| Summary: | Although long regarded as providing a passive physical barrier to the contents of
the lumen, a large body of evidence implicates intestinal epithelial cells (lECs) as playing a central role in the regulation of the gut immune response. One of the major immune functions of intestinal epithelial cells is the production of chemotactic cytokines, known as chemokines, which are responsible for the recruitment and activation of the underlying mononuclear cells, neutrophils, and dendritic cells. Several soluble agonists, including bacterial products as well as host-derived cytokines, are capable of driving this
response. The prototypic proinflammatory agonist interleukin-1β (IL-1β), signals
through the interleukin-1 receptor (IL-1R), the founding member of the IL-1R superfamily, and robustly activates this pro-inflammatory cascade. Synthesis of many
downstream chemokines in response to IL-1β, requires the activation of the transcription factor NFKB. The objective of the work presented here is to characterize
the regulatory pathways responsible for chemokine synthesis in lECs in response to IL-1β and to examine their relationship with NFKB. Three important pathways in IL-1β
signaling were examined. Initially the MAPK family of pathways was examined. IL-1β
treatment led to activation of ERK, p38 MAPK, as well as JNK. Inhibition of ERK had no effect on either chemokine synthesis, or NFKB activation. Inhibition of p38 MAPK
caused a 50% reduction in IL-1β induced chemokine release. This effect was due to
regulation of the IL-8 promoter, and independent of NFKB and posttranscriptional
regulation. JNK inhibition, using curcumin or a pharmacological inhibitor, attenuated NFKB activation and IL-8 promoter activation; however, only curcumin was able to inhibit chemokine release. The next chapter focuses on regulation by protein kinase CK2. CK2 was found increased in tissues of ulcerative colitis patients when compared to normal uninflamed tissue. Its activity was required for the activation of NFKB, and this was through regulation of p65 transactivation in the nucleus of the cell, via serine 529. As a result, CK2 inhibition leads to attenuation of chemokine synthesis. Finally the phosphatidylinositol-3 kinase (PI3K) pathway was examined. Activation of PI3K, protein kinase B (PKB) and 3'-phosphoinositide dependant kinase (PDK1) were all required for chemokine synthesis as well as NFKB activation, although they had
differing regulatory roles. PI3K and PKB regulated NFKB transactivation, through an
undetermined mechanism. PDK1 regulated the IKK complex, potentially through a
phosphorylation at serine residues 180/181 on IKKa and IKK(3. This resulted in PDK1
regulation of NFKB DNA binding, as well as p65 transactivation through phosphorylation
of serine 536. The work presented here has examined the signaling pathways important
in the regulation of IEC chemokine release. This may provide potential targets for in
vivo studies of chronic inflammation, and thus have implications for chronic
inflammatory conditions such as IBD.
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