Wang X, Skelley L, Cade R, Sun Z
Department of Medicine, College of Medicine, University of Florida, Gainesville, 32610-0274, USA.
Gene Ther. 2006 Jul;13(14):1097-103. doi: 10.1038/sj.gt.3302768. Epub 2006 Mar 23.
The aim of this study was to determine the effect of RNA interference inhibition of mineralocorticoid receptor (MR) on cold-induced hypertension (CIH) and renal damage. Recombinant adeno-associated virus (AAV) carrying short hairpin small interference (si)RNA for MR (AAV.MR-shRNA) was constructed and tested for the ability to inhibit renal MR and to control CIH. Three groups of rats with CIH received AAV.MR-shRNA (1.25 x 10(9) particles/rat, intravenous), AAV carrying scrambled shRNA (AAV.Control-shRNA) (1.25 x 10(9) particles/rat, intravenous) and phosphate buffer solution (PBS), respectively. All rats were kept in a cold chamber (6.7 degrees C) throughout the experiment. Adeno-associated virus delivery of MR-shRNA prevented progression of CIH. Blood pressure (BP) of the AAV.MR-shRNA-treated group did not increase and remained at 145+/-3 mm Hg, whereas BP of the AAV.Control-shRNA-treated and PBS-treated group increased to 167+/-4 and 161+/-3 mm Hg, respectively, at 3 weeks after gene delivery. Thus, the antihypertensive effect of a single injection of AAV.MR-shRNA lasted for at least 3 weeks (length of the study). Adeno-associated virus carrying short hairpin siRNA for MR significantly increased urinary sodium excretion and decreased proteinuria. It also decreased serum creatinine and blood urea nitrogen, suggesting enhanced renal function. Both Western blot and immunohistochemical analysis showed that MR expression was decreased significantly in the kidney in the AAV.MR-shRNA-treated rats, confirming that renal MR is effectively inhibited by AAV.MR-shRNA. Adeno-associated virus carrying short hairpin siRNA for MR also significantly attenuated renal hypertrophy. In addition, AAV delivery of MR-shRNA prevented atrophy and dilation of renal tubules and abolished tubular deposition of proteinaceous material seen in CIH rats.
(1) AAV delivery of MR-shRNA effectively silenced MR in vivo. (2) RNA interference inhibition of MR may open a new avenue for the long-term control of hypertension and renal damage.
本研究的目的是确定RNA干扰抑制盐皮质激素受体（MR）对冷诱导高血压（CIH）和肾损伤的影响。构建携带针对MR的短发夹小干扰（si）RNA的重组腺相关病毒（AAV）（AAV.MR-shRNA），并测试其抑制肾MR和控制CIH的能力。三组CIH大鼠分别接受AAV.MR-shRNA（1.25×10⁹颗粒/大鼠，静脉注射）、携带乱序shRNA的AAV（AAV.Control-shRNA）（1.25×10⁹颗粒/大鼠，静脉注射）和磷酸盐缓冲溶液（PBS）。在整个实验过程中，所有大鼠均饲养在冷室（6.7℃）中。腺相关病毒递送MR-shRNA可防止CIH进展。基因递送3周后，AAV.MR-shRNA治疗组的血压（BP）未升高，维持在145±3 mmHg，而AAV.Control-shRNA治疗组和PBS治疗组的BP分别升至167±4和161±3 mmHg。因此，单次注射AAV.MR-shRNA的降压作用持续至少3周（研究时长）。携带针对MR的短发夹siRNA的腺相关病毒显著增加尿钠排泄并减少蛋白尿。它还降低了血清肌酐和血尿素氮，提示肾功能增强。蛋白质印迹和免疫组织化学分析均显示，AAV.MR-shRNA治疗的大鼠肾脏中MR表达显著降低，证实AAV.MR-shRNA可有效抑制肾MR。携带针对MR的短发夹siRNA的腺相关病毒也显著减轻了肾肥大。此外，腺相关病毒递送MR-shRNA可防止CIH大鼠肾小管萎缩和扩张，并消除肾小管中蛋白质物质的沉积。
（1）腺相关病毒递送MR-shRNA可在体内有效沉默MR。（2）RNA干扰抑制MR可能为高血压和肾损伤的长期控制开辟一条新途径。
