中国肿瘤临床
中國腫瘤臨床
중국종류림상
CHINESE JOURNAL OF CLINICAL ONCOLOGY
2014年
12期
753-757
,共5页
韩康%赵廷宝%卞娜%蔡成魁%颜世举%王鑫%马琼%沙浩%董川%杨彤涛%周勇%马保安
韓康%趙廷寶%卞娜%蔡成魁%顏世舉%王鑫%馬瓊%沙浩%董川%楊彤濤%週勇%馬保安
한강%조정보%변나%채성괴%안세거%왕흠%마경%사호%동천%양동도%주용%마보안
骨肉瘤%miR-194%慢病毒%成骨肉瘤细胞系U2-OS%转移
骨肉瘤%miR-194%慢病毒%成骨肉瘤細胞繫U2-OS%轉移
골육류%miR-194%만병독%성골육류세포계U2-OS%전이
osteosarcoma%microRNA-194%lentivirus%U2-OS cell line%metastasis
目的：通过构建携带针对miR-194的慢病毒沉默及过表达载体，研究miR-194对骨肉瘤细胞转移的影响。方法：以miR-194前体及成熟体序列为模板，设计引物，进行PCR扩增，酶切后克隆于慢病毒载体plent-i GFP中，转染293T细胞，包装慢病毒颗粒，检测滴度，最终对人骨肉瘤细胞系U2-OS进行感染。获取稳定感染细胞后，进行Transwell、划痕等实验。结果：1）慢病毒表达质粒构建成功，沉默表达及过表达重组慢病毒的滴度分别为4×108TU/mL及1.5×108TU/mL；2）稳定过表达miR-194的人骨肉瘤细胞系U2-OS的转移能力较其他组有明显降低（P<0.01）。结论：成功构建了miR-194慢病毒表达载体，miR-194的表达水平对U2-OS细胞的转移能力具有显著影响，miR-194可能成为骨肉瘤治疗的潜在新靶点。
目的：通過構建攜帶針對miR-194的慢病毒沉默及過錶達載體，研究miR-194對骨肉瘤細胞轉移的影響。方法：以miR-194前體及成熟體序列為模闆，設計引物，進行PCR擴增，酶切後剋隆于慢病毒載體plent-i GFP中，轉染293T細胞，包裝慢病毒顆粒，檢測滴度，最終對人骨肉瘤細胞繫U2-OS進行感染。穫取穩定感染細胞後，進行Transwell、劃痕等實驗。結果：1）慢病毒錶達質粒構建成功，沉默錶達及過錶達重組慢病毒的滴度分彆為4×108TU/mL及1.5×108TU/mL；2）穩定過錶達miR-194的人骨肉瘤細胞繫U2-OS的轉移能力較其他組有明顯降低（P<0.01）。結論：成功構建瞭miR-194慢病毒錶達載體，miR-194的錶達水平對U2-OS細胞的轉移能力具有顯著影響，miR-194可能成為骨肉瘤治療的潛在新靶點。
목적：통과구건휴대침대miR-194적만병독침묵급과표체재체，연구miR-194대골육류세포전이적영향。방법：이miR-194전체급성숙체서렬위모판，설계인물，진행PCR확증，매절후극륭우만병독재체plent-i GFP중，전염293T세포，포장만병독과립，검측적도，최종대인골육류세포계U2-OS진행감염。획취은정감염세포후，진행Transwell、화흔등실험。결과：1）만병독표체질립구건성공，침묵표체급과표체중조만병독적적도분별위4×108TU/mL급1.5×108TU/mL；2）은정과표체miR-194적인골육류세포계U2-OS적전이능력교기타조유명현강저（P<0.01）。결론：성공구건료miR-194만병독표체재체，miR-194적표체수평대U2-OS세포적전이능력구유현저영향，miR-194가능성위골육류치료적잠재신파점。
Objective:This study aimed to construct a lentiviral expression vector for microRNA-194 and investigate its effect on the metastasis of human osteosarcoma cell line U2-OS. Methods:Pri-and mature miR-194 amplified by PCR were inserted into the plenty-GFP vector and identified by restriction endonuclease digestion and nucleotide sequencing. The osteosarcoma cell line U2-OS was transfected with the lentivirus. Then, the stable transfected cells were used in Transwell and wound healing assay. Results:Restric-tion analysis and sequencing showed that the recombinant lentiviral expression vector was constructed correctly. The titers of obtained overexpression and suppression expression recombinant lentivirus were 1.5*108 and 4*108 TU/ml. Cell metastasis ability was signifi-cantly different in different experimental groups (P<0.01). Conclusion:The lentiviral expression vector for microRNA-194 was suc-cessfully constructed. MicroRNA-194 could influence the metastasis of the osteosarcoma cell line U2-OS;thus, it could be further ex-plored as a potential target in osteosarcoma therapy.
