Negative Regulation of hTERT Expression in Cervical Cancer Hela Cell Line by miR-21
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摘要:
目的 通过观察在miR-21作用下hTERT的表达情况，探讨在宫颈癌HeLa细胞中miR-21对hTERT的调控作用。 方法 利用RT-PCR 法及West Blot 法分别检测HeLa中miR-21 的表达，以及hTERT蛋白的表达情况；通过实验设计，生成miR-21模拟物和 miR-21抑制剂，并分别设立两组的空白对照序列；将空转liposome2000 （lipo2000）设为对照组，利用脂质体 liposome2000包裹宫颈癌细胞株 HeLa并予以转染。在转染后24 h及时检测hTERT及蛋白的表达量。 结果 HeLa细胞株中miR-21及hTERT表达阳性；转染后24 h模拟物组、模拟物对照组、抑制剂组、抑制剂对照组、脂质体组 hTERT mRNA相对表达水平分别为（0.51±0.33）、（0.69±0.19）、（1.33±0.39）、（0.83±0.26）和（0.58±0.16）；蛋白相对表达水平分别为（0.41±0.18）、（0.62±0.18）、（1.45±0.60）、（0.83±0.15）和（0.82±0.30）；模拟物组与其对照组及脂质体组相比较，hTERT mRNA及蛋白表达情况均较低且差异有统计学意义（P < 0.05）；相反，在抑制剂组与其对照组及脂质体组相比较中，hTERT mRNA及蛋白表达情况明显升高，差异有统计学意义（P < 0.05）。 结论 通过检测发现miR-21及hTERT基因在HeLa细胞株中均呈现高表达状态；miR- 21可以调节hTERT mRNA 和蛋白表达水平，从而发挥负向调控hTERT的作用。 Abstract:Objective To observe the effect of miR-21 on hTERT expression in human cervical cancer Hela cell line, explore the regulatory effect of miR-21 on hTERT. Methods RT-PCR was used to detect the expressions of miR-21 and hTERT mRNA in human cervical cancer Hela cell line. West Blot was used to detect the expression of hTERT protein; miR-21 mimics, miR-21 inhibitor, mimics control and inhibitor control were designed and synthesized, then they were transfected into human cervical cancer Hela cell line after parceled by liposome 2000, blank liposome 2000 was designed as control group. The changes of expressions of hTERT mRNA and protein at 24 hours after transfection were detected. Results miR-21 and hTERT were positive in human cervical cancer Hela cell line; the relative expression levels of hTERT mRNA in mimics group, mimics control group, inhibitor group, inhibitor control group and liposome group at 24 hours after transfection were （0.51±0.33）, （0.69±0.19）, （1.33±0.39）, （0.83±0.26） and （0.58±0.16）, respectively; the relative expression levels of hTERT protein were （0.41±0.18）, （0.62±0.18）, （1.45±0.60）, （0.83±0.15） and （0.82±0.30）, respectively. The relative expression levels of hTERT mRNA and protein in mimics group were statistically significantly lower than those in mimics control group and liposome group （P < 0.05）. The relative expression levels of hTERT mRNA and protein in inhibitor group were statistically significantly higher than those in inhibitor control group and liposome group （P < 0.05）. Conclusions miR-21 and hTERT genes are highly expressed in HeLa cell lines. MiR-21 can regulate hTERT mRNA and protein expression levels, thus negatively regulating hTERT. -
Key words:
- miR-21 /
- hTERT /
- Cervical carcinoma /
- Hela /
- Negative regulation
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图 1 miR-21及hTERTmRNA在HeLa细胞株中的表达（×100）
A：模拟物组；B：模拟物对照组；C：抑制剂组；D：抑制剂对照组；E：脂质体组
Figure 1. Expression of miR-21 and hTERTmRNA in HeLa cell line （×100）
图 2 hTERT mRNA 在各组别细胞中的表达情况
Figure 2. Expression of hTERT mRNA in different groups of cells
表 1 hTERT mRNA 在各组别细胞中的表达情况（
$\bar{x}\pm s$ ）Table 1. Expression of hTERT mRNA in different groups of cells （
$\bar{x}\pm s$ ）组别 hTERT mRNA t P 模拟物组 0.51 ± 0.33 4.478 0.011 模拟物对照组 0.69 ± 0.19 抑制剂组 1.33 ± 0.39 5.387 0.005 抑制剂对照组 0.83 ± 0.26 脂质体组 0.58 ± 0.16 注：单因素多组间比较（F = 5.403，P = 0.002）。 
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表 2 hTERT 蛋白在各组别细胞中的表达情况（
$\bar{x}\pm s$ ）（1）Table 2. hTERT protein expression in each group of cells （
$\bar{x}\pm s$ ）（1）组别 实验结果 t P 模拟物组 0.41 ± 0.18 5.072 0.0071 模拟物对照组 0.62 ± 0.18 抑制剂组 1.45 ± 0.60 5.85 0.0043 抑制剂对照组 0.83 ± 0.15 脂质体组 0.82 ± 0.30
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表 2 hTERT 蛋白在各组别细胞中的表达情况（
$\bar{x}\pm s$ ）（2）Table 2. hTERT protein expression in each group of cells （
$\bar{x}\pm s$ ）（2）组别 hTERT蛋白 t P 模拟物组 0.41 ± 0.18 5.072 0.007 模拟物对照组 0.62 ± 0.18 抑制剂组 1.45 ± 0.60 5.85 0.004 抑制剂对照组 0.83 ± 0.15 脂质体组 0.82 ± 0.30
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