中华泌尿外科杂志
中華泌尿外科雜誌
중화비뇨외과잡지
CHINESE JOURNAL OF UROLOGY
2010年
9期
604-607
,共4页
膀胱肿瘤%中期因子%化疗敏感性
膀胱腫瘤%中期因子%化療敏感性
방광종류%중기인자%화료민감성
Bladder neoplasrns%Midkine%Chemosensitivity
目的 观察中期因子(MK)基因小干扰RNA(siRNA)对膀胱癌细胞化疗药物敏感性的影响.方法 设计合成3个MK siRNA,转染人膀胱癌RT4细胞株,定量RT-PCR方法筛选下调MK mRNA效果最好的siRNA;以该siRNA转染RT4细胞后分组:空白对照组(Con-A)、空载对照组(Con-B)、错配对照组及MK siRNA 3.125、6.25、12.50、25.00、50.00、100.00、200.00 nmol/L组,实时定量RT-PCR和蛋白质印迹方法检测各组细胞MK表达情况;噻唑盐法检测紫杉醇(PTX)对各组细胞生长的影响;通过检测caspase-3活性和TUNEL方法观察癌细胞凋亡情况.结果 3个siRNA均明显下调MK mRNA水平,以S3效果最好.PTX组、Con-A十PTX组、Con-B+PTX组、siRNA 12.50 nmol/L组、3.125 nmol/L+PTX组、siRNA 6.25 nmol/L+PTX组、siRNA 12.50nmol/L+PTX组肿瘤细胞抑制率分别为(18.21±0.36)%、(18.19±0.29)%、(17.89±0.33)%、(1.86±0.52)%、(32.56±0.53)%、(53.83±0.38)%、(78.95±0.55)%.TUNEL结果显示,ConA、Con-B、siRNA 3.125、6.25、12.50 nmol/L组凋亡指数分别为(1.81±0.36)%、(1.89±0.38)%、(5.56±0.58)%、(9.68±0.55)%和(15.25±0.56)%.结果 MK基因siRNA可增强膀胱癌细胞化疗敏感性,诱导凋亡是其重要机制之一.
目的 觀察中期因子(MK)基因小榦擾RNA(siRNA)對膀胱癌細胞化療藥物敏感性的影響.方法 設計閤成3箇MK siRNA,轉染人膀胱癌RT4細胞株,定量RT-PCR方法篩選下調MK mRNA效果最好的siRNA;以該siRNA轉染RT4細胞後分組:空白對照組(Con-A)、空載對照組(Con-B)、錯配對照組及MK siRNA 3.125、6.25、12.50、25.00、50.00、100.00、200.00 nmol/L組,實時定量RT-PCR和蛋白質印跡方法檢測各組細胞MK錶達情況;噻唑鹽法檢測紫杉醇(PTX)對各組細胞生長的影響;通過檢測caspase-3活性和TUNEL方法觀察癌細胞凋亡情況.結果 3箇siRNA均明顯下調MK mRNA水平,以S3效果最好.PTX組、Con-A十PTX組、Con-B+PTX組、siRNA 12.50 nmol/L組、3.125 nmol/L+PTX組、siRNA 6.25 nmol/L+PTX組、siRNA 12.50nmol/L+PTX組腫瘤細胞抑製率分彆為(18.21±0.36)%、(18.19±0.29)%、(17.89±0.33)%、(1.86±0.52)%、(32.56±0.53)%、(53.83±0.38)%、(78.95±0.55)%.TUNEL結果顯示,ConA、Con-B、siRNA 3.125、6.25、12.50 nmol/L組凋亡指數分彆為(1.81±0.36)%、(1.89±0.38)%、(5.56±0.58)%、(9.68±0.55)%和(15.25±0.56)%.結果 MK基因siRNA可增彊膀胱癌細胞化療敏感性,誘導凋亡是其重要機製之一.
목적 관찰중기인자(MK)기인소간우RNA(siRNA)대방광암세포화료약물민감성적영향.방법 설계합성3개MK siRNA,전염인방광암RT4세포주,정량RT-PCR방법사선하조MK mRNA효과최호적siRNA;이해siRNA전염RT4세포후분조:공백대조조(Con-A)、공재대조조(Con-B)、착배대조조급MK siRNA 3.125、6.25、12.50、25.00、50.00、100.00、200.00 nmol/L조,실시정량RT-PCR화단백질인적방법검측각조세포MK표체정황;새서염법검측자삼순(PTX)대각조세포생장적영향;통과검측caspase-3활성화TUNEL방법관찰암세포조망정황.결과 3개siRNA균명현하조MK mRNA수평,이S3효과최호.PTX조、Con-A십PTX조、Con-B+PTX조、siRNA 12.50 nmol/L조、3.125 nmol/L+PTX조、siRNA 6.25 nmol/L+PTX조、siRNA 12.50nmol/L+PTX조종류세포억제솔분별위(18.21±0.36)%、(18.19±0.29)%、(17.89±0.33)%、(1.86±0.52)%、(32.56±0.53)%、(53.83±0.38)%、(78.95±0.55)%.TUNEL결과현시,ConA、Con-B、siRNA 3.125、6.25、12.50 nmol/L조조망지수분별위(1.81±0.36)%、(1.89±0.38)%、(5.56±0.58)%、(9.68±0.55)%화(15.25±0.56)%.결과 MK기인siRNA가증강방광암세포화료민감성,유도조망시기중요궤제지일.
Objective To study the effects of knock down midkine(MK)by siRNA on chemosinsitivity in bladder cancer cells. Methods Three MK siRNAs were designed and constructed. After transfected with MK siRNA or scrambled siRNA for different time, cultured cells were harvested to carry on the next experiments. Expression of MK was determined by real-time RT-PCR and western blot, and apoptosis were evaluated by caspase-3 activity and TUNEL assay. MTT was performed to examine the inhibition effect of Paclitaxel (PTX) on cells. Results MK siRNA could down-regulate the MK expression in a dose- and time-dependent manner. The MTT results showed that the inhibit rates were (18.21±0.36)%, (18.19±0.29)%, (17.89±0.33)%, (1.86±0.52)%, (32.56±0.53) %, (53. 83±0.38) % and (78. 95±0.55) % in different groups PTX alone(0.2μmol/L), ConA +PTX(0.2 μmol/L), Con-B +PTX(0.2 μmol/L), siRNA alone(12. 50 nmol/L), siRNA(3. 125nmol/L) + PTX (0. 2 μmol/L), siRNA (6. 25 nmol/L) + PTX (0. 2 μmol/L) and siRNA (12. 50nmol/L)+PTX(0.2 μmol/L), respectively. The TUNEL results showed that apoptosis index was (1.81 ±0. 36)%, (1. 89±0. 38)%, (5. 56±0. 58)%, (9. 68±0.55)% and (15. 25±0.56)% in different groups (Con-A, Con-B, siRNA (3. 125 nmol/L), siRNA (6. 25 nmol/L) and siRNA ( 12. 5nmol/L), respectively. The activity of caspase-3 increased significantly in transefected cells with a dose-and time-dependent manner. Conclusion MK siRNA could sensitize human bladder cancer cells to chemotherapy which might be through the apotosis.
