PURPOSE: Design short hairpin RNA (shRNA) interference sequence to silence glutathione S-transferase P1 (GSTP1) gene of androgen-independent prostate cancer cell line DU145, and then to explore its effect on sensitivity to chemotherapeutics. METHODS: Target sequence was picked up to form the shRNA. DU145 cell was divided into five groups according to the shRNA added for transfection: shRNA255, shRNA554, shRNA593, negative-shRNA and blank group. Fluorescence microscope was used to pick up the shRNA with the highest transfection ratio. Western blotting and RT-PCR were taken to pick up the shRNA with the best gene silencing result. 3-(4,5-Dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide assay and terminal de-oxynucleotidyl transferase-mediated dUTP nick end-labeling assay were used to detect survival ratio and apoptosis ratio of DU145 administered of fluorouracil (5-FU) or paclitaxel (PA) at different concentrations before and after shRNA transfection. RESULTS: Three different shRNA oligonucleotides (shRNA255; shRNA554; shRNA593) targeting the coding sequence of GSTP1 mRNA and one negative control shRNA were constructed. The transfection ratio of shRNA554 (76.2 ± 0.68 %) was higher than that of shRNA255 (63.3 ± 1.04 %) (P < 0.01) or shRNA593 (72.7 ± 0.33 %) (P < 0.01). After transfection of shRNA554, the mRNA and protein of level were the lowest, P < 0.01. The survival ratio of DU145 administered with 5-FU of different concentrations (30, 60, 120, 240 μg/ml) declined after transfection (P < 0.01). Besides, the apoptosis ratio increased after transfection (P < 0.01). Similarly the survival ratio of DU145 administered with PA of different concentrations (0.2, 2, 10, 20 μg/ml) declined (P < 0.01) and the apoptosis ratio increased (P < 0.01) after transfection. CONCLUSIONS: The gene GSTP1 silence via shRNA transfection to androgen-independent prostate cancer cell line DU145 enhances the sensitivity to chemotherapeutics.
PURPOSE: Design short hairpin RNA (shRNA) interference sequence to silence glutathione S-transferase P1 (GSTP1) gene of androgen-independent prostate cancer cell line DU145, and then to explore its effect on sensitivity to chemotherapeutics. METHODS: Target sequence was picked up to form the shRNA. DU145 cell was divided into five groups according to the shRNA added for transfection: shRNA255, shRNA554, shRNA593, negative-shRNA and blank group. Fluorescence microscope was used to pick up the shRNA with the highest transfection ratio. Western blotting and RT-PCR were taken to pick up the shRNA with the best gene silencing result. 3-(4,5-Dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide assay and terminal de-oxynucleotidyl transferase-mediated dUTP nick end-labeling assay were used to detect survival ratio and apoptosis ratio of DU145 administered of fluorouracil (5-FU) or paclitaxel (PA) at different concentrations before and after shRNA transfection. RESULTS: Three different shRNA oligonucleotides (shRNA255; shRNA554; shRNA593) targeting the coding sequence of GSTP1 mRNA and one negative control shRNA were constructed. The transfection ratio of shRNA554 (76.2 ± 0.68 %) was higher than that of shRNA255 (63.3 ± 1.04 %) (P < 0.01) or shRNA593 (72.7 ± 0.33 %) (P < 0.01). After transfection of shRNA554, the mRNA and protein of level were the lowest, P < 0.01. The survival ratio of DU145 administered with 5-FU of different concentrations (30, 60, 120, 240 μg/ml) declined after transfection (P < 0.01). Besides, the apoptosis ratio increased after transfection (P < 0.01). Similarly the survival ratio of DU145 administered with PA of different concentrations (0.2, 2, 10, 20 μg/ml) declined (P < 0.01) and the apoptosis ratio increased (P < 0.01) after transfection. CONCLUSIONS: The gene GSTP1 silence via shRNA transfection to androgen-independent prostate cancer cell line DU145 enhances the sensitivity to chemotherapeutics.
Authors: David I Quinn; Catherine M Tangen; Maha Hussain; Primo N Lara; Amir Goldkorn; Carol M Moinpour; Mark G Garzotto; Philip C Mack; Michael A Carducci; J Paul Monk; Przemyslaw W Twardowski; Peter J Van Veldhuizen; Neeraj Agarwal; Celestia S Higano; Nicholas J Vogelzang; Ian M Thompson Journal: Lancet Oncol Date: 2013-07-17 Impact factor: 41.316
Authors: J Ferlay; E Steliarova-Foucher; J Lortet-Tieulent; S Rosso; J W W Coebergh; H Comber; D Forman; F Bray Journal: Eur J Cancer Date: 2013-02-26 Impact factor: 9.162
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