下调泛素样含PHD和环指域1基因的表达对喉癌Hep-2细胞增殖侵袭能力的影响

Effect of down-regulation of UHRF1 on proliferation and invasive capacity in human laryngeal carcinoma Hep-2 cells

目的探讨下调泛素样含PHD和环指域1(ubiquitin-like protein containing PHD and RING finger domains 1,UHRF1)基因在喉癌Hep-2细胞中的表达后对其增殖,凋亡,侵袭迁移能力的影响。方法实验分为空白组,阴性对照组和干扰组。利用UHRF1-si RNA干扰UHRF1在喉癌Hep-2细胞中的表达,采用q RT-PCR、Western blot检测UHRF1的表达变化。采用MTT法、流式细胞术、Transwell实验检测下调UHRF1表达后Hep-2细胞的增殖、凋亡、侵袭、迁移能力的变化。Western blot检测下调UHRF1表达后Hep-2细胞内Bax、Bcl-2、MMP-2、MMP-9蛋白表达的变化情况。结果与空白组和阴性对照组相比干扰组下调UHRF1的表达后可明显抑制喉癌Hep-2细胞的增殖能力(P<0.001)。空白组,阴性对照组,干扰组凋亡率分别为(5.47±2.77)%,(3.95±0.66)%,(18.95±1.10)%,干扰组凋亡率明显提高(P<0.001)。空白组,阴性对照,干扰组每视野侵袭细胞数分别为(213.7±13.1),(221.3±10.3),(97.7±7.5),干扰组侵袭细胞数明显降低(P<0.001)。空白组,阴性对照,干扰组每视野迁移细胞数分别为(230.7±5.5),(222.7±11.2),(111.7±7.6),干扰组迁移细胞数明显降低(P<0.001)。干扰组细胞Bax蛋白表达上调(P<0.001),Bcl-2蛋白表达下调(P<0.001),MMP-2蛋白、MMP-9蛋白表达下调(P<0.001)。结论在喉癌Hep-2细胞中下调UHRF1的表达后可明显抑制细胞的增殖,促进细胞凋亡,抑制细胞的侵袭迁移能力。

Objective To determine the effect of down-regulation of ubiquitin-like protein containing PHD and ring finger domains 1 （UHRF1） on the proliferation, apoptosis, invasion and migration in human laryngeal carcinoma Hep-2 celt line. Methods The Hep-2 cells were randomly divided into 3 groups： blank control group, negative control group and tranfected group. UHRFI-siRNA was used to down-regulate the expression of UHRF1 in the Hep-2 cells, and quantitative real-time PCR （qRT-PCR） and Western blotting were used to detect the mRNA and protein expression of UHRF1 after down-regulation. The proliferative, invasive and migratory capabilities of Hep-2 cells transfeeted with UHRFI-siRNA were detected by MT]＂ assay, flow cytometry and Transwell chamber assay. Western blotting was used to measure the protein levels of Bel-2, Bax, MMP-2 and MMP-9. Results Compared with the blank control group and negative controlgroup, down-regulation of UHRF1 significantly inhibited the proliferative capabilities of Hep-2 cells （P 〈 O. 001 ）. The apoptotic rate was （5.47± 2.77）%, （3.95 ±0. 66）% and （18.95 ± 1.10）%, respectively in the blank control group, negative control group and transfected group, and the rate was the highest in the transfected group. The number of invasive cells was （213.7 ± 13.1 ）, （221.3 ± 10.3） and （97.7 ±7.5） respectively, and that of migratory cells was （230.7 ±5.5）, （222.7 ± 11.2） and （ 111.7 ± 7.6） respectively in the above 3 groups. The numbers of invasive and migratory cells were both obviously lower in the transfeeted group （P 〈0. 001 ）. Western blotting results showed that Bax was up-regulated （P 〈0O. 001 ） , and Bcl-2, MMP-2 and MMP-9 were down-regulated （P 〈 0. 001 ）. Conclusion Down-regulation of UHRF1 inhibits the proliferation, induces the apoptosis, and reduces the invasive and migratory capability in Hep-2 cells.