Sphingomyelin Synthase 2 Deficiency Attenuates NFκB Activation

Abstract

Background-NFB has long been regarded as a proatherogenic factor, mainly because of its regulation of many of the proinflammatory genes linked to atherosclerosis. Metabolism of sphingomyelin (SM) has been suggested to affect NFB activation, but the mechanism is largely unknown. SMS2 regulates SM levels in cell plasma membrane and lipid rafts and has a potential to regulate NFB activation. Methods and Results-To investigate the role of SMS2 in NFB activation we used macrophages from SMS2 knockout (KO) mice and SMS2 siRNA-treated HEK 293 cells. We found that NFB activation and its target gene expression are attenuated in macrophages from SMS2 KO mice in response to lipopolysaccharide (LPS) stimulation and in SMS2 siRNA-treated HEK 293 cells after tumor necrosis factor (TNF)-␣ simulation. In line with attenuated NFB activation, we found that SMS2 deficiency substantially diminished the abundance of toll like receptor 4 (TLR4)-MD2 complex levels on the surface of macrophages after LPS stimulation, and SMS2 siRNA treatment reduced TNF-␣-stimulated lipid raft recruitment of TNF receptor-1 (TNFR1) in HEK293 cells. SMS2 deficiency decreased the relative amounts of SM and diacylglycerol (DAG) and increased ceramide, suggesting multiple mechanisms for the decrease in NFB activation. Conclusions-SMS2 is a modulator of NFB activation, and thus it could play an important role in NFB-mediated proatherogenic process. Key Words: sphingomyelin synthase 2 Ⅲ sphingomyelin Ⅲ lipid rafts Ⅲ NFB Ⅲ atherosclerosis A therosclerosis is an inflammatory disease. The accumulation of macrophage-derived foam cells in the vessel wall is always accompanied by the production of a wide range of chemokines, cytokines, and growth factors. 1 These factors regulate the turnover and differentiation of immigrating and resident cells, eventually influencing plaque development. One of the key regulators of inflammation is NFB, 2 which has long been regarded as a proatherogenic factor, mainly because of its regulation of many of the proinflammatory genes linked to atherosclerosis. 3,4 Sphingomyelin (SM) is one of the major lipids on the plasma membrane and is enriched in lipid rafts, which are considered microdomains of plasma membrane critical for signal transduction. 5,6 Depletion of cholesterol from rafts causes a redistribution of TNF-␣ receptor 1 to nonraft plasma membrane, preventing NFB activation 7 or ligand-induced RhoA activation, 8 and such treatment also inhibits proinflammatory signals mediated by TLRs. 9 Studies also suggest that NFB activation is triggered by SM-derived ceramide. 10,11 On the contrary, it has been shown that ceramide is not necessary or even inhibits NFB activation. 12 SM biosynthesis might also affect NFB activation. SM is synthesized by sphingomyelin synthase (SMS), which transfers the phosphorylcholine moiety from phosphatidylcholine (PC) onto ceramide, producing SM and diacylglycerol (DAG). 13 Luberto et al 14 found that D609, a nonspecific SMS inhibitor, blocks TNF-␣-and phorbol ester-mediated NFB activation that was c...

Tales and Mysteries of the Enigmatic Sphingomyelin Synthase Family

“…Confirmation of the involvement of SMS2 in the physiological maintenance of plasma membrane SM has come from studies using macrophages from SMS2 knock out (KO) mice. 76 Macrophages from SMS2 KO mice showed diminished recruitment of the Toll Like Receptor 4-MD2 complex on the cell surface in response to lypopolysaccharide (LPS) treatment, consistent with the reduction of LPS-mediated apoptosis observed in THP-1-derived macrophages after siRNA-mediated down-regulation of SMS1 or SMS2. 77 Likewise, siRNA-mediated down-regulation of SMS2 in HEK 293 cells partly inhibited recruitment of the tumor necrosis factor (TNF) _ receptor 1 to detergent-resistant microdomains in response to stimulation with TNF, decreasing TNF-mediated down-stream signaling.…”

“…Moreover, SMS1 and a pool of SMS2 are both localized to the Golgi. 58,66,72,74,76 Why is this organelle equipped with two different SMS enzymes? What is the relevance of the reverse activity of SMS enzymes?…”

“…In support of this conclusion, over-expression of either SMS1 or SMS2 in HeLa cells comparably increased de novo SM biosynthesis, known to occur in the Golgi, 74 whereas down-regulation of either one significantly inhibited it in HeLa, 74,80 Huh, 72 HEK 293 cells and macrophages isolated from KO mice. 76 In HeLa cells down-regulation of either SMS1 or SMS2 inhibited TNF-mediated NF-gB activation by altering plasma membrane receptor activation, similarly to HEK 293 cells (Luberto and Marimuthu, unpublished observations).…”

Sphingosine-1–Phosphate, FTY720, and Sphingosine-1–Phosphate Receptors in the Pathobiology of Acute Lung Injury

“…Recent studies demonstrated that SMS2 deficiency attenuates NF-kB signaling, thereby suggesting a role for ceramide in NF-kB signal transduction (63). However, ceramide also inhibits NF-kB activation (64)…”

Caspase-mediated inhibition of sphingomyelin synthesis is involved in FasL-triggered cell death

“…29 Protection from LPS-induced cell death has been further shown to be mediated by an impaired membrane recruitment of TLR4 signaling complexes in macrophages of SMS2 knockout mice. 30 This study aimed at investigating the function of SM biosynthesis in the regulation of FasL-induced cell death. We show that Fas-mediated apoptosis is associated with a profound inhibition of SM synthesis, which likely involved the caspase-mediated cleavage and relocation of SMS1.…”