[Down-regulation of Twist1 increases the sensitivity of nasopharyngeal carcinoma cell lines HNE1 to taxol].
OBJECTIVE
To investigate whether down-regulation of Twist1 could change sensitivity of nasopharyngeal carcinoma cell line HNE1 to taxol.


METHODS
HNE1 cells were transfected with the small interfering RNA (siRNA) expression vector pSuppressor-Retro-Si-Twist, containing the short hairpin RNA (shRNA) sequence targeting the Twist gene-coding region by Fugene 6. Positive clones were then selected in Neomycin (400 microg/ml) for 21 days. The low expressions of Twist1 were examined by real-time reverse-transcription polymerase chain reaction (RT-PCR) and Western blot. Drug sensitivity of si-Twist1 HNE1 to taxol was determined by Annexin V-fluorescein isothiocyanate( FITC)/propidium lodide (PI) double-labeled flow cytometry and detection of DNA ladder. The Effect of Twist1 inactivation on HNE1 cell proliferation was observed by MTT assay and flow cytometry.


RESULTS
Annexin V- FITC-PI assay showed that apoptosis ratio was 40.2% in si-Twist HNE1 after treated with 10 ng/ml taxol, significantly higher than that in the control siRNA group 24.3%. The deference had statistic meaning. After the re-expression of HNE1, apoptosis ratio was 44.80% +/- 4.80% (x +/- s) in low Twist1 protein expression group and that was 27.00% +/- 2.91% in high expression group. The deference had statistic meaning (t = 4.374, P = 0.049). Real time PCR test revealed apoptosis protein bcl-2 expression in si-Twist HNE1 was 0.28 +/- 0.05, significantly lower than that in the control siRNA HNE1 (0.57 +/- 0.08, t = 6.710, P = 0.021), nevertheless, significant bax and bcl-XL changes were not observed (t = 2.000, P = 0.184 and t = 1.502, P = 0.272). MTT and FCM showed that down-regulation of Twist1 did not alter cell proliferation rate (P>0.05).


CONCLUSIONS
Down-regulation of Twist1 could increase drug sensitivity of nasopharyngeal carcinoma cell line HNE1 to taxol by inducing apoptosis. These results suggested that Twist1 may be a promising treatment target for nasopharyngeal carcinoma therapy.
Meng
Da-wei
DW
Department of Otorhinolaryngology, Jinqiu Hospital, Shenyang 110016, China.
Bao
Ji-min
JM
Ma
Yun-peng
YP
Li
Zhe
Z
Li
Su-jie
SJ
Li
Dan
D
D017209:Apoptosis / Q000187:drug effects*; D045744:Cell Line, Tumor; D015536:Down-Regulation; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D009303:Nasopharyngeal Neoplasms / Q000188:drug therapy*; D009687:Nuclear Proteins / Q000235:genetics*; D017239:Paclitaxel / Q000494:pharmacology*; D034622:RNA Interference; D034741:RNA, Small Interfering; D051879:Twist-Related Protein 1 / Q000235:genetics*
D009687:Nuclear Proteins; D034741:RNA, Small Interfering; C103173:TWIST1 protein, human; D051879:Twist-Related Protein 1; D017239:Paclitaxel