Fig 1.
Shigella-Salmonella agar and Congo-red binding assay.
A) Shigella-Salmonella agar showed pink colour colonies in S.dysenteriae inoculated plate (lower panel) and no colonies were found in the control plate (upper panel). B) Appearance of orange/pink colour colonies in S.dysenteriae inoculated plate (lower Panel) and no colonies were observed in the control plate (upper panel).
Fig 2.
Histology of rat ileal loop infected with S.dysenteriae.
A) Control rat ileal loop showed normal and elongated villi. B) S.dysenteriae infected rat ileal loop showed ulceration, inflammatory infiltration, broadening and congestion of the villi. Scale bar represents 50μM.
Fig 3.
Activation of proinflammatory cytokines during S.dysenteriae infection.
A) S.dysenteriae infected rat ileal loop protein lysates were analysed by western blots for the indicated proteins. B) IL-8 and TNF-α mRNA expression in S.dysenteriae infected rat ileal intestinal tissue lysate. IL-8 and TNF-α mRNA levels were normalized with GAPDH. Data are the mean ± SD. (n = 3). *, p <0.05.
Fig 4.
Differential expression of NF-κB and subcellular localization of β-catenin in rat ligated ileal loop model during S.dysenteriae infection.
A) Mild expression of NF-κB was observed in lower part of the crypts in untreated tissue sections whereas increased expression of NF-κB was found in S.dysenteriae infected rat intestinal tissue sections by immunohistochemistry. Scale bar represents 50μM. B) Immunohistochemical analysis of β-catenin showed mild membranous expression in untreated sections whereas weak cytoplasmic expression was found in S.dysenteriae infected tissue sections. Scale bar represents 50μM.
Fig 5.
S.dysenteriae infection promotes binding of NF-κB and β-catenin through GSK-3β.
A) S.dysenteriae infected rat ileal loop protein lysates were analysed by western blots for the indicated proteins. B) S.dysenteriae infected rat intestinal protein lysate were immunoprecipitated with β-catenin and immunoblotted with NF-κB and β-catenin. Nonspecific IgG was used as negative control for immunoprecipitation. C) S.dysenteriae infected rat ileal loop protein lysates analysed by western blots for the indicated proteins. D) Protein lysate from S.dysenteriae infected rat intestinal tissues were immunoprecipitated with GSK-3β and immunoblotted with NF-κB, β-catenin and GSK-3β. Nonspecific IgG was used as negative control for immunoprecipitation.
Fig 6.
Proposed model depicting β-catenin/NF-κB role in regulating Shigella-induced proinflammatory cascade.
S.dysenteriae stimulated phosphorylation of β-catenin through GSK-3β and β-catenin is subsequently degraded in the proteosome, simultaneously NF- κB is liberated from IκBα and it is degraded in the proteosome as that of β-catenin and NF- κB freely translocate to the nucleus. NF- κB activates transcription of IL-8 and other proinflammatory genes in the nucleus, thus leading to inflammation.