HOXB2反义寡核苷酸对原代脐静脉内皮细胞生物学特性的影响

Effects of HOXB2 antisense oligodeoxynucleotides on the biological properties of primary human umbilical vein endothelial cells

目的 探讨同源盒基因B2 (HOXB2 )反义硫代寡核苷酸对原代脐静脉内皮细胞生物学特性的影响。方法 采用荧光标记观察HOXB2反义寡核苷酸 (Asodn)在内皮细胞中的分布 ;观察不同浓度HOXB2反义寡核苷酸对内皮细胞DNA合成的影响 ;应用流式细胞仪和RT PCR观察其对细胞周期及HOXB2表达的影响。结果 经脂质体介导 ,转染 15min胞核有较弱荧光染色 ,4～ 8h胞核荧光最强 ,16h荧光减弱、消散 ;HOXB2反义寡核苷酸能抑制内皮细胞DNA的合成 ,表现出浓度依赖效应 ,能延缓内皮细胞由G1期进入S期 ;同时HOXB2mRNA表达水平在 2 4～ 4 8h明显下降。结论 HOXB2在内皮细胞增殖中起重要作用 。

Objective To explore the effects of HOXB2 antisense oligodeoxynucleotides (Asodn) on the biological properties of primary human umbilical vein endothelial cells (ECs). Methods Fluorescent labelled Asodn was transfected into the endothelial cells of human unbilical vein mediated liposome and its distribution within endothelia was observed. 3H TdR incorporation test was employed to determine its effects on the DNA synthesis. Flow cytometry was applied to determine the change of the cell cycle. In the same time, RT PCR was adopted to study the influence of Asodn on the expression of target genes. Results Fifteen minutes after the transfection, weak nucleic staining was observed. The fluorescent staining was the strongest 4～8 hours after the transfection and began to weaken in 16 hours. The proportion of cells in G1/0 phase in Asodn group was 53.4±3.1, significantly higher than that in control group (35.8±7.3, P <0.01), and the proportion of cells in S phase in Asodn group was 42.2±3.5, significantly lower than that in control group (60.8±6.2, P <0.01). The expression of HOXB2 mRNA was remarkably decreased during 24 to 48 hours. Conclusion HOXB2 Asodn exerts inhibitory effects on EC proliferation dose dependently, delays the conversion of G1 phase to S Phase, and inhibits the expression of HOXB2 mRNA. HOXB2 gene plays an important role in proliferation of endothelial cells and the mechanism is related to cell cycle.