Discriminators of mouse bladder response to intravesical Bacillus Calmette-Guerin (BCG).
BACKGROUND
Intravesical Bacillus Calmette-Guerin (BCG) is an effective treatment for bladder superficial carcinoma and it is being tested in interstitial cystitis patients, but its precise mechanism of action remains poorly understood. It is not clear whether BCG induces the release of a unique set of cytokines apart from its pro-inflammatory effects. Therefore, we quantified bladder inflammatory responses and alterations in urinary cytokine protein induced by intravesical BCG and compared the results to non-specific pro-inflammatory stimuli (LPS and TNF-alpha). We went further to determine whether BCG treatment alters cytokine gene expression in the urinary bladder.


METHODS
C57BL/6 female mice received four weekly instillations of BCG, LPS, or TNF-alpha. Morphometric analyses were conducted in bladders isolated from all groups and urine was collected for multiplex analysis of 18 cytokines. In addition, chromatin immune precipitation combined with real-time polymerase chain reaction assay (CHIP/Q-PCR) was used to test whether intravesical BCG would alter bladder cytokine gene expression.


RESULTS
Acute BCG instillation induced edema which was progressively replaced by an inflammatory infiltrate, composed primarily of neutrophils, in response to weekly administrations. Our morphological analysis suggests that these polymorphonuclear neutrophils are of prime importance for the bladder responses to BCG. Overall, the inflammation induced by BCG was higher than LPS or TNF-alpha treatment but the major difference observed was the unique granuloma formation in response to BCG. Among the cytokines measured, this study highlighted the importance of IL-1beta, IL-2, IL-3, IL-4, IL-6, IL-10, IL-17, GM-CSF, KC, and Rantes as discriminators between generalized inflammation and BCG-specific inflammatory responses. CHIP/Q-PCR indicates that acute BCG instillation induced an up-regulation of IL-17A, IL-17B, and IL-17RA, whereas chronic BCG induced IL-17B, IL-17RA, and IL-17RB.


CONCLUSIONS
To the best of our knowledge, the present work is the first to report that BCG induces an increase in the IL-17 family genes. In addition, BCG induces a unique type of persisting bladder inflammation different from TNF-alpha, LPS, and, most likely, other classical pro-inflammatory stimuli.
Saban
Marcia R
MR
College of Medicine, Department of Physiology, Oklahoma University Health Sciences Center, Ohlahoma City, OK 73104, USA. marcia-saban@ouhsc.edu <marcia-saban@ouhsc.edu>
Simpson
Cindy
C
Davis
Carole
C
Wallis
Gemma
G
Knowlton
Nicholas
N
Frank
Mark Barton
MB
Centola
Michael
M
Gallucci
Randle M
RM
Saban
Ricardo
R
D000283:Administration, Intravesical; D000818:Animals; D001500:BCG Vaccine / Q000008:administration & dosage*; D047369:Chromatin Immunoprecipitation; D003556:Cystitis / Q000139:chemically induced* / Q000473:pathology / Q000652:urine*; D016207:Cytokines / Q000235:genetics / Q000378:metabolism* / Q000652:urine; D004195:Disease Models, Animal; D005260:Female; D015870:Gene Expression / Q000187:drug effects; D006099:Granuloma / Q000139:chemically induced / Q000473:pathology / Q000652:urine; D007150:Immunohistochemistry; D020381:Interleukin-17 / Q000235:genetics / Q000378:metabolism / Q000652:urine*; D008070:Lipopolysaccharides / Q000008:administration & dosage; D051379:Mice; D008810:Mice, Inbred C57BL; D009504:Neutrophils / Q000473:pathology / Q000648:ultrastructure; D014409:Tumor Necrosis Factor-alpha / Q000008:administration & dosage; D015854:Up-Regulation; D001743:Urinary Bladder / Q000187:drug effects* / Q000276:immunology / Q000648:ultrastructure
D001500:BCG Vaccine; D016207:Cytokines; D020381:Interleukin-17; D008070:Lipopolysaccharides; D014409:Tumor Necrosis Factor-alpha