人类心房利钠肽基因在自发性高血压大鼠体内的表达及其影响

Expression and effects of human atrial natriuretic peptide gene in spontaneously hypertensive rats

目的研究人类心房利钠肽(hANP)基因在自发性高血压大鼠(SHRs)体内表达、持续时间和对血压、心、血管及肾脏的影响。方法12只8周龄雄性SHRs,随机分为两组,于左腿股四头肌注射pcDNA3.1-hANP质粒者为实验组,在同一部位注射pcDNA3.1空质粒者为对照组,每周测量大鼠尾动脉收缩压,并利用放射免疫方法监测血中hANP水平;10周后处死动物,RT-PCR和Western印迹杂交技术检测hANP基因表达情况;分别称量体重、全心脏和左心室重量,大体评价对心肌重构的影响;苏木素-伊红(HE)染色和胶原组织学(VG)染色观察对心、血管及肾脏组织形态学的影响。结果转基因后第1周起与对照组比较,实验组血压开始下降,两组动物一直相差(12±3.1)mmHg(P<0.05),其作用可持续10周;实验组血中hANP水平较高;实验组hANP基因在肌肉组织中高效表达,对照组则未见表达;实验组心脏重量/体重比和左心室重量/心脏重量比明显低于对照组;组织形态学分析显示实验组心肌、血管及肾脏组织细胞形态基本正常,胶原沉积明显减少。结论肌肉注射法将hANP基因导入SHRs后,hANP基因在肌肉组织中高效表达,且hANP可释放入血,降低SHRs的血压并对心脏、血管及肾脏等高血压靶器官的损害起到了预防作用。显示了hANP基因对高血压患者治疗的可能性。

Objectives To investigate the expression of human atrial natriuretic peptide （hANP） gene and its effects on blood pressure and functions of heart, blood vessel and kidney in spontaneously hypertensive rats （SHRs）. Methods Twelve eight-week-old male SHRs were divided into two groups randomly. DNA plasmid pcDNA3.1-CMV-hANP was injected to the left quadriceps femoris muscle of the SHRs in the test group （n = 6）. Controls （n = 6） were injected with DNA plasmid pcDNA3.1-CMV into the same site. After the gene delivery, the caudal artery systolic pressure was measured in SHRs and the plasma level of hANP was detected by means of radioimmunoassay every week. After 10 weeks, all the animals were sacrificed. The expression of hANP gene was measured by RT-PCR and Western blotting. Body weight （BW）, heart weight （HW） and left ventricle weight （LVW） were measured to evaluate the reconstitution of myocardium. Hematoxylin-Eosin （HE） and Van Gieson （VG） staining were used to determine the effect of hANP on blood vessel and kidney. Results Compared with the control group, blood pressure of the test group began to decrease during the first week after the injection of plasmid pcDNA3.1-CMV-hANP. The significant blood pressure difference （up to 12.0 ＋ 2.55 mmHg, P 〈 0.05） lasted for 10 weeks. The results of RT-PCR and Western blotting showed that hANP gene was highly expressed in the muscles, while the no expression was observed in the control group. The ratio of HW/BW and LVW/HW decreased markedly in the test group. Morphological analysis showed that cardiovascular hypertrophy and renal injury were attenuated in the test group. Conclusions hANP gene could be expressed efficiently in the muscles of SHRs through muscle injections, hANP protein released into blood circulation could not only stably reduce blood pressure, but also prevent the hypertension induced damage to target organs, such as kidney and cardiovascular system. This study suggested the possibility of hANP gene therapy for human hypertension.