Chinese Journal of Tissue Engineering Research ›› 2012, Vol. 16 ›› Issue (15): 2704-2708.doi: 10.3969/j.issn.1673-8225.2012.15.010

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Transfection and expression of human apM1 gene into the aorta of rabbits mediated by using ultrasound with SonoVue   

Yang Wen-kai1, 2, Chen Qing3, Yang Chun-li4, Wang Xian-guo1, Xie Ting1, Dong Nian-guo1, Chen Xin-zhong1   

  1. 1Department of Cardiosurgery, 3Department of General Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan  430022, Hubei Province, China; 2Department of Cardiothoracic Surgery, 4ICU Department, Jiangxi Provincial People’s Hospital, Nanchang  330006, Jiangxi Province, China
  • Received:2011-12-21 Revised:2012-01-27 Online:2012-04-08 Published:2012-04-08
  • Contact: author: Chen Xin-zhong, Doctor, Attending physician, Department of Cardiosurgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, Hubei Province, China xinzhongchen@yahoo.com.cn
  • About author:Yang Wen-kai☆, Studying for doctorate, Attending physician, Department of Cardiosurgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, Hubei Province, China; Department of Cardiothoracic Surgery, Jiangxi Provincial People’s Hospital, Nanchang 330006, Jiangxi Province, China doctor_wenkaiyang@yahoo.com.cn

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Abstract:

BACKGROUND: Gene delivery using ultrasound microbubbles is an emerging directional transmission gene technology.
OBJECTIVE: To construct a recombinant plasmid containing human apM1 gene and delivery apM1 gene into the aorta of rabbits by using ultrasound with SonoVue. 
METHODS: Twenty-one rabbits were randomized into control group (n=6) and apM1 group (n=15). apM1 mRNA from epicardial adipose tissue was amplified by reverse transcription-PCR and cloned into plasmid pcDNA3.1 to generate the recombinant plasmid pcDNA3.1-apM1, which was confirmed by nucleotide sequencing and transiently transfected into human umbilical vein endothelial cells (HUVECs). While the mixture of SonoVue and pcDNA3.1-apM1 was injected though helix vein, the thoracoabdominal aorta of rabbits were sonicated by ultrasound for 3 minutes. The arterial vessels and blood serum of rabbits were harvested at 2, 7and 14 days respectively after delivery to detect apM1 expression by Western blot analyses and enzyme linked immunosorbent assay (ELISA).
RESULTS AND CONCLUSION: The identification of pcDNA3.1-apM1 recombinant by DNA sequencing ws correct. The expression of apM1 gene could be detected by Western blot in HUVECs after pcDNA3.1-apM1 transient transfection. apM1 gene could be effectively transfered into the aorta wall of rabbit by using ultrasound with SonoVue. The high expression of apMl gene was detected at 2 days after delivery and until to 14 days. ELISA results showed that the concentration of apM1 protein in serum of rabbit could be significantly increased by delivery of apM1 gene using ultrasound with SonoVue (P < 0.01). These findings indicate that the eukaryotic expression recombinant containing apM1 gene can be successfully constructed. apM1 gene can be transferred into the deep area of rabbit aorta vessel wall by ultrasound with SonoVue and expressed and secreted effectively. 

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