Related Experiment Video
Updated: Apr 11, 2026
Urinary Bladder Distention Evoked Visceromotor Responses as a Model for Bladder Pain in Mice
Published on: April 27, 2014
Macrophage Migration Inhibitory Factor Mediates PAR-Induced Bladder Pain
Dimitrios E Kouzoukas1, Katherine L Meyer-Siegler2, Fei Ma3
1Research and Development, Lexington Veterans Affairs Medical Center, Lexington, Kentucky, United States of America; Saha Cardiovascular Research Center, University of Kentucky, Lexington, Kentucky, United States of America.
Introduction:
Macrophage migration inhibitory factor (MIF), a pro-inflammatory cytokine, is constitutively expressed in urothelial cells that also express protease-activated receptors (PAR). Urothelial PAR1 receptors were shown to mediate bladder inflammation. We showed that PAR1 and PAR4 activator, thrombin, also mediates urothelial MIF release. We hypothesized that stimulation of urothelial PAR1 or PAR4 receptors elicits release of urothelial MIF that acts on MIF receptors in the urothelium to mediate bladder inflammation and pain. Thus, we examined the effect of activation of specific bladder PAR receptors on MIF release, bladder pain, micturition and histological changes.
Methods:
MIF release was measured in vitro after exposing immortalized human urothelial cells (UROtsa) to PAR1 or PAR4 activating peptides (AP). Female C57BL/6 mice received intravesical PAR1- or PAR4-AP for one hour to determine: 1) bladder MIF release in vivo within one hour; 2) abdominal hypersensitivity (allodynia) to von Frey filament stimulation 24 hours after treatment; 3) micturition parameters 24 hours after treatment; 4) histological changes in the bladder as a result of treatment; 5) changes in expression of bladder MIF and MIF receptors using real-time RT-PCR; 6) changes in urothelial MIF and MIF receptor, CXCR4, protein levels using quantitative immunofluorescence; 7) effect of MIF or CXCR4 antagonism.
Results:
PAR1- or PAR4-AP triggered MIF release from both human urothelial cells in vitro and mouse urothelium in vivo. Twenty-four hours after intravesical PAR1- or PAR4-AP, we observed abdominal hypersensitivity in mice without changes in micturition or bladder histology. PAR4-AP was more effective and also increased expression of bladder MIF and urothelium MIF receptor, CXCR4. Bladder CXCR4 localized to the urothelium. Antagonizing MIF with ISO-1 eliminated PAR4- and reduced PAR1-induced hypersensitivity, while antagonizing CXCR4 with AMD3100 only partially prevented PAR4-induced hypersensitivity.
Conclusions:
Bladder PAR activation elicits urothelial MIF release and urothelial MIF receptor signaling at least partly through CXCR4 to result in abdominal hypersensitivity without overt bladder inflammation. PAR-induced bladder pain may represent an interesting pre-clinical model of Interstitial Cystitis/Painful Bladder Syndrome (IC/PBS) where pain occurs without apparent bladder injury or pathology. MIF is potentially a novel therapeutic target for bladder pain in IC/PBS patients.
Insights
Activation of bladder protease-activated receptors (PAR) triggers the release of macrophage migration inhibitory factor (MIF), leading to hypersensitivity and potential therapeutic targets for bladder pain.
Area of Science:
- Urology
- Immunology
- Pain Research
Background:
- Macrophage migration inhibitory factor (MIF) is a pro-inflammatory cytokine found in urothelial cells.
- Protease-activated receptors (PARs) are present in urothelial cells and implicated in bladder inflammation.
- Thrombin, a PAR1 and PAR4 activator, is known to induce urothelial MIF release.
Purpose of the Study:
- To investigate the hypothesis that urothelial PAR1 or PAR4 receptor stimulation releases MIF, activating MIF receptors and mediating bladder inflammation and pain.
- To examine the effects of specific bladder PAR receptor activation on MIF release, bladder pain, micturition, and histological changes.
Main Methods:
- In vitro assessment of MIF release from human urothelial cells (UROtsa) exposed to PAR1 or PAR4 activating peptides (AP).
- In vivo studies in mice involving intravesical administration of PAR1- or PAR4-AP to evaluate MIF release, abdominal hypersensitivity, micturition parameters, and bladder histology.
- Analysis of bladder MIF and MIF receptor (CXCR4) expression via real-time RT-PCR and quantitative immunofluorescence.
- Investigation of the effects of MIF and CXCR4 antagonism.
Main Results:
- PAR1- and PAR4-AP stimulated MIF release in both in vitro and in vivo models.
- Intravesical PAR1- or PAR4-AP induced abdominal hypersensitivity 24 hours post-treatment, without altering micturition or bladder histology.
- PAR4-AP demonstrated greater efficacy, increasing bladder MIF and urothelial CXCR4 expression.
- MIF antagonism (ISO-1) abolished PAR4-induced hypersensitivity and reduced PAR1-induced hypersensitivity, while CXCR4 antagonism (AMD3100) only partially inhibited PAR4-induced hypersensitivity.
Conclusions:
- Bladder PAR activation leads to urothelial MIF release and signaling, partly via CXCR4, causing abdominal hypersensitivity without apparent bladder inflammation.
- This PAR-induced bladder pain model may be relevant for understanding Interstitial Cystitis/Painful Bladder Syndrome (IC/PBS).
- MIF presents a potential novel therapeutic target for managing bladder pain in IC/PBS patients.
