Cystitis-Related Bladder Pain Involves ATP-Dependent HMGB1 Release from Macrophages and Its Downstream H2S/Cav3.2 Signaling in Mice

• Published in: Cells (Basel, Switzerland) Vol. 9; no. 8; p. 1748
• Main Authors: Hiramoto, Shiori, Tsubota, Maho, Yamaguchi, Kaoru, Okazaki, Kyoko, Sakaegi, Aya, Toriyama, Yuki, Tanaka, Junichi, Sekiguchi, Fumiko, Ishikura, Hiroyasu, Wake, Hidenori, Nishibori, Masahiro, Nguyen, Huy Du, Okada, Takuya, Toyooka, Naoki, Kawabata, Atsufumi
• Format: Journal Article
• Language: English
• Published: Basel MDPI AG 22-07-2020; MDPI
• Subjects: Acetylcysteine; Acrolein; Antagonists; Antibodies; Antioxidants; ATP; Behavior; Bladder; Calcium channels; Calcium channels (T-type); Calcium channels (voltage-gated); Cav3.2 T-type Ca2+ channel; Channel gating; Cyclophosphamide; cyclophosphamide (CPA); Cystitis; Experiments; Females; high mobility group box 1 (HMGB1); HMGB1 protein; Hydrogen sulfide; hydrogen sulfide (H2S); interstitial cystitis/bladder pain syndrome (IC/BPS); Laboratory animals; Leukocyte migration; Macrophages; Metabolites; NF-κB protein; Pain; Pain perception; Proteins; Reactive oxygen species; receptor for advanced glycation end products (RAGE); United States > US; Japan
• ISSN: 2073-4409; 2073-4409
• DOI: 10.3390/cells9081748

Cystitis-related bladder pain involves RAGE activation by HMGB1, and increased Cav3.2 T-type Ca2+ channel activity by H2S, generated by upregulated cystathionine-γ-lyase (CSE) in mice treated with cyclophosphamide (CPA). We, thus, investigated possible crosstalk between the HMGB1/RAGE and CSE/H2S/Cav3.2 pathways in the bladder pain development. Bladder pain (nociceptive behavior/referred hyperalgesia) and immuno-reactive CSE expression in the bladder were determined in CPA-treated female mice. Cell signaling was analyzed in urothelial T24 and macrophage-like RAW264.7 cells. The CPA-induced bladder pain was abolished by pharmacological inhibition of T-type Ca2+ channels or CSE, and genetic deletion of Cav3.2. The CPA-induced CSE upregulation, as well as bladder pain was prevented by HMGB1 inactivation, inhibition of HMGB1 release from macrophages, antagonists of RAGE or P2X4/P2X7 receptors, and N-acetylcysteine, an antioxidant. Acrolein, a metabolite of CPA, triggered ATP release from T24 cells. Adenosine triphosphate (ATP) stimulated cell migration via P2X7/P2X4, and caused HMGB1 release via P2X7 in RAW264.7 cells, which was dependent on p38MAPK/NF-κB signaling and reactive oxygen species (ROS) accumulation. Together, our data suggest that CPA, once metabolized to acrolein, causes urothelial ATP-mediated, redox-dependent HMGB1 release from macrophages, which in turn causes RAGE-mediated CSE upregulation and subsequent H2S-targeted Cav3.2-dependent nociceptor excitation, resulting in bladder pain.