Joo Young Lee

Results from our previous studies demonstrated that activation of Toll-like receptor 4 (Tlr4), the lipopolysaccharide (LPS) receptor, is sufficient to induce nuclear factor kappaB activation and expression of inducible cyclooxygenase (COX-2) in macrophages. Saturated fatty acids (SFAs) acylated in lipid A moiety of LPS are essential for biological activities of LPS. Thus, we determined whether these fatty acids modulate LPS-induced signaling pathways and COX-2 expression in monocyte/macrophage cells (RAW 264.7). Results show that SFAs, but not unsaturated fatty acids (UFAs), induce nuclear factor kappaB activation and expression of COX-2 and other inflammatory markers. This induction is inhibited by a dominant-negative Tlr4. UFAs inhibit COX-2 expression induced by SFAs, constitutively active Tlr4, or LPS. However, UFAs fail to inhibit COX-2 expression induced by activation of signaling components downstream of Tlr4. Together, these results suggest that both SFA-induced COX-2 expression and its inhibition by UFAs are mediated through a common signaling pathway derived from Tlr4. These results represent a novel mechanism by which fatty acids modulate signaling pathways and target gene expression. Furthermore, these results suggest a possibility that propensity of monocyte/macrophage activation is modulated through Tlr4 by different types of free fatty acids, which in turn can be altered by kinds of dietary fat consumed.

Human subjects consuming fish oil showed a significant suppression of cyclooxygenase-2 (COX-2) expression in blood monocytes when stimulated in vitro with lipopolysaccharide (LPS), an agonist for Toll-like receptor 4 (TLR4). Results with a murine monocytic cell line (RAW 264.7) stably transfected with COX-2 promoter reporter gene also demonstrated that LPS-induced COX-2 expression was preferentially inhibited by docosahexaenoic acid (DHA, C22:6n-3) and eicosapentaenoic acid (EPA, C20:5n-3), the major n-3 polyunsaturated fatty acids (PUFAs) present in fish oil. Additionally, DHA and EPA significantly suppressed COX-2 expression induced by a synthetic lipopeptide, a TLR2 agonist. These results correlated with the preferential suppression of LPS- or lipopeptide-induced NFκB activation by DHA and EPA. The target of inhibition by DHA is TLR itself or its associated molecules, but not downstream signaling components. In contrast, COX-2 expression by TLR2 or TRL4 agonist was potentiated by lauric acid, a saturated fatty acid. These results demonstrate that inhibition of COX-2 expression by n-3 PUFAs is mediated through the modulation of TLR-mediated signaling pathways.Thus, the beneficial or detrimental effects of different types of dietary fatty acids on the risk of the development of many chonic inflammatory diseases may be in part mediated through the modulation of TLRs. Human subjects consuming fish oil showed a significant suppression of cyclooxygenase-2 (COX-2) expression in blood monocytes when stimulated in vitro with lipopolysaccharide (LPS), an agonist for Toll-like receptor 4 (TLR4). Results with a murine monocytic cell line (RAW 264.7) stably transfected with COX-2 promoter reporter gene also demonstrated that LPS-induced COX-2 expression was preferentially inhibited by docosahexaenoic acid (DHA, C22:6n-3) and eicosapentaenoic acid (EPA, C20:5n-3), the major n-3 polyunsaturated fatty acids (PUFAs) present in fish oil. Additionally, DHA and EPA significantly suppressed COX-2 expression induced by a synthetic lipopeptide, a TLR2 agonist. These results correlated with the preferential suppression of LPS- or lipopeptide-induced NFκB activation by DHA and EPA. The target of inhibition by DHA is TLR itself or its associated molecules, but not downstream signaling components. In contrast, COX-2 expression by TLR2 or TRL4 agonist was potentiated by lauric acid, a saturated fatty acid. These results demonstrate that inhibition of COX-2 expression by n-3 PUFAs is mediated through the modulation of TLR-mediated signaling pathways. Thus, the beneficial or detrimental effects of different types of dietary fatty acids on the risk of the development of many chonic inflammatory diseases may be in part mediated through the modulation of TLRs. Toll-like receptors (TLRs) play a critical role in the detection of microbial infection and the induction of inflammatory and immune responses against conserved microbial structures, called pathogen-associated molecular patterns (PAMPs) (1Medzhitov R. Janeway Jr., C. The Toll receptor family and microbial recognition.Trends Microbiol. 2000; 8: 452-456Google Scholar). The activation of TLRs leads to the induction of nuclear factor κB (NFκB) activation and the expression of inflammatory cytokines (2Aderem A. Ulevitch R.J. Toll-like receptors in the induction of the innate immune response.Nature. 2000; 406: 782-787Google Scholar, 3Heldwein K.A. Golenbock D.T. Fenton M.J. Recent advances in the biology of Toll-like receptors.Mod. Asp. Immunobiol. 2001; 1: 249-252Google Scholar). Ten members of the TLR family have so far been identified in human and mouse, and these TLRs are ubiquitously expressed in human tissues (4Medzhitov R. Preston-Hurlburt P. Janeway Jr, C.A. A human homologue of the Drosophila Toll protein signals activation of adaptive immunity.Nature. 1997; 388: 394-397Google Scholar, 5Akira S. Toll-like receptors and innate immunity.Adv. Immunol. 2001; 78: 1-56Google Scholar, 6Zarember K.A. Godowski P.J. Tissue expression of human Toll-like receptors and differential regulation of Toll-like receptor mRNAs in leukocytes in response to microbes, their products, and cytokines.J. Immunol. 2002; 168: 554-561Google Scholar). However, endogenous ligands for these TLRs have not been fully identified. Genetic and biochemical evidence demonstrated that TLR4 confers the responsiveness to lipopolysaccharide (LPS) derived from gram-negative bacteria (7Poltorak A. He X. Smirnova I. Liu M.Y. Huffel C.V. Du X. Birdwell D. Alejos E. Silva M. Galanos C. Freudenberg M. Ricciardi-Castagnoli P. Layton B. Beutler B. Defective LPS signaling in C3H/HeJ and C57BL/10ScCr mice: mutations in Tlr4 gene.Science. 1998; 282: 2085-2088Google Scholar, 8Qureshi S.T. Lariviere L. Leveque G. Clermont S. Moore K.J. Gros P. Malo D. Endotoxin-tolerant mice have mutations in Toll-like receptor 4 (Tlr4).J. Exp. Med. 1999; 189: 615-625Google Scholar, 9Rhee S.H. Hwang D. Murine TOLL-like receptor 4 confers lipopolysaccharide responsiveness as determined by activation of NF kappa B and expression of the inducible cyclooxygenase.J. Biol. Chem. 2000; 275: 34035-34040Google Scholar), whereas TLR2 recognizes other bacterial cell wall components, including bacterial lipoproteins (1Medzhitov R. Janeway Jr., C. The Toll receptor family and microbial recognition.Trends Microbiol. 2000; 8: 452-456Google Scholar, 2Aderem A. Ulevitch R.J. Toll-like receptors in the induction of the innate immune response.Nature. 2000; 406: 782-787Google Scholar, 3Heldwein K.A. Golenbock D.T. Fenton M.J. Recent advances in the biology of Toll-like receptors.Mod. Asp. Immunobiol. 2001; 1: 249-252Google Scholar). Other agonists for TLR4 from nonmicrobial origins include heat shock protein 60, fibronectin, taxol, respiratory syncytical virus coat protein, and saturated fatty acids (10Ohashi K. Burkart V. Flohe S. Kolb H. Cutting edge: heat shock protein 60 is a putative endogenous ligand of the toll-like receptor-4 complex.J. Immunol. 2000; 164: 558-561Google Scholar, 11Okamura Y. Watari M. Jerud E.S. Young D.W. Ishizaka S.T. Rose J. Chow J.C. Strauss 3rd, J.F. The extra domain A of fibronectin activates Toll-like receptor 4.J. Biol. Chem. 2001; 276: 10229-10233Google Scholar, 12Yoshimura A. Lien E. Ingalls R.R. Tuomanen E. Dziarski R. Golenbock D. Cutting edge: recognition of Gram-positive bacterial cell wall components by the innate immune system occurs via Toll-like receptor 2.J. Immunol. 1999; 163: 1-5Crossref Google Scholar, 13Kurt-Jones E.A. Popova L. Kwinn L. Haynes L.M. Jones L.P. Tripp R.A. Walsh E.E. Freeman M.W. Golenbock D.T. Anderson L.J. Finberg R.W. Pattern recognition receptors TLR4 and CD14 mediate response to respiratory syncytial virus.Nat. Immunol. 2000; 1: 398-401Google Scholar, 14Lee J.Y. Sohn K.H. Rhee S.H. Hwang D. Saturated fatty acids, but not unsaturated fatty acids, induce the expression of cyclooxygenase-2 mediated through Toll-like receptor 4.J. Biol. Chem. 2001; 276: 16683-16689Google Scholar). Such a broad spectrum of TLR4 agonists implies the promiscuous nature of ligand specificity for this receptor. This leads to the speculation that TLRs have much broader roles than we currently understand. Lipid A, which possesses most of the biological activities of LPS, is acylated with hydroxy saturated fatty acids. The 3-hydroxyl groups of these saturated fatty acids are further 3-O-acylated by saturated fatty acids. Removal of these O-acylated saturated fatty acids from lipid A not only results in complete loss of endotoxic activity, but also makes the lipid A act as an antagonist to the native lipid A (15Munford R.S. Hall C.L. Detoxification of bacterial lipopolysaccharides (endotoxins) by a human neutrophil enzyme.Science. 1986; 234: 203-205Google Scholar, 16Kitchens R.L. Ulevitch R.J. Munford R.S. Lipopolysaccharide (LPS) partial structures inhibit responses to LPS in a human macrophage cell line without inhibiting LPS uptake by a CD14- mediated pathway.J. Exp. Med. 1992; 176: 485-494Google Scholar). Lipid A(s) containing unsaturated fatty acids are also known to be nontoxic or act as an antagonist against endotoxin (17Krauss J.H. Seydel U. Weckesser J. Mayer H. Structural analysis of the nontoxic lipid A of Rhodobacter capsulatus 37b4.Eur. J. Biochem. 1989; 180: 519-526Google Scholar, 18Qureshi N. Takayama K. Kurtz R. Diphosphoryl lipid A obtained from the nontoxic lipopolysaccharide of Rhodopseudomonas sphaeroides is an endotoxin antagonist in mice.Infect. Immun. 1991; 59: 441-444Google Scholar). It was also demonstrated that the deacylated bacterial lipoproteins were unable to activate TLR2 and to induce cytokine expression in monocytes (19Brightbill H.D. Libraty D.H. Krutzik S.R. Yang R.B. Belisle J.T. Bleharski J.R. Maitland M. Norgard M.V. Plevy S.E. Smale S.T. Brennan P.J. Bloom B.R. Godowski P.J. Modlin R.L. Host defense mechanisms triggered by microbial lipoproteins through toll-like receptors.Science. 1999; 285: 732-736Google Scholar). These results suggest that the fatty acids acylated on lipid A or bacterial lipoproteins play a critical role in ligand recognition and receptor activation for TLR2 and TLR4. Indeed, it was suggested that the rapid interaction of bacterial lipopeptides with plasma membrane of macrophages occurs via insertion of their acylated saturated fatty acids as determined by electron energy loss spectroscopy and freeze-fracture techniques (20Wolf B. Hauschildt S. Uhl B. Metzger J. Jung G. Bessler W.G. Localization of the cell activator lipopeptide in bone marrow-derived macrophages by electron energy loss spectroscopy (EELS).Immunol. Lett. 1989; 20: 121-126Google Scholar, 21Uhl B. Speth V. Wolf B. Jung G. Bessler W.G. Hauschildt S. Rapid alterations in the plasma membrane structure of macrophages stimulated with bacterial lipopeptides.Eur. J. Cell Biol. 1992; 58: 90-98Google Scholar). Results from our previous studies (9Rhee S.H. Hwang D. Murine TOLL-like receptor 4 confers lipopolysaccharide responsiveness as determined by activation of NF kappa B and expression of the inducible cyclooxygenase.J. Biol. Chem. 2000; 275: 34035-34040Google Scholar) demonstrated that the ligand independent activation of TLR4 is sufficient to induce the activation of NFκB and the expression of the (COX-2) in saturated fatty acids induce NFκB activation and COX-2 but unsaturated fatty acids inhibit saturated fatty and LPS-induced NFκB and the expression of COX-2 and other inflammatory in a murine monocytic cell line (RAW 264.7) J.Y. Sohn K.H. Rhee S.H. Hwang D. Saturated fatty acids, but not unsaturated fatty acids, induce the expression of cyclooxygenase-2 mediated through Toll-like receptor 4.J. Biol. Chem. 2001; 276: 16683-16689Google Scholar). The inhibition of LPS-induced NFκB activation and COX-2 expression by unsaturated fatty acids was mediated through the suppression of signaling J.Y. Sohn K.H. Rhee S.H. Hwang D. Saturated fatty acids, but not unsaturated fatty acids, induce the expression of cyclooxygenase-2 mediated through Toll-like receptor 4.J. Biol. Chem. 2001; 276: 16683-16689Google Scholar). It was demonstrated that consuming n-3 polyunsaturated fatty acids (PUFAs) leads to the suppression of the of LPS-induced cytokines in blood in S. R. K. G. The of dietary with n-3 polyunsaturated fatty acids on the of and factor by J. Med. 1989; Scholar, S. R. R. C.A. with n-3 fatty acids and cell Biol. Scholar). However, the is not inducible (COX-2) and cytokines to a family of response The expression of response not protein response induced by and Biochem. 1991; Scholar). This that the suppressed expression of LPS-induced cytokines by n-3 PUFAs may be mediated by modulation of LPS signaling pathways. Thus, we determined COX-2 expression is inhibited in monocytes derived from human subjects consuming fish and this inhibition is mediated through the modulation of TLR4 signaling by n-3 the activation of TLRs is by the types of fatty acids, signaling downstream of target gene and responses also be by different types of fatty acids. This modulation for the role of dietary with of fatty acids on inflammatory and immune responses induced by the activation of TLRs that are ubiquitously expressed in human of unsaturated and saturated fatty acids were from LPS was from A synthetic bacterial was from of other were from The reporter containing the promoter of the murine COX-2 gene was by reporter was from and for shock protein reporter was from Modlin of The expression for a TLR2 and a were from C. B. of was obtained from C. A. The of differential factor and the were by of The and the of were from M. were in for murine monocytic cell and by were in containing and and in a stably transfected with murine COX-2 promoter were as stably transfected with a containing were a from were in and for an to the of to were in the for to the with were in and transfected with murine COX-2 promoter to the was to transfected the of the containing was and for the that were and of the activities were determined for with LPS The that showed the response to LPS was These were as in our previous studies (9Rhee S.H. Hwang D. Murine TOLL-like receptor 4 confers lipopolysaccharide responsiveness as determined by activation of NF kappa B and expression of the inducible cyclooxygenase.J. Biol. Chem. 2000; 275: 34035-34040Google Scholar, 14Lee J.Y. Sohn K.H. Rhee S.H. Hwang D. Saturated fatty acids, but not unsaturated fatty acids, induce the expression of cyclooxygenase-2 mediated through Toll-like receptor 4.J. Biol. Chem. 2001; 276: 16683-16689Google Scholar). or were in and with a containing murine COX-2 promoter or and as an to the expression or for signaling components were The of transfected was by with the in to the from and activities were determined the and to the was by These were the as D. G. M. of induced by through protein and signaling in 1997; Scholar, J.H. J.H. J.Y. Hwang D.H. effects of on the expression of the inducible through different signaling Biol. Chem. 2000; 275: Scholar). COX-2 and were to the was to a membrane in the The membrane was to of in containing COX-2 was by with to in in for COX-2 were and as S.H. E. P. S. H. S. D. Hwang D. expression of in macrophages stimulated with Biol. Chem. 1992; Scholar, D. D. J. E. of and cyclooxygenase-2 in human 1998; Scholar). the membrane for was in the P. L. S. M. G. J.H. M. Y. B. Hwang D. a protein the expression of in macrophages stimulated with lipopolysaccharide in an Chem. Scholar) for with of and by with to in in The membrane was on an detection and to were from studies in human subjects the of dietary n-3 and PUFAs were with fish oil or oil The was by the and subjects In subjects to of fish oil with of acid for whereas in subjects to of fish oil with a of acid for 4 The acid without fish oil The studies and were to a broader of to the of fish oil. The for the in and were to the in The for of fish oil with different of acid in were with the for the of fish oil in Thus, of for the and the of of fish oil were than for the groups of or of fish oil The of the was on the that the of acid not the of for the and of the are D. P. D.H. M. M. oil the beneficial effects to fish oil risk for J. 1997; Scholar). monocytes from blood A for from of human Immunol. Scholar) were with to endogenous expressed in response to LPS was determined by the of in the of acid but not is expressed in macrophages S.H. E. P. S. H. S. D. Hwang D. expression of in macrophages stimulated with Biol. Chem. 1992; Scholar). as by these of COX-2 D. G. M. of induced by through protein and signaling in 1997; Scholar, S.H. E. P. S. H. S. D. Hwang D. expression of in macrophages stimulated with Biol. Chem. 1992; Scholar). we determined the of fish a major dietary of docosahexaenoic acid (DHA, C22:6n-3) and eicosapentaenoic acid (EPA, leads to suppression of COX-2 expression in human monocytes to LPS in was as a for COX-2 The of was significantly suppressed by of fish oil but not by the of that COX-2 expression by LPS in human monocytes was suppressed by the fish oil It also been demonstrated by other that the suppression of the of cytokines and in human by fish oil the of for S. R. K. G. The of dietary with n-3 polyunsaturated fatty acids on the of and factor by J. Med. 1989; Scholar, S. R. R. C.A. with n-3 fatty acids and cell Biol. Scholar). These results that the suppression of COX-2 and cytokine expression by fish oil occurs the were it is that the suppression of the expression of COX-2 and cytokines of fish oil the were the by which n-3 PUFAs inhibit LPS-induced COX-2 expression in human blood we determined the of unsaturated fatty acids in inhibiting the signaling and the target gene expression in murine monocytic cell line (RAW which is stably transfected with NFκB or COX-2 promoter reporter These stably transfected cell the of for the reporter gene and the Thus, or effects of fatty acids on TLR activation be determined in a that LPS-induced NFκB activation and COX-2 expression in is mediated through TLR4 was (9Rhee S.H. Hwang D. Murine TOLL-like receptor 4 confers lipopolysaccharide responsiveness as determined by activation of NF kappa B and expression of the inducible cyclooxygenase.J. Biol. Chem. 2000; 275: 34035-34040Google Scholar). we NFκB activation and COX-2 expression as for TLR activation and its suppression or by fatty acids in in these unsaturated fatty acids inhibit LPS-induced NFκB activation and COX-2 expression as determined by reporter gene unsaturated fatty acids, DHA and EPA are the most This the results from the human studies and that n-3 PUFAs and as with PUFAs acid and are much of TLR4 In contrast, a saturated fatty acid, lauric acid LPS-induced NFκB activation and COX-2 expression Results from previous studies showed that saturated fatty acids without other induce NFκB activation and COX-2 expression in J.Y. Sohn K.H. Rhee S.H. Hwang D. Saturated fatty acids, but not unsaturated fatty acids, induce the expression of cyclooxygenase-2 mediated through Toll-like receptor 4.J. Biol. Chem. 2001; 276: 16683-16689Google Scholar). also showed that LPS-induced COX-2 expression is suppressed by DHA but potentiated by the saturated fatty acid unsaturated fatty acids inhibit the activation of TLR4 in a human were transfected with a of TLR4 to activate signaling in a (4Medzhitov R. Preston-Hurlburt P. Janeway Jr, C.A. A human homologue of the Drosophila Toll protein signals activation of adaptive immunity.Nature. 1997; 388: 394-397Google Scholar). DHA but NFκB activation in These results are with the results the of modulation by fatty acids for the activation of TLR4 in acid a TLR4 but not a differential factor or NFκB activation in were with reporter and the expression of human TLR4 and with DHA or were with reporter and the expression of or and with are different from the determined the target of inhibition by DHA is TLR4 or its downstream signaling components. of the components of the downstream signaling of TLRs is known to be an protein, (1Medzhitov R. Janeway Jr., C. The Toll receptor family and microbial recognition.Trends Microbiol. 2000; 8: 452-456Google Scholar, 2Aderem A. Ulevitch R.J. Toll-like receptors in the induction of the innate immune response.Nature. 2000; 406: 782-787Google Scholar, 3Heldwein K.A. Golenbock D.T. Fenton M.J. Recent advances in the biology of Toll-like receptors.Mod. Asp. Immunobiol. 2001; 1: 249-252Google Scholar, 5Akira S. Toll-like receptors and innate immunity.Adv. Immunol. 2001; 78: 1-56Google Scholar). The activation of NFκB mediated through is of the major downstream signaling derived from TLRs. DHA not inhibit NFκB activation induced by the activation of downstream or of TLR signaling unsaturated fatty acids inhibit NFκB activation induced by TLR4 agonist (LPS) and TLR4 These results suggest that the molecular target of inhibition by DHA is TLR itself or its associated molecules, but not the downstream components. by saturated fatty acids of bacterial lipopeptides is also for the activation of TLR2 (19Brightbill H.D. Libraty D.H. Krutzik S.R. Yang R.B. Belisle J.T. Bleharski J.R. Maitland M. Norgard M.V. Plevy S.E. Smale S.T. Brennan P.J. Bloom B.R. Godowski P.J. Modlin R.L. Host defense mechanisms triggered by microbial lipoproteins through toll-like receptors.Science. 1999; 285: 732-736Google Scholar). we determined unsaturated fatty acids the activation of TLR2 as TLR4 our we demonstrated that a synthetic of bacterial lipopeptides that are known agonists of activates TLR2 in a system that not TLR2 (4Medzhitov R. Preston-Hurlburt P. Janeway Jr, C.A. A human homologue of the Drosophila Toll protein signals activation of adaptive immunity.Nature. 1997; 388: 394-397Google Scholar, A. A. Cutting edge: toll-like receptor and or in response to Immunol. 2001; Scholar). activates as determined by NFκB activation and its inhibition by a of TLR2 or downstream signaling or in The murine monocytic cell line (RAW 264.7) TLR2 and TLR4 K.A. Godowski P.J. Tissue expression of human Toll-like receptors and differential regulation of Toll-like receptor mRNAs in leukocytes in response to microbes, their products, and cytokines.J. Immunol. 2002; 168: 554-561Google Scholar). Thus, we determined activates endogenous TLR2 in NFκB activation and expression of and this induction was inhibited by a of TLR2 or These results demonstrate that activates expressed TLR2 in and endogenous TLR2 in to the results with TLR4 agonist n-3 and EPA are the most the unsaturated fatty acids for NFκB activation and COX-2 expression in The saturated fatty acid potentiated NFκB activation and COX-2 expression in In DHA but lauric acid NFκB activation induced by in These results demonstrate the effects of the fatty acids on the activation of TLR2 and TLR4. fatty acids be major saturated fatty acids and fatty acids acid PUFAs and n-3 PUFAs The unsaturated fatty acids be to polyunsaturated fatty acids through a of and It is that is the groups of unsaturated fatty acids the H. The effects of of fatty acids the fatty acid of the Lipid Scholar, B. A. P. D. J.H. of of in of human plasma in response to dietary fatty 1992; Scholar, H. of unsaturated n-3 and fatty 2000; Scholar). as acid and EPA be to B. and of S. Scholar, P. A. H. and and biological Scholar, and mechanisms of J. 1992; Scholar). of of our of fatty acids evidence that fatty acids not only are the of and other lipid but also signaling and D. acids and immune in for to 2000; 20: Scholar, G. but not fatty acids inhibit and cell in 2001; Scholar, The of n-3 polyunsaturated fatty Biol. Chem. 2002; Scholar). the of this modulation to different types of dietary the of many DHA and EPA are the major n-3 PUFAs present in and biochemical studies have demonstrated beneficial effects of these n-3 PUFAs in of inflammatory and J. fatty acids. in Scholar, A. effects of n-3 fatty J. Med. Scholar, fatty acids in and and in and J. 1991; Google Scholar, M.J. of and to the risk of in a J. Med. Scholar, K.A. and and risk for in Scholar, M.J. n-3 fatty acids and human and J. Scholar, M. G. A. P. P. C. N. I. G. of different of fish oil on cell in with Scholar, M. G. R. A. I. G. L. N. E. G. of fatty acids on cell in subjects risk for 1992; Google Scholar). However, the mechanisms by which dietary n-3 PUFAs beneficial effects are not It been demonstrated that consuming n-3 PUFAs in fish oil suppressed of cytokines and in blood in response to the TLR4 agonist LPS S. R. K. G. The of dietary with n-3 polyunsaturated fatty acids on the of and factor by J. Med. 1989; Scholar, S. R. R. C.A. with n-3 fatty acids and cell Biol. Scholar, A. R. acid cell and of inflammatory in 1999; Scholar). of from by the dietary of n-3 PUFAs also been demonstrated in human subjects and in mice C.A. R.S. polyunsaturated fatty acids murine and the of and 1997; Scholar, G. B. R. of responses and in and in vitro by fish oil and Immunol. 1989; Scholar). the cytokines and COX-2 to a family of response response induced by and Biochem. 1991; Scholar), we that n-3 PUFAs also the expression of COX-2 through modulation of the signaling to its Indeed, our results demonstrate that the of n-3 PUFAs leads to suppression of LPS-induced COX-2 expression in human The was the of the molecular that the suppression of cytokine or COX-2 expression by n-3 PUFAs as with this we the molecular through which PUFAs signaling pathways. The results in demonstrate that unsaturated fatty acids inhibit LPS-induced COX-2 The results in suggest that the molecular target for the inhibition by DHA is TLR itself or its associated molecules, but not the components of the downstream pathways. we the of n-3 PUFAs in the molecular LPS-induced NFκB activation and COX-2 expression were preferentially inhibited by n-3 PUFAs in in a and This the results of the studies the suppression of LPS-induced COX-2 expression in blood monocytes by of fish oil bacterial lipopeptides also fatty acid for the activation of TLR2 (19Brightbill H.D. Libraty D.H. Krutzik S.R. Yang R.B. Belisle J.T. Bleharski J.R. Maitland M. Norgard M.V. Plevy S.E. Smale S.T. Brennan P.J. Bloom B.R. Godowski P.J. Modlin R.L. Host defense mechanisms triggered by microbial lipoproteins through toll-like receptors.Science. 1999; 285: 732-736Google Scholar), we fatty acids also TLR2 signaling pathways. to the results obtained with TLR4 agonist unsaturated fatty acids inhibited but the saturated fatty acid lauric acid TLR2 NFκB activation and COX-2 these results a by which n-3 PUFAs inhibit the expression of COX-2 that is in of and in many types of tissues D. D. J. E. of and cyclooxygenase-2 in human 1998; Scholar, Jones N. J. R.L. is expressed in human evidence for a Scholar, R.J. A. S. of gene expression in human and Google Scholar, H. Y. R.L. A. S. K. S. H. M. of and in human Scholar, P.J. J.F. S. of and protein in human Scholar). these results suggest that the effects of dietary n-3 PUFAs are mediated in part through the inhibition of signaling and target gene and are for the development of many chonic diseases G. to the of the 2002; Scholar, diseases of an of a 2001; Scholar, C. diseases have an 2001; Scholar). Thus, our results suggest the that the beneficial and detrimental effects of different dietary fatty acids on the risk of chonic inflammatory diseases may in part be mediated through the modulation of Toll-like The for the This was by from the of and and for acid docosahexaenoic acid eicosapentaenoic acid acid lipopolysaccharide nuclear factor κB differential factor acid polyunsaturated fatty acid Toll-like receptors

Toll-like receptor-4 (TLR4) can be activated by nonbacterial agonists, including saturated fatty acids. However, downstream signaling pathways activated by nonbacterial agonists are not known. Thus, we determined the downstream signaling pathways derived from saturated fatty acid-induced TLR4 activation. Saturated fatty acid (lauric acid)-induced NFkappaB activation was inhibited by a dominant-negative mutant of TLR4, MyD88, IRAK-1, TRAF6, or IkappaBalpha in macrophages (RAW264.7) and 293T cells transfected with TLR4 and MD2. Lauric acid induced the transient phosphorylation of AKT. LY294002, dominant-negative (DN) phosphatidylinositol 3-kinase (PI3K), or AKT(DN) inhibited NFkappaB activation, p65 transactivation, and cyclooxygenase-2 (COX-2) expression induced by lauric acid or constitutively active (CA) TLR4. AKT(DN) blocked MyD88-induced NFkappaB activation, suggesting that AKT is a MyD88-dependent downstream signaling component of TLR4. AKT(CA) was sufficient to induce NFkappaB activation and COX-2 expression. These results demonstrate that NFkappaB activation and COX-2 expression induced by lauric acid are at least partly mediated through the TLR4/PI3K/AKT signaling pathway. In contrast, docosahexaenoic acid (DHA) inhibited the phosphorylation of AKT induced by lipopolysaccharide or lauric acid. DHA also suppressed NFkappaB activation induced by TLR4(CA), but not MyD88(CA) or AKT(CA), suggesting that the molecular targets of DHA are signaling components upstream of MyD88 and AKT. Together, these results suggest that saturated and polyunsaturated fatty acids reciprocally modulate the activation of TLR4 and its downstream signaling pathways involving MyD88/IRAK/TRAF6 and PI3K/AKT and further suggest the possibility that TLR4-mediated target gene expression and cellular responses are also differentially modulated by saturated and unsaturated fatty acids.

Toll-like receptor 4 (TLR4) and TLR2 agonists from bacterial origin require acylated saturated fatty acids in their molecules. Previously, we reported that TLR4 activation is reciprocally modulated by saturated and polyunsaturated fatty acids in macrophages. However, it is not known whether fatty acids can modulate the activation of TLR2 or other TLRs for which respective ligands do not require acylated fatty acids. A saturated fatty acid, lauric acid, induced NFkappaB activation when TLR2 was co-transfected with TLR1 or TLR6 in 293T cells, but not when TLR1, 2, 3, 5, 6, or 9 was transfected individually. An n-3 polyunsaturated fatty acid (docosahexaenoic acid (DHA)) suppressed NFkappaB activation and cyclooxygenase-2 expression induced by the agonist for TLR2, 3, 4, 5, or 9 in a macrophage cell line (RAW264.7). Because dimerization is considered one of the potential mechanisms for the activation of TLR2 and TLR4, we determined whether the fatty acids modulate the dimerization. However, neither lauric acid nor DHA affected the heterodimerization of TLR2 with TLR6 as well as the homodimerization of TLR4 as determined by co-immunoprecipitation assays in 293T cells in which these TLRs were transiently overexpressed. Together, these results demonstrate that lauric acid activates TLR2 dimers as well as TLR4 for which respective bacterial agonists require acylated fatty acids, whereas DHA inhibits the activation of all TLRs tested. Thus, responsiveness of different cell types and tissues to saturated fatty acids would depend on the expression of TLR4 or TLR2 with either TLR1 or TLR6. These results also suggest that inflammatory responses induced by the activation of TLRs can be differentially modulated by types of dietary fatty acids.

Atopic dermatitis (AD) is a common and relapsing skin disease that is characterized by skin barrier dysfunction, inflammation, and chronic pruritus. While AD was previously thought to occur primarily in children, increasing evidence suggests that AD is more common in adults than previously assumed. Accumulating evidence from experimental, genetic, and clinical studies indicates that AD expression is a precondition for the later development of other atopic diseases, such as asthma, food allergies, and allergic rhinitis. Although the exact mechanisms of the disease pathogenesis remain unclear, it is evident that both cutaneous barrier dysfunction and immune dysregulation are critical etiologies of AD pathology. This review explores recent findings on AD and the possible underlying mechanisms involved in its pathogenesis, which is characterized by dysregulation of immunological and skin barrier integrity and function, supporting the idea that AD is a systemic disease. These findings provide further insights for therapeutic developments aiming to repair the skin barrier and decrease inflammation.