Wang H, Katovich M J, Gelband C H, Reaves P Y, Phillips M I, Raizada M K
Department of Physiology, Colleges of Medicine and Pharmacy, and University of Florida Brain Institute, Gainesville, FL 32610, USA.
Circ Res. 1999 Oct 1;85(7):614-22. doi: 10.1161/01.res.85.7.614.
Angiotensin I-converting enzyme (ACE) inhibitors have been proven to be highly effective and are for the most part the drugs of choice in the treatment and control of hypertension, congestive heart failure, and left ventricular dysfunction. Despite this, questions regarding side effects and compliance with this traditional pharmacological strategy remain. In view of these observations, coupled with recent advances in gene-transfer technology, our objective in this study was to determine whether the expression of ACE could be controlled on a permanent basis at a genetic level. We argued that the introduction of ACE antisense to inhibit the enzyme would be a prerequisite in considering the antisense gene therapy for the control of hypertension and other related pathological states. Retroviral vectors (LNSV) containing ACE sense (LNSV-ACE-S) and ACE antisense (LNSV-ACE-AS) sequences were constructed and were used in rat pulmonary artery endothelial cells (RPAECs) to determine the feasibility of this approach. Infection of rat RPAECs with LNSV-ACE-S and LNSV-ACE-AS resulted in a robust expression of transcripts corresponding to ACE-S and ACE-AS, respectively, for the duration of these experiments, ie, 8 consecutive passages. The expression of ACE-AS but not of ACE-S was associated with a permanent decrease of approximately 70% to 75% in ACE expression and a 50% increase in the B(max) for the AT(1)s. Although angiotensin II caused a concentration-dependent stimulation of intracellular Ca(2+) levels in both ACE-S- and ACE-AS-expressing cells, the stimulation was significantly higher in ACE-AS-expressing RPAECs. In vivo experiments demonstrated a prolonged expression of ACE-AS transcripts in cardiovascularly relevant tissues of rats. This was associated with a long-term reduction in blood pressure by approximately 15 mm Hg, exclusively in the spontaneously hypertensive rat. These observations demonstrate that delivery of ACE-AS by retroviral vector results in a permanent inhibition of ACE and a long-term reduction in high blood pressure in the spontaneously hypertensive rat.
血管紧张素I转换酶（ACE）抑制剂已被证明非常有效，并且在很大程度上是治疗和控制高血压、充血性心力衰竭及左心室功能障碍的首选药物。尽管如此，关于副作用以及对这种传统药理学策略的依从性问题仍然存在。鉴于这些观察结果，再加上基因转移技术的最新进展，我们在本研究中的目标是确定是否可以在基因水平上永久性地控制ACE的表达。我们认为，引入ACE反义序列以抑制该酶将是考虑采用反义基因疗法来控制高血压及其他相关病理状态的一个先决条件。构建了含有ACE正义序列（LNSV-ACE-S）和ACE反义序列（LNSV-ACE-AS）的逆转录病毒载体（LNSV），并将其用于大鼠肺动脉内皮细胞（RPAECs），以确定这种方法的可行性。用LNSV-ACE-S和LNSV-ACE-AS感染大鼠RPAECs，在这些实验期间，即连续8代，分别导致了与ACE-S和ACE-AS相对应的转录物的强劲表达。ACE-AS而非ACE-S的表达与ACE表达永久性降低约70%至75%以及AT(1)s的B(max)增加50%相关。尽管血管紧张素II在表达ACE-S和ACE-AS的细胞中均引起细胞内Ca(2+)水平的浓度依赖性刺激，但在表达ACE-AS的RPAECs中这种刺激明显更高。体内实验证明ACE-AS转录物在大鼠心血管相关组织中持续表达。这仅在自发性高血压大鼠中与血压长期降低约15 mmHg相关。这些观察结果表明，通过逆转录病毒载体递送ACE-AS可导致ACE的永久性抑制以及自发性高血压大鼠的高血压长期降低。
