经心包腔转染腺病毒血管内皮生长因子165基因对猪冠脉血管形成的影响

Effects of adenovirus-mediated vascular endothelial growth factor 165 gene on angiogenesis of pigs via pericardial cavity

目的:探讨以腺病毒为载体的血管内皮生长因子165(Ad VEGF165)基因经心包腔转染的表达规律、合适的剂量及对缺血心肌血管生成的作用。方法:随机将20头中华小型猪分为实验组和对照组,每组10头。采用球囊堵塞前降支第一对角支远端建立心肌梗死模型。构建后即刻,实验组采用经皮剑突下穿刺,中心静脉导管插入心包腔内,以含胶原酶1200u及透明质酸酶3000u的生理盐水2ml预处理心包后,在心包腔内注入含Ad VEGF165基因2.0×109pfu的生理盐水1ml;对照组同样方法预处理心包后,在心包腔内注射生理盐水1ml。注射后3d(n=2)、7d(n=2)及28d(n=6)分别用免疫组化、超声心动图检测缺血心肌血管新生情况及心功能,并以酶联免疫吸附试验(ELISA法)检测血浆及心肌组织中Ad VEGF165的表达。结果:Ad VEGF165基因经心包腔转染缺血心肌组织后,在心肌组织中呈高表达,于7d达到高峰,28d降至基线水平,血浆中无目的基因的表达;28d时,实验组缺血心肌微血管密度(MVD)、心功能均明显高于对照组(MVD(517.00±75.70)mm-2vs(226.50±54.10)mm-2,P=0.009;LVEF(%)(72.11±5.20)vs(55.14±4.37),P=0.005)。结论:用胶原酶及透明质酸酶预处理心包后,经其转染Ad VEGF165可以诱导急性心肌梗死模型局部VEGF蛋白表达,促进缺血心肌组织血管新生并能改善心功能。

Aim ： To explore the effects of adenovirus-mediated vascular endothelial growth factor 165 gene on angiogenesis of pigs via perieardial cavity. Methods ：Twenty healthy pigs were randomly divided into two groups, experimental group（n = 10） and control group（ n = 10）. AMI model was constructed, then the intra-perieardial injections were performed through a small incision of the abdominal wall below the xyphoid appendix using center venous catheter. The pericardium was pretreated by injecting a mixture of eollagenase（ 1 200 U） and hyaluronidase（3 000 U）. Then 2.0×10^9 pfu Ad-VEGF165 was injected into perieardial cavity in experimental group, while physiological saline was injected into control group. Imrnunohistoehemistry assay and eehoeardiography were carried out to evaluate the angiogenesis in the isehemie region and eardial function at the 3rd d （n = 2） , 7th d （n = 2） and 28th d （n = 6） after injection, respectively, and enzyme-linked immunoassay（ELISA） was used to investigate the local expression of Ad-VEGF165 in plasma and myoeardium. Results： The peak expression of Ad-VEGF165 gene in myocardium occurred at the 7th d after the injection and decreased gradually to baseline level at the 28th d. Microvenous density and cardiac function of the experimental group significantly increased and had the advantage over the control group（ at 28th d, MVD （517. 00 ± 75.70） ram-2 vs （226.50 ± 54.10） mm^-2, p =0. 009 ; LVEF （72.11 ± 5.20） % vs （55.14 ± 4.37） % , P = 0. 005）. Conclusion ： Gene transfer of Ad-VEGF165 via pericardial cavity can induce VEGF protein expression and enhance angiogenesis in ischemic region and improve cardial function of patients with AMI after being pre-treated by collagenase and hyaluronidase.