| 于云祥,龚泰芳,刘小涛,等.微小 RNA-335-3p靶向小鼠双微体基因调控骨髓瘤细胞的增殖和凋亡[J].安徽医药,2021,25(12):2491-2495. |
| 微小 RNA-335-3p靶向小鼠双微体基因调控骨髓瘤细胞的增殖和凋亡 |
| MiR-335-3p regulates proliferation and apoptosis of myeloma cells by targeting MDM2 gene |
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| DOI：10.3969/j.issn.1009-6469.2021.12.037 |
| 中文关键词: 多发性骨髓瘤 微小 RNA-335-3p 小鼠双微体基因 细胞增殖 细胞凋亡 |
| 英文关键词: Multiple myeloma MicroRNA-335-3p Mouse double minute 2 Cell proliferation Apoptosis |
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| 中文摘要: |
| 目的探讨微小 RNA(miR)-335-3p对骨髓瘤细胞增殖和凋亡的影响及作用机制。方法 2018年 5月至 2019年 1月，培养正常骨髓细胞 MNC和骨髓瘤细胞 KM3和 U266，实时荧光定量逆转录聚合酶链反应( qRT-PCR)检测细胞中 miR-335-3p表达水平。将 KM3细胞分为 miR-NC组(转染模拟对照序列)、 miR-335-3p组［转染 miR-335-3p模拟物( miR-335-3p mimics)］、 pcDNA3.1+miR-335-3p组(共转染空载体和 miR-335-3p mimics)和 pcDNA3.1-MDM2+miR-335-3p组［共转染小鼠双微体基因(MDM2)过表达载体和 miR-335-3p mimics］，MTT法检测细胞增殖，流式细胞仪检测细胞凋亡的影响，蛋白质印迹法( Western blotting)检测细胞周期蛋白 D1(cyclin D1)、细胞周期蛋白依赖性激酶抑制剂 1A(P21)、 B淋巴细胞瘤 -2(Bcl-2)蛋白和 Bcl-2相关 X(Bax)蛋白表达。双荧光素酶报告基因实验验证 KM3细胞中 miR-335-3p与 MDM2调控关系。结果骨髓瘤细胞 KM3和 U266中 miR-335-3p表达水平分别为 0.24±0.01、0.38±0.02，显著低于正常骨髓细胞 MNC中的 miR-335-3p表达水平 0.77±0.03(均 P<0.05)。与 miR-NC组比较， miR-335-3p组 KM3细胞吸光度［48 h:(0.60±0.03)比( 0.99±0.06)； 72 h:(0.82±0.05)比( 1.67±0.07)］、细胞中 cyclin D1和 Bcl-2蛋白表达均降低(均 P<0.05)细胞凋亡率［(24.31±0.99)%比( 8.13±0.83)%］、细胞中 P21和 Bax |
| 英文摘要: |
| Objective To explore the effects of microRNA (miR)-335-3p on proliferation and apoptosis of myeloma cells and its pos. sible mechanism.Methods The study started from May 2018 and ended in January 2019. Normal bone marrow cells MNC and myelo.ma cells KM3 and U266 were cultured, and the levels of miR-335-3p were detected by real-time fluorescent quantitative reverse tran. scription polymerase chain reaction (qRT-PCR). The KM3 cells were assigned into miR-NC group (mimics control), miR-335-3p group (miR-335-3p mimics), pcDNA3.1+miR-335-3p group (miR-335-3p mimics+empty vector) and pcDNA3.1-MDM2+miR-335-3p group [mouse double minute 2 (MDM2) overexpression vector +miR-335-3p mimics]. And then the cell proliferation was detected by MTT,the apoptosis was detected by flow cytometry. The protein expressions of cyclin D1, P21, Bcl-2 and Bax in cells were detected by West. ern blotting. The dual luciferase reporter gene assay verified the relationship between miR-335-3p and MDM2 in KM3 cells.Results The expression levels of miR-335-3p in myeloma cells KM3 and U266 were 0.24±0.01 and 0.38±0.02, respectively, which were signifi.cantly lower than that in normal bone marrow cells 0.77±0.03 (both P<0.05). Compared with the miR-NC group, the absorbance of KM3 cells in the miR-335-3p group [48 h: (0.60±0.03) vs. (0.99±0.06); 72 h: (0.82±0.05) vs. (1.67±0.07)] and the protein expressions of cy. clin D1 and Bcl-2 were decreased (all P<0.05), but the apoptosis rates [(24.31±0.99)% vs. (8.13±0.83)%] and the protein expressions of P21 and Bax were increased (all P<0.05). miR-335-3p negatively regulated the expression of MDM2 expression in KM3 cells. Com. pared with the pcDNA3.1+miR-335-3p group, the absorbance of KM3 cells in the pcDNA3.1-MDM2+miR-335-3p group [48 h: (0.80± 0.05) vs. (0.63±0.04); 72 h: (1.28±0.06) vs. (0.88±0.05)] and the protein expressions of cyclin D1 and Bcl-2 were increased (all P<0.05), but the apoptosis rate [(15.34±0.66)% vs. (23.98±1.41)%] and the protein expressions of P21 and Bax were decreased (all P<0.05).Con. clusion miR-335-3p may inhibit the proliferation of myeloma cells and promote its apoptosis by down-regulating MDM2 expression. |
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