人arresten基因转染对自体移植静脉内膜增生的影响
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郑启昌

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The effect of human arresten gene transfection on intimal hyperplasia of autogenous vein graft in rats
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    摘要:

    目的:探讨体内转染arresten基因对自体移植静脉内膜增生的影响。
    方法:建立大鼠自体静脉移植模型。血管吻合术前,用脂质体介导重组质粒pSecTag2-AT(Ⅰ组),空载体pSecTag2转染(Ⅱ组)移植血管,等体积脂质体溶液处理移植血管(空白对照组,Ⅲ组)。各组动物均于4周后切取移植血管,RT-PCR检测移植血管中arresten mRNA的表达;常规HE,Verhoeff弹力纤维染色;计算机图象分析检测移植静脉血管内膜及中膜面积、厚度;免疫组化检测移植血管内膜α-SMA及PCNA的表达;Western blot检测TGF-β1蛋白的表达。
    结果:Ⅰ组转染的移植静脉中有目的基因mRNA的表达, 而Ⅱ、Ⅲ组无Ⅰ组内膜、中膜面积小均于Ⅱ组和Ⅲ组,差异有显著性(P<0.05)。而内膜面积/中膜面积3组间无统计学差异(P>0.05);Ⅰ组内膜厚度小于Ⅱ组和Ⅲ组,组间比较有统计学差异(P<0.01);α-SMA染色表明增生内膜中的细胞是血管平滑肌细胞;PCNA阳性细胞数及表达指数Ⅰ组均低于Ⅱ组和Ⅲ组(P<0.05);Ⅰ组TGF-β1蛋白的表达明显低于Ⅱ组和Ⅲ组。
    结论:移植血管转染arresten基因,可有效抑制自体移植静脉内膜的增生,在防治血管移植术后再狭窄方面显示出良好的临床应用前景。

    Abstract:

    Abstract:Objective:To investigate the effects of human arresten gene transfection on the intimal hyperplasia of venous autografts in vivo.
    Methods :Autogenous vein graft model was established in male Sprague-Dawley rats. The transplanted veins were immersed in the liposome mediated recombinant eukaryotic expression plasmid pSecTag2-AT solution(group Ⅰ) or the liposome mediated control plasmid pSecTag2 solution(group Ⅱ) for 30minutes just before vascular anastomosis. Normal control group(group Ⅲ) was only treated with isovolume of liposome. Vein graft samples were harvested at 4 weeks after operation. RT-PCR was used to detect the expression of arresten mRNA in blood vessel; the surface area and thickness of the intima and media were measured by computerized planimetry under a light microscope to compare the degree of neointimal hyperplasia by the calculated ratio between intima (I) and media (M) after staining with hematoxylin-eosin and Verhoeff(elastic fibers). Immunohistochemical labeling and morphologic analysis of vein graft sections were used to identify PCNA positive cells and α-SMA positive cells; Western blot was used to detect the protein of TGF-β1.
    Results:The genome of arresten-transferred tissue contained a 449bp specific fragment of arresten gene was expressed in group Ⅰ, but was not expressed in group Ⅱ or group Ⅲ; the surface area of the intima and media of group Ⅰ was less than that of group Ⅱ and group Ⅲ, and the difference were statistically significant (P<0.05), while I/M had no statistical difference (P>0.05). A less intimal thickness of groupⅠ was seen compared with group Ⅱ and group Ⅲ (P<0.01). α-SMA staining suggested that VSMC was present in the hyperplastic intimal. The number of PCNA-positive-stained cells and expression index of group Ⅰ was lower as compared with that of group Ⅱ and group Ⅲ (P<0.05). Protein level of TGF-β1 of group Ⅰ decreased obviously compared with group Ⅱ and group Ⅲ.
    Conclusions:Transfection of human arresten gene can effectively inhibit the intimal hyperplasia of venous autograft. Arresten has a potential perspective for clinic application in prevention and treatment of restenosis after vascular transplantation.

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尚丹, 宋自芳, 夏印, 李毅清, 胡青钢, 郑启昌.人arresten基因转染对自体移植静脉内膜增生的影响[J].中国普通外科杂志,2008,17(2):13-158.
DOI:10.7659/j. issn.1005-6947.2008.02.013

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  • 在线发布日期: 2008-02-25