A Marine Natural Product CMDD8299 Attenuates Atherosclerosis by Anti-inflammatory Effect through Inducing MKP-1

Abstract

Atherosclerosis is known as a chronic inflammatory disease and numerous attempts to improve atherosclerosis by anti-inflammatory molecules were conducted. Marine sponges are rich sources of anti-inflammatory natural products. To find the anti-atherosclerotic molecule, anti-inflammatory effects of marine natural products against cytokines such as interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α) and interleukin-1 β (IL-1β) were evaluated. I identified several marine natural products including phorbaketal A, which showed anti-inflammatory effects. Phorbaketal A is a marine sponge-derived bioactive sesterterpenoid, which isolated from Phorbas sp. and Monanchora sp. Byun MR et al. demonstrated that phorbaketal A stimulates osteogenic differentiation and inhibits adipogenic differentiation through activation of transcriptional coactivator with PDZ-binding motif (TAZ) and extracellular signal-regulated kinase (ERK) in C3H10T1/2 and human mesenchymal stem cell. In the present study, phorbaketal A significantly reduced IL-6, TNF-α and IL-1β mRNA expression in lipopolysaccharide (LPS)-induced raw 264.7 macrophages and inhibited IL-6 and TNF-α secretion. Phorbaketal A reduced nitric oxide (NO) secretion through inhibition of inducible nitric oxide synthase (iNOS) expression. In addition, pre-incubation of human umbilical vein endothelial cells (HUVECs) with phorbaketal A abrogated TNF-α-induced expression of VCAM-1 and MCP-1. To demonstrate anti-atherogenic effect of phorbaketal A in vivo, 5 mg/kg of phorbaketal A administered to three types of atherosclerosis in vivo models with atherogenic diet (21% fat, 1.25% cholesterol). The severity of atherosclerosis was analyzed by en face staining of aorta. Phorbaketal A treated mice had less atherosclerotic lesions than control group in progression models. MAPKs regulate numerous fundamental cellular processes especially inflammation and immune response. TLR agonists activate MAPKs signals in innate immune cells and contribute to pathogenesis of atherosclerosis. To identify the molecular mechanisms of phorbaketal A, effects of phorbaketal A on MAPK activation were estimated. Phorbaketal A inhibited LPS-induced phosphorylation of p38 MAPK but it has no effect on ERK or JNK phosphorylation in raw264.7 macrophage. In addition, phorbaketal A abrogated p38 phosphorylation in TNF-α stimulated HUVECs. The inhibitory effect of phorbaketal A recovered by MAPK phosphatase 1 (MKP-1) inhibitor, triptolide. MKP-1 is known to inhibit inadequate inflammatory response through negative feedback of MAPKs, especially p38 and JNK. Phorbaketal A induced MKP-1 mRNA and protein expression in concentration-dependent manners. Transcription factors which regulate MKP-1 expression, such as cAMP response element binding protein (CREB) and activating transcription factor 2 (ATF-2) were phosphorylated at early time (5 min) by phorbaketal A. The phosphorylation of ERK pathway signaling molecules were suppressed by U0126 (ERK inhibitor). These suggest that phorbaketal A induced MKP-1 through ERK-activated CREB and ATF-2 signals. In this study, I identified a novel mechanism of phorbaketal A, and demonstrated its anti-atherogenic effects on model of atherosclerosis in vitro and in vivo. These results suggest that phorbaketal A could be a drug candidate for treatment of inflammatory disease, such as atherosclerosis.