Silencing of heat shock protein 70 expression enhances radiotherapy efficacy and inhibits cell invasion in endometrial cancer cell line.
OBJECTIVE
To investigate the role of heat shock proteins 70 (HSP70) in radiosensitivity and invasiveness of endometrial cancer in vitro.


METHODS
HSP70 expression was silenced in relatively radioresistant, well-differentiated human endometrial cancer cell line ISK, using small interference RNA method, or by HSP70 overexpression after transfecting a HSP70-expressing vector. The effect of HSP70 on ISK cell line response to irradiation was evaluated. The surviving fraction was measured using colony-formation assay. Apoptosis was detected by flow cytometry and HSP70 expression was determined by quantitative real-time polymerase chain reaction, western-blot, and/or immunocytochemistry. Cell invasiveness was measured using transwell invasion assay.


RESULTS
HSP70 silencing caused a significant increase in irradiation-induced cell killing in comparison with control cells, with an enhancement factor of 1.27, and in the percentage of apoptotic cells (14.22% vs 6.74%, P = 0.021). After 4 Gy irradiation, mean +/- standard deviation survival fraction in ISK cells was reduced to 0.32 +/- 0.04 in comparison with control values but in ISK/siRNA-HSP70 cells the survival fraction was higher and amounted to 0.51 +/- 0.08 (P = 0.026). Silencing HSP70 significantly inhibited cell invasion before and after irradiation (106 +/- 19 vs 219 +/- 18 and 119 +/- 16 vs 256 +/- 31, P = 0.007). On the contrary, ectopic overexpression of HSP70 attenuated irradiation-induced apoptosis (7.15% vs 4.08%, P = 0.043) and induced more ISK/HSP70 cells invaded through the filters than mock-infected cells before and after irradiation (274 +/- 21 vs 194 +/- 16 before irradiation, and 298 +/- 24 vs 227 +/- 19 after irradiation, respectively, P = 0.032).


CONCLUSIONS
Disruption of HSP70-induced cytoprotection during irradiation enhances therapeutic effect of irradiation, which makes HSP70 a promising target in the research of endometrial cancer.
Du
Xue-lian
XL
Department of Gynecologic Oncology, Shandong Tumor Hospital, Shandong University, No. 440, Jiyan Road, Jinan, China 250117. melody23800@yahoo.com.cn
Jiang
Tao
T
Wen
Ze-qing
ZQ
Gao
Rong
R
Cui
Min
M
Wang
Fei
F
D000704:Analysis of Variance; D017209:Apoptosis / Q000235:genetics / Q000502:physiology; D015153:Blotting, Western; D045744:Cell Line, Tumor / Q000378:metabolism / Q000528:radiation effects*; D002470:Cell Survival / Q000528:radiation effects; D016889:Endometrial Neoplasms / Q000235:genetics / Q000378:metabolism / Q000532:radiotherapy*; D005260:Female; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic; D020868:Gene Silencing; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D011336:Probability; D012333:RNA, Messenger / Q000032:analysis; D011829:Radiation Dosage; D011836:Radiation Tolerance / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D014162:Transfection
D018840:HSP70 Heat-Shock Proteins; D012333:RNA, Messenger