Prolongation of carrageenan-induced inflammation in human colonic epithelial cells by activation of an NFκB‐BCL10 loop

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Abstract

Carrageenan, a sulfated polysaccharide that is widely used as a food additive, induces inflammatory responses in animal models and human cells. The carrageenan-induced inflammatory cascades involve toll-like receptor (TLR)4- and B-cell leukemia/lymphoma (BCL)10-dependent activation of NF-κB, leading to increased IL-8 production. Translocations involving BCL10 in the mucosa-associated lymphoid tissue (MALT) lymphomas are associated with constitutive activation of NF-κB. This report presents a mechanism by which carrageenan exposure leads to prolonged activation of both BCL10 and NF-κB in human colonic epithelial cells. Study findings demonstrate that nuclear RelA and RelB bind to an NF-κB binding motif in the BCL10 promoter in human colonic epithelial NCM460 and HT-29 cells. In vitro oligonucleotide binding assay, non-radioactive gel shift assay, and chromatin immunoprecipitation (ChIP) indicate binding of RelA and RelB to the BCL10 promoter. Prolonged inflammation follows activation of the BCL10-NFκB inflammatory loop in response to carrageenan, shown by increased BCL10, RelA, and IL-8 for 36 to 48 h and increased RelB for 24 h following withdrawal of carrageenan after 12 h. In contrast, exposure to dextran sulfate sodium, which does not cause inflammation through TLR4 and BCL10 in the colonic epithelial cells, did not provoke prolonged activation of inflammation. The carrageenan-enhanced BCL10 promoter activity was blocked by caffeic acid phenethyl ester (CAPE) and MB-132 which inhibit NF-κB activation. These results indicate that NF-κB binding to the BCL10 promoter can lead to prolonged activation of the carrageenan-induced inflammatory cascade by a transcriptional mechanism involving an NF-κB‐BCL10 loop.

Highlights

► Exposure to common food additive carrageenan initiates protracted inflammation. ► The presence of NF-κB binding site in the BCL10 promoter sets up inflammatory loop. ► Both RelA and RelB bind to the NF-κB binding site in the BCL10 promoter. ► In contrast to carrageenan, DSS exposure does not initiate prolonged inflammation.

Abbreviations

BCL10
B-cell leukemia/lymphoma 10
CAPE
caffeic phenethyl ester
CARMA
caspase recruitment domain membrane-associated guanylate kinase
CBM
CARMA-BCL10-MALT complex
CGN
carrageenan
ChIP
chromatic immunoprecipitation
DSS
dextran sulfate sodium
Hsp
heat-shock protein
MALT
mucosa-associated lymphoid tissue
IκB
inhibitor of κB
IKK
inhibitor of IκB kinase
IL-8
Interleukin-8
IRAK
Interleukin-β receptor associated kinase
NF
NF-κB consensus oligonucleotide
NF-κB
nuclear factor kappaB
NFE
experimental NF-κB binding site in BCL10 promoter
NFEM
mutated experimental NF-κB binding site in BCL10 promoter
NIK
NF-κB inducing kinase
PAF
platelet-activating factor
ROS
reactive oxygen species
TLR4
Toll-like receptor 4
Ub
ubiquitin

Keywords

NF-kappaB
BCL10
Carrageenan
Inflammation
IL-8
DSS

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Co-first authors.