• Asian Pacific Journal of Cancer Prevention
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• v.14 no.4
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• pp.2387-2391
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• 2013

Resistance to apoptosis is a major obstacle preventing effective therapy for malignancies. Bcl-2 plays a significant role in inhibiting apoptosis. We reconstructed a stable human Bcl-2 transfected cell line, BIU87-Bcl-2, that was derived from the transfection of human bladder carcinoma cell line BIU87 with a plasmid vector containing recombinant Bcl-2 [pcDNA3.1(+)-Bcl-2]. A cell line transfected with the plasmid alone [pcDNA3.1(+)-neo] was also established as a control. BIU87 and BIU87-neo proved sensitive to adriamycin induced apoptosis, while BIU87-Bcl-2 was more resistant. In view of the growing evidence that NF-${\kappa}B$ may play an important role in regulating apoptosis, we determined whether Bcl-2 could modulate the activity of NF-${\kappa}B$ in bladder carcinoma cells. Stimulation of BIU87, BIU87-neo and BIU87-Bcl-2 with ADR resulted in an increase expression of NF-${\kappa}B$ (p<0.001). The expression of NF-${\kappa}B$ in BIU87-Bcl-2 was higher than in the other two cases, with a concomitant reduction in the $I{\kappa}B{\kappa}$ protein level. These results suggest that the overexpression of Bcl-2 renders human bladder carcinoma cells resistant to adriamycin-induced cytotoxicity and there is a link between Bcl-2 and the NF-${\kappa}B$ signaling pathway in the suppression of apoptosis.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.2
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• pp.895-901
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• 2013

Resistance to induction of apoptosis is a major obstacle for bladder cancer treatment. Bcl-2 is thought to be involved in anti-apoptotic signaling. In this study, we investigated the effect of Bcl-2 overexpression on apoptotic resistance and intracellular reactive oxygen species (ROS) generation in bladder cancer cells. A stable Bcl-2 overexpression cell line, BIU87-Bcl-2, was constructed from human bladder cancer cell line BIU87 by transfecting recombinant Bcl-2 [pcDNA3.1(+)-Bcl-2]. The sensitivity of transfected cells to adriamycin (ADR) was assessed by MTT assay. Apoptosis was examined by flow cytometry and acridine orange fluorescence staining. Intracellular ROS was determined using flow cytometry, and the activities of superoxide dismutase (SOD) and catalase (CAT) were also investigated by the xanthinoxidase and visible radiation methods using SOD and CAT detection kits. The susceptibility of BIU87-Bcl-2 cells to ADR treatment was significantly decreased as compared with control BIU87 cells. Enhanced expression of Bcl-2 inhibited intracellular ROS generation following ADR treatment. Moreover, the suppression of SOD and CAT activity induced by ADR treatment was blocked in the BIU87-Bcl-2 case but not in their parental cells. The overexpression of Bcl-2 renders human bladder cancer cells resistant to ADR-induced apoptosis and ROS might act as an important secondary messenger in this process.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.4
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• pp.1517-1520
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• 2014

Aim: To investigate effects of sulforaphane on the BIU87 cell line and underlying mechanisms involving IGFBP-3. Methods: Both BIU87 and IGFBP-3-silenced BIU87 cells were treated with sulforaphane. Cell proliferation was detected by MTT assay. Cell cycle and apoptosis were determined via flow cytometry. Quantitative polymerase chain reaction and Western blotting were applied to analyze the expression of IGFBP-3 and NF-${\kappa}B$ at both mRNA and protein levels. Results: Sulforaphane (80 ${\mu}M$) treatment could inhibit cell proliferation, inducing apoptosis and cell cycle arrest at G2/M phase. All these effects could be antagonized by IGFBP-3 silencing. Furthermore, sulforaphane (80 ${\mu}M$) could down-regulate NF-${\kappa}B$ expression while elevating that of IGFBP-3. Conclusions: Sulforaphane could suppress the proliferation of BIU87 cells via enhancing IGFBP-3 expression, which negatively regulating the NF-${\kappa}B$ signaling pathway.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.1
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• pp.135-138
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• 2012

Objective: To investigate whether apoptin is a apoptosis-inducing protein with a potential for bladder cancer therapy. Methods: We constructed a PCDNA3/Apoptin eukaryotic expression vector, and transfected this vector into bladder cancer cell lines BIU-87 and EJ, then observed the results by RT-PCR, transmission electron microscopy, MTT assay and the flow cytometry (TUNEL method). Results: PCDNA3/Apoptin successfully induced a high level apoptosis in both bladder cancer cell lines, compared with the controls (p<0.05). Conclusions: Apoptin can induce high level apoptosis in human bladder cancer EJ and BIU-87 cells, which suggests a potential for human bladder cancer therapy.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.8
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• pp.3471-3476
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• 2014

Background: Aberrant expression of the microRNA-29 family is associated with tumorigenesis and cancer progression. As transport carriers, tumor-derived exosomes are released into the extracellular space and regulate multiple functions of target cells. Thus, we assessed the possibility that exosomes could transport microRNA-29c as a carrier and correlations between microRNA-29c and apoptosis of bladder cancer cells. Materials and Methods: A total of 28 cancer and adjacent tissues were examined by immunohistochemistry to detect BCL-2 and MCL-1 expression. Disease was Ta-T1 in 12 patients, T2-T4 in 16, grade 1 in 8, 2 in 8 and 3 in 12. The expression of microRNA-29c in cancer tissues was detected by quantitative reverse transcriptase PCR (QRT-PCR). An adenovirus containing microRNA-29c was used to infect the BIU-87 human bladder cancer cell line. MicroRNA-29c in exosomes was measured by QRT-PCR. After BIU-87 cells were induced by exosomes-derived microRNA-29c, QRT-PCR was used to detect the level of microRNA-29c. Apoptosis was examined by flow cytometry and BCL-2 and MCL-1 mRNA expressions were assessed by reverse transcription-polymerase chain reaction. Western blotting was used to determine the protein expression of BCL-2 and MCL-1. Results: The expressions of BCL-2 and MCL-1 protein were remarkably increased in bladder carcinoma (p<0.05), but was found mainly in the basal and suprabasal layers in adjacent tissues. The expression of microRNA-29c in cancer tissues was negatively correlated with the BCL-2 and MCL-1. The expression level of microRNA-29c in exosomes and BIU-87 cells from the experiment group was higher than that in control groups (p<0.05). Exosome-derived microRNA-29c induced apoptosis (p<0.01). Although only BCL-2 was reduced at the mRNA level, both BCL-2 and MCL-1 were reduced at the protein level. Conclusions: Human bladder cancer cells infected by microRNA-29c adenovirus can transport microRNA-29c via exosomes. Moreover, exosome-derived microRNA29c induces apoptosis in bladder cancer cells by down-regulating BCL-2 and MCL-1.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.6
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• pp.2799-2806
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• 2012

The aim of this study was to elucidate the role of the integrin-linked kinase (ILK) gene in development of human bladder transitional cell carcinoma (BTCC). Expression of ILK protein and ILK mRNA in 56 cases of human BTCC tissue and in 30 cases of adjacent normal bladder tissue was detected by immunohistochemistry S-P and reverse transcription polymerase chain reaction (RT-PCR), respectively. Four specific miRNA RNAi vectors targeting human ILK were synthesized and transfected into BIU-87 cells by liposome to obtain stable expression cell strains. The influence of ILK on proliferation of BTCC was detected by MTT, FCM on athymic mouse tumorigenesis. The positive rate of ILK protein in BTCC tissue (53.6%) was much higher than adjacent normal bladder tissue (10.0%) (p<0.05). Similarly, expression of ILK mRNA in BTCC tissue ($0.540{\pm}0.083$) was significantly higher than in adjacent normal bladder tissue ($0.492{\pm}0.070$) (p<0.05). MTT showed that the proliferation ability of miRNA-ILK transfected group was clearly decreased (p<0.05), the cell cycle being arrested in G0/G1-S, an tumorigenesis in vivo was also significantly reduced (p<0.05). ILK gene transcription and protein expression may be involved in the development of BTCC, so that ILK might be the new marker for early diagnosis and the new target for gene treatment.