cytochrome-c-t and Carcinoma--Transitional-Cell

Intravesical chemotherapy is often used to prevent the recurrence of superficial bladder cancer after transurethral resection. A search for more effective and less toxic intravesical agents is urgently needed. We previously found the in vitro apoptotic effects of silibinin, a natural flavonoid, on high-risk bladder carcinoma cells. Here, we further explored the underlying mechanisms and examined the intravesical efficacy in the prevention and treatment of bladder cancer. Human bladder carcinoma cell line 5637, which has the same molecular features of high-risk superficial bladder cancer, was used as the model system in vitro and in vivo. Autochthonous rat model of bladder cancer induced by intravesical N-methyl-N-nitrosourea (MNU) was used to investigate its intravesical efficacy. Exposure of 5637 cells to silibinin resulted in growth inhibition and induction of caspase-dependent and -independent apoptosis, which was associated with disruption of mitochondrial membrane potential and selective release of cytochrome c, Omi/HtrA2, and apoptosis-inducing factor (AIF) from mitochondria. Silibinin also downregulated survivin and caused nuclear translocation of AIF. Oral silibinin suppressed the growth of 5637 xenografts, which was accompanied with the activation of caspase-3, downregulation of survivin, and increased translocation of AIF. Furthermore, intravesical silibinin effectively inhibited the carcinogenesis and progression of bladder cancer in rats initiated by MNU by reducing the incidence of superficial and invasive bladder lesions without any side effects, which was accompanied with proapoptotic effects. These findings identify the in vitro and in vivo antitumor efficacy of silibinin, and suggest silibinin as an effective and novel intravesical agent for bladder cancer.

Topics: Administration, Intravesical; Animals; Antioxidants; Apoptosis; Apoptosis Inducing Factor; Carcinoma, Transitional Cell; Cell Line, Tumor; Cell Survival; Cytochromes c; Down-Regulation; Female; High-Temperature Requirement A Serine Peptidase 2; Humans; Inhibitor of Apoptosis Proteins; Membrane Potential, Mitochondrial; Mice; Mice, Inbred BALB C; Mice, Nude; Mitochondria; Mitochondrial Proteins; Rats; Rats, Sprague-Dawley; Serine Endopeptidases; Silybin; Silymarin; Survivin; Urinary Bladder Neoplasms; Xenograft Model Antitumor Assays

This study investigated the anticancer activity of Magnolia officinalis on urinary bladder cancer in vitro and in vivo, and elucidated the mechanism of its activity. An aqueous extract of M. officinalis inhibited cell viability and DNA synthesis in cultured human urinary bladder cancer 5637 cells. Inhibition of proliferation was the result of apoptotic induction, because FACS analyses of 5637 cells treated with M. officinalis showed a sub-G1 phase accumulation. M. officinalis extract also increased cytoplasmic DNA-histone complex dose-dependently. These inhibitory effects were associated with the upregulation of proapoptotic molecules Bax, cytochrome c and caspase 3. Treatment of 5637 cells with M. officinalis extract suppressed the expression of matrix metalloproteinase 2 (MMP-2) and MMP-9, as revealed by zymographic and immunoblot analyses. When M. officinalis extract was given to mice simultaneously with the carcinogen N-butyl-N-(4-hydroxybutyl) nitrosamine, which induces urinary bladder tumors, the size of the induced tumors was smaller. Finally, histological data indicated that the histological grade of carcinoma and the depth of invasion were dramatically decreased by treatment with M. officinalis extract in mice with N-butyl-N-(4-hydroxybutyl) nitrosamine-induced urinary bladder tumors. In conclusion, the findings showed that M. officinalis extract exhibited potential chemopreventive activity against urinary bladder tumor in vitro and in vivo.

Topics: Animals; Anticarcinogenic Agents; Apoptosis; bcl-2-Associated X Protein; Butylhydroxybutylnitrosamine; Carcinoma, Transitional Cell; Caspase 3; Cell Line, Tumor; Cell Proliferation; Cytochromes c; DNA; Dose-Response Relationship, Drug; Humans; Magnolia; Male; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Mice; Mice, Inbred C57BL; Nucleic Acid Synthesis Inhibitors; Plant Extracts; Urinary Bladder Neoplasms

Taxol is the most commonly used agent for salvage chemotherapy in transitional cell carcinoma of the urothelium. We examined mechanisms responsible for taxol resistance by using T24 human bladder carcinoma cells.. We used an inhibitor and an activator of the phosphatidylinositol-3 kinase-Akt pathway in cell survival and caspase-3 assays, an HPLC method for determining released cytochrome c and immunoblotting for detecting protein phosphorylation.. Activation of Akt increased paclitaxel resistance by increasing Bad phosphorylation, leading to decreased release of mitochondrial cytochrome c and caspase-3-mediated apoptosis. On the other hand, inhibition of Akt prevented paclitaxel resistance by enhancing the effects of paclitaxel on Bad phosphorylation, mitochondrial cytochrome c release and caspase-3-mediated apoptosis, besides diminishing or abolishing the opposing effects of Akt activation.. Akt-mediated Bad phosphorylation plays an important role in preservation of mitochondrial membrane systems leading to paclitaxel resistance in T24 cells.

Topics: Antineoplastic Agents, Phytogenic; bcl-Associated Death Protein; Carcinoma, Transitional Cell; Caspase 3; Chromones; Cytochromes c; Drug Resistance, Neoplasm; Enzyme Activation; Humans; Mitochondrial Membranes; Morpholines; Oncogene Protein v-akt; Paclitaxel; Phosphatidylinositol 3-Kinases; Phosphorylation; Urinary Bladder Neoplasms

Consumption of the traditional kava preparation was reported to correlate with low and uncustomary gender ratios (more cancer in women than men) of cancer incidences in three kava-drinking countries: Fiji, Vanuatu, and Western Samoa. We have identified flavokawain A, B, and C but not the major kavalactone, kawain, in kava extracts as causing strong antiproliferative and apoptotic effect in human bladder cancer cells. Flavokawain A results in a significant loss of mitochondrial membrane potential and release of cytochrome c into the cytosol in an invasive bladder cancer cell line T24. These effects of flavokawain A are accompanied by a time-dependent decrease in Bcl-x(L), a decrease in the association of Bcl-x(L) to Bax, and an increase in the active form of Bax protein. Using the primary mouse embryo fibroblasts Bax knockout and wild-type cells as well as a Bax inhibitor peptide derived from the Bax-binding domain of Ku70, we showed that Bax protein was, at least in part, required for the apoptotic effect of flavokawain A. In addition, flavokawain A down-regulates the expression of X-linked inhibitor of apoptosis and survivin. Because both X-linked inhibitor of apoptosis and survivin are main factors for apoptosis resistance and are overexpressed in bladder tumors, our data suggest that flavokawain A may have a dual efficacy in induction of apoptosis preferentially in bladder tumors. Finally, the anticarcinogenic effect of flavokawain A was evident in its inhibitory growth of bladder tumor cells in a nude mice model (57% of inhibition) and in soft agar.

Topics: Animals; Apoptosis; bcl-2-Associated X Protein; Carcinoma, Transitional Cell; Caspase 3; Caspase 9; Caspases; Cell Growth Processes; Chalcone; Cytochromes c; Flavonoids; Humans; Inhibitor of Apoptosis Proteins; Kava; Membrane Potentials; Mice; Mice, Nude; Microtubule-Associated Proteins; Mitochondria; Neoplasm Proteins; Plant Extracts; Poly(ADP-ribose) Polymerases; Proteins; Proto-Oncogene Proteins c-bcl-2; Survivin; Urinary Bladder Neoplasms; X-Linked Inhibitor of Apoptosis Protein; Xenograft Model Antitumor Assays