中华肿瘤杂志
中華腫瘤雜誌
중화종류잡지
CHINESE JOURNAL OF ONCOLOGY
2013年
12期
886-891
,共6页
郁佳佳%申娴娟%王旭东%鞠少卿
鬱佳佳%申嫻娟%王旭東%鞠少卿
욱가가%신한연%왕욱동%국소경
MiR-202%多发性骨髓瘤%B细胞活化因子%细胞增殖%细胞凋亡
MiR-202%多髮性骨髓瘤%B細胞活化因子%細胞增殖%細胞凋亡
MiR-202%다발성골수류%B세포활화인자%세포증식%세포조망
MiR-202%Multiple myeloma%B cell-activating factor%Cell proliferation%Cell apoptosis
目的 研究miR-202对多发性骨髓瘤U266细胞中B细胞活化因子(BAFF)的调节作用及其机制.方法 通过生物信息学软件预测miR-202与BAFF的潜在结合位点,构建荧光素酶报告基因载体.将人类miR-202-3P模拟物(has-miR-202-3 P-mimics)、人类miR-202-3P抑制物(has-miR-202-inhibitor)、siBAFF及各自阴性对照转染U266细胞,采用实时荧光定量PCR和Western blot法验证miR-202对BAFF的调控作用,采用细胞增殖实验和流式细胞仪检测转染后U266细胞的增殖能力和凋亡情况.结果 未转染组、has-miR-202-3 P-mimics转染组、has-miR-202-3 P-inhibitor转染组和siBAFF转染组U266细胞中BAFF mRNA的相对表达量分别为1.040±0.057、0.573±0.073、1.205±0.097和0.368±0.052.与未转染组比较,has-miR-202-3 P-mimics转染组和siBAFF转染组U266细胞中BAFF mRNA的表达明显下调(P＜0.05).Western blot法检测BAFF蛋白的表达结果与mRNA基本一致.未转染组、has-miR-202-3 P-mimics转染组、has-miR-202-3 P-inhibitor转染组和siBAFF转染组U266细胞的A450nm值分别为1.063±0.052、0.714 ±0.045、0.936±0.066和0.764±0.053.与未转染组比较,has-miR-202-3 P-mimics转染组和siBAFF转染组U266细胞的A450nm值明显降低(P＜0.05).未转染组、has-miR-202-3 P-mimics转染组、has-miR-202-3 P-inhibitor转染组和siBAFF转染组U266细胞的凋亡率分别为26.2％、49.6％、21.1％和30.7％.与未转染组比较,has-miR-202-3P-mimics转染组U266细胞的凋亡率明显增加(P＜0.05).has-miR-202DP-mimics转染组U266细胞中p-JNK蛋白的表达水平降低.结论 miR-202通过调节BAFF的表达对U266细胞发挥抑制增殖和诱导凋亡的作用.JNK/SAPK信号通路参与了miR-202对BAFF的调控作用.
目的 研究miR-202對多髮性骨髓瘤U266細胞中B細胞活化因子(BAFF)的調節作用及其機製.方法 通過生物信息學軟件預測miR-202與BAFF的潛在結閤位點,構建熒光素酶報告基因載體.將人類miR-202-3P模擬物(has-miR-202-3 P-mimics)、人類miR-202-3P抑製物(has-miR-202-inhibitor)、siBAFF及各自陰性對照轉染U266細胞,採用實時熒光定量PCR和Western blot法驗證miR-202對BAFF的調控作用,採用細胞增殖實驗和流式細胞儀檢測轉染後U266細胞的增殖能力和凋亡情況.結果 未轉染組、has-miR-202-3 P-mimics轉染組、has-miR-202-3 P-inhibitor轉染組和siBAFF轉染組U266細胞中BAFF mRNA的相對錶達量分彆為1.040±0.057、0.573±0.073、1.205±0.097和0.368±0.052.與未轉染組比較,has-miR-202-3 P-mimics轉染組和siBAFF轉染組U266細胞中BAFF mRNA的錶達明顯下調(P＜0.05).Western blot法檢測BAFF蛋白的錶達結果與mRNA基本一緻.未轉染組、has-miR-202-3 P-mimics轉染組、has-miR-202-3 P-inhibitor轉染組和siBAFF轉染組U266細胞的A450nm值分彆為1.063±0.052、0.714 ±0.045、0.936±0.066和0.764±0.053.與未轉染組比較,has-miR-202-3 P-mimics轉染組和siBAFF轉染組U266細胞的A450nm值明顯降低(P＜0.05).未轉染組、has-miR-202-3 P-mimics轉染組、has-miR-202-3 P-inhibitor轉染組和siBAFF轉染組U266細胞的凋亡率分彆為26.2％、49.6％、21.1％和30.7％.與未轉染組比較,has-miR-202-3P-mimics轉染組U266細胞的凋亡率明顯增加(P＜0.05).has-miR-202DP-mimics轉染組U266細胞中p-JNK蛋白的錶達水平降低.結論 miR-202通過調節BAFF的錶達對U266細胞髮揮抑製增殖和誘導凋亡的作用.JNK/SAPK信號通路參與瞭miR-202對BAFF的調控作用.
목적 연구miR-202대다발성골수류U266세포중B세포활화인자(BAFF)적조절작용급기궤제.방법 통과생물신식학연건예측miR-202여BAFF적잠재결합위점,구건형광소매보고기인재체.장인류miR-202-3P모의물(has-miR-202-3 P-mimics)、인류miR-202-3P억제물(has-miR-202-inhibitor)、siBAFF급각자음성대조전염U266세포,채용실시형광정량PCR화Western blot법험증miR-202대BAFF적조공작용,채용세포증식실험화류식세포의검측전염후U266세포적증식능력화조망정황.결과 미전염조、has-miR-202-3 P-mimics전염조、has-miR-202-3 P-inhibitor전염조화siBAFF전염조U266세포중BAFF mRNA적상대표체량분별위1.040±0.057、0.573±0.073、1.205±0.097화0.368±0.052.여미전염조비교,has-miR-202-3 P-mimics전염조화siBAFF전염조U266세포중BAFF mRNA적표체명현하조(P＜0.05).Western blot법검측BAFF단백적표체결과여mRNA기본일치.미전염조、has-miR-202-3 P-mimics전염조、has-miR-202-3 P-inhibitor전염조화siBAFF전염조U266세포적A450nm치분별위1.063±0.052、0.714 ±0.045、0.936±0.066화0.764±0.053.여미전염조비교,has-miR-202-3 P-mimics전염조화siBAFF전염조U266세포적A450nm치명현강저(P＜0.05).미전염조、has-miR-202-3 P-mimics전염조、has-miR-202-3 P-inhibitor전염조화siBAFF전염조U266세포적조망솔분별위26.2％、49.6％、21.1％화30.7％.여미전염조비교,has-miR-202-3P-mimics전염조U266세포적조망솔명현증가(P＜0.05).has-miR-202DP-mimics전염조U266세포중p-JNK단백적표체수평강저.결론 miR-202통과조절BAFF적표체대U266세포발휘억제증식화유도조망적작용.JNK/SAPK신호통로삼여료miR-202대BAFF적조공작용.
Objective To explore the regulating effect of miR-202 on B cell-activating factor,and check whether the regulation influences the growth of multiple myeloma cells.Methods The potential binding sites of BAFF for miR-202 were predicted using bioinformatics software.Luciferase reporter gene analysis was used to evaluate the regulatory effect of miR-202 on BAFF.Human multiple myeloma U266 cells were transfected with has-miR-202-mimics,has-miR-202-inhibitor,siBAFF and their negative controls,respectively.After above treatments,BAFF mRNA and protein levels were detected by real-time PCR and Western blot analysis,and the proliferation and apoptosis in the multiple myeloma (MM) cells were examined by WST-1 and annexin V-FLUOS assay,respectively.Results The BAFF mRNA expression levels in the untransfected group,has-miR-202-3P-mimics transfected group,has-miR-202-3P-inhibitor transfected group and siBAFF transfected group were 1.040 ± 0.057,0.573 ± 0.073,1.205 ± 0.097 and 0.368 ± 0.052,respectively.BAFF mRNA expressions in U266 cells transfected with has-miR-202-3P-mimics and siBAFF were significantly decreased compared with that in the untransfected group (P ＜ 0.05).The BAFF protein expression level of each group was consistent with the mRNA assay result.The absorbance value in 450 nm of the untransfected group,has-miR-202-3P-mimics transfected group,hasmiR-202-3P-inhibitor transfected group and siBAFF transfected group were 1.063 ±0.052,0.714 ±0.045,0.936 ± 0.066 and 0.764 ± 0.053,respectively.In comparison with the untransfected group,the absorbance value at 450 nm of has-miR-202-3P-mimics and siBAFF transfected groups was significantly reduced (P ＜ 0.05).The cell apoptosis rates of untransfected group,has-miR-202-3P-mimics transfected group,has-miR-202-3P-inhibitor transfected group and siBAFF transfected group were 26.2％,49.6％,21.1％ and 30.7％,respectively.Therefore,the cell apoptosis rate of has-miR-202-3P-mimics transfected group was significantly increased than that of the untransfected group (P ＜ 0.05).p-JNK protein expression level was decreased in the has-miR-202-3P-mimics transfected cells.Conclusions MiR-202 can inhibit the proliferation and induce apoptosis in MM cells via regulating BAFF.JNK/SAPK signaling pathway is involved in the regulation of BAFF by miR-202.
