RESUMEN
The aim of this study was to determine how the function of human stromal antigen 2 (STAG2) plays an important role in proper chromosome separation. STAG2 mRNA in normal bladder cells and bladder tumor cells was evaluated by RT-PCR. The protein levels of STAG2 in normal bladder cells and bladder tumor cells were determined by western blot. A cell proliferation assay was used to measure the growth of tumor cells and STAG2-inhibited normal cells, and STAG2- inhibited normal cells were subjected to karyotype analysis. Both STAG-2 mRNA and protein expression levels were lower in bladder cancer cells compared to the controls. Knockdown of STAG2 caused aneuploidy in normal bladder cells, leading to a decreased expression of the cohesin complex components SMC1, SMC3 and RAD21, but there was no obvious effect of STAG2 knockdown on cell proliferation. Our study indicated that abnormal expression of STAG2 could cause aneuploidy in normal bladder cells.
Asunto(s)
Aneuploidia , Antígenos Nucleares/genética , Expresión Génica , Interferencia de ARN , Antígenos Nucleares/metabolismo , Western Blotting , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Línea Celular , Línea Celular Tumoral , Proteoglicanos Tipo Condroitín Sulfato/genética , Proteoglicanos Tipo Condroitín Sulfato/metabolismo , Proteínas Cromosómicas no Histona/genética , Proteínas Cromosómicas no Histona/metabolismo , Proteínas de Unión al ADN , Humanos , Cariotipificación , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Vejiga Urinaria/citología , Vejiga Urinaria/metabolismo , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/metabolismo , Neoplasias de la Vejiga Urinaria/patologíaRESUMEN
OBJECTIVE: Targeted down-regulation of TGF-ß expression inhibits invasion and metastasis in breast cancer cells. However, the mechanism that TGF-ß functions by remains largely unknown. In the present study we report the mechanism of ERK1/2 dependant S100A4 regulation by TGF-ß and its possible role in TGF-ß-mediated tumour invasion in vitro. MATERIALS AND METHODS: Small interfering RNA targeting TGF-ß1 (TGF-ß1 siRNA) were stably transfected into the breast cancer cell line MDA231. The TGF-ß1 siRNA/9MDA231 cells were then treated with TGF-ß1 (5 ng/ml) or treated with PD98059 (25 µM) or transfected into S100A4 siRNA before TGF-ß1 treatment. The cells were used in several in vitro analyses, including migration, invasion, angiogenesis, and signaling assays. A wound-healing assay was used to determine migration of the cells in culture and a Boyden chamber transwell assay was used for invasion. In vitro angiogenesis studies using conditioned medium in HMEC-1 cells. RESULTS: Inhibition of TGF-ß1 expression by TGF-ß1 siRNA transfection in MDA231 cells showed significant decrease migration, invasion and angiogenesis in vitro. TGF-ß1 siRNA/MDA231 cells treated with 5 ng/ml TGF-ß1 for 24 hs restored the invasive ability of TGF-ß1 siRNA/MDA231 cells. TGF-ß1 treatment could not increase migration, invasion and angiogenesis in TGF-ß1 siRNA/MDA231 cells when treated with 25 µM PD98059 or transfected with S100A4 siRNA before TGF-ß1 treatment. Analysis of TGF-ß1 signaling pathways showed a decrease in p-ERK1/2 activation and an decrease in S100A4 expression. Interestingly, TGF-ß1 regulated S100A4 via ERK1/2 signalling. CONCLUSIONS: Our findings showed that blocking TGF-ß inhibits breast cancer cell invasiveness, migration and angiogenesis via ERK/S100A4 signalling. Therapies targeting the TGF-ß signaling pathway may be more effective to prevent progression in breast cancer.