Anti-Inflammatory Properties of the Enaminone E121 in the Dextran Sulfate Sodium (DSS) Colitis Model

Abstract

BackgroundEnaminones are synthetic compounds with an established role in the prevention of various forms of seizures. Recent evidence suggests potent anti-tussive, bronchodilation and anti-inflammatory properties. Pre-treatment with particularly E121 compound resulted in a decrease in leukocyte recruitment in the ovalbumin induced-model of asthma, immune cell proliferation and cytokine release in vitro. We hypothesize that E121 might serve as a therapeutic potential in intestinal inflammation through modulating immune cell functions.MethodsColitis was induced by daily dextran sulfate sodium (DSS) administration for 5 days, and its severity was determined by gross and histological assessments. The plasma level of various cytokines was measured using flow cytometry-based assay. The colonic expression/ phosphorylation level of various molecules was determined by immunofluorescence and western blotting. The effects of E121 treatment on in vitro neutrophil chemotaxis (under-agarose assay), superoxide release (luminol oxidation assay) and apoptosis (annexin V/7AAD) were also determined.ResultsDSS-induced colitis in mice was significantly reduced by daily E121 treatment (30–100 mg/kg) at gross and histological levels. This effect was due to modulated plasma levels of interleukin (IL-2) and colonic expression levels of various signaling molecules and proteins involved in apoptosis. In vitro neutrophil survival, chemotaxis, and superoxide release were also reduced by E121 treatment.ConclusionOur results indicate important anti-inflammatory actions of E121 in the pathogenesis of IBD.

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“…Bcl-2 is regarded as a prosurvival molecule, whereas Bax is a proapoptotic molecule as it binds to and antagonizes the effects of Bcl-2 [ 42 ]. Thus, Caspase-3 activation is an important event in cell death [ 43 ]. SCB showed substantial downregulation of proapoptotic genes, such as Bax and Caspase-3 ( P < 0.001 and P < 0.05, resp.).…”

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“…Also, the enhanced total and phosphorylated form of AKT in the colon of DSS‐treated mice were reduced back to expression control levels by E121 treatment, and the enhanced colonic expression levels of p‐ERK1/2 and p38 MAPK post‐DSS treatment were reduced to control levels by E121 treatment. Thus, evidence was shown that the anti‐inflammatory property of E121 is in part mediated through modulating the activity of pro‐inflammatory signalling pathways, colonic epithelial cell apoptosis, neutrophil apoptosis, chemotaxis and superoxide release . Given its potential to decrease pro‐inflammatory cytokines such as TNF‐alpha and IL‐6 in vitro as well as in vivo , E121 may play a role in mitigating cytokines in IBD or RA, without the same adverse effects as glucocorticoids, given their structural differences.…”

“…Further, results indicate that E121 is able to suppress the release of proinflammatory cytokines TNF-alpha and IL-6, as well as moderately decrease total nitric oxide after stimulation with LPS, but not IFN-gamma, indicating that there is a target for this compound in the TLR4 signalling pathway, potentially p38 MAPK. [42] Studies of p38 MAPK inhibition alongside glucocorticoid treatment have suggested that inhibiting p38 can possibly increase glucocorticoid effectiveness by increasing GR nuclear translocation. [43,44] Thus, inhibiting kinases may increase the efficacy of the glucocorticoid in decreasing pro-inflammatory cytokine release.…”

“…With regard to E121 inhibition of cellular signalling, Khajah et al [42] recently reported the enhancement of tumour necrosis factor receptor-associated factor-2 (TRAF-2) which acts as an adaptor that transmits signals through the TNF receptor superfamily, IL-1 and Toll-like receptors [46,47] resulting in NF-kB activation (that is required to prevent epithelial cell apoptosis). The TRAF-2 expression in the colon of dextran sulfate sodium (DSS)treated mice was enhanced, and E121 treatment reduced its expression back to control level.…”

“…Thus, evidence was shown that the anti-inflammatory property of E121 is in part mediated through modulating the activity of pro-inflammatory signalling pathways, colonic epithelial cell apoptosis, neutrophil apoptosis, chemotaxis and superoxide release. [42] Given its potential to decrease pro-inflammatory cytokines such as TNF-alpha and IL-6 in vitro as well as in vivo, E121 may play a role in mitigating cytokines in IBD or RA, without the same adverse effects as glucocorticoids, given their structural differences.…”