功能化氧化石墨烯的制备及其介导HIF-1α基因RNA干扰的实验研究

Construction and effects of functionalized graphene oxide nonparticles-mediated RNA interference of hypoxia-inducible factor-1α gene

目的探讨功能化氧化石墨烯(GO)-聚乙二醇(PEG)-叶酸(FA)-1-芘甲胺盐酸盐(PyNH2)介导的乏氧诱导因子-1α(HIF-1α)基因的RNA干扰(RNAi)对胰腺癌Patu8988细胞生物学行为的影响。方法制备GO-PEG-FA-PyNH2载体以及介导HIF-1α基因RNAi的转染微粒(GO-PEG-FA-PyNH2-HIF-1α-RNAi),检测其转染Patu8988细胞后HIF-1α和葡萄糖转运蛋白-1(Glut-1)的表达情况,Patu8988细胞增殖、侵袭能力和细胞周期变化,以及对Patu8988细胞摄取18F-脱氧葡萄糖(FDG)的影响。多组间比较采用单因素方差分析,两两比较行最小显著差异t检验。结果成功制得GO-PEG-FA-PyNH2载体,未发现细胞毒性。乏氧或常氧状态下,GO-PEG-FA-PyNH2-HIF-1α-RNAi转染(实验组)的细胞的HIF-1α mRNA和蛋白表达以及Glut-1 mRNA表达均低于GO-PEG-FA-PyNH2(阴性组)和去血清培养基(Opti-MEM,空白组)转染的细胞(F=30.25~32.58,t=3.66~5.81,均P<0.05);实验组迁移的细胞数远少于阴性组和空白组(F=38.63和41.35,t=20.51~35.25,均P<0.01);实验组细胞增殖低于阴性组和空白组(F=35.19和38.11,t=15.11~22.19,均P<0.05);实验组G0/G1细胞比例高于阴性组和空白组(F=36.40和34.83,t=11.55~34.56;均P<0.05);实验组对18F-FDG摄取少于阴性组和空白组(F=29.85和31.69,t=3.35~4.35,均P<0.05)。结论GO-PEG-FA-PyNH2靶向介导的HIF-1α RNAi可抑制胰腺癌细胞HIF-1α的表达,导致相关生物学变化。

Objective To explore the effects of graphene oxide (GO)-polyethylene glycol (PEG)-folic acid (FA)-pyrenemethylamine hydrochloride (PyNH2)-mediated RNA interference (RNAi) of hypoxia-inducible factor-1α(HIF-1α) on the biological behaviors of human pancreatic cancer Patu8988 cells. Methods GO-PEG-FA-PyNH2 and RNAi targeting HIF-1α gene (GO-PEG-FA-PyNH2-HIF-1α-RNAi) was constructed. The expressions of HIF-1α and glucose transporter 1 (Glut-1) in Patu8988 cells were determined after knockdown of HIF-1α by RNAi. The invasive ability, the proliferation and the cell cycle of Patu8988 cells were investigated. The effect of HIF-1α knockdown on the uptake of 18F-fluorodeoxyglucose (FDG) in Patu8988 cells was also detected. Comparison of data was conducted by one-way analysis of variance and least significant difference t test. Results The GO-PEG-FA-PyNH2 was successfully constructed, and no cytotoxicity was found. Under the hypoxia or normoxia state, the mRNA and protein levels of HIF-1α and mRNA level of Glut-1 in cells transfected with GO-PEG-FA-PyNH2-HIF-1α-RNAi (study group) were lower than those in cells transfected with GO-PEG-FA-PyNH2(negative group) and cells transfected with Opti-minimal essential medium (Opti-MEM, control group;F=30.25-32.58, t=3.66-5.81, all P<0.05);the numbers of migrated cells in the study group were much lower than those in the negative group and the control group (F=38.63 and 41.35, t=20.51-35.25, all P<0.01);the cell proliferation in the study group was significantly lower than that in the negative group and the control group (F=35.19 and 38.11, t=15.11-22.19, all P<0.05). The proportions of G0/G1 cells in the study group were higher than those in the negative group and the control group (F=34.60 and 34.83, t=11.55-34.56, all P<0.05);the 18F-FDG uptake in the study group was lower than that in the negative group and control group (F=29.85 and 31.69, t=3.35-4.35, all P<0.05). Conclusion GO-PEG-FA-PyNH2-mediated HIF-1α RNAi inhibits the expression of HIF-1α in pancreatic cancer cells, leading to changes in related biological behaviors.