title: Bacille Calmette-Guerin can induce cellular apoptosis of urothelial cancer directly through toll-like receptor 7 activation.
abstract: Immunotherapy using bacille Calmette-Guerin (BCG) instillation is the mainstay treatment modality for superficial urothelial cancer (UC) through toll-like receptor (TLR) activation of cognitive immune response. We investigated the roles of TLR7 in the activation of apoptosis in UC cells after BCG treatment. The in vitro cytotoxicity effect of BCG on UC cells was measured by a modified 3-(4,5-dimethylthiazo-2-yl)-2,5-diphenyl tetrazolium assay. Expressions of TLR7 mRNA and protein in native UC cells prior to and after BCG treatment were analyzed using real-time quantitative polymerase chain reaction and western blot methods. Phagocytotic processes after BCG treatment in UC cells were observed microscopically using a specific immunostain, subsequent cellular apoptosis-related signals induced by TLR7 were analyzed by western blot. Low-grade UC cells, TSGH8301, showed significant cellular death (4.23-fold higher than the high-grade UC cells T24 and J82) when treated with BCG and the BCG cytotoxicity was displayed in a dose-time-dependent manner. TSGH8301 cells had the highest content of TLR7 mRNA, 7.2- and 4.5-fold higher than that of T24 and J82 cells, respectively. TLR7 protein expression was also significantly increased in TSGH8301 cells. Phagocytosis-related markers, including beclin 1, ATG2, and LC3, were increased when TSGH8301 cells were treated by BCG. Interleukin-1 receptor-associated kinases 2 and 4 were also increased markedly in TSGH8301 cells. On the contrary, cellular apoptosis of TSGH8301 cells decreased by 34% when TLR7 activation was suppressed by the TLR antagonist IRS661 after BCG treatment. Our findings suggest that well differentiated TCC cells have higher expression of TLR7 and BCG can drive cellular death of TCC cells directly via TLR7 activation and related apoptotic pathway.
authors.affiliation: Division of Urology, Department of Surgery, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan. Electronic address: yuds@ms21.hinet.net.
authors.forename: Dah-Shyong
authors.initials: DS
authors.lastname: Yu
authors.affiliation: Graduate Institute of Life Science, National Defense Medical Center, Taipei, Taiwan.
authors.forename: Chia-Lun
authors.initials: CL
authors.lastname: Wu
authors.affiliation: Division of Urology, Department of Surgery, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan.
authors.forename: Szu-Yuan
authors.initials: SY
authors.lastname: Ping
authors.affiliation: Division of Urology, Department of Surgery, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan.
authors.forename: Cheng
authors.initials: C
authors.lastname: Keng
authors.affiliation: Division of Urology, Department of Surgery, Chi-Mei Medical Center, Tainan, Taiwan; Department of Optometry, Chung Hwa University of Medical Technology, Tainan, Taiwan.
authors.forename: Kun-Hung
authors.initials: KH
authors.lastname: Shen
mesh_terms: D017209:Apoptosis / Q000187:drug effects*; D001500:BCG Vaccine / Q000494:pharmacology*; D002454:Cell Differentiation / Q000187:drug effects; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D010587:Phagocytosis / Q000187:drug effects; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051199:Toll-Like Receptor 7 / Q000819:agonists* / Q000235:genetics / Q000378:metabolism*; D001749:Urinary Bladder Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology*; D019459:Urothelium / Q000187:drug effects / Q000378:metabolism / Q000473:pathology*
chemical_list: D001500:BCG Vaccine; D012333:RNA, Messenger; C495348:TLR7 protein, human; D051199:Toll-Like Receptor 7
keywords: Apoptosis; Bacillus Calmette-Guerin; Toll-like receptors; Urothelial cancer