摘要
目的探讨hsa—miR-206对乳腺癌增殖、侵袭及迁移的影响。方法用脂质体转染法将hsa-miR-206模拟体（206m组）与阴性对照（NC组）、抑制体（206i组）及阴性对照（INC组）转染入人乳腺癌细胞株MDA—MB-231，通过CCK-8法、Transwell试验检测细胞增殖和侵袭力；用分子生物学方法检测基质金属蛋白酶（MMP）-2、MMP-9、乳腺癌转移抑制基因（BRMS）-1和细胞间隙蛋白43（Cx43）的表达；检测临床样本内miR-206及Cx43表达；双荧光素酶报告基因验证miR-206和Cx43的结合。结果（1）206m组细胞活性（0．74±0．16）较对照组（1．12±0．23）低（t=-3．066，P=0．037），2061组细胞活性（1．43±0．26）比INC组（0．98±0．14）高（t=3．635，P=0．022）。（2）Transwell实验证明较对照组，206m组细胞迁移和侵袭明显降低（迁移：0．56±0．01与0．63±0．01，t=-23．000，P=0．002；侵袭：0．79±0．01与0．99±0．01，t=-21．200，P=0．002），而206i组细胞迁移和侵袭明显增加（迁移：0．97±0．11与0．61±0．09，t=32．787，P=0．001；侵袭：1．10±0．01与0．93±0．05，t=5．167，P=0．035）。（3）分子生物学检测发现206m组MMP-2、MMP-9和Cx43的表达降低，BRMS-1表达增高，而206i组MMP-2、MMP-9和Cx43的表达增高，BRMS—1表达降低。（4）临床样本检测发现miR-206在淋巴结阴性组中表达高于淋巴结阳性组（Z=-2．098，P=0．036），而Cx43的表达则相反。（5）双荧光素酶报告基因验证miR-206和Cx43问存在特异性结合位点（1．004-0．04与0．74±0．04，t=26．911，P=0．001）。结论Hsa—miR-206对乳腺癌增殖、侵袭及迁移能力存在负调控。这一调控能力通过Cx43实现。
Objective To explore the effects of hsa-miR-206 on the proliferation, migration and invasion of breast cancer. Methods The hsa-miR-206 mimics, inhibitors and their paired negative controls were transfected into human breast cancer cell line MDA-MB-231 by liposome. The proliferation of cell was evaluated by CCK-8 and the migration and invasion was detected by Transwell. Matrix metalloproteinase 2 （ MMP-2）, matrix metalloproteinase 9 （ MMP-9）, breast cancer metastasis-suppressor 1 （ BRMS-1 ） and connexin 43 （Cx43） were detected by both quantitative polymerase chain reaction （qPCR） and Western blot. The expression of miR-206 was detected by qPCR. Dual luciferase assay was detected to confirm the specific binding sites of miR-206 and Cx43. Results （ 1 ） The proliferation activity of 206m-group cell （0. 74 ±0. 16） was significantly lower than that of control group （ 1.12 ±0. 23） （t = -3. 066, P =0. 037） while that of 206i-group cell （ 1.43 ±0. 26 ） was higher than that of control group （ 0. 98 ±0. 14 ） （ t = 3. 635, P = 0. 022）. （2） Transwell tests showed the migration and invasion of 206m-group cell decreased significantly （ migration ：0. 56 ±0. 01 vs 0. 63 ±0. 01, t = - 23.00, P = 0. 002 ; invasion ： 0. 79 ±0. 01 vs 0. 99±0. 01, t = -21. 200, P =0. 002）, but that of 206i-group cell increased significantly （migration： 0. 97 ±0. 11 vs 0. 61 ±0. 09,t =32. 787,P =0. 001 ; invasion： 1.10 ±0. 01 vs 0. 93 ±0. 05, t =5. 167, P = 0. 035）. （3） The expressions of MMP-2, MMP-9 and Cx43 decreased and the expression of BRMS-1 increased in 206m-group cell and vice versa in 206i group. （4） The expression of miR-206 in lymph node- negative group of clinical breast cancer sample was higher than that of lymph node-positive one. And there was statistical difference （ Z = - 2. 098, P = 0. 003 ）. And the expression of Cx43 was opposite. （ 5 ） Dual luciferase reporter assay confirmed the specific binding sites of hsa-miR-206 and Cx43. Conclusio
出处
《中华医学杂志》
CAS
CSCD
北大核心
2013年第36期2890-2894,共5页
National Medical Journal of China
基金
上海市卫生局青年项目（20114y155）
上海市申康前沿课题（SHDC12010116）
上海市卫生局项目（20124164）
上海市科委项目（12411%4700）
