, leading to reduced expression of pro-inflammatory and pro-fibrotic molecules in aortas. Similar results were observed in oxLDL-stimulated mouse primary peritoneal macrophages (MPMs) in vitro. We showed that pretreatment with CTD (2.5, 5. 10 mu M) restrained oxLDL-induced inflammatory responses in MPMs by blocking pro-inflammatory NF-kappa B/p65 signaling pathway. We further demonstrated that CTD inactivated NF-kappa B via covalent binding to cysteine 179 on IKK beta, a canonical upstream regulator of NF-kappa B, reducing its phosphorylation and leading to conformational change in the active loop of IKK beta. Our results discover IKK beta as the target of CTD for its anti-inflammatory activity and elucidate a molecular mechanism underlying the anti-atherosclerosis effect of CTD. CTD is a potentially therapeutic candidate for retarding inflammatory atherosclerotic diseases.