Transcription factor network downstream of protease activated receptors (PARs) modulating mouse bladder inflammation

<p>Abstract</p> <p>Background</p> <p>All four PARs are present in the urinary bladder, and their expression is altered during inflammation. In order to search for therapeutic targets other than the receptors themselves, we set forth to determine TFs downstream of PAR activation in the C57BL/6 urinary bladders.</p> <p>Methods</p> <p>For this purpose, we used a protein/DNA combo array containing 345 different TF consensus sequences. Next, the TF selected was validated by EMSA and IHC. As mast cells seem to play a fundamental role in bladder inflammation, we determined whether c-kit receptor deficient (Kit^w/Kit^w-v) mice have an abrogated response to PAR stimulation. Finally, TFEB antibody was used for CHIP/Q-PCR assay and revealed up-regulation of genes known to be downstream of TFEB.</p> <p>Results</p> <p>TFEB, a member of the MiTF family of basic helix-loop-helix leucine zipper, was the only TF commonly up-regulated by all PAR-APs. IHC results confirm a correlation between inflammation and TFEB expression in C57BL/6 mice. In contrast, Kit^w/Kit^w-vmice did not exhibit inflammation in response to PAR activation. EMSA results confirmed the increased TFEB binding activity in C57BL/6 but not in Kit^w/Kit^w-vmice.</p> <p>Conclusion</p> <p>This is the first report describing the increased expression of TFEB in bladder inflammation in response to PAR activation. As TFEB belongs to a family of TFs essential for mast cell survival, our findings suggest that this molecule may influence the participation of mast cells in PAR-mediated inflammation and that targeting TFEB/MiTF activity may be a novel approach for the treatment of bladder inflammatory disorders.</p

Partial Protection against Helicobacter pylori in the Absence of Mast Cells in Mice▿

The goal of this study is to evaluate the contribution of mast cells to Helicobacter pylori immunity in a model of vaccine-induced protection. Mast cell-deficient KitlSl/KitlSl-d and control mice were immunized with H. pylori sonicate plus cholera toxin and challenged with H. pylori, and the bacterial loads, inflammatory infiltrates, and cytokine responses were evaluated and compared at 1, 2, and 4 weeks postchallenge. In vitro stimulation assays were performed using bone marrow-derived mast cells, and recall assays were performed with spleen cells of immunized mast cell-deficient and wild-type mice. Bacterial clearance was observed by 2 weeks postchallenge in mast cell-deficient mice. The bacterial load was reduced by 4.0 log CFU in wild-type mice and by 1.5 log CFU in mast cell-deficient mice. Neutrophil numbers in the gastric mucosa of immune KitlSl/KitlSl-d mice were lower than those for immune wild-type mice (P < 0.05). Levels of gastric interleukin-17 (IL-17) and tumor necrosis factor alpha (TNF-α) were also significantly lower in immune KitlSl/KitlSl-d mice than in wild-type mice (P < 0.001). Immunized mast cell-deficient and wild-type mouse spleen cells produced IFN-γ and IL-17 in response to H. pylori antigen stimulation. TNF-α and CXC chemokines were detected in mast cell supernatants after 24 h of stimulation with H. pylori antigen. The results indicate that mast cells are not essential for but do contribute to vaccine-induced immunity and that mast cells contribute to neutrophil recruitment and inflammation in response to H. pylori