中华医学杂志
中華醫學雜誌
중화의학잡지
National Medical Journal of China
2013年
36期
2890-2894
,共5页
傅韵%蒋蓓琦%武羿%李正东%庄志刚
傅韻%蔣蓓琦%武羿%李正東%莊誌剛
부운%장배기%무예%리정동%장지강
乳腺肿瘤%hsa-miR-206%迁移%侵袭%细胞间隙蛋白43
乳腺腫瘤%hsa-miR-206%遷移%侵襲%細胞間隙蛋白43
유선종류%hsa-miR-206%천이%침습%세포간극단백43
Breast neoplasms%hsa-miR-206%Migration%Invasion%Connexin43
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),206i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,=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间存在特异性结合位点(1.00±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.
目的 探討hsa-miR-206對乳腺癌增殖、侵襲及遷移的影響.方法 用脂質體轉染法將hsa-miR-206模擬體(206m組)與陰性對照(NC組)、抑製體(206i組)及陰性對照(INC組)轉染入人乳腺癌細胞株MDA-MB-231,通過CCK-8法、Transwell試驗檢測細胞增殖和侵襲力；用分子生物學方法檢測基質金屬蛋白酶(MMP)-2、MMP-9、乳腺癌轉移抑製基因(BRMS)-1和細胞間隙蛋白43(Cx43)的錶達；檢測臨床樣本內miR-206及Cx43錶達；雙熒光素酶報告基因驗證miR-206和Cx43的結閤.結果 (1) 206m組細胞活性(0.74 ±0.16)較對照組(1.12±0.23)低(t=-3.066,P=0.037),206i組細胞活性(1.43±0.26)比INC組(0.98 ±0.14)高(t=3.635,P=0.022).(2)Transwell實驗證明較對照組,206m組細胞遷移和侵襲明顯降低(遷移:0.56±0.01與0.63±0.01,t=-23.000,P=0.002；侵襲:0.79±0.01與0.99±0.01,t=-21.200,P=0.002),而206i組細胞遷移和侵襲明顯增加(遷移:0.97±0.11與0.61 ±0.09,t=32.787,P=0.001;侵襲:1.10±0.01與0.93 ±0.05,=5.167,P=0.035).(3)分子生物學檢測髮現206m組MMP-2、MMP-9和Cx43的錶達降低,BRMS-1錶達增高,而206i組MMP-2、MMP-9和Cx43的錶達增高,BRMS-1錶達降低.(4)臨床樣本檢測髮現miR-206在淋巴結陰性組中錶達高于淋巴結暘性組(Z=-2.098,P=0.036),而Cx43的錶達則相反.(5)雙熒光素酶報告基因驗證miR-206和Cx43間存在特異性結閤位點(1.00±0.04與0.74±0.04,t =26.911,P=0.001).結論 Hsa-miR-206對乳腺癌增殖、侵襲及遷移能力存在負調控.這一調控能力通過Cx43實現.
목적 탐토hsa-miR-206대유선암증식、침습급천이적영향.방법 용지질체전염법장hsa-miR-206모의체(206m조)여음성대조(NC조)、억제체(206i조)급음성대조(INC조)전염입인유선암세포주MDA-MB-231,통과CCK-8법、Transwell시험검측세포증식화침습력；용분자생물학방법검측기질금속단백매(MMP)-2、MMP-9、유선암전이억제기인(BRMS)-1화세포간극단백43(Cx43)적표체；검측림상양본내miR-206급Cx43표체；쌍형광소매보고기인험증miR-206화Cx43적결합.결과 (1) 206m조세포활성(0.74 ±0.16)교대조조(1.12±0.23)저(t=-3.066,P=0.037),206i조세포활성(1.43±0.26)비INC조(0.98 ±0.14)고(t=3.635,P=0.022).(2)Transwell실험증명교대조조,206m조세포천이화침습명현강저(천이:0.56±0.01여0.63±0.01,t=-23.000,P=0.002；침습:0.79±0.01여0.99±0.01,t=-21.200,P=0.002),이206i조세포천이화침습명현증가(천이:0.97±0.11여0.61 ±0.09,t=32.787,P=0.001;침습:1.10±0.01여0.93 ±0.05,=5.167,P=0.035).(3)분자생물학검측발현206m조MMP-2、MMP-9화Cx43적표체강저,BRMS-1표체증고,이206i조MMP-2、MMP-9화Cx43적표체증고,BRMS-1표체강저.(4)림상양본검측발현miR-206재림파결음성조중표체고우림파결양성조(Z=-2.098,P=0.036),이Cx43적표체칙상반.(5)쌍형광소매보고기인험증miR-206화Cx43간존재특이성결합위점(1.00±0.04여0.74±0.04,t =26.911,P=0.001).결론 Hsa-miR-206대유선암증식、침습급천이능력존재부조공.저일조공능력통과Cx43실현.
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 vs0.61 ±0.09,t=32.787,P=0.001; invasion:l.10 ± 0.01 vs0.93±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 nodenegative 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.Conclusion HsamiR-206 has negative controls of proliferation,migration and invasion of breast cancer cell by targeting Cx43.
