Linoleic acid induces interleukin-8 production by Crohn's human intestinal smooth muscle cells via arachidonic acid metabolites

Abstract

Previously we reported that linoleic acid (LA), but not oleic acid, caused a marked increase in the secretion of IL-8 by Crohn's human intestinal smooth muscle (HISM) cells. Antioxidants inhibited this response, implicating a role for oxidative stress and NF-κB, a transcription factor for IL-8 that is activated by oxidative stress. In this study, we examined two mechanisms whereby LA, the dietary precursor for arachidonic acid (AA), could increase the production of IL-8 via activation of AA pathways: 1) by generation of reactive oxygen species by the AA-pathway enzymes to activate NF-κB or 2) by AA metabolites. Normal and Crohn's HISM cells were exposed to LA, oxidizing solution (Ox), or oxidizing solution enriched with LA (OxLA). Exposure of cells to Ox or OxLA induced oxidative stress as determined by thiobarbituric acid reactive substances. In normal cells, Ox but not LA activated NF-κB as determined by transfection experiments and Western blot. In Crohn's cells, NF-κB was spontaneously activated and was not further activated by Ox or LA. In contrast, TNF-α markedly increased activation of NF-κB in both normal and Crohn's cells. These results indicated that LA did not increase IL-8 by activating NF-κB, so we evaluated the second mechanism of an effect of AA metabolites. In normal cells, OxLA, but not LA, markedly stimulated IL-8, whereas in Crohn's cells, both OxLA and LA stimulated IL-8. OxLA, also stimulated production of AA metabolites leukotriene B4(LTB4), PGE2, and thromboxane B2(TXB2) by normal and Crohn's cells. To determine whether AA metabolites mediated the IL-8 response, cells were treated with OxLA plus indomethacin (Indo), a cyclooxygenase inhibitor, and nordihydroguaiaretic acid (NDGA), a lipoxygenase inhibitor. Both Indo and NDGA blocked the IL-8 response to OxLA. To determine more specifically a role for AA metabolites, AA was used. Similar to OxLA, OxAA stimulated production of IL-8 and AA metabolites. Pinane thromboxane, a selective thromboxane synthase inhibitor and receptor blocker, inhibited OxAA stimulation of TXB2and IL-8 in a dose-response manner. MK886, a selective 5-lipoxygenase inhibitor, inhibited OxAA stimulation of LTB4and IL-8 also in a dose-response manner. Analysis of specific gene products by RT-PCR demonstrated that HISM cells expressed receptors for both thromboxane and LTB4. We conclude that AA metabolites mediated the IL-8 response to LA in HISM cells. Both cyclooxygenase and lipoxygenase pathways were involved. LA did not increase IL-8 by activating NF-κB, but NF-κB appeared to be involved, because LA increased IL-8 only in situations where NF-κB was activated, either spontaneously in Crohn's cells or by Ox in normal cells. We speculate that AA metabolites increased IL-8 production by enhancing NF-κB-dependent transcription of IL-8.

Group VIA Phospholipase A2 (iPLA2β) Participates in Angiotensin II-induced Transcriptional Up-regulation of Regulator of G-protein Signaling-2 in Vascular Smooth Muscle Cells

“…To determine whether AA metabolites are involved in Ang II-induced RGS2 mRNA up-regulation in VSMC, cells were incubated with the lipoxygenase inhibitor nordihydroguaiaretic acid, the cyclooxygenase inhibitor indomethacin, or the cytochrome P450-dependent epoxygenases inhibitor 17-octadecynoic acid, respectively, prior to Ang II or AA stimulation. Interestingly, we found that nordihydroguaiaretic acid (30 M) (60, 61) completely and indomethacin (50 M) (61,62) partially, but significantly, inhibited Ang II or AA-stimulated RGS2 mRNA up-regulation, whereas 17-octadecynoic acid (10 M) (63) had no inhibitory effect (Fig. 8E).…”

Heme induces DNA damage and hyperproliferation of colonic epithelial cells via hydrogen peroxide produced by heme oxygenase: A possible mechanism of heme‐induced colon cancer

“…IL-8 production in intestinal epithelial cells increases in response to pro-inflammatory cytokines such as tumor necrosis factor-a [37,38] and IL-1 [32], or oxidative stress such as that by H 2 O 2 [32,39,40]. To determine whether We next treated Caco-2 cells with hemin at various concentrations for 24 h, and the culture medium was collected for IL-8 determination by ELISA.…”

Omega-3 fatty acids exacerbate DSS-induced colitis through decreased adiponectin in colonic subepithelial myofibroblasts

“…6,12 Among the various unsaturated fatty acids, many studies have shown that -3 polyunsaturated fatty acids have a better effect than that of -6 polyunsaturated fatty acids. 7,8,[13][14][15][16] However, the effects of specific forms of dietary fat on IBD remain controversial. 2,3 Attention has recently been paid to adiponectin, a substance released from adipocytes, because of its involvement in the pathophysiology of many lifestyle-related diseases such as diabetes mellitus, 17 atherosclerosis, 17 and nonalcoholic steatohepatitis.…”