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Transcription factor network downstream of protease activated receptors (PARs) modulating mouse bladder inflammation
Ricardo Saban1, Cindy Simpson, Carole A Davis
1Department of Physiology, The University Oklahoma Health Sciences Center, Oklahoma City, OK 73104, USA. ricardo-saban@ouhsc.edu
Background:
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.
Methods:
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.
Results:
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-v mice 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-v mice.
Conclusion:
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.
Insights
We identified Transcription Factor EB (TFEB) as a key molecule in PAR-mediated bladder inflammation. Targeting TFEB may offer a new therapeutic strategy for inflammatory bladder disorders.
Area of Science:
- Urology
- Immunology
- Molecular Biology
Background:
- Protease-activated receptors (PARs) are present in the urinary bladder and their expression is altered during inflammation.
- Identifying transcription factors (TFs) downstream of PAR activation is crucial for finding new therapeutic targets beyond the receptors themselves.
Purpose of the Study:
- To determine transcription factors (TFs) downstream of protease-activated receptor (PAR) activation in C57BL/6 mouse urinary bladders.
- To investigate the role of mast cells in PAR-mediated bladder inflammation.
Main Methods:
- Utilized a protein/DNA combo array with 345 TF consensus sequences.
- Validated the identified TF using electrophoretic mobility shift assay (EMSA) and immunohistochemistry (IHC).
- Assessed PAR activation response in c-kit receptor deficient (Kit w/Kit w-v) mice and used ChIP/Q-PCR for TFEB target gene analysis.
Main Results:
- Transcription Factor EB (TFEB) was the sole TF commonly upregulated by all PAR-activating peptides (PAR-APs).
- IHC confirmed a correlation between bladder inflammation and TFEB expression in C57BL/6 mice.
- EMSA demonstrated increased TFEB binding activity in C57BL/6 mice but not in Kit w/Kit w-v mice, which showed no inflammation response to PAR activation.
Conclusions:
- This study is the first to report increased TFEB expression in bladder inflammation following PAR activation.
- TFEB, essential for mast cell survival, may influence mast cell participation in PAR-mediated inflammation.
- Targeting TFEB/MiTF activity presents a potential novel therapeutic approach for inflammatory bladder disorders.
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