ABSTRACT
We investigated how the dual inhibition of the molecular mechanism of the mammalian target of the rapamycin (mTOR) downstreams, P70S6 kinase (P70S6K) and eukaryotic initiation factor 4E (eIF4E), can lead to a suppression of the proliferation and progression of urothelial carcinoma (UC) in an orthotopic mouse non-muscle invasive bladder tumor (NMIBT) model. A KU-7-luc cell intravesically instilled orthotopic mouse NMIBC model was monitored using bioluminescence imaging (BLI) in vivo by interfering with different molecular components using rapamycin and siRNA technology. We then analyzed the effects on molecular activation status, cell growth, proliferation, and progression. A high concentration of rapamycin (10 microM) blocked both P70S6K and elF4E phosphorylation and inhibited cell proliferation in the KU-7-luc cells. It also reduced cell viability and proliferation more than the transfection of siRNA against p70S6K or elF4E. The groups with dual p70S6K and elF4E siRNA, and rapamycin reduced tumor volume and lamina propria invasion more than the groups with p70S6K or elF4E siRNA instillation, although all groups reduced photon density compared to the control. These findings suggest that both the mTOR pathway downstream of eIF4E and p70S6K can be successfully inhibited by high dose rapamycin only, and p70S6K and Elf4E dual inhibition is essential to control bladder tumor growth and progression.
Subject(s)
Animals , Female , Mice , Cell Line , Cell Proliferation/drug effects , Cell Survival/drug effects , Disease Progression , Eukaryotic Initiation Factor-4E/antagonists & inhibitors , Mice, Nude , Mucous Membrane/pathology , Phosphorylation/drug effects , RNA Interference , RNA, Small Interfering , Ribosomal Protein S6 Kinases, 70-kDa/antagonists & inhibitors , Signal Transduction/drug effects , Sirolimus/pharmacology , TOR Serine-Threonine Kinases/antagonists & inhibitors , Urinary Bladder Neoplasms/genetics , Urothelium/pathologyABSTRACT
We investigated how the dual inhibition of the molecular mechanism of the mammalian target of the rapamycin (mTOR) downstreams, P70S6 kinase (P70S6K) and eukaryotic initiation factor 4E (eIF4E), can lead to a suppression of the proliferation and progression of urothelial carcinoma (UC) in an orthotopic mouse non-muscle invasive bladder tumor (NMIBT) model. A KU-7-luc cell intravesically instilled orthotopic mouse NMIBC model was monitored using bioluminescence imaging (BLI) in vivo by interfering with different molecular components using rapamycin and siRNA technology. We then analyzed the effects on molecular activation status, cell growth, proliferation, and progression. A high concentration of rapamycin (10 microM) blocked both P70S6K and elF4E phosphorylation and inhibited cell proliferation in the KU-7-luc cells. It also reduced cell viability and proliferation more than the transfection of siRNA against p70S6K or elF4E. The groups with dual p70S6K and elF4E siRNA, and rapamycin reduced tumor volume and lamina propria invasion more than the groups with p70S6K or elF4E siRNA instillation, although all groups reduced photon density compared to the control. These findings suggest that both the mTOR pathway downstream of eIF4E and p70S6K can be successfully inhibited by high dose rapamycin only, and p70S6K and Elf4E dual inhibition is essential to control bladder tumor growth and progression.
Subject(s)
Animals , Female , Mice , Cell Line , Cell Proliferation/drug effects , Cell Survival/drug effects , Disease Progression , Eukaryotic Initiation Factor-4E/antagonists & inhibitors , Mice, Nude , Mucous Membrane/pathology , Phosphorylation/drug effects , RNA Interference , RNA, Small Interfering , Ribosomal Protein S6 Kinases, 70-kDa/antagonists & inhibitors , Signal Transduction/drug effects , Sirolimus/pharmacology , TOR Serine-Threonine Kinases/antagonists & inhibitors , Urinary Bladder Neoplasms/genetics , Urothelium/pathologyABSTRACT
We established an orthotopic non-muscle invasive bladder cancer (NMIBC) mouse model expressing the mammalian target of the rapamycin (mTOR) signaling pathway. After intravesical instillation of KU-7-lucs (day 0), animals were subsequently monitored by bioluminescence imaging (BLI) on days 4, 7, 14, and 21, and performed histopathological examination. We also validated the orthotopic mouse model expressing the mTOR signaling pathway immunohistochemically. In vitro BLI photon density was correlated with KU-7-luc cell number (r2 = 0.97, P < 0.01) and in vivo BLI photon densities increased steadily with time after intravesical instillation. The tumor take rate was 84.2%, formed initially on day 4 and remained NMIBC up to day 21. T1 photon densities were significantly higher than Ta (P < 0.01), and histological tumor volume was positively correlated with BLI photon density (r2 = 0.87, P < 0.01). The mTOR signaling pathway-related proteins were expressed in the bladder, and were correlated with the western blot results. Our results suggest successful establishment of an orthotopic mouse NMIBC model expressing the mTOR signaling pathway using KU-7-luc cells. This model is expected to be helpful to evaluate preclinical testing of intravesical therapy based on the mTOR signaling pathway against NMIBC.
Subject(s)
Animals , Female , Humans , Mice , Blotting, Western , Cell Line, Tumor , Disease Models, Animal , Genes, Reporter , Green Fluorescent Proteins/genetics , Immunohistochemistry , Luciferases, Firefly/genetics , Luminescent Measurements , Mice, Nude , Neoplasm Staging , Signal Transduction , TOR Serine-Threonine Kinases/metabolism , Transplantation, Heterologous , Urinary Bladder Neoplasms/metabolismABSTRACT
We found an error in our published article.