Transfered Human Thrombomodulin Gene into Artery Wall of Rabbit by High-Pressure Injection in Vivo_CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY

Authors：

LI Jingdong ¹ , DAI Yi ¹ , CHEN Kai ¹ , ZHOU Lin ²

1.Department of Hepatobiliary Surgery, The Affiliated Hospital of North Sichuan Medical College, Nanchong 637000, Sichuan Province, China;;
2.Center for Clinical Molecular Medicine of Children’s Hospital of Chongqing Medical University, Chongqing 400014, China;

Corresponding author：

Keywords：

Human thrombomodulin; Gene transfection; Artery; Rabbit

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Objective To explore the feasibility of high-pressure injection to transfer human thrombomodulin (hTM) gene into arterial wall of rabbits.
Methods Eighty-four healthy New Zealand rabbits were randomly divided into three groups: pcDNA3.1/hTM plasmid group (n=28), pcDNA3.1(+)/neo plasmid group (n=28) and untransfected group (n=28). After gene transfection, the model of arterial injury-blocking was established. Then, the expressions of hTM mRNA and protein in arterial wall were examined by RT-PCR and immunohistochemistry at 3 d, 7 d, 14 d and 28 d after operation.
Results Seventeen rabbits died accidentally from the day of operation to 3 d after operation. The expressions of hTM mRNA of different time points in pcDNA3.1/hTM plasmid group were significantly higher than that in pcDNA3.1(+)/neo plasmid group and untransfected group (P lt;0.01). For the expressions of hTM mRNA at different time points in pcDNA3.1(+)/neo plasmid group and untransfected group, the difference of inter-group and intra-group was not significant (P gt;0.05). hTM protein was expressed in every group and mainly localized in the inner lining of arterial wall. The expressions of hTM protein at different time points in pcDNA3.1/hTM plasmid group were significantly higher than that in pcDNA3.1(+)/neo plasmid group and untransfected group (P lt;0.05). The expression of hTM protein at different time points in pcDNA3.1(+)/neo plasmid group and untransfected group kept relative constancy, the difference of inter-group and intra-group was also not significant (P gt;0.05). Conclusion High-pressure injection is feasible to transfer pcDNA3.1/hTM plasmid into arterial wall of live animals.

Citation： LI Jingdong ,DAI Yi,CHEN Kai,ZHOU Lin. Transfered Human Thrombomodulin Gene into Artery Wall of Rabbit by High-Pressure Injection in Vivo. CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY, 2009, 16(8): 628-632. doi: Copy

1.	戴毅，陈开，邹琳，等. 人血栓调节蛋白基因编码片段的克隆及在真核细胞中的表达 [Ｊ].中国普外基础与临床杂志， 2005; 12（6）: 577-580.
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6.	Forte A, Finicelli M, Mattia M, et al. Mesenchymal stem cells effectively reduce surgically induced stenosis in rat carotids [Ｊ]. J Cell Physiol, 2008; 217(3): 789-799.
7.	陈华，景在平，包俊敏，等. RNA干扰对兔血管平滑肌细胞bcl-2基因表达的影响 [Ｊ]. 中国普外基础与临床杂志， 2007; 14（1）: 56-58.
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10.	Brunetti-Pierri N, Stapleton GE, Law M, et al. Efficient, long-term hepatic gene transfer using clinically relevant HDAd doses by balloon occlusion catheter delivery in nonhuman primates [Ｊ]. Mol Ther, 2009; 17(2): 327-333.
11.	Morishita R. Recent progress in gene therapy for cardiovascular disease [Ｊ]. Circ J, 2002; 66(12): 1077-1086.
12.	Li W, Tanaka K, Morioka K, et al. Long-term effect of gene therapy for chronic ischemic myocardium using platelet-derived endothelial cell growth factor in dogs [Ｊ]. J Gene Med, 2008; 10(4): 412-420.
13.	罗涛，张强，张建，等. 转染单纯疱疹病毒胸苷激酶基因抑制移植静脉内膜增生 [Ｊ]. 中国普外基础与临床杂志， 2008; 15（10）: 743-747.
14.	Wu S, Wang X, Guo L, et al. Adenovirus mediated endothelial nitric oxide synthase gene transfer prevents restenosis of vein grafts [Ｊ]. ASAIO J, 2004; 50(3): 272-277.
15.	梁宏立, 朱洪生, 黄忠耀，等. 兔自体移植静脉基因转移模型的建立 [Ｊ]. 上海实验动物科学， 2000; 20（1）: 3-6.
16.	曲乐丰, 景在平, 曹贵松，等. 手术缝线携载反义基因预防血管吻合口内膜增生的实验研究 [Ｊ]. 中华普通外科杂志， 2002; 17（1）: 36-37.
17.	Wen DZ, Dittman WA, Ye RD, et al. Human thrombomodulin complete cDNA sequence and chromosome localization of the gene [Ｊ]. Biochemistry, 1987; 26(14): 4350-4357.
18.	Hirsch T, Spielmann M, Velander P, et al. Insulin-like gr-owth factor-1 gene therapy and cell transplantation in diabetic wounds [Ｊ]. J Gene Med, 2008; 10(11): 1247-1252.
19.	Kishida A, Nakashima M, Sakamoto N, et al. Study on complex formation between recombinant human thrombomodulin fragment and thrombin using surface plasmon resonance [Ｊ]. Am J Hematol, 2000; 63(3): 136-140.
20.	Yeh HY, Lin JC. Bioactivity and platelet adhesion study of a human thrombomodulin-immobilized nitinol surface [Ｊ]. J Biomater Sci Polym Ed, 2009; 20(5-6): 807-819.

1. 戴毅，陈开，邹琳，等. 人血栓调节蛋白基因编码片段的克隆及在真核细胞中的表达 [Ｊ].中国普外基础与临床杂志， 2005; 12（6）: 577-580.
2. 戴毅，陈开，乔正荣，等. 凝血酶调节蛋白基因对脐静脉内皮细胞抗凝功能的调控 [Ｊ]. 中华血液学杂志， 2006; 27（3）:162-165.
3. Dulak J, Schwarzacher SP, Zwick RH, et al. Effects of local gene transfer of VEGF on neointima formation after balloon injury in hypercholesterolemic rabbits [Ｊ]. Vasc Med, 2005; 10(4): 285-291.
4. J.萨姆布鲁克， D.W.拉塞尔主编. 黄培堂主译. 分子克隆实验指南 [M]. 上册. 第3版. 北京: 科学出版社， 2002: 128-129.
5. Liu Q, Chen ZQ, Bobustuc GC, et al. Local gene transduction of cyclooxygenase-1 increases blood flow in injured atherosclerotic rabbit arteries [Ｊ]. Circulation, 2005; 111(14): 1833-1840.
6. Forte A, Finicelli M, Mattia M, et al. Mesenchymal stem cells effectively reduce surgically induced stenosis in rat carotids [Ｊ]. J Cell Physiol, 2008; 217(3): 789-799.
7. 陈华，景在平，包俊敏，等. RNA干扰对兔血管平滑肌细胞bcl-2基因表达的影响 [Ｊ]. 中国普外基础与临床杂志， 2007; 14（1）: 56-58.
8. Nicklin SA, White SJ, Nicol CG, et al. In vitro and in vivo characterisation of endothelial cell selective adenoviral vectors [Ｊ]. J Gene Med, 2004; 6(3): 300-308.
9. Kallenbach K, Salcher R, Heim A, et al. Inhibition of sm-ooth muscle cell migration and neointima formation in vein grafts by overexpression of matrix metalloproteinase-3 [Ｊ]. J Vasc Surg, 2009; 49(3): 750-758.
10. Brunetti-Pierri N, Stapleton GE, Law M, et al. Efficient, long-term hepatic gene transfer using clinically relevant HDAd doses by balloon occlusion catheter delivery in nonhuman primates [Ｊ]. Mol Ther, 2009; 17(2): 327-333.
11. Morishita R. Recent progress in gene therapy for cardiovascular disease [Ｊ]. Circ J, 2002; 66(12): 1077-1086.
12. Li W, Tanaka K, Morioka K, et al. Long-term effect of gene therapy for chronic ischemic myocardium using platelet-derived endothelial cell growth factor in dogs [Ｊ]. J Gene Med, 2008; 10(4): 412-420.
13. 罗涛，张强，张建，等. 转染单纯疱疹病毒胸苷激酶基因抑制移植静脉内膜增生 [Ｊ]. 中国普外基础与临床杂志， 2008; 15（10）: 743-747.
14. Wu S, Wang X, Guo L, et al. Adenovirus mediated endothelial nitric oxide synthase gene transfer prevents restenosis of vein grafts [Ｊ]. ASAIO J, 2004; 50(3): 272-277.
15. 梁宏立, 朱洪生, 黄忠耀，等. 兔自体移植静脉基因转移模型的建立 [Ｊ]. 上海实验动物科学， 2000; 20（1）: 3-6.
16. 曲乐丰, 景在平, 曹贵松，等. 手术缝线携载反义基因预防血管吻合口内膜增生的实验研究 [Ｊ]. 中华普通外科杂志， 2002; 17（1）: 36-37.
17. Wen DZ, Dittman WA, Ye RD, et al. Human thrombomodulin complete cDNA sequence and chromosome localization of the gene [Ｊ]. Biochemistry, 1987; 26(14): 4350-4357.
18. Hirsch T, Spielmann M, Velander P, et al. Insulin-like gr-owth factor-1 gene therapy and cell transplantation in diabetic wounds [Ｊ]. J Gene Med, 2008; 10(11): 1247-1252.
19. Kishida A, Nakashima M, Sakamoto N, et al. Study on complex formation between recombinant human thrombomodulin fragment and thrombin using surface plasmon resonance [Ｊ]. Am J Hematol, 2000; 63(3): 136-140.
20. Yeh HY, Lin JC. Bioactivity and platelet adhesion study of a human thrombomodulin-immobilized nitinol surface [Ｊ]. J Biomater Sci Polym Ed, 2009; 20(5-6): 807-819.

CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY

Transfered Human Thrombomodulin Gene into Artery Wall of Rabbit by High-Pressure Injection in Vivo

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