中华神经医学杂志
中華神經醫學雜誌
중화신경의학잡지
CHINESE JOURNAL OF NEUROMEDICINE
2009年
1期
5-9
,共5页
神经胶质瘤%Polo样激酶1%RNA干扰%小干扰RNA%端粒酶
神經膠質瘤%Polo樣激酶1%RNA榦擾%小榦擾RNA%耑粒酶
신경효질류%Polo양격매1%RNA간우%소간우RNA%단립매
Glioma%Polo-like kinase-1%RNA interference%Small interfering RNA%Telomerase
目的 探讨Polo样激酶1(PLK1)基因对胶质瘤细胞增殖的影响和可能机制. 方法 根据PLK1基因特点,设计并用化学方法合成了5个小干扰核糖核酸分子(siRNA)(P1、P2、P3、P4和P5).以这5个siRNA转染人胶质瘤TJ905细胞后.分别采用荧光实时定量RT-PCR和Western blot检测PLK1 mRNA和蛋白表达水平.分别采用MTT法和Western blot方法检测癌细胞增殖和增殖细胞核抗原(PCNA)蛋白水平,用TRA-.ELISA方法检测胶质瘤细胞端粒酶活性. 结果 所设计的5个siRNA均能明显抑制胶质瘤TJ905细胞PLK1 mRNA水平,以P4效果最好.以P4转染处理胶质瘤细胞后与脂质体对照组比较,PLK1基因mRNA水平和蛋白水平明显下调,差异有统计学意义(P均=0.000).MTT结果显示.与脂质体对照组比较P4 siRNA转染组癌细胞生长明显受到抑制,且呈浓度依赖性(r=0.868,P=0.000).Western blot结果显示,与脂质体对照组比较P4 siRNA转染组PCNA蛋白水平明显下降,差异有统计学意义(F=181.36,P=0.000).TRAP-ELISA结果显示,与脂质体对照组比较P4 siRNA转染组胶质瘤细胞端粒酶活性明显受到抑制,且呈浓度和时间依赖性(P=0.000). 结论 PLK1基因对胶质瘤细胞增殖具有重要的调控作用;以PLK1 siRNA转染处理胶质瘤细胞,可明显抑制胶质瘤细胞的恶性增殖,其机制可能与抑制端粒酶活性有关.
目的 探討Polo樣激酶1(PLK1)基因對膠質瘤細胞增殖的影響和可能機製. 方法 根據PLK1基因特點,設計併用化學方法閤成瞭5箇小榦擾覈糖覈痠分子(siRNA)(P1、P2、P3、P4和P5).以這5箇siRNA轉染人膠質瘤TJ905細胞後.分彆採用熒光實時定量RT-PCR和Western blot檢測PLK1 mRNA和蛋白錶達水平.分彆採用MTT法和Western blot方法檢測癌細胞增殖和增殖細胞覈抗原(PCNA)蛋白水平,用TRA-.ELISA方法檢測膠質瘤細胞耑粒酶活性. 結果 所設計的5箇siRNA均能明顯抑製膠質瘤TJ905細胞PLK1 mRNA水平,以P4效果最好.以P4轉染處理膠質瘤細胞後與脂質體對照組比較,PLK1基因mRNA水平和蛋白水平明顯下調,差異有統計學意義(P均=0.000).MTT結果顯示.與脂質體對照組比較P4 siRNA轉染組癌細胞生長明顯受到抑製,且呈濃度依賴性(r=0.868,P=0.000).Western blot結果顯示,與脂質體對照組比較P4 siRNA轉染組PCNA蛋白水平明顯下降,差異有統計學意義(F=181.36,P=0.000).TRAP-ELISA結果顯示,與脂質體對照組比較P4 siRNA轉染組膠質瘤細胞耑粒酶活性明顯受到抑製,且呈濃度和時間依賴性(P=0.000). 結論 PLK1基因對膠質瘤細胞增殖具有重要的調控作用;以PLK1 siRNA轉染處理膠質瘤細胞,可明顯抑製膠質瘤細胞的噁性增殖,其機製可能與抑製耑粒酶活性有關.
목적 탐토Polo양격매1(PLK1)기인대효질류세포증식적영향화가능궤제. 방법 근거PLK1기인특점,설계병용화학방법합성료5개소간우핵당핵산분자(siRNA)(P1、P2、P3、P4화P5).이저5개siRNA전염인효질류TJ905세포후.분별채용형광실시정량RT-PCR화Western blot검측PLK1 mRNA화단백표체수평.분별채용MTT법화Western blot방법검측암세포증식화증식세포핵항원(PCNA)단백수평,용TRA-.ELISA방법검측효질류세포단립매활성. 결과 소설계적5개siRNA균능명현억제효질류TJ905세포PLK1 mRNA수평,이P4효과최호.이P4전염처리효질류세포후여지질체대조조비교,PLK1기인mRNA수평화단백수평명현하조,차이유통계학의의(P균=0.000).MTT결과현시.여지질체대조조비교P4 siRNA전염조암세포생장명현수도억제,차정농도의뢰성(r=0.868,P=0.000).Western blot결과현시,여지질체대조조비교P4 siRNA전염조PCNA단백수평명현하강,차이유통계학의의(F=181.36,P=0.000).TRAP-ELISA결과현시,여지질체대조조비교P4 siRNA전염조효질류세포단립매활성명현수도억제,차정농도화시간의뢰성(P=0.000). 결론 PLK1기인대효질류세포증식구유중요적조공작용;이PLK1 siRNA전염처리효질류세포,가명현억제효질류세포적악성증식,기궤제가능여억제단립매활성유관.
Objective To investigate the regulatory role of polo-like kinase-1 (PLK1) gene in the proliferation of human glioma cells. Methods Five small interfering RNAs (siRNAs) targeting PLK1 gene were designed and synthesized according to PLK1 mRNA sequence. After transfection of human glioma TJ905 cells with the siRNAs, real-time RT-PCR and Western blotting were performed to examine the changes in PLK1 gene expression in the cells. The growth of the transfected cells was evaluated by MTT assay and proliferating cell nuclear antigen (PCNA) protein expression determined using Western blotting. Telomeric repeat amplification protocol-enzyme-linked immunosorbent assay (TRAP-ELISA) was used to detect the changes in telomerase activity of the transfected cells. Results All the five siRNAs were capable of suppressing PLK1 mRNA expression in TJ905 cells, among which the P4 siRNA showed the strongest effect by reducing the PLK1 mRNA level by 93% 48 h after transfection at the concentration of 100 nmol/L. Compared with the oligofecamine control group cells the protein expression of PLK1 in TJ905 cancer cells transfected with P4 siRNA was also siguifieantly down-regulated. Transfection with P4 siRNA resulted in significant dose-dependent inhibitory effects on the proliferation and PCNA protein expression of TJ905 cells as compared to oligofecamine control group. The results of TRAP-ELISA showed obvious time- and dose-dependent inhibition of telomerase activity in the transfected cells as compared to oligofecamine control group. Conclusion PKL1 gene plays an important regulatory role in the proliferation of human glioma cells, and RNA interference of PLK1 gene can inhibit the cell proliferation possibly by suppressing the telomerase activity.
