Gyurko R, Tran D, Phillips M I
Department of Physiology, College of Medicine, University of Florida, Gainesville 32610, USA.
Am J Hypertens. 1997 May;10(5 Pt 2):56S-62S.
Antisense oligodeoxynucleotides (AS-ODN) can be designed to provide inhibition of a specific protein. Since angiotensin receptors are involved in blood pressure regulation we constructed AS-ODN to angiotensin II type-1 receptor (AT1) mRNA. When given centrally, the AS-ODN reduces blood pressure in spontaneously hypertensive rats (SHR) 24 h after injection. To study the time course of a single AS-ODN injection on blood pressure and heart rate, groups of SHR were injected intracerebroventricularly (icv) with either single dose of AS-ODN or scrambled (SC) ODN and blood pressure was recorded through implanted catheters daily for up to 9 days. Blood pressure decreased significantly in the AS-ODN treated rats compared to the SC-ODN rats for up to 7 days. The maximum decrease (38 mm Hg) occurred at 3 days. There appeared to be no toxic reaction or side effects and the blood pressure level had recovered by days 8 and 9. Heart rate was not altered by AS-ODN treatment. To test that the ODN was entering the brain tissue, fluorescein-isothiocyanate labelled (FITC) ODN was injected in Sprague-Dawley rats and the fluorescence detected 1 h later by confocal microscopy. Within 1 h there was rapid uptake into cells close to the site of injection and into brain parenchyma around the third and lateral ventricles. To test that the AS-ODN had reduced AT1 receptors, binding studies were carried out on membranes from hypothalamic tissue. There was a modest (approximately 20%) but significant (P < .05) decrease in the AT1 receptor binding after 25 microm or 50 microm AS-ODN. AT2 receptors were not altered by the AS-ODN, indicating its specificity for the AT1 receptor. The small decrease in receptor binding, relative to its large effect on blood pressure, is discussed in terms of the AT1 receptor life cycle. The mechanism for the long action of a single AS-ODN injection is hypothesized as resulting from the persistence of AS-ODN in the nucleus, preventing transport of the mRNA into the cytoplasm.
反义寡脱氧核苷酸（AS - ODN）可被设计用于抑制特定蛋白质。由于血管紧张素受体参与血压调节，我们构建了针对血管紧张素II 1型受体（AT1）mRNA的AS - ODN。当经中枢给药时，AS - ODN在注射后24小时可降低自发性高血压大鼠（SHR）的血压。为研究单次注射AS - ODN对血压和心率的时间进程，将SHR组经脑室内（icv）注射单剂量的AS - ODN或乱序（SC）ODN，通过植入的导管每天记录血压，最长记录9天。与SC - ODN大鼠相比，AS - ODN处理的大鼠血压在长达7天的时间里显著下降。最大降幅（38 mmHg）出现在第3天。似乎没有毒性反应或副作用，并且血压水平在第8天和第9天已恢复。AS - ODN处理未改变心率。为检测ODN是否进入脑组织，将异硫氰酸荧光素标记（FITC）的ODN注射到Sprague - Dawley大鼠体内，1小时后通过共聚焦显微镜检测荧光。在1小时内，注射部位附近的细胞以及第三脑室和侧脑室周围的脑实质中迅速摄取了荧光。为检测AS - ODN是否减少了AT1受体，对下丘脑组织的膜进行了结合研究。在给予25微米或50微米的AS - ODN后，AT1受体结合有适度（约20%）但显著（P <.05）的下降。AS - ODN未改变AT2受体，表明其对AT1受体具有特异性。相对于其对血压的显著影响，受体结合的小幅下降根据AT1受体生命周期进行了讨论。单次注射AS - ODN产生长效作用的机制被推测是由于AS - ODN在细胞核中持续存在，阻止了mRNA转运到细胞质中。
