AAV delivery of mineralocorticoid receptor shRNA prevents progression of cold-induced hypertension and attenuates renal damage

Abstract

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 Â 10 9 particles/rat, intravenous), AAV carrying scrambled shRNA (AAV.Control-shRNA) (1.25 Â 10 9 particles/rat, intravenous) and phosphate buffer solution (PBS), respectively. All rats were kept in a cold chamber (6.71C) 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 14573 mm Hg, whereas BP of the AAV.Control-shRNA-treated and PBStreated group increased to 16774 and 16173 mm Hg, respectively, at 3 weeks after gene delivery. Thus, the antihypertensive effect of a single injection of AAV.MRshRNA 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. Conclusions: (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.

Adeno-associated virus-mediated knockdown of melanocortin-4 receptor in the paraventricular nucleus of the hypothalamus promotes high-fat diet-induced hyperphagia and obesity

“…In this study, we demonstrate that selective MC4R knockdown in PVN neurons of the adult wild-type rat results in no changes in food intake and body weight when animals are fed with a regular diet within 2 weeks, but causes increased feeding and excessive weight gain in response to a high-fat diet. It is unlikely that the unaltered feeding and body weight in response to the regular diet is due to insufficient gene suppression because AAV-mediated gene expression occurs within 3 days (Flotte et al 1993, Takahashi et al 2002 and AAV-mediated effective gene silencing has been reported at days 6-10 after injection of AAV-shRNA (Hommel et al 2003, Wang et al 2006, Paskowitz et al 2007). Thus, our results suggest that the disruption of MC4R in PVN neurons increases the sensitivity to diet-induced hyperphagia and obesity.…”

AAV vectors for RNA-based modulation of gene expression

“…This led to a delayed onset of hypertension and an attenuated disease. 51 Cold-induced hypertension was attenuated when a U6-driven shRNA against the mineralocorticoid receptor was systemically delivered by an AAV2 vector in rats 52 and mice. 53 A 50% reduction in expression of the receptor was documented in the kidney where hypertrophy was attenuated, but not in the heart or aorta.…”

“…The precise cellular targets of the active vector remain to be characterized. 52 In a rat model of salt hypertension induced by salt loading, an AAV expressing an antisense sequence against the central a2B-adrenergic receptor was administered by intracerebroventricular injection. It resulted in a twofold decrease in protein levels in the cerebellum and hippocampus, and a long-lasting fall in blood pressure.…”

Remedial applications of silencing ribonucleic acids and modalities for its delivery to the kidneys - A review

“…This approach resulted in specific targeted proteins knockdown. The antibodies were also detected in colon tissues, spleen, liver, lung and muscles (Wang et al, 2006). Complex of silencing RNA to streptavidin conjugated antibodies was developed as alternative to protamine-silencing RNA complex, PEGylated liposomes as carrier for silencing RNA and antibodies conjugation with cationic lipid-silencing RNA complex (Shou et al, 2009).…”